Staphylococcus

Life Sci, 1998, 62(22), 2005 - 14
The effect of LHRH and TRH on human interferon-gamma production in vivo and in vitro; Grasso G et al.; Accumulating evidence suggests that hypothalamic luteinizing hormone-releasing hormone (LHRH) and thyrotropin-releasing hormone (TRH) are two hypophysiotropic factors which modulate the immune response . The aim of the present study was to determine the in vivo effects of an intravenous bolus of LHRH and TRH on plasma interferon (IFN)-gamma production in five normoprolactinemic women with irregular menstrual cycles . We also determined prolactin (PRL), thyrotropin (TSH), follicle stimulating hormone (FSH), and luteinizing hormone (LH) levels before and after intravenous administration of LHRH and TRH . The results demonstrate that intravenous bolus of LHRH/TRH increases plasma IFN-gamma levels, with the maximum response 45 min after in vivo administration of hypothalamic peptides and after peak levels of adenohypophyseal hormones (PRL: 15 min; TSH: 30 min; FSH: 30 min; LH: 30 min) . In order to investigate a possible direct action of hypothalamic hormones on immune cells, we also evaluated, in the same subjects, the influence of LHRH and TRH on IFN-gamma production by human peripheral blood mononuclear cells (PBMCs), collected before the intravenous administration of the peptides and stimulated in vitro with bacterial superantigen staphylococcal enterotoxin A (SEA) and concanavalin A (Con A) . LHRH and TRH, separately and together, significantly enhanced in vitro IFN-gamma production by SEA- and ConA-activated PBMCs . The present results suggest that hypothalamic peptides (LHRH and TRH) directly, and/or indirectly pituitary hormones (PRL, TSH, FSH, and LH) or IL-2, have stimulatory effect on IFN-gamma producing cells and are further evidence of interactions between the neuroendocrine and immune systems.

FEMS Immunol Med Microbiol, 1998 Apr, 20(4), 301 - 10
Differentiation and distribution of three types of exfoliative toxin produced by Staphylococcus hyicus from pigs with exudative epidermitis; Andresen LO; Exfoliative toxins of approximately 30 kDa produced by Staphylococcus hyicus strains NCTC 10350, 1289D-88 and 842A-88 were purified and specific polyclonal antisera were raised against each of the toxins . It was shown by immunoblot analysis and ELISA that three exfoliative toxins from S . hyicus were antigenically distinct . The three toxins were designated ExhA, ExhB and ExhC . From 60 diseased pigs, each representing an outbreak of exudative epidermitis, a total of 584 isolates of S . hyicus were phage typed and tested for production of exfoliative toxin . ExhA-, ExhB- and ExhC-producing S . hyicus isolates were found in 12 (20%), 20 (33%) and 11 (18%), respectively, of the 60 pig herds investigated . Production of the different types of exfoliative toxin was predominantly associated with certain phage groups . However, toxin production was found in all of the six phage groups defined by the phage typing system . Some changes in the distribution of isolates between phage groups were observed when the results of this study were compared to previous investigations . In this study two new antigenically distinct exfoliative toxins were isolated and tools for in vitro detection of toxin producing S . hyicus isolates and for further studies on the exfoliative toxins from S . hyicus have been provided.

J Clin Invest, 1998 Jun 1, 101(11), 2406 - 14
In vivo tumor transfection with superantigen plus cytokine genes induces tumor regression and prolongs survival in dogs with malignant melanoma; Dow SW et al.; In vivo transfection of established tumors with immunostimulatory genes can elicit antitumor immunity . Therefore, we evaluated the safety and efficacy of intratumoral injections of a bacterial superantigen with a cytokine gene in dogs with malignant melanoma, a spontaneous and highly malignant canine tumor . 26 dogs with melanoma were treated with lipid-complexed plasmid DNA encoding staphylococcal enterotoxin B and either GM-CSF or IL-2 . Dogs were evaluated for treatment-associated toxicity, tumor responses, immunologic responses, and survival times . The overall response rate (complete or partial remissions) for all 26 dogs was 46% (12 of 26), and was highest in patients with smaller tumors . Toxicity was minimal or absent in all dogs . Injected tumors developed marked infiltrates of CD4+ and CD8+ T cells and macrophages, and tumor regression was associated with development of high levels of antitumor cytotoxic T lymphocyte activity in peripheral blood lymphocytes . Survival times for animals with stage III melanomas treated by intratumoral gene therapy were prolonged significantly compared with animals treated with surgical tumor excision only . Thus, local tumor transfection with superantigen and cytokine genes was capable of inducing both local and systemic antitumor immunity in an outbred animal with a spontaneously developing malignant tumor.

Antimicrob Agents Chemother, 1998 Jun, 42(6), 1417 - 23
Pharmacokinetics of sparfloxacin in the serum and vitreous humor of rabbits: physicochemical properties that regulate penetration of quinolone antimicrobials; Liu W et al.; We have used a recently described animal model to characterize the ocular pharmacokinetics of sparfloxacin in vitreous humor of uninfected albino rabbits following systemic administration and direct intraocular injection . The relationships of lipophilicity, protein binding, and molecular weight to the penetration and elimination of sparfloxacin were compared to those of ciprofloxacin, fleroxacin, and ofloxacin . To determine whether elimination was active, elimination rates following direct injection with and without probenecid or heat-killed bacteria were compared . Sparfloxacin concentrations were measured in the serum and vitreous humor by a biological assay . Protein binding and lipophilicity were determined, respectively, by ultrafiltration and oil-water partitioning . Pharmacokinetic parameters were characterized with RSTRIP, an iterative, nonlinear, weighted, least-squares-regression program . The relationship between each independent variable and mean quinolone concentration or elimination rate in the vitreous humor was determined by multiple linear regression . The mean concentration of sparfloxacin in the vitreous humor was 59.4% +/- 12.2% of that in serum . Penetration of sparfloxacin, ciprofloxacin, fleroxacin, and ofloxacin into, and elimination from, the vitreous humor correlated with lipophilicity (r2 > 0.999) . The linear-regression equation describing this relationship was not improved by including the inverse of the square root of the molecular weight and/or the degree of protein binding . Elimination rates for each quinolone were decreased by the intraocular administration of probenecid . Heat-killed Staphylococcus epidermidis decreased the rate of elimination of fleroxacin . Penetration of sparfloxacin into the noninflamed vitreous humor was greater than that of any quinolone previously examined . There was an excellent correlation between lipophilicity and vitreous entry or elimination for sparfloxacin as well as ciprofloxacin, fleroxacin, and ofloxacin . There are two modes of quinolone translocation into and out of the vitreous humor: diffusion into the eye and both diffusion and carrier-mediated elimination out of the vitreous humor.

Lik Sprava, 1998 Jan-Feb, (1), 109 - 12
{Antibody production in the blood of donors immunized with staphylococcal anatoxin}; Dyzyk HM et al.; Blood serum content was studied of specific antistaphylococcal antibodies (staphylolysins) in 576 donors immunized with staphylococcal anatoxin with the purpose of obtaining an antistaphylococcal plasma and antistaphylococcal immunoglobulin to be used in clinical settings . 292 donors had been immunized and examined prior to 1986, 284--after 1986 (before 1994) . Comparison of the immune responses in the above periods of time permitted finding out that 13.03% of immunized donors responded to the antigenic stimulus by such paradoxical reaction as disappearance of specific antibodies; the number of persons-active respondents has gotten reduced from 17.12% to 5.98% as has the number of individuals having the baseline level of staphylolysins (1-2 ME/ml) . The changes were at their greatest in donors with group A (II) blood.

J Biol Chem, 1998 Jun 5, 273(23), 14085 - 9
Inactivation of human manganese-superoxide dismutase by peroxynitrite is caused by exclusive nitration of tyrosine 34 to 3-nitrotyrosine; Yamakura F et al.; Peroxynitrite has recently been implicated in the inactivation of many enzymes . However, little has been reported on the structural basis of the inactivation reaction . This study proposes that nitration of a specific tyrosine residue is responsible for inactivation of recombinant human mitochondrial manganese-superoxide dismutase (Mn-SOD) by peroxynitrite . Mass spectroscopic analysis of the peroxynitrite-inactivated Mn-SOD showed an increased molecular mass because of a single nitro group substituted onto a tyrosine residue . Single peptides that had different elution positions between samples from the native and peroxynitrite-inactivated Mn-SOD on reverse-phase high performance liquid chromatography were isolated after successive digestion of the samples by staphylococcal serine protease and lysylendopeptidase and subjected to amino acid sequence and molecular mass analyses . We found that tyrosine 34 of the enzyme was exclusively nitrated to 3-nitrotyrosine by peroxynitrite . This residue is located near manganese and in a substrate O-2 gateway in Mn-SOD.

J Mol Biol, 1998 May 15, 278(4), 879 - 94
Anion-induced folding of Staphylococcal nuclease: characterization of multiple equilibrium partially folded intermediates; Uversky VN et al.; The refolding of acid-unfolded staphylococcal nuclease (SNase) induced by anions was characterized, and revealed the existence of three different partially folded intermediates (A states) . The three intermediates lack the rigid tertiary structure characteristic of native states, and differ in their degree of folding as measured by probes of secondary structure, size, stability and globularity . The least structured conformation, A1, is stabilized by chloride (600 mM) or sulfate (100 mM) . It is about 50% folded (based on circular dichroism and small angle X-ray scattering (SAXS) data) . The next most structured intermediate, A2, is induced by trifluoroacetate (300 mM) and has approximately 70% native-like secondary structure . The most structured intermediate, A3, is stabilized by trichloroacetate (50 mM) and has native-like secondary structure content and is almost as compact as the native state . The stability toward urea denaturation increases with increasing structure of the intermediates . Moreover, ureainduced unfolding studies show that these partially folded species are separated from each other, and from the unfolded state, by significant free energy barriers, suggesting that they are distinct conformational states . Kratky plots, based on the SAXS data, indicate that the two more structured intermediates have significant globularity (i.e . a tightly packed core), whereas the less structured intermediate has very little globularity . These observations support a model of protein folding in which certain conformations are of particularly low free energy and hence populated under conditions which differentially destabilize the native state . These partially folded intermediates probably consist of ensembles of substates with a common core of native-like secondary structure, which is responsible for their stability . Consequently, it is likely that the intermediates observed here represent the equilibrium counterparts of transient kinetic intermediates.

Nutrition, 1998 May, 14(5), 427 - 32
Elimination of intraluminal colonization by antibiotic lock in silicone vascular catheters; Andris DA et al.; An in vitro model was designed to evaluate the efficacy of instilled antimicrobials to reduce or eliminate intraluminal microbial colonization . Minimal inhibitory concentration and minimal bactericidal concentration activity of appropriate test anti-infectives were determined using standard methodology against clinically derived and reference test strains commonly associated with catheter-related infection . Drug activity was validated by bioassay for the test anti-infectives . Reference and clinical test strains were inoculated to the intraluminal surface of silicone catheter segments and incubated for 30 min, after which the inoculum was replaced with total parenteral nutrition (TPN) solution and reincubated for 12 h . For 7 d, instillation of antibiotic and TPN solution was alternated every 12 h to simulate clinical conditions . On days 1, 4, and 7, catheter segments were rinsed, bisected, and sonicated for quantitative plate count to determine mean microbial counts per centimeter of catheter surface . Catheter segments were also prepared for scanning electron microscopy . A significant decrease in staphylococcal intraluminal colonization after instillation of nafcillin, ceftriaxone, gentamicin, and vancomycin was demonstrated (P < 0.001) . Aztreonam, ceftriaxone, and gentamicin completely eliminated gram-negative catheter colonization (P < 0.001) . Yeast was eradicated from the internal catheter surface after treatment with amphoteracin B, and fluconazole significantly decreased intraluminal colonization (P < 0.001) . Results show a significant decrease in staphylococcal, gram-negative, and fungal intraluminal colonization after instillation of appropriate antimicrobial . In vitro results support early clinical success using this technique . Future studies are warranted to identify optimal drug concentrations and dosing intervals.

Biochemistry, 1998 May 12, 37(19), 6939 - 48
Stability effects of increasing the hydrophobicity of solvent-exposed side chains in staphylococcal nuclease; Schwehm JM et al.; A total of fifty single site surface phenylalanine substitution mutants have been made in the model protein staphylococcal nuclease . The fifty residues that were replaced with phenylalanine were chosen to give a broad sampling of solvent accessibility, secondary structure, and backbone conformations . The change in the stability of each mutant protein relative to wild type was measured by guanidine hydrochloride denaturation . These results were compared to previous results obtained when these same sites were substituted with an alanine and a glycine . By this means, changes in the stability due to the loss of interactions of the wild-type side chain can be separated from the effects of introducing the bulky, hydrophobic phenylalanine in these solvent-exposed positions . In general, our results agree with the conventional wisdom that placing a hydrophobic residue in a solvent-exposed position is destabilizing in most cases, but less destabilizing than most changes in the hydrophobic core of the protein . However, the degree to which a hydrophobic surface substitution destabilizes or stabilizes a globular protein is highly context-dependent, with some mutations being as destabilizing as those in the core . This indicates that steric and packing considerations are also important on the surface of a globular protein but generally are not as important as in the interior . No evidence for the widespread occurrence of the so-called reverse hydrophobic effect at solvent-exposed sites was found . In addition, this survey of numerous sites suggests that previous measurements of alpha-helix "propensities" often seriously underestimate the importance of the environment of the side chain.

J Exp Med, 1998 May 4, 187(9), 1427 - 38
A role for Fas in negative selection of thymocytes in vivo; Kishimoto H et al.; To seek information on the role of Fas in negative selection, we examined subsets of thymocytes from normal neonatal mice versus Fas-deficient lpr/lpr mice injected with graded doses of antigen . In normal mice, injection of 1-100 microg of staphylococcal enterotoxin B (SEB) induced clonal elimination of SEB-reactive Vbeta8+ cells at the level of the semi-mature population of HSAhi CD4+ 8- cells found in the thymic medulla; deletion of CD4+ 8+ cells was minimal . SEB injection also caused marked elimination of Vbeta8+ HSAhi CD4+ 8- thymocytes in lpr/lpr mice . Paradoxically, however, elimination of these cells in lpr/lpr mice was induced by low-to-moderate doses of SEB (</=1 microg) but not by high doses (100 microg) . Similar findings applied when T cell receptor transgenic mice were injected with specific peptide . These findings suggest that clonal elimination of semi-mature medullary T cells is Fas independent at low doses of antigen but Fas dependent at high doses . Previous reports documenting that negative selection is not obviously impaired in lpr/lpr mice could thus reflect that the antigens studied were expressed at only a low level.

Z Gastroenterol, 1998 Apr, 36(4), 287 - 93
Short-term efficacy and long-term outcome of cyclosporine treatment in patients with severe ulcerative colitis; Wenzl HH et al.; Cyclosporine A (CyA) has been recommended for the treatment of severe steroid-resistant ulcerative colitis, however, long-term results are scarce . We prospectively followed a treatment plan in 14 patients with severe ulcerative colitis receiving intravenous CyA after failure to respond to at least eight days of standard therapy with prednisolone (1-1.5 mg/kg/day) . CyA was delivered in a daily dose of 5 mg/kg i.v . for a mean of 14 days (range 7-28) in addition to ongoing medical therapy . CyA whole blood levels were monitored by HPLC and maintained between 100 ng/ml and 400 ng/ml . Responders were switched to oral CyA (5-7.5 mg/kg/day) for a mean of two months, and steroids were gradually tapered . Eleven patients (79%) initially responded to i.v . CyA, three patients failed to respond and underwent urgent colectomy . Time until response averaged seven days (range 3-13) . Four of the eleven responders underwent colectomy because of severe relapse after one, eleven, twelve and 13 months of follow-up . The remaining seven patients were followed for a median of 48 months . During the first year of follow-up three out of seven had a severe relapse and responded to steroids (two patients) or to a further course of i.v . CyA (one patient) . During CyA therapy one patient developed staphylococcal sepsis, other adverse events were mild and reversible . The results confirm that CyA is effective in severe steroid-refractory ulcerative colitis . Severe relapse and colectomy are uncommon after the first year of follow-up and the colon preserving effect of CyA can be maintained in up to 50% of patients over a period of four years.

Am J Physiol, 1998 Jun, 274(6 Pt 1), C1583 - 91
Endothelin-1 activates phospholipases and channels at similar concentrations in porcine coronary arteries; Jones AW et al.; Sensitivity of endothelin-1 (ET-1)-ion channel interactions has been proposed to exceed that of ET-1-phospholipase activation in vascular smooth muscle . We wanted to determine whether short-circuiting ion channels with staphylococcal alpha-toxin pores would shift the ET-1-force relation to the right as predicted from the above proposal . Medium size porcine coronary arteries (outer diameter 0.7-1.5 mm) were mounted on isometric force transducers . ET-1 concentration response curves were compared between intact rings and those subjected to alpha-toxin treatment with Ca buffered at 0.1 microM . The EC50 for treated rings (1.5 +/- 1.0 nM, n = 5 pigs) was similar to that for intact rings (1.9 +/- 0.4 nM) . The Ca sensitivity of the alpha-toxin-treated rings (EC50 = 0.43 +/- 0.08 microM) was similar to that reported by other laboratories for intact and alpha-toxin-treated arteries and was shifted eightfold to the left by a high concentration of ET-1 (10 nM) . Measurements of {32P}phosphatidic acid ({32P}PA) levels were used to evaluate phospholipase activity in intact arteries . The time courses for {32P}PA production and contraction were similar in response to high (100 nM) and to low (1 nM) ET-1 . Significant increases in both steady-state contraction and {32P}PA occurred over a wide range of ET-1 concentrations tested (0.3-100 nM) . Our findings support the concept that ET-1-phospholipase coupling is operative over the whole concentration range that induces contractile responses . It is suggested that both Ca entry and Ca sensitization processes are activated by ET-1 at low concentrations (<EC50) and that both processes contribute significantly to the integrated response.

J Neuroimmunol, 1998 Mar 15, 83(1-2), 102 - 15
Marijuana, immunity and infection; Klein TW et al.; The influence of marijuana cannabinoids on immune function has been examined extensively over the last 25 yr . Various experimental models have been used employing drug-abusing human subjects, experimental animals exposed to marijuana smoke or injected with cannabinoids, and in vitro models employing immune cell cultures treated with various cannabinoids . For the most part, these studies suggest that cannabinoids modulate the function of T and B lymphocytes as well as NK cells and macrophages . In addition to studies examining cannabinoid effects on immune cell function, other reports have documented that these substances modulate host resistance to various infectious agents . Viruses such as herpes simplex virus and murine retrovirus have been studied as well as bacterial agents such as members of the genera Staphylococcus, Listeria, Treponema, and Legionella . These studies suggest that cannabinoids modulate host resistance, especially the secondary immune response . Finally, a third major area of host immunity and cannabinoids is that involving drug effects on the cytokine network . Employing in vivo and in vitro models, it has been determined that cannabinoids modulate the production and function of acute phase and immune cytokines as well as modulate the activity of network cells such as macrophages and T helper cells, Th1 and Th2 . These results are intriguing and demonstrate that under certain conditions, cannabinoids can be immunomodulatory and enhance the disease process . However, more studies are needed to determine both the health risk of marijuana abuse and the role of the cannabinoid receptor/ligand system in immune regulation and homeostasis.

Z Kardiol, 1998 Apr, 87(4), 276 - 82
{Surgery of acute aortic valve endocarditis: prognosis in paravalvular abscess}; Bauernschmitt R et al.; The occurrence of paravalvular abscesses in the course of an acute endocarditis of the aortic valve indicates an advanced stadium of the disease . The infection has spread beyond the limits of the valve leaflets, and ongoing destruction of the paravalvular tissue is to be expected, if the endocarditis is continually treated by antibiotics alone . Surgery of acute endocarditis with paravalvular abscess, however, supposedly carries an increased risk of early mortality and late morbidity . The following prospective study was carried out to determine whether a radical surgical approach together with aggressive postoperative antibiotic therapy could help to improve results . Between 1988 and 1995, 138 patients were operated during the acute phase of infective endocarditis; in 102 the aortic valve was involved . Among these, 44 had paravalvular abscesses at the time of surgery . The mean age of both groups was the same, but there was a higher rate of concomitant coronary artery disease, multiple valve involvement, advanced NYHA-class, and staphylococcal disease among the patients with abscesses . All interventions were carried out with cardiopulmonary bypass and cardioplegic arrest . The aortic valve was resected, abscesses were removed, and each part of potentially infected or necrotic tissue was resected as complete as possible, irrespective of the possibility to jeopardize the conduction system or to create large tissue defects . The aortic valve was replaced with a mechanical prosthesis in each case . The postoperative antibiotic regimen was specifically directed against the microorganisms isolated preoperatively; therapy was only modified, if signs of systemic infection did not disappear three days after surgery . The operative mortality was 10% among patients without an abscess and 11% in patients with a paravalvular abscess . Early recurrent endocarditis was recorded in two patients without and in only one patient with an abscess . Late recurrent endocarditis was noted in three patients; none of them had abscesses at the time of surgery . We conclude that the operative risk of acute endocarditis of the aortic valve with a paravalvular abscess does not have to be inevitably higher compared to cases without paravalvular involvement . To achieve these results, it is necessary to use a radical surgical approach and to adjust postoperative antibiotic therapy, if infectious signs do not disappear shortly after surgery.

Biochemistry (Mosc), 1998 Feb, 63(2), 212 - 8
Site-specific endonuclease SscL1 I from strain Staphylococcus species L1; Vasiljeva LY et al.; A site-specific endonuclease SscL1 I preparation has been isolated and purified to near homogeneity from the strain Staphylococcus sp . L1 without admixtures of other nuclease activity . DNA cleavage proceeds according to the scheme: 5'-G down arrow ANTC-3' 3'-CTNA up arrow G-5', and thus the isolated enzyme is an isoschizomer of restriction endonuclease HinfI and belongs to the second class of restriction endonucleases . SscL1 I works over a broad range of temperature and pH . The enzyme is characterized by high stability during storage.

JAMA, 1998 May 20, 279(19), 1537 - 41
Role of rifampin for treatment of orthopedic implant-related staphylococcal infections: a randomized controlled trial . Foreign-Body Infection (FBI) Study Group; Zimmerli W et al.; CONTEXT: Rifampin-containing regimens are able to cure staphylococcal implant-related infections based on in vitro and in vivo observations . However, this evidence has not been proven by a controlled clinical trial . OBJECTIVE: To evaluate the clinical efficacy of a rifampin combination in staphylococcal infections associated with stable orthopedic devices . DESIGN: A randomized, placebo-controlled, double-blind trial conducted from 1992 through 1997 . SETTING: Two infectious disease services in tertiary care centers in collaboration with 5 orthopedic surgeons in Switzerland . PATIENTS: A total of 33 patients with culture-proven staphylococcal infection associated with stable orthopedic implants and with a short duration of symptoms of infection (exclusion limit <1 year; actual experience 0-21 days) . INTERVENTION: Initial debridement and 2-week intravenous course of flucloxacillin or vancomycin with rifampin or placebo, followed by either ciprofloxacin-rifampin or ciprofloxacin-placebo long-term therapy . MAIN OUTCOME MEASURES: Cure was defined as (1) lack of clinical signs and symptoms of infection, (2) C-reactive protein level less than 5 mg/L, and (3) absence of radiological signs of loosening or infection at the final follow-up visit at 24 months . Failure was defined as (1) persisting clinical and/or laboratory signs of infection or (2) persisting or new isolation of the initial microorganism . RESULTS: A total of 18 patients were allocated to ciprofloxacin-rifampin and 15 patients to the ciprofloxacin-placebo combination . Twenty-four patients fully completed the trial with a follow-up of 35 and 33 months . The cure rate was 12 (100%) of 12 in the ciprofloxacin-rifampin group compared with 7 (58%) of 12 in the ciprofloxacin-placebo group (P=.02) . Nine of 33 patients dropped out due to adverse events (n=6), noncompliance (n=1), or protocol violation (n=2) . Seven of the 9 patients who dropped out were subsequently treated with rifampin combinations, and 5 of them were cured without removal of the device . CONCLUSION: Among patients with stable implants, short duration of infection, and initial debridement, patients able to tolerate long-term (3-6 months) therapy with rifampin-ciprofloxacin experienced cure of the infection without removal of the implant.

J Immunol, 1998 Jun 1, 160(11), 5309 - 13
Perforin and IFN-gamma are involved in the antitumor effects of antibody-targeted superantigens; Rosendahl A et al.; The bacterial superantigen staphylococcal enterotoxin A (SEA) is a potent inducer of cytokine production and cytotoxic T cell responses . To target a T cell attack against tumor cells we have genetically engineered a fusion protein of SEA and the Fab part of the tumor-reactive mAb C215 . Injection of this Fab-SEA fusion protein to mice carrying lung metastases of the poorly immunogenic B16 melanoma transfected with the C215 Ag resulted in infiltration of cytokine-producing T cells, perforin-containing CTL, and a marked tumor elimination . Fab-SEA therapy induced substantial levels of IFN-gamma and TNF-alpha in serum . In the present study we have characterized the molecular mechanisms of the antitumor effect induced by Fab-SEA treatment in vivo . Neutralization of cytokines by specific Abs demonstrated a major role for IFN-gamma in the suppression of tumor growth . In addition, a minor contribution of TNF-alpha was recorded . Injections of Fab-SEA into normal mice induced strong CTL activity but failed to promote cytotoxic function in perforin knockout mice . Also, a markedly reduced therapy was noted in perforin knockout mice, implicating a role for CTL in Fab-SEA-mediated tumor eradication . The data suggest that Fab-SEA-targeted T cells may suppress tumor growth by both perforin-dependent cytotoxicity and local release of cytokines such as IFN-gamma . The latter mechanism may have an important role in cytostatic effects against Ag-negative bystander tumor cells.

J Immunol, 1998 Jun 1, 160(11), 5246 - 52
In vivo inflammatory response to a prototypic B cell superantigen: elicitation of an Arthus reaction by staphylococcal protein A; Kozlowski LM et al.; Staphylococcal protein A (SpA) is representative of a new class of Ags, the B cell superantigens (SAgs) . These SAgs, unlike conventional Ags, bind to the Fab regions of Ig molecules outside their complementarity-determining regions . In addition, B cell SAgs can react with a substantial amount of a host's serum Igs by virtue of their ability to interact with many members of an entire variable heavy chain (VH) or variable light chain gene family . For example, SpA reacts with the Fabs of most human Igs using heavy chains from the VH3 gene family (VH3+) . Members of this gene family are expressed on 30 to 60% of human peripheral B cells . We sought to determine whether the interaction of a B cell SAg with its reactive Igs can elicit immune complex-mediated tissue injury . Using the Arthus reaction in rabbits as an in vivo model of immune complex-mediated tissue inflammation, we demonstrated that untreated rabbits, which were administered SpA intradermally (i.d.), do not develop a cutaneous inflammatory response . However, when rabbits were pretreated i.v . with human IgG (hIgG), i.d . injections of SpA induced an inflammatory response with the classical histologic features of an Arthus reaction . To determine whether this Arthus-like response occurred via a B cell superantigenic mechanism, the rabbits were pretreated with VH3-depleted hIgG and then were administered SpA i.d . We found that the induction of a prominent inflammatory response by SpA was dependent upon the presence of VH3+ molecules in the hIgG pretreatment . These results provide compelling evidence that an interaction of the B cell SAg, SpA, with its reactive (VH3+) IgGs leads to an immune complex-mediated inflammatory response in vivo.

J Immunol, 1998 Jun 1, 160(11), 5231 - 8
CD95 (Fas)-based, superantigen-dependent, CD4+ T cell-mediated down-regulation of human in vitro immunoglobulin responses; Stohl W et al.; Naturally occurring microbial superantigens (SAg) have been implicated in several human idiopathic disorders, and a compelling argument for the role of SAg in autoantibody-associated disorders, such as systemic lupus erythematosus, has been proposed . To test the effects of SAg on human in vitro Ig responses, CD4+ T cell + B cell cultures were stimulated with graded doses of staphylococcal enterotoxin B (SEB) . Ig-secreting cell (IgSC) responses were very weak in CD4+ T cell + B cell cultures stimulated with SEB at the optimal mitogenic concentration (high dose SEB; 100 ng/ml) but were strong in parallel cultures stimulated with low dose SEB (0.01 ng/ml) . High dose SEB actually enhanced B cell differentiation in the presence of CD4+ T cell soluble helper factors as long as the B cells were prevented from physically contacting the CD4+ T cells . However, when cell-cell contact between CD4+ T cells and B cells was permitted, high dose, but not low dose, SEB promoted increased CD4+ T cell-mediated B cell apoptosis with resulting decreases in viable CD20+ B cells and IgSC . High dose, but not low dose, SEB triggered increased levels of soluble CD95 ligand, and down-regulation of IgSC responses and incremental apoptosis of activated B cells were prevented by antagonist anti-CD95 mAb . This strongly suggests that CD4+ T cell-mediated CD95-based killing of activated B cells plays a major role in controlling SEB-driven IgSC responses . Defects in SAg-based down-regulation may contribute to autoimmune disorders such as SLE.

Cornea, 1998 May, 17(3), 335 - 7
Staphylococcal endophthalmitis following cataract extraction in a patient with Darier's disease; Macsai MS et al.; PURPOSE: To report staphylococcal endophthalmitis following cataract extraction in a patient with Darier' s disease . METHODS: A 67-year-old man presented with decreased visual acuity OD and hypopyon 3 days status post-cataract extraction with intraocular lens placement . The patient was hospitalized and placed on topical and intravenous antibiotics . A diagnostic vitreous tap, pars plana vitrectomy, and intravitreal antibiotic installation were performed . RESULTS: Vitreal tap cultures indicated Staphylococcus epidermidis . His clinical status improved after vitrectomy and antibiotic therapy . The same bacteria was cultured from the patient's eyelids . CONCLUSIONS: The source of the S . epidermidis was the skin lesions on the patient's face and eyelids . Darier's disease is an exfoliative hyperkeratotic skin disease that affects all areas on the body except the buttocks . Although clean, sterile surgical techniques were followed, the risk of endophthalmitis following intraocular surgery in a patient with Darier's disease may be increased due to his or her dermatologic condition.

Infect Immun, 1998 Jun, 66(6), 2778 - 81
The abilities of a Staphylococcus epidermidis wild-type strain and its slime-negative mutant to induce endocarditis in rabbits are comparable; Perdreau-Remington F et al.; The abilities of a parent and mutant pair of Staphylococcus epidermidis strains, the slime-producing parent RP62A and its slime-negative mutant, to establish endocarditis in a rabbit model of aortic valve endocarditis and to accumulate and adhere to surfaces in vitro were compared . Vegetation titer and infection rate depended on the presence or absence of a catheter (P = 0.020) and on inoculum size (P < 0.001) but not on the infecting strain . The ability of the parent strain vis-a-vis its mutant to accumulate in vitro on surfaces as demonstrated in a slime test did not correlate with any enhancement in the development of endocarditis in the rabbit model . In vitro initial adherence rates were identical . Both isolates accumulated to the same reduced extent in vitro in the presence of serum, albumin, or gelatin . Adhesion was equally promoted by addition of fibronectin . These data suggest that the in vitro phenomenon of accumulation described as slime production in the absence of serum may not be an important virulence determinant in vivo.

Biochemistry, 1998 Apr 28, 37(17), 5785 - 90
Pressure denaturation of proteins: evaluation of compressibility effects; Prehoda KE et al.; One of the key pieces of information from pressure denaturation experiments is the standard volume change for unfolding (Delta V(o)) . The pressure dependence of the volume change, the standard compressibility change (Delta K(o)T), is typically assumed to be zero in the analysis of these experiments . We show here that this assumption can be incorrect and that the neglect of compressibility differences can skew the interpretation of experimental results . Analysis of experimental, variable-pressure NMR data for bovine pancreatic ribonuclease A in 2H2O at pH 2.0 and 295 K yielded the following statistically significant, non-zero values: Delta K(o) T = 0.015 +/- 0.002 mL mol-1 bar-1, Delta V(o) = -21 +/- 2 mL mol-1, and Delta G(o) = 2.8 +/- 0.3 kcal mol-1 . The experimental protein stability is in good agreement with one (Delta G(o) = 2.5 kcal mol-1) determined independently for the same protein by calorimetry at atmospheric pressure under equivalent conditions {Makhatadze, G . I., Clore, G . M., and Gronenborn, A . M . (1995) Nat . Struct . Biol . 2, 852-855} . The positive value for Delta K(o)T indicates that the denatured form of ribonuclease A is more compressible than the native form; this is explained in terms of an interplay between the intrinsic compressibility of the protein and solvation effects . When the same data were fitted to a model that assumes a zero compressibility change, the Delta G(o) value of 4 . 0 +/- 0.1 kcal mol-1 returned by the model no longer agreed with the independent measurement, and the Delta V(o) returned by the model was a very different -59 +/- 1 mL mol-1 . By contrast, it was not possible to carry out a similar thermodynamic analysis of fluorescence spectroscopic data for the denaturation of staphylococcal nuclease to yield well-defined values of Delta G(o), Delta V(o), and Delta K(o)T . This limitation was shown by evaluation of synthetic data to be intrinsic to spectroscopic data whose analysis requires fitting of the plateaus at either side of the transition . Because NMR data do not have this requirement, they can be analyzed more rigorously.

Protein Eng, 1997 Dec, 10(12), 1433 - 43
Spontaneous oligomerization of a staphylococcal alpha-hemolysin conformationally constrained by removal of residues that form the transmembrane beta-barrel; Cheley S et al.; Staphylococcal alpha-hemolysin is a water soluble, monomeric, bacterial exotoxin, which forms heptameric pores in membranes . The rate determining step in assembly is the conversion of a heptameric prepore to the fully assembled pore in which the central glycine-rich domain of each subunit inserts into the membrane to form a 14 strand beta barrel . Barrel formation is accompanied by a conformational change in which each N terminus latches onto an adjacent subunit . In the monomer in solution, the central domain is loosely organized and exposed to solvent . In this study, 25 amino acids of the central domain were removed and replaced with the sequence Asp-Gly, which favors the formation of a type I' beta-turn, to yield a mutant devoid of hemolytic activity . Within minutes after synthesis in the absence of membranes, the mutant polypeptide spontaneously assembled into heptamers, as demonstrated by atomic force microscopy . Limited proteolysis suggested that the N termini of the subunits in the heptamers were in the fully assembled pore conformation rather than the prepore conformation . Based on these findings, the deletion is proposed to constrain the central domain and thereby force the creation of a shortened beta barrel, which in turn induces the additional structural changes that normally accompany pore formation . The truncated pore might make a useful framework for the construction of designed membrane active macromolecules.

J Mol Biol, 1998 Apr 3, 277(3), 733 - 45
Refolding kinetics of staphylococcal nuclease and its mutants in the presence of the chaperonin GroEL; Tsurupa GP et al.; We have analyzed the effect of the chaperonin GroEL on the refolding kinetics of staphylococcal nuclease and its three mutants by stopped-flow fluorescence measurements . It was found that a transient folding intermediate of staphylococcal nuclease was tightly bound to GroEL and refolded in the GroEL-bound state without releasing the non-native protein in solution, and the refolding rate in the GroEL-bound state was 0.01 s-1 . The GroEL-affected refolding of the nuclease appears to be in decided contrast to that of apo-alpha-lactalbumin reported in our previous study, wherein alpha-lactalbumin was shown to be more weakly bound by GroEL and to refold in the free state in solution . In spite of the apparent difference between the proteins, the GroEL-affected refolding reactions of both the proteins can be represented by a common unified reaction scheme . On the basis of this scheme, the binding constant between the nuclease intermediate and GroEL was estimated to be larger than 10(9) M-1 . The stoichiometry of binding of the nuclease and its mutants to GroEL was found to be two (nuclease/GroEL 14-mer) . The increase in ionic strength resulted in a weakening of the interaction between the nuclease and GroEL, which was attributed to a weakening of the electrostatic attraction between the two proteins as a result of electrostatic screening by ions . Although ATP was found to accelerate the GroEL-affected refolding of the nuclease, the refolding rate was still far from the rate of the free refolding . The free refolding behavior of the nuclease and its mutants was restored in the presence of the cochaperonin GroES and ATP .

No Shinkei Geka, 1998 Apr, 26(4), 357 - 62
{Multiple bacterial aneurysms: case report}; Suzuki Y et al.; A 59-year-old man presented with an internal carotid artery (ICA) bacterial aneurysm which ruptured during surgery for treatment of another bacterial aneurysm . He had been admitted to our hospital because of the recurrence of colon cancer . He had undergone aortic and mitral valve replacement because of closure incompetence due to bacterial endocarditis two months previously . Two months after treatment for colon cancer, he developed fever, and arterial blood culture demonstrated . Staphylococcus epidermidis . A few days later, he suddenly suffered severe headache and vomiting, followed by deterioration of consciousness . CT showed subarachnoid hemorrhage and angiography showed a saccular aneurysm at the opercular portion of the left middle cerebral artery (MCA) . Immediate clipping of the aneurysm was attempted . The carotid cistern was opened via a left frontotemporal craniotomy, but an ICA aneurysm, which had not been previously recognized, ruptured suddenly . The ICA aneurysm was wrapped with Vascwrap with some difficulty . The MCA aneurysm was then trapped . Postoperatively, the patient continued to be stuporous for a few days . Two weeks later, he died of complications caused by pneumonia . Bacterial aneurysm is more likely to be located in the distribution of the distal arterial tree, mainly in the distribution of the MCA . The difficulty of preoperative diagnosis and the unpredictable clinical course of bacterial aneurysms are emphasized.

J Biochem (Tokyo), 1998 Apr, 123(4), 760 - 5
Presence of conserved domains in the C-terminus of MARCKS, a major in vivo substrate of protein kinase C: application of ion trap mass spectrometry to the elucidation of protein structures; Yamauchi E et al.; MARCKS, the major protein kinase C substrate in various cells and tissues, binds to calmodulin, acidic membrane phospholipids, and actin filaments, and these interactions are regulated by protein phosphorylation . We have previously analyzed MARCKS purified from bovine brain using capillary liquid chromatography/electrospray mass spectrometry and found that the protein structure differed significantly from that deduced from cDNA sequences {Taniguchi, H., Manenti, S., Suzuki, M., and Titani, K . (1994) J . Biol . Chem . 269, 18299-18302} . Moreover, the alignment of the protein from various species showed a lack of any conserved sequences in the C-terminal half of the molecule . This prompted us to reexamine the C-terminal amino-acid sequence of bovine MARCKS . The purified protein was digested with lysyl endoprotease, and the obtained C-terminal peptide was further digested with either Staphylococcus V8 protease or NTCB . The small peptides thus obtained were analyzed by liquid chromatography/electrospray/tandem mass spectrometry . This combined with gas-phase Edman sequencing allowed us to determine the C-terminal primary structure . The sequence obtained differed significantly from that reported previously, and the comparison with other species revealed the presence of a novel conserved domain in the C-terminal region of MARCKS.

J Immunol, 1998 May 15, 160(10), 5170 - 80
In vivo effects of a bacterial superantigen on macaque TCR repertoires; Kou ZC et al.; A macaque model was employed to explore staphylococcal enterotoxin B (SEB) superantigen-driven T lymphocyte responses . The SEB-reactive Vbeta+ cell subpopulations demonstrated a striking tri-phase response in rhesus monkeys following an SEB challenge in vivo . The hyperacute down-regulation, seen as early as 2 h through 2 days after SEB injection, was characterized by a disappearance of the reactive Vbeta-restricted PBL subpopulations from the circulation and decreased expression of these cell subpopulations in lymphoid tissues . Following this, a dominant expansion of reactive Vbeta-expressing CD4+ cell subpopulations occurred in lymph nodes and spleens, whereas in the peripheral blood a preferential expansion of reactive Vbeta-expressing CD8+ cell subpopulations was seen . An exhaustion of this response was then seen, with a prolonged decrease in the number of the reactive Vbeta+ CD4+ lymphocyte subpopulations . Interestingly, monoclonal or oligoclonal dominance was seen in the reactive Vbeta+ cell subpopulations in the period of the transition from the polyclonal cellular expansion to the exhaustion of the response, suggesting that some Vbeta+ cell clones may be more resistant than others to superantigen-mediated depletion . These results indicate that in vivo SEB superantigen-mediated effect on lymphocyte subpopulations in macaques is complex, suggesting that profound dynamics in the TCR repertoires may in part account for the susceptibility of higher primates to SEB-induced diseases.

J Heart Valve Dis, 1998 Mar, 7(2), 235 - 9
Repair of fungal aortic prosthetic valve endocarditis associated with periannular abscess; Remsey ES et al.; The incidence of prosthetic valve endocarditis is 2-4%; in most cases the involved organisms are Staphylococcus epidermidis and Staph . aureus . Fungal endocarditis is much less common (incidence < 0.1%), but it is often fatal, with a long-term mortality rate of 90-100% . Most fungal endocarditis cases occur after aortic valvular surgery, due to Candida sp . Late-onset symptoms, long-term development and aggressive nature of the fungus makes its eradication complicated and treatment difficult . Fungal valvular mycoses produce systemic embolization and cause serious perioperative bleeding on resection of infected tissue . Usually surgery includes aortic root replacement with an aortic homograft conduit after radical debridement, to attain local sterilization . This report describes a patient with complex infection, requiring replacement of an infected prosthetic valve with an aortic homograft conduit, aggressive and radical debridement of infected tissue, and reconstruction using biologic tissues . The case demonstrates the importance of perioperative and long-term antifungal treatment and presents a modified 'Cabrol procedure' to prevent critical intraoperative hemorrhage.

Proc Natl Acad Sci U S A, 1998 May 12, 95(10), 5480 - 3
Association-induced folding of globular proteins; Uversky VN et al.; It has generally been assumed that the aggregation of partially folded intermediates during protein refolding results in the termination of further protein folding . We show here, however, that under some conditions the association of partially folded intermediates can induce additional structure leading to soluble aggregates with many native-like properties . The amount of secondary structure in a monomeric, partially folded intermediate of staphylococcal nuclease was found to double on formation of soluble aggregates at high protein or salt concentrations . In addition, more globularity, as determined from Kratky plots of small-angle x-ray scattering data, was also noted in the associated states.

Eur J Immunol, 1998 Apr, 28(4), 1417 - 25
The crucial role of IL-10 in the suppression of the immunological response in mice exposed to staphylococcal enterotoxin B; Hasko G et al.; Staphylococcal enterotoxin B (SEB), a bacterial superantigen, activates the immune system resulting in a burst of pro- and anti-inflammatory cytokines . A central anti-inflammatory mediator in this process is IL-10 . Using IL-10-deficient C57BL/6 (IL-10 KO) mice, we studied the role of endogenous IL-10 in the regulation of the immune response to SEB . SEB (100 microg) induced the release of IL-10 in control C57BL/6 {IL-10 wild type (WT)} mice, but not in their IL-10 KO counterparts . SEB-evoked plasma levels of TNF-alpha, IL-1beta, IL-2, IL-6, IL-12 and IFN-gamma were significantly higher in the IL-10 KO mice than in the WT animals . The release of macrophage inflammatory proteins-1alpha and -2 was also enhanced in the IL-10 KO mice . Further, upon SEB challenge, mice deficient in IL-10 produced higher levels of nitric oxide than the WT animals . IL-10 deficiency resulted in a marked enhancement of the SEB-induced apoptosis of thymocytes . Finally, IL-10 KO mice were more susceptible to SEB-induced lethal shock than their WT controls . These results show that IL-10 plays an important immunoregulatory role in the response to a superantigenic stimulus, by dampening of the shock-inducing inflammatory response and early activation-induced cell death elicited by SEB.

Eur J Haematol, 1998 Apr, 60(4), 233 - 9
Antibody-directed superantigen-mediated T-cell killing of myeloid leukaemic cell line cells; Gidlof C et al.; Bacterial superantigens (SAgs) bound to MHC class II molecules on target cells are efficient activators of cytotoxic T cells expressing certain T cell receptor (TCR) Vbeta regions We described earlier that the specificity of the SAg Staphylococcus enterotoxin A (SEA) can be changed by introducing a D227A point mutation in the major MHC class II binding site and by genetically fusing the SEA mutant (SEAm) to protein A (PA) . This SEAm-PA fusion protein can then be used to direct cytotoxic T cells to tumour cells coated with monoclonal antibodies (mAbs) . In this communication, we tested the PA-SEAm fusion protein together with mAbs against the myeloid cell surface antigens CD13, CD15 and CD33 . A SEA-reactive T cell line was used as effector cells against 10 different myeloid leukaemic cell lines . Optimal lysis of antigen positive leukaemic cells was obtained at a PA-SEAm concentration of 1 ng/ml and effector : target cell ratios of 15 : 1 . No correlation between target cell sensitivity and the level of surface antigen expression could be seen . The 6 acute myeloid leukaemia (AML) cell lines tested appeared to be more sensitive than the 4 chronic myeloid leukaemia (CML) cell lines . The sensitivity of the AML cell line HL-60 could be improved further by stimulation with TNFalpha . This was accompanied by increased surface ICAM-1 expression whereas specific target molecule expression (CD13, CD33) was unchanged . This suggests that sensitivity to lysis is related to the leukaemic subtype and ICAM-1 expression but not to the tumour antigen density . Our results show that it is possible to direct cytotoxic T cells to myeloid leukaemia cells by using SAgs linked to mAbs, and encourage the construction and testing of a recombinant direct SAg-mAb fusion protein as a candidate drug for therapy of myeloid leukaemias.

Cell Immunol, 1998 Jan 10, 183(1), 42 - 51
Quantitative defect in staphylococcal enterotoxin A binding and presentation by HLA-DM-deficient T2.Ak cells corrected by transfection of HLA-DM genes; Albert LJ et al.; HLA-DM facilitates peptide acquisition by MHC class II proteins within the endosomes of APC by facilitating release of invariant chain peptide intermediates (CLIP) from the class II molecules . T2 cells have a deletion in the MHC II region which deletes HLA-DM and MHC II genes . T2 cells transfected with MHC class II proteins are defective in protein presentation, a defect that is corrected by HLA-DM transfection . Here we show that T2 cells transfected with Ak are also impaired in binding and presentation of the superantistaphylococcal enterotoxin A and that HLA-DM transfection corrects this defect . The poor ability of SEA to bind to Ak on DM-deficient cells is somewhat surprising since Ak has a low affinity for CLIP and is not predominantly occupied with CLIP on T2 cells compared to wide-type APC . These data suggest an influence of HLA-DM on the structure or composition of the Ak/peptide complex beyond its role in the release of invariant chain peptides.

J Clin Pathol, 1998 Jan, 51(1), 62 - 7
Identification of multiresistant Staphylococcus epidermidis in neonates of a secondary care hospital using pulsed field gel electrophoresis and quantitative antibiogram typing; Sloos JH et al.; AIMS: To determine the diversity of types of Staphylococcus epidermidis in a neonatal care unit of a secondary care hospital in the Netherlands . METHODS: In a prospective study, specimens from nose, ear, axilla, umbilicus, and groin were taken from patients twice a week during a period of up to two weeks . All isolates were typed by both pulsed field gel electrophoresis (PFGE) and antibiogram analysis . RESULTS: Fifty three S epidermidis isolates from 15 of 24 patients were obtained in one to four surveys . Fourteen isolates from six patients had a common PFGE pattern and were of one multiresistant antibiogram type . The remaining 39 isolates were allocated to 24 sporadic PFGE types and were more susceptible to antibiotics . Colonisation with the multiresistant strain correlated with a long period of stay and with the use of specific antibiotics . The multiresistant isolates were related closely to isolates of S epidermidis found in a recent study in a teaching hospital in the vicinity of the secondary care hospital . CONCLUSION: Repeated sampling and the use of two typing methods allowed the identification of two closely related multiresistant S epidermidis strains in two hospitals in the same area.

J Vasc Surg, 1998 Apr, 27(4), 689 - 98
Treatment of vascular graft infection by in situ replacement with cryopreserved aortic allografts: an experimental study; Knosalla C et al.; PURPOSE: The purposes of this study were to prove the efficacy of cryopreserved aortic allografts to treat an established vascular graft infection by in situ replacement in an animal model and to evaluate the role of the antibiotics normally used to decontaminate the allografts . METHODS: Twenty-three dogs underwent infrarenal aortic replacement with a gelatin-sealed knitted polyester graft contaminated in vitro by Staphylococcus epidermidis RP-62 . One week later, the 18 surviving animals underwent reoperation for graft removal and were randomized into three groups for in situ replacement: group I (control, n = 6) received a new gelatin-sealed graft; group II (n = 6) received a non-antibiotic-treated cryopreserved allograft; and group III (n = 6) received an antibiotic-treated cryopreserved allograft . Control grafts and allografts were removed 4 weeks after the initial intervention for quantitative bacteriologic analysis and histologic analysis . Bacteriologic results were expressed as colony-forming units per square centimeter of graft material . Qualitative bacteriologic analysis was also obtained from perigraft fluid and tissue . RESULTS: All of the initially implanted grafts and all of the control grafts (group I) were infected at the time of removal . In group II, three out of six allografts were not totally incorporated, whereas in group III incorporation was always complete, with a significantly decreased inflammatory reaction . All of the antibiotic-treated allografts were sterile, whereas three untreated allografts grew bacteria . CONCLUSIONS: In this model, cryopreserved aortic allografts were more resistant to reinfection than synthetic grafts after in situ replacement of an infected prosthetic graft . However, the antibiotic loading of the cryopreserved aortic allograft appears to be essential to obtain optimal therapeutic effects.

Klin Lab Diagn, 1998 Mar, (3), 36 - 8
{Method of quantitative determination of staphylococcal hyaluronidase activity}; Generalov II; The proposed method for measuring hyaluronidase activity of microorganism is based on prevention of hyaluronic acid clot formation with rivanol under the effect of hyaluronidase . This made possible the quantitative and qualitative detection of hyaluronidase activities of different staphylococcus species and strains . The maximum level of the enzyme and highest rate of its detection were typical of St . aureus . Its strains producing hyaluronidase in quantities of at least 0.5 IU are significantly (p < 0.01) more often isolated from medical staff.

J Bacteriol, 1998 May, 180(9), 2273 - 9
Regulation of lactose utilization genes in Staphylococcus xylosus; Bassias J et al.; The lactose utilization genes of Staphylococcus xylosus have been isolated and characterized . The system is comprised of two structural genes, lacP and lacH, encoding the lactose permease and the beta-galactosidase proteins, respectively, and a regulatory gene, lacR, coding for an activator of the AraC/XylS family . The lactose utilization genes are divergently arranged, the lacPH genes being opposite to lacR . The lacPH genes are cotranscribed from one promoter in front of lacP, whereas lacR is transcribed from two promoters of different strengths . Lactose transport as well as beta-galactosidase activity are inducible by the addition of lactose to the growth medium . Primer extension experiments demonstrated that regulation is achieved at the level of lacPH transcription initiation . Inducibility and efficient lacPH transcription are dependent on a functional lacR gene . Inactivation of lacR resulted in low and constitutive lacPH expression . Expression of lacR itself is practically constitutive, since transcription initiated at the major lacR promoter does not respond to the availability of lactose . Only the minor lacR promoter is lactose inducible . Apart from lactose-specific, LacR-dependent control, the lacPH promoter is also subject to carbon catabolite repression mediated by the catabolite control protein CcpA . When glucose is present in the growth medium, lacPH transcription initiation is reduced . Upon ccpA inactivation, repression at the lacPH promoter is relieved . Despite this loss of transcriptional regulation in the ccpA mutant strain, beta-galactosidase activity is still reduced by glucose, suggesting another level of control.

Intensive Care Med, 1998 Mar, 24(3), 258 - 61
Fibrinolytic changes in a patient with toxic shock syndrome; release of active u-PA; Haj MA et al.; OBJECTIVE: Definition of the changes in the activators of plasminogen, u-PA and t-PA, and examination of the possible generation of plasmin in the circulation in overwhelming sepsis . DESIGN: Serial blood analysis starting 4 h after development of symptoms of toxic shock syndrome . SETTING: Intensive care unit . PATIENT: A previously healthy woman underwent endometrial ablation and rapidly thereafter developed staphylococcal toxic shock syndrome with organ failure . MEASUREMENT AND RESULT: t-PA, PAI-1, t-PA-PAI-1 complexes, plasminogen, fibrinogen and plasmin-alpha 2-antiplasmin complexes were measured serially by ELISA and free u-PA by SDS-PAGE with zymography . The onset of symptoms was accompanied by a rise of t-PA antigen-followed rapidly by PAI-1 antigen . Plasmin-alpha 2-antiplasmin complex was generated in large amounts but disappeared within hours . From day 3, free u-PA was detectable in the circulation without plasmin generation . CONCLUSION: Despite the sustained presence of active u-PA in the circulation and of t-PA antigen at the onset of symptoms, plasmin-alpha 2-antiplasmin generation was largely suppressed by high levels of PAI-1 . The suppression of plasmin generation by u-PA and t-PA may ensure the persistence of fibrin in the microcirculation and so contribute to organ failure.

J Bone Joint Surg Am, 1998 Apr, 80(4), 481 - 91
Infection after total elbow arthroplasty; Yamaguchi K et al.; The purpose of this study was to review our experience with the treatment of twenty-five infections (in twenty-five patients) after total elbow arthroplasty and to examine indications for salvage of the prosthesis compared with those for resection arthroplasty . The patients were divided into three groups on the basis of treatment . Group I comprised fourteen patients who were managed with multiple, extensive irrigation and debridement procedures with retention of the original components . The primary indication for retention of the prosthesis was evidence that it was well fixed as determined both radiographically and intraoperatively . Group II comprised six patients who had removal of the prosthesis and debridement followed by immediate or staged reimplantation . Group III comprised five patients who were managed with resection arthroplasty . The infection was successfully eradicated in seven of the fourteen elbows that had salvage of the prosthesis with irrigation and debridement . The results were strongly dependent on the causative organism; attempts at debridement failed in the four elbows that were infected with Staphylococcus epidermidis compared with three of the ten that were infected with another organism . Four of the six patients in Group II had successful reimplantation of a prosthesis; in three, the infection had been caused by an organism other than Staphylococcus epidermidis . Only one of the three patients who had a Staphylococcus epidermidis infection had a successful reimplantation . None of the five patients who had a resection arthroplasty had signs of infection at the latest follow-up examination . We concluded that salvage of the prosthesis with extensive irrigation and debridement in the presence of an infection about the elbow can be reasonably successful if the infecting organism is not Staphylococcus epidermidis and if the components are well fixed . When removal of the components is warranted, staged reimplantation can also be highly successful when the infecting organism is not Staphylococcus epidermidis . However, the repeated operations necessary to retain a prosthesis and the high rates of complications seen with this approach--and the relatively good rates of satisfaction obtained with resection arthroplasty--suggest that resection arthroplasty remains the procedure of choice in medically frail patients or in patients for whom function of the elbow is less of a concern.

Ophthalmologica, 1998, 212(3), 184 - 7
In vitro Staphylococcus epidermidis growth in some viscoelastic substances containing sodium hyaluronate; Gallenga PE et al.; The aim of our study was to verify the in vitro growth of Staphylococcus epidermidis in various dilutions of some viscoelastic substances containing hyaluronic acid (Healon and Healon GV, IAL, Biolon) . Serial twofold dilutions of each sterile viscoelastic substance, prepared so as to obtain a final concentration ranging from 50 to 0.78% of the product in sterile saline solution (0.85% NaCl), were taken out with a pipette that delivered 1.0 ml/tube . One hundred microliters of the S . epidermidis inocula, used for the evaluation of the positive control of the test organism, was dispensed into each tube . After 24 h of aerobic incubation at 37 degrees C, 100 microl of sample was taken out from each tube and plated into the specific medium for the growth of the test organism . After 24 h of incubation at 37 degrees C, these agar plates were examined and the colony-forming unit count of the test organism was compared to the corresponding total colony count, acting as a positive control, in order to determine the quantitative variation of the test organism grown in the presence of the viscoelastic compounds . For the lowest dilutions (from 1:2 to 1:8) statistically significant bacterial growth was detected in all tested viscoelastic substances . For the highest dilutions (1:64 and 1:128) Biolon and Healon GV showed a significant inhibition of S . epidermidis growth . A significant inhibition was also observed in the highest dilution (1:128) of Healon . In every dilution of IAL a statistically significant increase in bacterial growth was observed . It remains to be carefully considered whether S . epidermidis, accidentally penetrating the eye via the intraocular lens, could find a culture medium in a small amount of sodium hyaluronate left in the capsular bag behind the optic.

Immunol Invest, 1998 Jan-Feb, 27(1-2), 73 - 88
T cell dependent B cell activation occurs during the induction of T cell anergy by staphylococcal enterotoxin B in mice; Abdul-Majid KB et al.; Staphylococcal enterotoxin B (SEB) can activate specific T cell clones bearing specific TcR V beta domains together with MHC class II ligands on accessory cells . The release of proinflammatory cytokines is the consequence of this activation as well as the main pathological aspect involved in SEB infection . This current study looked at the active role of both T and B cells during the induction of anergy by SEB in vivo . Euthymic and nude BALB/c mice were injected with SEB and over a period of 8 days, cells from the spleen and sera from the blood were collected . After a single injection with SEB (50 micrograms/mouse), a transient increase of CD4+V beta 8+ T cells were detected after 2 days followed by a decrease after 4 days, which persisted until day 8 . These clones were rendered anergic upon restimulation in vitro with SEB . Interestingly, cells taken out 2 days after SEB injection, exhibited reduced proliferation in response to Con A . However, this response gradually recovered on days 4, 6 and 8 . Furthermore, early IgM antibody production (day 2) was observed after SEB injection . SEB-induced IgM antibody production in euthymic BALB/c was found to have specificity against SEB, cardiolipin (CL) and phosphatidylethanolamine (PE) . SEB-treated nude mice did not produce antibody secreting cells in response to SEB, indicating that this process is T cell dependent.

Eur J Immunol, 1998 Mar, 28(3), 874 - 80
Ceramide-independent CD28 and TCR signaling but reduced IL-2 secretion in T cells of acid sphingomyelinase-deficient mice; Stoffel B et al.; Ceramide generated by lysosomal acid sphingomyelinase (aSMase) has been proposed to contribute to CD28 co-stimulatory signaling pathways . We used an aSMase-deficient mouse line (asmase-/-) to elucidate the role of the aSMase in splenocytes stimulated with either a combination of anti-CD3 and anti-CD28 antibodies, the lectin concanavalin A (Con A) or the superantigen staphylococcal enterotoxin B . All stimuli were shown to induce IL-2 expression, Con A additionally triggered the expression of high-affinity IL-2 receptor . However, in asmase-/- mice secretion of IL-2 was significantly reduced, whereas the intracellular IL-2 levels were elevated . Proliferation of anti-CD3/anti-CD28 or Con A-stimulated aSMase-deficient splenocytes was reduced up to 50% after 72 h in comparison to wild-type cells . We conclude that ceramide generated by aSMase is not involved in CD28 signal transduction, but rather a perturbation of the secretory system is responsible for the impaired proliferation of aSMase-deficient splenocytes.

Acta Paediatr, 1998 Mar, 87(3), 349 - 50
Coexistence of acquired protein S and protein C deficiency and the Arg506Gln mutation in factor Va in a child with severe thromboembolic disease; Shavit I et al.; An 11-y-old girl who presented with cellulitis and clinical signs of deep vein thrombosis (DVT) is reported here . She developed staphylococcal sepsis, recurrent septic emboli and a large vegetation on the tricuspid valve . The patient was found to be heterozygous for the Arg506Gln mutation in factor Va and had low levels of protein C and protein S during the sepsis . The coexistence of the two thrombophilic states may explain the severe thromboembolic manifestations.

Gen Pharmacol, 1998 May, 30(5), 777 - 82
Cyclosporin A and FK-506 inhibit development of superantigen-potentiated collagen-induced arthritis in mice; Takaoka Y et al.; 1 . Staphylococcal enterotoxine B (SEB; superantigen) accelerated the onset of arthritis in mice preimmunized with type II collagen (SEB-potentiated collagen-induced arthritis) . Cyclosporin A and FK-506 inhibited the induction and development of clinical signs and histopathological changes of SEB-potentiated collagen-induced arthritis in mice . 2 . Simultaneously, both cyclosporin A and FK-506 inhibited the development of humoral and cellular immunity to type II collagen . 3 . The expression of IL-2 receptor (CD25) by SEB on splenocyte T cells from collagen-preimmunized mice was inhibited by both agents in ex vivo experimentation.

Bull Soc Pathol Exot, 1998, 91(1), 104 - 8
{A great Franco-Mauritian epidemiologist: Joseph Désiré Tholozan (1820-18970}; Theodorides J; Born in 1820 from French parents in Diego Garcia, an islet then linked to Mauritius where he started in Port-Louis his school years, Joseph Desire Tholozan was an original personality . He undertook medical studies in France (M . D . thesis, Paris, 1843) after having joined the military Health Service (1841) as a surgeon serving in various garrisons in the country and later at the Hospital of the Valde-Grace in Paris (1849) . Successful at the "agregation" of Medicine in 1853, he later participated to the Crimean War (1854-1855) where he performed-interesting medical observations . In 1858, he was appointed personal physician to Nasreddin Shah and remained in Persia until his death in Teheran (1897) where he is buried . Tholozan published between 1847 and 1892 over fifty articles and books dealing chiefly with infectious pathology and epidemiology, written at a time when microbial etiology and specificity of such diseases were wholly unknown . He considered chiefly bubonic plague, studying as soon as 1871 the focus of the Iranian Kurdistan, a research which will be resumed by M . Baltazard and his collaborators between 1947 and 1971, i.e . a century later . He was also deeply interested by the "oriental" cholera of which he recalled masterly the history and geography in the Near and Middle East . He also performed, while in Crimea and Persia, personal observations on tuberculosis, diptheria, remittent fever, acrodynia and had studied in France in his early years various other diseases such as cutaneous staphylococcic infections, glanders, pulmonary haemorrhagies, etc . In Persia, he reorganized Public Health and medical teaching and educated many local physicians and surgeons . Being assured of the unlimited confidence of the Shah, he played an important cultural role, promoting French influence in Persia . Holder of many French and foreign decorations, Tholozan was Fellow of the French Academies of Sciences and Medicine . His name was given by Laboulbene to Ornithodoros tholozani, a tick vector of a recurrent fever (spirochetosis due to Borrelia persica), of which he had described both the symptoms and the vector in 1882.

Jpn J Thorac Cardiovasc Surg, 1998 Feb, 46(2), 170 - 4
{Postoperative infections related to pacing wires, pulmonary arterial catheters, and drainage tubes temporarily inserted during open-heart surgery}; Kanoh M et al.; Bacterial examinations of temporary pacing wires (P-wires), pulmonary arterial (P-A) catheters, and drainage tubes temporarily inserted during open-heart surgery were performed in 213 patients . Bacteria were detected in 19 (2.8%) of 672 specimens gathered from the subject patients, with coagulase-negative Staphylococcus (CNS) being most frequently observed . P-wires accounted for 17 out of 19 of the culture-positive specimens, and 7 of the P-wires remained in place for more than two weeks . The frequency of infection with the P-wires was significantly higher than with the P-A catheters or drainage tubes . The period of time that the P-wire was left in place significantly longer than for P-A catheter or drainage tube . There was, however, no statistically significant difference between the culture-positive and negative groups in respect to age, detention periods, operation times, CPB times, or length of ICU stay . As a result of these findings, we have concluded that P-wires should be removed as soon as possible following surgery, and in any case, a meticulous care should be taken to prevent transcutaneous infection.

J Immunol, 1998 Apr 15, 160(8), 3855 - 60
CTLA-4 regulates tolerance induction and T cell differentiation in vivo; Walunas TL et al.; Cytotoxic T lymphocyte Ag-4 (CTLA-4; CD152) is an important T cell regulatory molecule . In vitro experiments have shown that the blockade of signals through CTLA-4 augments T cell expansion, while CTLA-4 cross-linking results in decreased T cell proliferation due to decreased IL-2 production . However, less is known about the role of CTLA-4 in regulating an ongoing immune response . In this study, we examined the role of CTLA-4 in the expansion, decline, tolerization, and differentiation of T cells following treatment with staphylococcal enterotoxin B (SEB) . Anti-CTLA-4 treatment resulted in increased numbers of SEB-reactive T cells and blockade of subsequent tolerance induction . Further examination of the SEB-reactive cells from anti-CTLA-4-treated mice demonstrated that both the CD4+ and CD8+ Vbeta8+ T cells produced IL-4, providing evidence that not only do signals through CTLA-4 regulate T cell-tolerizing events, but they also play an important role in the differentiation of T cells in vivo.

Biochemistry (Mosc), 1998 Apr, 63(4), 470 - 5
Equilibrium unfolding of partially folded staphylococcal nuclease A2- and A3-forms is accompanied by the formation of an intermediate state; Uversky VN; Structural properties of an equilibrium intermediate formed upon urea-induced unfolding of more ordered staphylococcal nuclease A-forms (A2 and A3) are studied . The effect of association on the structural properties and conformational stability of this unfolding intermediate is also considered . A close structural similarity (including tendency for association) is shown between this intermediate and the least ordered A1-form, induced in the acid-unfolded nuclease by moderate sulfate or chloride concentrations.

Biochemistry (Mosc), 1998 Apr, 63(4), 463 - 9
Structural properties of staphylococcal nuclease in oligomeric A-forms; Uversky VN et al.; Association affects the structural properties of different partially folded conformations of staphylococcal nuclease induced by anions of different nature . It is shown that oligomerization induces new structural levels in non-native A-forms . A close structural similarity between the monomeric A2 and the dimeric (A1)2 forms as well as between the monomeric A3 and oligomeric {(A1)2}M and {A2}M forms is established . This suggests that association of a protein molecule in partially folded conformations can be an additional structure forming factor.

Biochemistry (Mosc), 1998 Apr, 63(4), 456 - 62
Structural effect of association on protein molecules in partially folded intermediates; Uversky VN et al.; Fluorescence and circular dichroism spectroscopy, small angle X-ray scattering and high performance liquid gel filtration have shown that oligomerization considerably affects the structural properties and conformational stability of partially folded intermediates of staphylococcal nuclease . Conformational transitions induced by different anions and association in the acid-unfolded protein are described . It is shown that association of non-native conformations of the protein molecule can be an additional structuring factor . The corresponding folding schemes and phase diagrams are suggested.

Zhonghua Min Guo Xiao Er Ke Yi Xue Hui Za Zhi, 1998 Jan-Feb, 39(1), 55 - 7
Pneumonia with pneumatocele formation caused by Mycobacterium tuberculosis: report of one case; Kao HC et al.; Pneumatoceles are usually characteristic of staphylococcal pneumonia . They are rarely formed as one of the complications of Mycobacterium tuberculosis pneumonia . We report a 1-year-and-5-month old male child with pneumonia who was confirmed to have the rare complication, pneumatocele formation, by chest radiography and computed tomography . Since the patient did not respond to empiric antibiotic therapy, gastric larvage through a nasogastric tube was performed on three consecutive mornings and, as a result, acid fast rods were found on the three specimens . The cultures subsequently yielded M . tuberculosis . He was finally cured with a 6-month course of antituberculous chemotherapy . We conclude that tuberculosis should be considered in infants or young children with pneumonia that presents radiologically as pneumatocele formation, especially in whom there has been no response to empiric antibiotic therapy.

J Immunol, 1998 Jan 1, 160(1), 225 - 32
Staphylococcal enterotoxin D is a promiscuous superantigen offering multiple modes of interactions with the MHC class II receptors; Al-Daccak R et al.; Dimerization of MHC class II molecules on the cell surface of human THP-1 monocytic cell line is a requirement for staphylococcal superantigen (SAG)-induced cytokine gene expression . The capacities of various SAG to induce this response are governed by their modes of interaction with MHC class II molecules . Staphylococcal enterotoxin A (SEA), with its two binding sites, dimerizes MHC class II molecules and subsequently induces cytokine gene expression in THP-1 cells . Here, we demonstrate that staphylococcal enterotoxin D (SED) and staphylococcal enterotoxin E (SEE) induce, similarly, IL-1beta and TNF-alpha gene expression in these cells . Using mutated toxins that lost their binding site with the MHC class II alpha- or beta-chain, we demonstrate that this response is also mediated by the dimerization of MHC class II molecules through two binding sites . Furthermore, SED forms Zn2+-dependent homodimers that allow multiple modes of MHC class II clustering, including ligation of alpha-chains (alpha/alpha), beta-chains (beta/beta), or the alpha- and beta-chains of two different class II molecules . The beta/beta interaction following Zn2+-dependent SED/SED homodimer formation seems to be mediated by the appearance of a novel binding site on SED that interacts with histidine 81 of the MHC class II beta-chain . The different modes of SED interactions also influence SED-induced T cell activation where simultaneous ligation of the alpha- and beta-chains is essential for optimal response . These various modes of SED binding may be used to preserve bivalency regardless of variability in the MHC class II alpha/beta/peptide complexes.

J Immunol, 1998 Jan 15, 160(2), 615 - 23
Intercellular adhesion molecule-1 and leukocyte function-associated antigen-3 provide costimulation for superantigen-induced T lymphocyte proliferation in the absence of a specific presenting molecule; Lamphear JG et al.; Bacterial superantigens can bind TCR in the absence of MHC class II molecules and activate T lymphocytes when cocultured with certain class II-deficient accessory cells . It has not been determined, however, whether these accessory cells provide direct costimulation to the T cell or serve to present superantigens via a nonconventional ligand . We have identified a human adenocarcinoma cell line, SW480, that assists in the activation of human T cells by the staphylococcal enterotoxins B (SEB), C1 (SEC1), and D (SED), but not SEA, SEC2, SEC3, or SEE . SW480 cells did not express class II molecules, and anti-class II mAbs did not inhibit T cell proliferation, supporting the hypothesis that class II is not absolutely required for enterotoxin-mediated T cell activation . The TCR Vbeta profile of T cells stimulated by SEB plus SW480 cells was similar to that of T cells stimulated by SEB plus class II+ APC, indicating that TCR-SEB interactions were preserved in the absence of class II molecules . Binding studies failed to detect specific association of SEB with SW480 cells, suggesting that SW480 cells do not express receptors for enterotoxin . SEB coupled to beads, however, stimulated T cell proliferation, but only in the presence of SW480 cells . SW480 cells express both ICAM-1 and LFA-3 molecules, and the addition of Abs to these receptors inhibited T cell proliferation . These findings support a model in which certain enterotoxins engage the TCR independent of MHC class II or other specific presenting molecules and induce T cell proliferation with signals provided by nonconventional accessory cells.

J Immunol, 1998 Jan 15, 160(2), 559 - 65
Suppression of immune responses by CD8 cells . I . Superantigen-activated CD8 cells induce unidirectional Fas-mediated apoptosis of antigen-activated CD4 cells; Noble A et al.; Stimulation of mature CD4 cells through the TCR induces cellular activation and expansion that are often followed by clonal elimination by a form of apoptosis termed activation-induced cell death . This process of CD4 cell apoptosis is generally thought to reflect clonal suicide and to be independent of other cell types . Here we show that during the response to the superantigen Staphylococcal enterotoxin A, activated CD8 cells, but not activated CD4 cells, suppress the CD4 proliferative response . Suppression by CD8 cells reflects their ability to induce CD4 cell apoptosis via ligation of Fas . Moreover, although activated CD8 cells that express Fas ligand and Fas eliminate CD4 cells through a Fas-dependent mechanism, they are themselves resistant to Fas-dependent apoptosis . These findings indicate a fundamental difference between the two major T cell subsets with regard to sensitivity to Fas-dependent apoptosis, expression of Fas ligand, and mediation of suppressive activity following immunization with superantigen.

J Immunol, 1997 Dec 15, 159(12), 5936 - 45
Maternal immunization with a soluble TCR-Ig chimeric protein: long term, V beta-8 family-specific suppression of T cells by maternally transferred antibodies; McKeever U et al.; Maternal transfer of TCR clonotypic Ab protected young NOD mice against the adoptive transfer of diabetes by the BDC 2.5 T cell clone . The effect of maternal anti-TCR Vbeta-8 Ab on T cell development and function has now been investigated . SJL/J mice, which lack TCR Vbeta-8, were immunized with soluble, chimeric D10 TCR-IgG1 containing Vbeta-8.2 . The (SJL/J x AKR/J) F1 offspring of immunized female SJL/J mice were severely depleted of peripheral T cells bearing Vbeta-8 until 11 to 17 wk of age . The loss of Vbeta-8 expression did not appear to be due to modulation of cell surface TCR . Since the Vbeta-8+ T cell population was unperturbed in the (AKR/J x SJL/J) F1 offspring of D10 TCR-IgG1-immunized AKR/J mothers making D10 clonotypic Ab, the effect was immunologically specific . The deletion of Vbeta-8+ T cells had functional consequences . In the in vitro response to the superantigen, staphylococcal enterotoxin B, the usually observed participation of Vbeta-8.2+ T cells was largely suppressed, whereas the recruitment of Vbeta-3+ T cells remained unaltered . In control mice, T cell responses to the 134- to 146-residue peptide of conalbumin (pCA(134-146)) were biased toward use of Valpha-2/Vbeta-8.2 TCR . In D10 TCR-IgG1 maternally immunized (SJL x AKR/J) F1 mice, the T cell responses to pCA(134-146) were suppressed, and T cell lines derived from these in vitro were devoid of Vbeta-8.2 expression . With an increased understanding of TCR V gene usage in autoimmune diseases, similar strategies for the depletion of autoreactive T cells may become feasible in humans.

J Biol Chem, 1998 Mar 27, 273(13), 7657 - 67
Construction and binding kinetics of a soluble granulocyte-macrophage colony-stimulating factor receptor alpha-chain-Fc fusion protein; Monfardini C et al.; Granulocyte-macrophage colony-stimulating factor (GM-CSF) activity is mediated by a cellular receptor (GM-CSFR) that is comprised of an alpha-chain (GM-CSFRalpha), which specifically binds GM-CSF, and a beta-chain (betac), shared with the interleukin-3 and interleukin-5 receptors . GM-CSFRalpha exists in both a transmembrane (tmGM-CSFRalpha) and a soluble form (sGM-CSFRalpha) . We designed an sGM-CSFRalpha-Fc fusion protein to study GM-CSF interactions with the GM-CSFRalpha . The construct was prepared by fusing the coding region of the sGM-CSFRalpha with the CH2-CH3 regions of murine IgG2a . Purified sGM-CSFRalpha-Fc ran as a monomer of 60 kDa on reducing SDS-polyacrylamide gel electrophoresis but formed a trimer of 160-200 kDa under nonreducing conditions . The sGM-CSFRalpha-Fc bound specifically to GM-CSF as demonstrated by standard and competitive immunoassays, as well as by radioligand assay with 125I-GM-CSF . The sGM-CSFRalpha-Fc also inhibited GM-CSF-dependent cell growth and therein is a functional antagonist . Kinetics of sGM-CSFRalpha-Fc binding to GM-CSF were evaluated using an IAsys biosensor (Affinity Sensors, Paramus, NJ) with two assay systems . In the first, the sGM-CSFRalpha-Fc was bound to immobilized staphylococcal protein A on the biosensor surface, and binding kinetics of GM-CSF in solution were determined . This revealed a rapid koff of 2.43 x 10(-2)/s . A second set of experiments was performed with GM-CSF immobilized to the sensor surface and the sGM-CSFRalpha-Fc in solution . The dissociation rate constant (koff) for the sGM-CSFRalpha-Fc trimer from GM-CSF was 1.57 x 10(-3)/s, attributable to the higher avidity of binding in this assay . These data indicate rapid dissociation of GM-CSF from the sGM-CSFRalpha-Fc and suggest that in vivo, sGM-CSFRalpha may need to be present in the local environment of a responsive cell to exert its antagonist activity.

J Mol Biol, 1998 Mar 20, 277(1), 61 - 79
Crystal structure of microbial superantigen staphylococcal enterotoxin B at 1.5 A resolution: implications for superantigen recognition by MHC class II molecules and T-cell receptors; Papageorgiou AC et al.; Staphylococcal enterotoxin B is a member of a family of toxins known as superantigens that activate a large number of T-cells (up to 20%) by cross-linking MHC class II molecules with T-cell receptors in a Vbeta-restricted fashion . The crystal structure of staphylococcal enterotoxin B presented here has been determined at 1.5 A resolution, the highest resolution so far for a superantigen . The final model contains 1948 protein atoms and 177 water molecules and has excellent geometry with root-mean-square (rms) deviation of 0.007 A and 1.73 degrees in bond lengths and bond angles, respectively . The overall fold is similar to that of other microbial superantigens, but as it lacks the zinc-binding site found in other members of this family, such as staphylococcal enterotoxin A, C2 and D, this enterotoxin possesses only one MHC class II binding site . Comparison of the crystal structure of free SEB and in complex with an MHC class II molecule revealed no major changes in the MHC-binding site upon complex formation . However, a number of water molecules found in the free SEB may be displaced in the complex or contribute further to its stability . Detailed analysis of the TcR-binding site of SEB, SEA and SEC2 shows significant differences which may account for the ability of each superantigen to bind specific Vbeta sequences .

J Immunol, 1997 Dec 1, 159(11), 5293 - 300
Acquisition of granzyme B and Fas ligand proteins by human keratinocytes contributes to epidermal cell defense; Berthou C et al.; In vertebrate tissues, cell integrity is maintained by at least three mechanisms . During an immune response, injured cells are eliminated by cytotoxic lymphoid cells that produce perforin, granzyme B, and Fas ligand (FasL) . Second, epithelial cells can produce FasL as an immunosuppressive protein, probably to protect the tissue against immune-mediated damage . Third, locally secreted antimicrobial peptides can be operative in the protection of animal and human epithelia . In this work, as another contribution to local mechanisms of host defense, the ability of human epidermal keratinocytes to produce cytotoxic proteins was investigated . To address this question, freshly isolated human epidermal cells and keratinocytes grown in vitro were studied . Freshly isolated epidermal cells did not express the cytolytic proteins . In contrast, keratinocyte growth to confluence was associated with granzyme B, perforin, and FasL mRNA and protein synthesis . These proteins were secreted in the culture medium . Further analysis showed that they were identical with the ones used by cytotoxic lymphocytes . Their function was then investigated with a view to a potential role in epidermal cell integrity . The data showed that activated human keratinocytes were able to protect against invading pathogens through granzyme B expression . This was demonstrated by the ability of granzyme B to greatly decrease the bacterial growth of Staphylococcus epidermidis . In addition, keratinocytes expressing FasL were found to prevent immune epidermal cell damage . Apoptosis of Fas-sensitive T cells occurred during coculture with confluent epidermal keratinocytes and was largely reduced by the addition of a FasL inhibitor . The data favor keratinocyte involvement in the regulation of dermal inflammatory responses.

Surg Today, 1998, 28(3), 325 - 7
Infected atherosclerotic ulcer of the abdominal aorta as a cause of mycotic aneurysm treated by in-situ prosthetic graft reconstruction: report of a case; Moriyama Y et al.; A 68-year-old man with an infrarenal mycotic aneurysm underwent successful in-situ graft reconstruction using a woven Dacron graft . A tissue culture taken from the excised aortic wall revealed Staphylococcus epidermidis, and histological study subsequently showed penetrating atherosclerotic ulcers (PAU) involving all layers of the aortic wall and marked neutrophilic infiltration with abscess formation inside the ulcer . Atherosclerotic aortic lesions such as PAU are considered susceptible to bacterial infection, which may lead to the formation of an aneurysm after destruction of the vessel wall . Hence, elderly hypertensive patients, being at high risk for such aortic pathology, require careful studies performed to assess the aorta . The usefulness of computed tomographic (CT) scans to determine the presence of PAU or surrounding inflammation should be borne in mind even when a small mycotic aneurysm exists.

J Pediatr, 1998 Mar, 132(3 Pt 1), 535 - 7
Successful treatment of a staphylococcal endocarditis vegetation with tissue plasminogen activator; Fleming RE et al.; A 930 gm premature infant had Staphylococcal endocarditis with a tricuspid valvular vegetation that was unresponsive to antibiotics and not amenable to resection . Infusion of tissue plasminogen activator over a 3-day period completely lysed the vegetation . The infection cleared with continued antibiotics, and the infant recovered without sequelae.

Arch Pediatr, 1997 Dec, 4(12), 1204 - 6
{Postoperative mediastinitis due to methicillin resistant Staphylococcus epidermidis with low sensitivity to vancomycin}; Hulin S et al.; BACKGROUND: Vancomycin is the drug of choice for methicillin-resistant Staphylococcus . Antibiotherapy failure is rarely clinically related to Staphylococcus with vancomycin low susceptibility . CASE REPORT: A surgical cure of an aortic stenosis in a neonate was complicated by a Staphylococcus mediastinitis . After initiation of antibiotherapy with vancomycin and rifampin and surgical debridement, there was a rapid improvement . Few days later, failure of therapy was obvious . Despite continuous infusion of vancomycin, with a serum level of 29 mg/L, blood cultures were positive again to Staphylococcus . There was no endocarditis or inadequate surgical drainage . Susceptibility of the Staphylococcus was tested, looking for a tolerant strain . The vancomycin minimum bactericidal concentration was 30 mg/L (above usual value 2 to 8 mg/L), while the minimum inhibitory concentration was 3.75 mg/L . A higher dosage of vancomycin associated with fusidic acid was rapidly efficient, and total recovery was achieved . CONCLUSION: In case of failure of vancomycin therapy, despite correct serum levels, the susceptibility of the Staphylococcus strain has to be determined . A low susceptibility strain prescribes more prolonged combination of two antibiotics.

Int J Cancer, 1998 Apr 13, 76(2), 274 - 83
Repeated treatment with antibody-targeted superantigens strongly inhibits tumor growth; Rosendahl A et al.; Superantigens (SAg) are microbial proteins with the capacity to activate a large proportion of T cells . We have developed a novel approach for cancer immunotherapy by genetically fusing the SAg staphylococcal enterotoxin A (SEA) to a Fab-fragment of a tumor-specific antibody . Repeated exposure to SEA induces a state of unresponsiveness including cell deletion and functional hyporesponsiveness, i.e., anergy . In this study we have developed improved therapeutic schedules to allow repeated injections of Fab-SEA, limit development of immunological unresponsiveness and promote maximal anti-tumor response . Four daily injections of Fab-SEA to mice carrying B 16-C215 lung metastases resulted in 90-95% reduction in the number of metastases . However, the animals did retain a minimal residual tumor disease . The immune system was in a hyporesponsive state after 4 daily Fab-SEA injections, and further injections did not improve therapy . Two repeated cycles, each comprising 4 daily injections of Fab-SEA, significantly prolonged the survival and resulted in complete cure of a fraction of the animals . A rest period of 10 days between the cycles was required to mount an efficient secondary anti-tumor response . This secondary immune response was characterized by partial recovery of cytokine production i.e., interleukin-2, interferon-gamma and tumor necrosis factor-alpha . Strong CTL activity was detected in animals that had rested for 8 weeks between the 2 cycles . Interestingly, irrespective of the resting period, the CD4+ SEA-reactive T cells expanded in response to all 4 additional Fab-SEA injections both locally and in spleen . In contrast, only marginal expansion of CD8+ T cells was seen if restimulation was given within 1 month . Our data show that potent anti-tumor effector functions can be induced after repeated stimulation cycles with a SAg-monoclonal antibody fusion protein resulting in a CD4+ T cell-dependent cytokine release, prolonged survival and induction of complete cures.

Pediatr Infect Dis J, 1998 Mar, 17(3), 179 - 83
Selective use of vancomycin to prevent coagulase-negative staphylococcal nosocomial bacteremia in high risk very low birth weight infants; Baier RJ et al.; OBJECTIVE: To determine whether vancomycin added to parental nutrition (PN) fluids could prevent nosocomial infections in very low birth weight newborns and which infants would benefit most from prophylaxis . DESIGN: Double blind, randomized controlled study . SETTING AND STUDY POPULATION: Very low birth weight infants receiving PN in a tertiary neonatal intensive care unit . METHODS: Thirty-eight infants with and without central vascular catheters were randomized to receive no medication or 25 microg/ml vancomycin added to PN for the duration of the infant's PN requirements . RESULTS: The addition of 25 microg/ml vancomycin to PN prevented bacteremia in very low birth weight infants receiving PN . There was a significant reduction in the number of coagulase-negative staphylococcal (CONS) bacteremias (defined as isolation of the same organism from two positive blood cultures) during PN (5 vs . 0; P = 0.037) as well as the total number of bacteremias and fungemias (9 vs . 1; P = 0.036) . The total number of hospital days (108 +/- 13 vs . 76 +/- 6; P = 0.039) were reduced in infants receiving vancomycin . Infants with birth weights of < 1000 g who received corticosteroids for treatment of chronic lung disease benefitted most from treatment . No vancomycin-resistant strains of CONS or enterococci were detected during the study period . CONCLUSIONS: Prophylactic treatment with vancomycin effectively prevented CONS bacteremia under the conditions of the study . Its use was most effective in infants with birth weights of <1000 g.

Intensive Crit Care Nurs, 1997 Oct, 13(5), 308 - 9
Teicoplanin (Targocid, Hoechst Marion Roussel): a new glycopeptide antibiotic; MacConnachie AM; The glycopeptides are important antibiotics in the management of Staphylococcal infections . Teicoplanin, the latest member of the group, may offer some advantages over vancomycin, the workhorse drug, which has retained its importance in the presence of increasing major resistance to other antistaphylococcal agents.

Scand J Immunol, 1980, 11(3), 253 - 60
Determination of the immunoglobulin class of complement-dependent cytotoxic antibodies in serum of D23 hepatoma-bearing rats; Lando P et al.; The immunoglobulin class of the complement-dependent cytotoxic antibodies in serum from D23 hepatoma-bearing rats (D23 TBS) for D23 hepatoma cells was analysed . When studied by affinity chromatography with concanavalin A, protamine, or staphylococcal protein A conjugated to Sepharose, the cytotoxic activity bound to the former two but not protein A . The binding fractions were further characterized by column chromatography on Sepharose CL-4B . The cytotoxic activity was recovered exclusively in the high molecular weight fractions corresponding to human IgM . Monitoring with IgG- or IgM-specific rabbit antibodies indicated that these high molecular weight cytotoxic fractions contained both IgG and IgM . However, fractionation of D23 TBS at low pH suggested that cytotoxicity was due to IgM antibodies rather than to immune-complexed IgG antibodies . This was supported by the findings that rabbit antirat IgM antibodies inhibited the cytotoxicity of TBS completely when added at high dilutions.

J Surg Res, 1998 Jan, 74(1), 17 - 22
Bacterial products primarily mediate fibroblast inhibition in biomaterial infection; Henke PK et al.; PURPOSE: The stimulation of fibroblast growth is essential for the normal healing and tissue integration of biomaterials . The local elevation of proinflammatory mediators in infected perigraft fluid (PGF) may inhibit this growth . We sought to determine whether infected PGF inhibited fibroblast growth, and, if so, whether this was primarily dependent on the biomaterial, bacteria, or host . METHODS: In vivo Dacron or expandable polytetra-fluoroethylene (ePTFE) grafts, sterile or colonized with slime-producing (RP-62A, viable or formalin-killed) or nonslime-producing (RP-62NA) Staphylococcus epidermidis (1 x 10(7) CFU/cm2), were implanted in Swiss Webster mice, and the PGF was harvested at 7 and 28 days . Antibodies to tumor necrosis factor alpha, interleukin 1 alpha, interferon gamma (7 micrograms/day), and indomethacin (50 micrograms/day) were administered by microinfusion pumps for 7 days and the PGF was harvested . Inhibition of the proinflammatory mediators was confirmed by enzyme-linked immunosorbant assay . The nontreated, heat-treated, or trypsin-digested in vivo PGF was incubated with an in vitro {3H}thymidine murine fibroblast (ATCC CCL-12) proliferation assay . RESULTS: Fibroblast inhibition was significant at 7 and 28 days with infected PGF incubation compared with sterile and was not dependent on bacterial slime production or viability . Dacron sterile PGF did not significantly inhibit fibroblasts compared with control, whereas sterile ePTFE stimulated (P < 0.05) fibroblasts . Treatment of the PGF with proinflammatory cytokines, heat, and trypsin failed to reverse fibroblast inhibition in the infected state . CONCLUSION: Biomaterial infection is associated with fibroblast inhibition that is dependent primarily on bacterial products and not the host or biomaterial . Conservative intervention strategies for graft infection need to address the problem of poor healing as well as bacterial clearance.

Immunology, 1998 Jan, 93(1), 20 - 5
Synovial fibroblasts as target cells for staphylococcal enterotoxin-induced T-cell cytotoxicity; Kraft M et al.; Rheumatoid arthritis (RA) is a chronic autoimmune disease of unknown aetiology . Recently, superantigens have been implied in the pathogenesis of RA . Superantigens activate a large fraction of T cells leading to the production of cytokines and proliferation . In addition, superantigens direct cellular cytotoxicity towards major histocompatibility complex (MHC) class II-expressing cells . There is now increasing evidence that cytotoxic T cells may be involved in the pathogenesis of RA . In the inflamed synovia class II-positive synovial fibroblasts (SFC) are found . In the present study it was tested whether MHC class II-positive SFC serve as target cells for superantigen-induced cellular cytotoxicity . SFC were stimulated with interferon-gamma to express class II antigens, then they were cultivated in the presence of CD4-positive T cells with or without staphylococcal enterotoxins (SE) . Cytotoxicity of T cells was measured as release of lactate dehydrogenase from SFC . Specific cytotoxicity was only found in the presence of class II-positive SFC depending on the dose of SE . Maximum lysis was seen after 20 hr . T-cell cytotoxicity was inhibited by antibodies to MHC class II antigens . The data suggest that class II-positive SFC not only function as accessory cells for SE-mediated T-cell proliferation and interleukin-2 production but may also be the targets of superantigen-mediated cellular cytotoxicity.

Mol Microbiol, 1998 Mar, 27(5), 967 - 76
Identification of novel staphylococcal virulence genes by in vivo expression technology; Lowe AM et al.; We have applied in vivo expression technology (IVET) to the study of staphylococcal virulence . Using a promoter trap that relies on genetic recombination as a reporter of gene expression, we identified 45 staphylococcal genes that are induced during infection in a murine renal abscess model . Of these, only six were known previously; 11 others have homology to known non-staphylococcal genes . The known staphylococcal genes include agrA, part of a key locus regulating numerous virulence products, and a glycerol ester hydrolase, which may enhance staphylococcal survival in abscesses . We constructed 11 strains containing mutations in previously unknown ivi genes . Of these strains, seven were significantly attenuated in virulence compared with the wild-type parent . The mutagenized ivi genes may encode novel staphylococcal virulence factors.

J Infect Dis, 1998 Apr, 177(4), 1013 - 22
Differential immune responses to staphylococcal enterotoxin B mutations in a hydrophobic loop dominating the interface with major histocompatibility complex class II receptors; Woody MA et al.; Bacterial superantigens, such as staphylococcal enterotoxin B (SEB), can trigger acute pathologic effects in humans . A hydrophobic loop on the surface of SEB and other bacterial superantigens, centered around a conserved leucine (L45) residue, is essential for binding to class II major histocompatibility complex molecules . Single residue changes of wild type SEB, designated Q43P, F44P, or L45R, resulted in nonlethal proteins at a dose equivalent to 30 murine LD50 of SEB . Relative to SEB, the mutant proteins did not elevate serum concentrations of proinflammatory cytokines in mice and caused minimal proliferation of human lymphocytes . Anti-SEB titers of mice immunized with Q43P, F44P, L45R, or SEB were similar and protected 77%-100% of animals against a lethal SEB challenge . Levels of toxin-specific IgG1, IgG2a, IgG2b, and IgG3 in mice immunized with SEB, Q43P, or F44P were equivalent, but animals immunized with L45R had significantly elevated levels of IgG2a and IgG2b . Vaccines against staphylococcal superantigens should focus on this critical leucine residue.

J Infect Dis, 1998 Apr, 177(4), 905 - 13
gp41 envelope protein of human immunodeficiency virus induces interleukin (IL)-10 in monocytes, but not in B, T, or NK cells, leading to reduced IL-2 and interferon-gamma production; Barcova M et al.; The effect of extracellular domain of human immunodeficiency virus (HIV-1) transmembrane glycoprotein gp41 on interleukin (IL)-10, IL-2, interferon (IFN)-y, IL-4, and tumor necrosis factor-alpha production by human peripheral blood mononuclear cells (PBMC) was assessed by ELISA . Rapid gp41-induced increase of IL-10 production was detected in resting PBMC and isolated monocytes but not in B, T, or NK cells . Furthermore, gp41 also enhanced IL-10 production in staphylococcal enterotoxin B-stimulated PBMC, while synthesis of IL-2, IFN-gamma, and IL-4 in these cells was down-modulated . Kinetic studies revealed that increased IL-10 production preceded reduction of IL-2, indicating the possible IL-10 regulatory role in the gp41-induced down-modulation of this cytokine . Anti-IL-10 antibody reversed almost completely the gp41 inhibitory effect on IL-2 production . In this study, HIV-1 gp41 was a potent modulator of cytokine production by PBMC, in particular by increasing IL-10 secretion from normal monocytes/macrophages and consequently down-regulating IL-2 and IFN-gamma.

Proteins, 1998 Mar 1, 30(4), 381 - 7
A fusion protein between serum amyloid A and staphylococcal nuclease--synthesis, purification, and structural studies; Meeker AK et al.; We developed a recombinant DNA system to overexpress a fusion protein between the small, minimally soluble acute phase serum protein, serum amyloid A (SAA), and the bacterial enzyme staphylococcal nuclease (SN) . This fusion protein is very soluble and is immunoreactive to polyclonal anti-SAA antibodies . Tryptophan fluorescence shows smooth denaturation curves for the fusion protein in guanidinium HCl or potassium thiocyanate . Fluorescence also indicates that only a single tryptophan residue (of the four present) is accessible to iodide quenching and, presumably, is exposed on the surface of the fusion protein . Circular dichroism (CD) shows a significant signal indicating alpha-helix, which can be attributed to the SAA portion of the molecule; these are the first CD spectral data available for SAA . pH titration shows persistence of helix domains for the fusion protein at pH 3.0, in contrast to the denaturation of SN under the same conditions . (The entire fusion protein shows a random coil pattern below pH 3.0.) By exploiting the structural and solubility properties of SN, this fusion protein has provided the first structural data about SAA-the precursor of the amyloid deposits in secondary amyloidosis . This fusion protein should be useful for further physical and physiologic studies of SAA.

Zentralbl Bakteriol, 1998 Jan, 287(1-2), 135 - 45
Effect of surface modifications of intraocular lenses on the adherence of Staphylococcus epidermidis; Schmidt H et al.; The development of polymers with different surface properties and surface modifications of intraocular lenses (IOL) should reduce foreign body reactions after implantation by reducing the surface hydrophobicity of the lenses . It was examined how far such surface variations influenced the adhesiveness of bacteria . The most common organism isolated from cases of postoperative endophthalmitis is Staphylococcus epidermidis . For this reason, three strains of this species, the type strain ATCC 14990 and two clinical isolates (8687, 6579 I), with different hydrophobic surfaces, were studied . IOL made of PMMA, silicone, and a copolymer as well as PMMA lenses with modified surfaces (unpolished, polished, silanized, and heparinized) were used . Bacteria were radiolabelled with 3H-thymidine and the adherent bacteria were calculated per mm2 of lens surface . The three strains adhered better to the unpolished surface of silicone than to PMMA . Treatment of PMMA surface by polishing diminished the differences between the strains . An influence of hydrophobic interactions on the adherence of S . epidermidis ATCC 14990 was demonstrated . The adherence of this hydrophobic type strain was clearly reduced by heparinization of the PMMA surface . In contrast, the hydrophilic catheter isolate 6579 I adhered better to modified surfaces . This strain differed clearly in its PFGE pattern from both hydrophobic strains . Hydrophobic interactions play a role in the bacterial adherence to intraocular lenses in vitro and in vivo . Modifications of polymer surfaces, however, can result in rather different effects depending on the bacterial surface composition and properties.

Zentralbl Bakteriol, 1998 Jan, 287(1-2), 19 - 31
In vitro anti-staphylococcal activity of heparinized biomaterials bonded with combinations of rifampicin; Fallgren C et al.; Biomaterial implants in various human body tissues are highly susceptible to bacterial colonization . We report here on the coating of heparinized biomaterials with heparin binding extracellular matrix proteins giving special regard to the efficient adsorption and slow release of antibiotics . Heparin was partially degraded and the resulting fragments were covalently end-point attached to 0.5 cm long silicone biomaterial surface . Collagen type I was immobilized on the heparinized biomaterials and then cross-linked with acyl-azide or carbodiimide . Finally, the resulting biosurfaces were exposed to antibiotics, i.e . rifampicin in combination with cefuroxime, fusidic acid, ofloxacin or vancomycin, respectively . The antibiotic bonded biomaterials were evaluated for their anti-staphylococcal activity after elution in NaCl, serum or blood by measuring the zones of inhibition for S . epidermidis strain RP12 . Furthermore, we examined the in-vitro colonization resistance to S . epidermidis RP12 for these combinations of rifampicin-bonded biomaterials by an ATP bioluminescence assay . The ATP measurements showed that initially adherent bacteria were eradicated from the polymer surface, for at least 24 or 48 h (fusidic acid > cefuroxime > vancomycin > ofloxacin) . The anti-staphylococcal activity of rifampicin-fusidic acid bonded heparinized biomaterials seems of sufficient duration and efficacy to merit testing in an animal model.

Zentralbl Bakteriol, 1998 Jan, 287(1-2), 7 - 18
Quantitation of bacterial adhesion to polymer surfaces by bioluminescence; Stollenwerk M et al.; Quantitation of microbes adhering to a surface is commonly used in studies of microbial adhesion to different surfaces . We have quantified different staphylococcal strains adhering to polymer surfaces by measuring bacterial ATP (adenosine triphosphate) by bioluminescence . The method is sensitive, having a detection limit of 10(4) bacterial cells . Viable counting of bacterial cells may yield falsely low results due to the presence of "dormant" and adherent bacteria . By using bioluminescence, this can be avoided . Cells of different bacterial species and cells of strains of the same species were shown to differ significantly in their basal ATP content (8.7 x 10(-13) - 5.2 x 10(-22) MATP) . The size of adherent and planktonic bacteria decreased with time (0.7 micron-->0.3 micron, 20 days) . During incubation in nutrient-poor buffer ("starvation"), the ATP content of adherent bacteria decreased after 24-96 h whereas that of planktonic bacteria was stable over 20 days . The presence of human serum or plasma did not interfere significantly with the test results . Since the ATP concentration of bacterial strains of different species varies and is also influenced by the growth conditions of bacteria (solid or liquid culture medium), a species-specific standard curve has to be established for bacteria grown under the same culture conditions . We conclude that the method is a sensitive tool to quantify adherent bacteria during experiments lasting for less than 6 h and constitutes a valuable method to be used in conjunction with different microscopical techniques.

Int Immunol, 1998 Feb, 10(2), 175 - 83
Regulation of programmed cell death following T cell activation in vivo; Yang Y et al.; Activation of T cell hybridomas in vitro induces rapid Fas-Fas ligand (FasL)-mediated programmed cell death (apoptosis) . In contrast, T cells activated by antigen or superantigen in vivo undergo a population expansion and then decline due to Fas-FasL-mediated activation-induced apoptosis (AIA) . We asked how T cells activated by antigen in vivo proliferated before undergoing apoptosis . Two possibilities were analyzed: either (i) the apoptosis program was not 'turned on' or (ii) was 'blocked' during the period of cellular proliferation in vivo . Data presented in this manuscript support the second of these possibilities . CD4+ T cells activated in vivo were resistant to anti-fas-mediated apoptosis until 48 h following staphylococcal enterotoxin B (SEB) administration, despite the fact that activated proliferating T cells expressed high levels of Fas (CD95) antigen and many 'apoptosis genes' were induced within 24 h of SEB administration . The analysis of the expression patterns of 'apoptosis genes' during the T cell activation further suggested that temporal blockade of AIA may be due to the induction of apoptosis-preventing genes, such as bag-1.

Infect Immun, 1998 Apr, 66(4), 1579 - 87
Characterization of leptospiral outer membrane lipoprotein LipL36: downregulation associated with late-log-phase growth and mammalian infection; Haake DA et al.; We report the cloning of the gene encoding a 36-kDa leptospiral outer membrane lipoprotein, designated LipL36 . We obtained the N-terminal amino acid sequence of a staphylococcal V8 proteolytic-digest fragment in order to design an oligonucleotide probe . A Lambda-Zap II library containing EcoRI fragments of Leptospira kirschneri DNA was screened, and a 2.3-kb DNA fragment which contained the entire structural lipL36 gene was identified . Several lines of evidence indicate that LipL36 is lipid modified in a manner similar to that of LipL41, a leptospiral outer membrane lipoprotein we described in a previous study (E . S . Shang, T . A . Summers, and D . A . Haake, Infect . Immun . 64:2322-2330, 1996) . The deduced amino acid sequence of LipL36 would constitute a 364-amino-acid polypeptide with a 20-amino-acid signal peptide, followed by an L-X-Y-C lipoprotein signal peptidase cleavage site . LipL36 is solubilized by Triton X-114 extraction of L . kirschneri; phase separation results in partitioning of LipL36 exclusively into the hydrophobic, detergent phase . LipL36 is intrinsically labeled during incubation of L . kirschneri in media containing {3H}palmitate . Processing of LipL36 is inhibited by globomycin, a selective inhibitor of lipoprotein signal peptidase . After processing, LipL36 is exported to the outer membrane along with LipL41 and lipopolysaccharide . Unlike LipL41, there appears to be differential expression of LipL36 . In early-log-phase cultures, LipL36 is one of the most abundant L . kirschneri proteins . However, LipL36 levels drop considerably beginning in mid-log phase . LipL36 expression in vivo was evaluated by examining the humoral immune response to leptospiral antigens in the hamster model of leptospirosis . Hamsters surviving challenge with culture-adapted virulent L . kirschneri generate a strong antibody response to LipL36 . In contrast, sera from hamsters surviving challenge with host-adapted L . kirschneri do not recognize LipL36 . These findings suggest that LipL36 expression is downregulated during mammalian infection, providing a marker for studying the mechanisms by which pathogenic Leptospira species adapt to the host environment.

J Exp Med, 1998 Mar 16, 187(6), 823 - 33
A mutational analysis of the binding of staphylococcal enterotoxins B and C3 to the T cell receptor beta chain and major histocompatibility complex class II; Leder L et al.; The three-dimensional structure of the complex between a T cell receptor (TCR) beta chain (mouse Vbeta8.2Jbeta2.1Cbeta1) and the superantigen (SAG) staphylococcal enterotoxin C3 (SEC3) has been recently determined to 3.5 resolution . To evaluate the actual contribution of individual SAG residues to stabilizing the beta-SEC3 complex, as well as to investigate the relationship between the affinity of SAGs for TCR and MHC and their ability to activate T cells, we measured the binding of a set of SEC3 and staphylococcal enterotoxin B (SEB) mutants to soluble recombinant TCR beta chain and to the human MHC class II molecule HLA-DR1 . Affinities were determined by sedimentation equilibrium and/or surface plasmon detection, while mitogenic potency was assessed using T cells from rearrangement-deficient TCR transgenic mice . We show that there is a clear and simple relationship between the affinity of SAGs for the TCR and their biological activity: the tighter the binding of a particular mutant of SEC3 or SEB to the TCR beta chain, the greater its ability to stimulate T cells . We also find that there is an interplay between TCR-SAG and SAG-MHC interactions in determining mitogenic potency, such that a small increase in the affinity of a SAG for MHC can overcome a large decrease in the SAG's affinity for the TCR . Finally, we observe that those SEC3 residues that make the greatest energetic contribution to stabilizing the beta-SEC3 complex ("hot spot" residues) are strictly conserved among enterotoxins reactive with mouse Vbeta8.2, thereby providing a basis for understanding why SAGs having other residues at these positions show different Vbeta-binding specificities.

Microbiol Immunol, 1998, 42(1), 33 - 40
The inhibitory effect of Staphylococcus epidermidis slime on the phagocytosis of murine peritoneal macrophages is interferon-independent; Shiau AL et al.; The extracellular slime produced by Staphylococcus epidermidis has been shown to interfere with several human neutrophil functions in vitro, such as chemotaxis, degranulation and phagocytosis . Slime production has been suggested as a useful marker for clinically significant infections with coagulase-negative Staphylococcus . Since the main role of macrophages in defense mechanisms is phagocytosis, the effect of slime on the phagocytic activity of macrophages was investigated . The phagocytic activity of murine peritoneal macrophages treated with slime in vitro decreased in a dose-dependent fashion . A similar decrease was also observed in macrophages isolated from mice that had previously received intraperitoneal injection of slime . To investigate whether interferon also plays a role in this process, mice were treated with interferon or an interferon inducer, polyinosinic-polycytidylic acid (poly I:C), together with slime before macrophage isolation . The slime-suppressed phagocytic activity of macrophages was partially relieved by both agents, and the recovery effect of poly I:C in slime-suppressed phagocytosis of macrophages in vivo might be attributed to the increased interferon level in peritoneal fluid and sera . However, when slime was given to poly I:C-pretreated mice, the phagocytic activity remained suppressed . Thus, it appears that slime is able to suppress the phagocytic activity of macrophages regardless of the state of macrophage activation by poly I:C . The results suggest that the inhibition of phagocytosis by S . epidermidis slime may be independent from the activation of interferon.

J Neurosurg, 1998 Apr, 88(4), 769 - 72
Acute quadriplegia with delayed onset and rapid recovery . Case report; Latronico N et al.; The authors describe a patient with severe head injury and sepsis who became acutely quadriplegic 3 days postinjury because of a critical illness polyneuropathy (CIP) and critical illness myopathy (CIM), which resolved rapidly after treatment of the underlying infection . In only 3 days the patient developed septic shock together with flaccid quadriplegia and absent deep tendon reflexes with no clinical or radiological evidence of central nervous system deterioration . Neurophysiological studies showed an acute axonal sensorimotor polyneuropathy, whereas the clinical course strongly suggested a concurrent myopathy . A severe Staphylococcus epidermidis infection accompanied by bacteremia was treated and the patient recovered fully within a few days . Although the case described here is unique because of its very early onset and rapid resolution, CIP and CIM are frequent complications of sepsis and multiple organ failure . The authors suggest that severely head injured patients with sepsis should be evaluated for CIP and CIM when presenting with unexplained muscle weakness or paralysis.

Exp Nephrol, 1998 Jan-Feb, 6(1), 67 - 73
Tubular epithelial cells as accessory cells for superantigen-induced T cell activation; Schulz H et al.; In various inflammatory kidney diseases, tubular epithelial cells (TEC) express major histocompatibility complex class II antigens . To assess whether they might have the capacity to directly activate T cells, human TEC in culture were treated with gamma interferon to induce class II expression . TEC were then cocultivated with staphylococcus enterotoxin and cloned T cells or highly purified peripheral T cells . After 1-2 days, release of interleukin 2 and of gamma interferon was seen; after 3-5 days T cell proliferation occurred . The proliferation could be inhibited by antibodies to class II antigens or by antibodies to ICAM-1; the latter is also expressed on TEC in inflammatory processes and on TEC in culture as well . In conclusion, human TEC might function as accessory cells for T cell activation and might support T cell dependent immune response.

Semin Hematol, 1998 Jan, 35(1 Suppl 1), 36 - 51
Management of chronic immune thrombocytopenic purpura in children and adults; Blanchette V et al.; Chronic immune thrombocytopenic purpura (ITP) is characterized by macrophage destruction of platelets, primarily in the spleen, and occurs in both adults and children . Historically, splenectomy is often required, especially in adults with chronic ITP; however, several medical options have been used successfully prior to splenectomy or when this procedure has either failed or been refused by the patient . These include corticosteroids, intravenous immunoglobulin (IVIg), intravenous anti-D, danazol, vinca alkaloids, and other immunosuppressive agents such as azathioprine and cyclophosphamide . Platelet transfusions have been used in emergency situations . Several therapies that have been reported, but that are rarely used in chronic ITP, include cyclosporine A, interferon-alpha (IFN-alpha), plasma exchange, staphylococcal protein A immunoadsorption, combination chemotherapy, dapsone, ascorbic acid, and colchicine . Each of these approaches to treatment of chronic ITP are discussed.

Eur J Immunol, 1998 Feb, 28(2), 540 - 7
Expression of B220 on activated T cell blasts precedes apoptosis; Renno T et al.; Superantigens are bacterial or viral products that polyclonally activate T cells bearing certain TCR beta chain variable elements . For instance, Vbeta8+ T cells proliferate in response to staphylococcal enterotoxin B (SEB) in vivo and then undergo Fas- and/or TNF-mediated apoptosis . We have recently shown that apoptotic SEB-reactive T cells express the B cell marker B220 . Here we report the identification of a novel subset of CD4+ B220+ T cell blasts that are the precursors of these apoptotic cells in SEB-immunized mice . Moreover, we show that the CD4- CD8- B220+ T cells that accumulate in the lymphoid organs of Fas ligand-defective gld mice stably express a form of the B220 molecule which exhibits biochemical similarities to that expressed by activated wild-type T cells, but is distinct from that displayed on the surface of B cells . Surprisingly, we also find a population of CD4+ B220+ pre-apoptotic T cells in FasL-defective gld mice, arguing that these cells can be generated in a Fas-independent fashion . Collectively, our data support a general model whereby upon activation, T cells up-regulate B220 before undergoing apoptosis . When the apoptotic mechanisms are defective, T cells presumably down-regulate their coreceptor molecules but retain expression of B220 as they accumulate in lymphoid organs.

Cancer Immunol Immunother, 1998 Mar, 46(1), 7 - 13
Immunization of mice with melanoma cells transfected to secrete the superantigen, staphylococcal enterotoxin A; Shrayer DP et al.; Immunization of mice with a melanoma vaccine coupled with staphylococcal enterotoxin A (SEA) inhibits the growth of primary melanoma tumors in mice . We have now successfully transfected B16 cells with the sea gene and have immunized C57BL/6 mice subcutaneously once per week for 4 weeks prior to tumor challenge with vaccines of irradiated B16 cells or, 4 weeks following tumor challenge of naive mice with B16 cells, with irradiated B16 cells transfected with the sea gene . Primary tumor growth following both types of treatments was inhibited significantly . To characterize immune responses to these immunogens, we examined the production of antibodies to the B700 melanoma antigen, the stimulation of endogenous IL-2 production, the expression of CD4, CD8, Vbeta and CD25 T cell markers, and the induction of NK activity . At 4 weeks following immunization of mice, there was a significant increase (P<0.05) in levels of interleukin-2 production by splenocytes from mice immunized with SEA-secreting B16 cells or with the parental B16 cells, compared to controls . Levels of antibodies to the B700 melanoma antigen were also significantly higher in mice immunized with the SEA-secreting B16 cells, as was expression of CD4, CD8, CD25 and Vbeta T cell antigens, particularly CD4 . Natural killer cell activity (at various E:T ratios) was tenfold higher in splenocytes of mice immunized with SEA-secreting B 16 cells, and fivefold higher in mice immunized with the parental B16 cells, compared to controls . These data confirm the possibility of using irradiated murine melanoma cells transfected to secrete SEA in vaccines targeted at preventing the development and growth of melanoma.

Cornea, 1998 Mar, 17(2), 233 - 5
Recurrent endophthalmitis after cataract surgery with a scleral-tunnel incision; Ayyala RS et al.; PURPOSE: To present a case of recurrent postoperative endophthalmitis with a scleral-tunnel abscess and adjacent microbial keratitis . METHODS: A 76-year-old woman with microbial keratitis and recurrent endophthalmitis after cataract surgery was referred to a tertiary care center for further management . The medical chart of the patient was reviewed . RESULTS: The patient was seen on the eighth postoperative day with endophthalmitis that responded to medical treatment . Initial vitreous cultures were negative . The endophthalmitis recurred after the medical treatment was discontinued . She subsequently developed microbial keratitis at 1 o'clock adjacent to the limbus . Cultures from the site of corneal abscess and vitreous grew coagulase-negative Staphylococcus . Gonioscopy revealed the presence of a scleral abscess, which responded to subconjunctival injection of vancomycin and an intense and prolonged course of topical antibiotics . CONCLUSION: A scleral abscess should be suspected in a patient with endophthalmitis or microbial keratitis or both after a scleral-tunnel incision for cataract surgery.

Biochem Biophys Res Commun, 1998 Mar 17, 244(2), 556 - 60
A monoclonal antibody selected for probing the folding of staphylococcal nuclease and its N-terminal fragments; Cheng P et al.; Monoclonal antibody McAb2C9 against Staphylococcal nuclease (SNase R) and its N-terminal fragments was produced and characterized . It was observed that the intact enzyme SNase R and its seven fragments (SNR141, SNR135, SNR121, SNR110, SNR102, SNR79 and SNR52) differed in their interactions with McAb2C9 . However, the fragments with weak immunoreactivity, such as SNR141 and SNR110, increased ability reacting with McAb2C9 in their partially unfolded state . It suggests that the differences of immunoreactivity among the fragments are due to diverse extent of the exposure of the specific epitope and the conformation of the peptide fragment . The monoclonal antibody McAb2C9 could be a useful probe to investigate the mechanism of folding of SNase R and its N-terminal fragments.

FEMS Immunol Med Microbiol, 1998 Jan, 20(1), 69 - 78
All individual domains of staphylococcal protein A show Fab binding; Jansson B et al.; The interactions between the individual domains (E, D, A, B and C) of staphylococcal protein A (SPA) and Fc and Fab regions of human immunoglobulins were studied using real-time biospecific interaction analysis . An engineered domain Z, similar to fragment B but with a single glycine to alanine amino acid substitution, was also included in the study . The domains were expressed in Escherichia coli, affinity purified and immobilised onto sensor chip surfaces in a directed manner using a unique C-terminal cysteine residue engineered into the recombinant proteins . All domains bound to a recombinant human IgG1 Fc fragment with similar strength . For the first time, binding to human Fab was demonstrated for all native SPA domains, using both polyclonal F(ab')2 and a recombinant scFv fragment as reagents . Interestingly, the engineered Z domain showed a considerably lower affinity for Fab as compared to the native domains.

Alcohol Clin Exp Res, 1998 Feb, 22(1), 211 - 6
Regulation of monocyte interleukin-12 production by acute alcohol: a role for inhibition by interleukin-10; Girouard L et al.; Acute ethanol treatment results in decreased antigen presentation capacity (Th1-type immunity) and elevated interleukin IL-10 (Th2 cytokine) production in human monocytes . Monocytes can contribute to both Th1 (IL-12) and Th2 (IL-10) immune responses via production of IL-12 and IL-10, respectively . Thus, we tested the hypothesis that acute alcohol treatment might affect Th1/Th2 immune balance by altering monocyte production of IL-12 and IL-10 . Neither acute ethanol treatment alone (25 to 100 mM) nor its combination with a bacterial challenge Staphylococcal enterotoxin B (SEB) induced IL-12 production in isolated blood monocytes . In contrast, the same physiological alcohol concentrations increased monocyte IL-10 levels, suggesting that ethanol can induce a dysbalance of monocyte-derived mediator production at the expense of Th1 cytokines . However, we found that monocyte activation with interferon-gamma (IFN-gamma) can prevent the preferential IL-10 induction by ethanol . IFN-gamma (100 units/ml) inhibited monocyte IL-10 production whether induced by 1 microg/ml of lipopolysaccharide (p < 0.01), 1 microg/ml of SEB (p < 0.02), or a combination of bacterial stimulation + ethanol (lipopolysaccharide: p < 0.01) . Furthermore, decreased IL-10 was concomitant to an increase in IL-12 production in IFN-gamma-treated monocytes . Moreover, acute ethanol treatment augmented IL-12 production in IFN-gamma-treated monocytes in response to SEB stimulation (25 mM ethanol, p < 0.01; 100 mM ethanol, p < 0.01) . Experiments with anti-IL-10 neutralizing antibody show that ethanol may prevent monocyte IL-12 induction via IL-10 . These results suggest that inhibition of ethanol-induced IL-10 production by IFN-gamma treatment is permissive for IL-12 induction by alcohol stimulation in monocytes . Thus, our results imply that the presence or absence of IFN-gamma is critical in determining the effect of acute ethanol treatment on monocyte IL-12 versus IL-10 induction.

Anaesth Intensive Care, 1998 Feb, 26(1), 51 - 5
Microbial contamination of three-way taps on arterial lines; Widdowson WM et al.; Arterial lines with three-way taps are used to measure blood pressure and aspirate blood, and are a potential source of catheter-related sepsis . Swabs were taken daily from 118 three-way taps on 98 arterial lines in a general intensive care unit . Infusion lines were changed weekly but arterial cannulae were not changed routinely . An overall contamination rate of 24.6% was found with the predominant organism being coagulase negative staphylococcus . The three-way taps became increasingly contaminated with time but this was shown to be unrelated to the manipulation rates . Blood culture organisms in those showing contamination of the three-way taps showed no relationship to the bacteria causing the contamination.

J Hosp Infect, 1998 Jan, 38(1), 27 - 35
Evaluation of clinical and laboratory findings in leukaemic patients with blood cultures positive for Staphylococcus epidermidis; Lyytikainen O et al.; It is not certain whether clinical or laboratory findings help to distinguish true bacteraemia from contamination among acute leukaemic patients with one or more blood cultures positive for Staphylococcus epidermidis . We studied 31 patients treated at the Haematological Unit between 1 January, 1992 and 30 June, 1995 who were considered to have 'true bacteraemia', indicated by at least two positive blood cultures, and 20 considered to have probable 'contamination', indicated by a single positive culture . Fever at onset of positive blood culture, level of C-reactive protein (CRP) one day after the first positive blood culture and mortality did not differ between the groups . However, the median increase in CRP over 24 h from the first positive blood culture was significantly higher in true bacteraemias than among contaminants (median 35 mg/L vs 5 mg/L, P < 0.05) . Patients with true bacteraemia were more likely than those with contaminants to have central catheters in situ (95 vs 75%, P < 0.05) and previous oral antibiotic prophylaxis (29 vs 5%, P < 0.05) . Also clinical signs of catheter infection (30 vs 7%) were more common in true bacteraemias . In conclusion, central catheterization, antibiotic prophylaxis and clinical signs of catheter infection increase the likelihood of true bacteraemia; however, these factors have limited clinical utility in differentiation of true bacteraemia from contamination . Daily monitoring of serum CRP levels may help in the clinical decision-making.

Can Fam Physician, 1998 Feb, 44, 317 - 22
Bacteremia in nursing home patients . Prevalence among patients presenting to an emergency department; Sinclair D et al.; OBJECTIVE: To measure the prevalence of bacteremia and any correlation between signs and symptoms, risk factors, and laboratory data in elderly patients . DESIGN: Prospective analysis . All patients were contacted by the study nurse at 48 hours and 7 days after study entry . SETTING: Adult tertiary care hospital with an emergency department managing 48,000 visits yearly in a metropolitan area of 250,000 . PARTICIPANTS: Members of the study population referred to the emergency department for medical or surgical problems . Of 113 nursing home patients, blood culture results were available for 111 . MAIN OUTCOME MEASURES: Blood cultures were obtained by standard protocol . Demographic and medical information was collected from the medical record . Three groups of patients were compared with respect to symptoms, risk factors, laboratory data, and outcome . RESULTS: Group 1 (n = 86) had two sets of negative blood cultures . Group 2 (n = 10) had true-positive cultures . Group 3 (n = 15) had false-positive cultures of Staphylococcus epidermidis . The prevalence of bacteremia was 9.8% in the study population . No risk factors were predictive of bacteremia . Great variation in signs and symptoms were noted in all three groups, none correlating with bacteremia . Although seven of the 10 patients with positive cultures were febrile, this association did not reach statistical significance . All groups had high admission (> 50%) and mortality (20% to 37%) rates . CONCLUSIONS: The prevalence of bacteremia in the nursing home population presenting to the emergency department was 9.8% . The symptoms and signs of bacteremia in this population were variable and nonspecific . The high rate of false-positive cultures in this setting is of concern.

J Mol Biol, 1998 Feb 20, 276(2), 325 - 30
Staphylococcal alpha-hemolysin can form hexamers in phospholipid bilayers; Czajkowsky DM et al.; Atomic force microscopy (AFM) was used to study the structure of the staphylococcal alpha-hemolysin (alpha HL) oligomer formed in supported phospholipid bilayers . In contrast to the recent X-ray crystallographic demonstration of a heptameric stoichiometry for the oligomer formed in deoxycholate (DOC) micelles, the high-resolution unprocessed AFM images unequivocally revealed a hexamer in these phospholipid bilayers . Independent support of this hexameric stoichiometry was obtained from the measurements of the lattice constant in the AFM images and from gel electrophoresis . Therefore, alpha HL can form two different, energetically stable oligomers, which differ in at least stoichiometry but perhaps subunit structure as well . Furthermore, stable, incomplete oligomers were observed in the AFM images, which may be of relevance to the mechanism by which alpha HL damages the cell.

Tissue Antigens, 1998 Feb, 51(2), 164 - 73
Staphylococcus enterotoxin A modulates interleukin 15-induced signaling and mitogenesis in human T cells; Gerwien J et al.; T cells expressing the appropriate T-cell receptor Vbeta chain proliferate in response to Staphylococcus enterotoxin A (SEA) pulsed antigen-presenting cells (APC), whereas other T cells do not (SEA "non-responders") . Activated human T cells express MHC class II molecules that are high affinity receptors for SEA . Here we show that, in the absence of APC, SEA induces a profound inhibition of IL-15-driven proliferation in MHC class II+, human SEA-"responder" T-cell lines . In contrast, proliferation induced by phorbol esther (PMA) was enhanced by SEA . The inhibitory effect on cytokine-mediated mitogenesis correlates with an inhibition of IL-2Rbeta expression and ligand-induced tyrosine phosphorylation of IL-2R . Cyclosporin A (CyA), an inhibitor of the protein phosphatase (PP2B) calcineurin, strongly inhibits the SEA-induced modulations of cytokine receptor expression . Moreover, CyA inhibits both the anti-mitogenic effect of SEA on cytokine-induced proliferation and the pro-mitogenic effect of PMA . In contrast, inhibitors of PP1, PP2A, protein kinase C (PKC), phosphatidyl-inositol-3-kinase (PI-3K) and mammalian target of rapamycin (mTOR) are unable to inhibit the effects of SEA . In a SEA "non-responder" T-cell clone obtained from the affected skin of a patient with psoriasis vulgaris, SEA does not inhibit IL-2Rbeta expression and IL-15-driven proliferation . On the contrary, SEA enhances IL-15- and IL-2-induced proliferation via a CyA-sensitive pathway in this T-cell clone . In conclusion, the present data show that (i) SEA selectively inhibits IL-15- (but not PMA-) mediated proliferation in SEA "responder" T cells, (ii) SEA enhances cytokine-driven growth in psoriasis T cells with a "non-responder" phenotype, and (iii) crosstalk between SEA receptors and the IL-15R (and IL-2R) pathway is mediated via a PP2B-dependent and PP1/PP2A-, PKC-, PI-3 kinase- and mTOR-independent pathway in human T-cell lines.

Br J Haematol, 1998 Mar, 100(3), 490 - 500
Stimulation of leukaemic cells from adult T-cell leukaemia patients with bacterial superantigens; Ogata M et al.; Bacterial superantigens stimulate T cells in a manner that is restricted to the Vbeta of the T-cell receptor . We examined the ability of adult T-cell leukaemia (ATL) cells to respond to these superantigens . Mononuclear cells from 10 patients were cultured with staphylococcal enterotoxin A (SEA), staphylococcal enterotoxin B (SEB) or toxic shock syndrome toxin-1 (TSST-1), and their response was determined by MTT assay and 3H-thymidine incorporation assay . Cells from six patients showed a specific response to a single superantigen . In two cases the cells responded to TSST-1 and bore Vbeta2, the known target of TSST-1 . In three cases the cells responded to SEA with one bearing Vbeta9, a target of SEA, and one bearing Vbeta16 . In one case the cells responded to SEB . Most of the cells which proliferated in response to superantigens were determined genetically to be leukaemic . The response to TSST-1 was inhibited by anti-Vbeta2 antibody . The responding cells showed a strongly enhancement expression of interleukin-2 receptor . These findings indicate that leukaemic cells from a proportion of ATL patients have an ability to respond to T-cell receptor-dependent superantigens . This suggests that bacterial infection in such patients may contribute to the expansion of ATL cells.

Probl Tuberk, 1997, (6), 61 - 2
{Systematic approach to detection of anti-idiotypic antibodies to antimycobacterial idiotype in man and animals}; Avdienko VG et al.; ELISA in the sandwich modification was developed to detect anti-idiotypic antibodies to the antituberculosis idiotype of monoclonal antibodies (MAb) S4C1G4 (anti 15-18 kDa H37Rv) . Sera immunoglobulins were determined by Staphylococcus protein A capture . F{ab+}2, the MAb fragments S4C1G4 (IgG2a, kappa) and 1.3.3.B5 (IgG2a, kappa), were used as a negative control, which were not recognized mycobacterial antigens and human immunoglobulins . The anti-idiotypic antibody titers were determined in the murine hyperimmune sera during the period of immunization with the MSAb SaC1G4 . The method detected anti-idiotypic antibodies in the sera of infected mice and in patients with pulmonary tuberculosis . It is suggested that this method is applicable while using diagnostic tools.

J Biol Chem, 1998 Feb 20, 273(8), 4323 - 8
Directing sequence-specific proteolysis to new targets . The influence of loop size and target sequence on selective proteolysis by tissue-type plasminogen activator and urokinase-type plasminogen activator; Coombs GS et al.; We have previously used substrate phage display to identify peptide sequences that are efficiently and selectively cleaved by tissue-type plasminogen activator (t-PA) or urokinase-type plasminogen activator (u-PA) . We demonstrate that this information can be used to direct selective proteolysis to new protein targets . Sequences that were labile to selective cleavage by t-PA or u-PA when in the context of a peptide were introduced into the 43-52 (or Omega) loop of staphylococcal nuclease . Both t-PA and u-PA hydrolyze the engineered proteins at the inserted target sequences, and Km values for protein cleavage were reduced up to 200-fold relative to values for cleavage of analogous sequences within 15 residue peptides . Variation of loop size surrounding a target sequence affects the efficiency of t-PA approximately 5-fold more strongly than that of trypsin, suggesting that cleavage by t-PA is more dependent on target site mobility . Cleavage of proteins by t-PA and u-PA is sequence selective . u-PA is 47-fold more active than t-PA for cleavage of a sequence known to be u-PA selective within small peptide substrates, whereas t-PA is 230-fold more active toward a t-PA-selective sequence.

Int J Tissue React, 1997, 19(3-4), 135 - 40
Cellular mechanisms of cold stress-related immunosuppression and the action of interleukin 1; Rybakina EG et al.; Recent studies of the immunomodulatory cytokines, especially interleukin 1 (IL-1), as the mediators of immunoneuroendocrine links have suggested their important physiological role in the development of host resistance during stress reaction . The present study examined the effects of cold stress (-20 degrees C, 20 min) in rats on the value of humoral immune response, lymphocyte activating factor (LAF) production by peritoneal macrophages, as well as IL-1 beta action on lymphocyte proliferation, and the level of general antibody titers . Cold stress exposure led to a considerable elevation of corticosterone (Cs) in rat blood serum and to pronounced suppression of humoral immune response . This kind of experimental stress induced peritoneal macrophages in rats to release LAF without additional stimulation within 24 h after termination of cooling . At the same time LAF production by macrophages after their additional stimulation by LPS in vivo and heat-killed staphylococcus in vitro was decreased during cold stress reaction . Cold stress exposure suppressed the ability of peripheral blood lymphocytes to proliferate in response to concomitant action of IL-1 beta . Immunoprotective effect of rat recombinant IL-1 beta administrated i.p . to rats before cooling was revealed . It is suggested that IL-1 beta modulates immunological and neuroendocrine responses to stress and plays a critical physiological role in realization of stress reaction.

Br J Ophthalmol, 1997 Nov, 81(11), 953 - 5
Peroperative microbial contamination of anterior chamber aspirates during extracapsular cataract extraction and phacoemulsification; Beigi B et al.; BACKGROUND: The normal conjunctival flora is one of the main sources of intraocular contamination during cataract surgery . The theory that the positive anterior chamber (AC) pressure during phacoemulsification (phaco), and the smaller wound utilised, might reduce the rate of contamination was studied . METHODS: The peroperative AC aspirates of 210 consecutive patients undergoing cataract surgery were assessed . In group 1, 100 patients underwent a standard extracapsular cataract extraction (ECCE) . In group 2, 110 patients underwent phacoemulsification of the crystal-line lens through a scleral tunnel . AC aspirates from the Simcoe irrigation/aspiration cannula (group 1) and phaco probe (group 2) were collected and microbiological studies performed after direct and enrichment cultures . RESULTS: There were 29 (29%) positives in the ECCE group compared with 22 (20%) positive cultures from AC aspirates in the phaco group . Coagulase negative staphylococcus (CNS) was the commonest contaminant in both groups . CONCLUSION: Although there was a higher rate of AC contamination during ECCE, the difference was not statistically significant (p > 0.10, chi 2 = 2.31).

J Immunol, 1998 Mar 1, 160(5), 2107 - 14
Structural dichotomy of staphylococcal enterotoxin C superantigens leading to MHC class II-independent activation of T lymphocytes; Lamphear JG et al.; We have recently characterized an MHC class II-deficient human cell line, SW480, that supports the proliferation of purified human T cells in the presence of the staphylococcal enterotoxin and superantigen SEC1, but not the closely related isotypes SEC2 or SEC3 . We now investigate the structural basis of this dichotomy and explore possible mechanisms that may account for it . Differences in activity between SEC1 and SEC2 were not attributable to differences in biochemical modification, to differences in Vbeta specificity, or to the potential to induce anergy . SEC2 inhibited SEC1-mediated T cell activation in the presence of SW480 cells, suggesting that SEC2 could compete with SEC1 for binding to the TCR but was unable to productively signal through the TCR . Utilizing a panel of hybrid enterotoxins we identified specific amino acids near the NH2-terminus of SEC1 that abrogated MHC class II-independent T cell activation, yet did not alter potency in the presence of class II+ APC . These residues mapped to the putative TCR binding domain of SEC1, and suggest that subtle differences in TCR binding affinity or the topology of the SEC1-TCR interaction can compensate for the lack of MHC class II and hence promote T cell proliferation.

J Immunol, 1998 Mar 1, 160(5), 2072 - 9
Activation of human T cells with superantigen (staphylococcal enterotoxin B) and CD28 confers resistance to apoptosis via CD95; McLeod JD et al.; Ag recognition is an essential component for an effective T cell response . However, T cell activation is also subject to additional regulation by accessory molecules . CD28 provides essential costimulatory signals that allow T cells to proliferate, whereas molecules such as CTLA-4 and CD95 (Fas) appear to be negative regulators . Currently, which outcome predominates under conditions of antigenic challenge is poorly understood . In particular it has been suggested that one consequence of antigenic activation of T cells is the up-regulation of both CD95 and CD95 ligand, thereby exposing activated T cells to apoptotic death . We have investigated this possibility in normal human peripheral blood T cells triggered by the superantigen SEB either in the presence of endogenous APCs or transfectants expressing DR4 and CD80 . In either case, we find that such activation does not expose the majority of T cells to anti-CD95-induced apoptosis as detected by annexin V externalization and DNA fragmentation . Furthermore, by phenotypically identifying, by flow cytometry, those cells that received both antigenic and costimulatory signals from those cells that did not, we observed that CD95-induced apoptosis was not seen in activated T cells receiving Ag and costimulatory signals via CD28 . However, while not all T cells were stimulated by superantigen, CD95 expression was found to be homogeneously up-regulated, suggesting a mechanism whereby bystander cells might be made susceptible to CD95-induced death . We conclude that antigenic activation of T cells via the TCR and CD28 engagement provides protection from CD95-induced apoptosis.

J Immunol, 1998 Mar 1, 160(5), 2059 - 64
IFN-gamma is critical for long-term allograft survival induced by blocking the CD28 and CD40 ligand T cell costimulation pathways; Konieczny BT et al.; It is postulated that IFN-gamma production hinders long-term acceptance of transplanted organs . To test this hypothesis, we compared survival of skin and heart allografts in wild-type (IFN-gamma+/+) mice to that in IFN-gamma gene knockout (IFN-gamma-/-) mice . We found that perioperative blockade of the CD28 and/or CD40 ligand T cell costimulation pathways induces long-term skin and heart allograft survival in IFN-gamma+/+ recipients but fails to do so in IFN-gamma-/- mice or in wild-type mice treated with IFN-gamma-neutralizing Ab at the time of transplantation . In vitro studies showed that endogenously produced IFN-gamma down-regulates T cell proliferation and CTL generation in MLCs . These actions of IFN-gamma were not mediated by TNF-alpha production or Fas-Fas ligand interactions . In vivo studies revealed exaggerated expansion and, subsequently, impaired deletion of superantigen-reactive T lymphocytes in IFN-gamma-/- mice injected with staphylococcal enterotoxin B . Taken together, our findings indicate that IFN-gamma does not hinder but instead facilitates induction of long-term allograft survival possibly by limiting expansion of activated T cells.

Microbiology, 1998 Feb, 144 ( Pt 2), 509 - 17
Physical and genetic map of the genome of Staphylococcus carnosus TM300; Wagner E et al.; A genome map of Staphylococcus carnosus TM300, an important micro-organism in the food industry and long used as a starter culture, was constructed by pulsed-field gel electrophoresis of DNA fragments obtained after digestion with NotI, SfiI and ApaI . The size of the chromosome was estimated to be 2590 kb . The fragments were assembled into a physical map using a combination of complementary methods including multiple and partial digests of genomic DNA, hybridization with homologous gene probes, and cross-Southern hybridization . Fifteen genes or gene clusters were positioned on the physical map by Southern hybridization analysis . The map provides a basis for further analysis of the S . carnosus chromosome.

J Chemother, 1997 Dec, 9(6), 420 - 6
Do vancomycin serum levels predict failures of vancomycin therapy or nephrotoxicity in cancer patients?
Kralovicova K, Spanik S, Halko J, Netriova J, Studena-Mrazova M, Novotny J, Grausova S, Koren P, Krupova I, Demitrovicova A, Kukuckova E, Krcmery V Jr.
The purpose of this study was to determine if patients with high vancomycin (VAN) serum levels experience more toxicity than underdosed patients with lower (VAN) levels, and whether low VAN serum levels cause therapeutic failures in patients with gram-positive bacteremia . In 198 cancer patients trough and peak serum levels of VAN were measured . Acute toxicity (Red Man syndrome) appeared in 3 patients (1.5%) . Patients previously or currently treated with other nephrotoxic compounds (134 patients) presented the same incidence of nephrotoxicity as those receiving VAN for the first time in monotherapy (64 patients) . VAN did not increase the toxicity when patients were dosed simultaneously or previously with aminoglycosides or amphotericin B . Our second observation, when studying serum levels in our 198 patients was that high VAN trough serum levels (trough > 15 microg/mL) were associated with significantly more nephrotoxicity (33.3% vs . 11.1%, P < 0.03) than low levels in the subgroups of either pretreated patients or unpretreated with other nephrotoxic drugs . None of 198 patients who had trough levels below 15 microg/mL had peak levels exceeding 40 microg/mL . This suggests that only serum monitoring of trough levels may predict nephrotoxicity . A case control study was conducted to compare a group of 22 VAN failures with 22 successfully treated patients matched in underlying disease and neutropenia who were treated in the same period, under the same antibiotic policy, at the same cancer center, for gram-positive bacteremia . Persisting, enterococcal, or mixed enterococcal plus staphylococcal bacteremia were the only statistically significant risk factors which predicted therapy failure in cancer patients . Neither peak nor trough VAN serum levels predicted failure or cure of gram-positive bacteremia in cancer patients.

Microbiol Immunol, 1997, 41(12), 933 - 8
Molecular typing of Staphylococcus epidermidis and other CNS with repetitive element sequence-based PCR; Hu X et al.; The chromosomal distribution of the repetitive DNA sequence found in Mycoplasma pneumoniae (REP-MP2) provides an ideal target for detecting DNA fragment patterns specific to individual Staphylococcus epidermidis and S . haemolyticus strains . A REP-MP2 sequence-based PCR (rep-PCR) was developed and applied to CNS isolates . We identified a 450 bp genomic DNA fragment which was common and specific to S . epidermidis isolates and not found in other CNS . In addition, S . epidermidis isolates showed several bands that could be grouped into 14 different fragment patterns . Similarly, S . haemolyticus isolates were classified into 10 groups . Significant correlations between the typing patterns of S . epidermidis and resistance to oxacillin (P< 0.05), gentamicin (P< 0.01), erythromycin (P< 0.02), and sulfamethoxazole-trimethoprim (P< 0.001) were found . The rep-PCR method is a rapid and reproducible discriminatory means for molecular typing of S . epidermidis and other CNS.

Lik Sprava, 1997 Sep-Oct, (5), 153 - 4
{The correction of immunity in adolescents and children with focal staphylococcal diseases}; Marushko IuV; The aim of the work done was to improve treatment options for focal Staphylococcus-induced diseases in adolescents and children with the aid of adsorbed staphylococcal anatoxin (ASA) concurrently with low-intensive EHF therapy . Overall fifty patients aged 3 to 17 years with Staphylococcus infection in tonsils, nose, ears were kept under medical surveillance . ASA and EHF therapies were instituted according to the developed schemes of such therapies . Positive dynamics was shown of clinical picture and parameters characterizing humoral and cell-mediated immunity . There were no unfavourable side-effects . The proposed mode of treatment can, we believe, be widely used in a clinical setting.

J Am Vet Med Assoc, 1998 Feb 15, 212(4), 534 - 8
Microbial flora and antimicrobial susceptibility patterns of isolated pathogens from the horizontal ear canal and middle ear in dogs with otitis media; Cole LK et al.; OBJECTIVE: To compare microbial flora and antimicrobial susceptibility patterns of isolated pathogens from the horizontal ear canal and middle ear in dogs with otitis media . DESIGN: Prospective study . ANIMALS: 23 dogs with chronic bilateral otitis externa . PROCEDURES: Swab specimens of the horizontal ear canal and middle ear were obtained for cytologic analysis, bacterial culture, and antimicrobial susceptibility testing . Integrity of the tympanic membrane was observed . If the tympanic membrane was intact, myringotomy was performed to collect specimens . RESULTS: Otitis media was diagnosed in 38 of 46 (82.6%) ears evaluated . The tympanic membrane was intact in 71.1% of the ears with otitis media . The 3 most common organisms isolated from the horizontal ear canal and middle ear were Staphylococcus intermedius, yeast, and Pseudomonas spp . A difference in total isolates or susceptibility patterns between the horizontal ear canal and middle ear was found in 34 (89.5%) ears . Compared with results of bacterial culture, cytologic examination of swab specimens was not as effective for detection of rods and cocci from the middle ear . CLINICAL IMPLICATIONS: In dogs with chronic otitis externa, otitis media often exists even when there is an intact tympanic membrane . In our study, the same isolates were rarely found in the horizontal ear canal and middle ear . Therefore, to choose appropriate antimicrobial agents, in addition to cytologic examination, bacterial culture and susceptibility testing of swab specimens from the horizontal ear canal and middle ear should be performed.

AJR Am J Roentgenol, 1998 Mar, 170(3), 753 - 7
Prostatic abscess: diagnosis and treatment; Barozzi L et al.; OBJECTIVE: Prostatic abscesses are uncommon in clinical practice because early antibiotic therapy has reduced complications of prostatitis . Prostatic abscess mainly affects diabetic and immunosuppressed patients . The organisms most frequently involved are Escherichia coli and Staphylococcus, whereas gonococcus is rarely encountered . The results in eight men with prostatic abscess, five of whom were treated with sonographically guided percutaneous drainage, are reported . The diagnosis, clinically suspected in only three patients, was confirmed by transrectal sonography . All patients experienced complete abscess resolution . CONCLUSION: Transrectal sonography is the most reliable imaging method to diagnose prostatic abscess . Percutaneous transperineal or transrectal drainage is the first choice for therapy because of the lower risk of complication compared with surgery.

Am Surg, 1998 Feb, 64(2), 165 - 70
Central line-related sepsis in acute burn patients; Still JM et al.; A retrospective review of all 443 burn patients admitted during a 13-month period from October 1, 1992 to October 31, 1993, was completed . Of these, 8 were transferred and eliminated from the study . Twenty-two patients who were felt to be terminal on admission and did not have blood cultures were included in the demographic data but were excluded from subsequent statistical analysis . One hundred ten patients had central venous lines (CVLs) . Three patients with CVLs were transferred, thus leaving 107 patients with CVLs for statistical analysis . Additionally, 17 of the aforementioned terminal patients who had CVLs and 1 patient with a CVL who had documented sepsis before CVL insertion were excluded, leaving 89 patients with CVLs used in statistical analysis . Mean burn surface for those with central lines was 35.8 per cent, and for those without, 10.9 per cent . Sixty-four patients (59.8%) with a central line had inhalation injuries, as did 18 patients (5.5%) without . The number of lines per patient varied from 1 to 7 . Sixty-one patients had one line, 46 had more than one . The total number of central line days for the entire group was 1749 . The mean number of central line days per patient was 16.3 . The mean number of line days per catheter was 8.48 . The mortality rate for the 107 patients with a central line was 34 (32.7%) . Mortality for all patients was 41 (9.4%) . The incidence of sepsis increased with increasing number of central line days and increasing number of central line changes, but the effect of these two factors on the incidence of sepsis could not be studied separately, as they are highly correlated with each other . The most commonly recovered organisms were various types of Staphylococcus . Polymicrobial infections were common . There were 51 subclavian, 17 internal jugular, and 135 femoral catheters inserted . By logistic regression analysis, there was no statistically significant difference in the incidence of sepsis between upper- and lower-body CVL sites . Twenty-four patients (22.4%) with a CVL and one or more positive blood cultures were felt to have demonstrated sepsis . Some had more than one septic episode while lines were in place, reported as separate patients but not as separate septic episodes.

Immunology, 1997 Nov, 92(3), 321 - 7
Anti-alpha 4 integrin antibody induces apoptosis in murine thymocytes and staphylococcal enterotoxin B-activated lymph node T cells; Tchilian EZ et al.; We have shown that an antibody (9C10) to the alpha 4 integrin induces apoptosis in murine immature CD4+ CD8+ thymocytes and in activated (but not resting) mature lymph node T cells . In both cases, apoptosis is blocked by the highly selective protein kinase C (PKC) inhibitor Ro31.8425, suggesting that 9C10 induces signalling through the alpha 4 integrin resulting in PKC activation leading to apoptosis . Overall, our results indicate the potential role of the alpha 4 integrin-mediated interactions in apoptosis induction during T-cell development and following mature T-cell activation.

Eur J Immunol, 1998 Jan, 28(1), 70 - 9
Down-modulation of CD2 delays deletion of superantigen-responsive T cells; Fortner KA et al.; CD2 is a cell surface glycoprotein expressed on most T lymphocytes that is generally viewed as a cell adhesion molecule and, in this capacity, contributes to T cell receptor (TCR) signaling . CD2 has a relatively long cytoplasmic tail which associates with the src family tyrosine kinases, p56(lck) and p59(fyn), and could potentially signal directly . Down-modulation of CD2 on T cells has been shown to result in diminished proliferative capacity and interleukin (IL)-2 production . Furthermore, re-expression of CD2 can result in the restoration of these functions . This suggests that CD2 can influence the intensity of TCR signaling . As TCR signal intensity is pivotal to the induction of T cell apoptosis, we considered the hypothesis that the level of CD2 on the T cell surface may influence its propensity toward apoptosis . Using an anti-CD2 antibody, CD2 was down-modulated in vivo on mouse T lymphocytes without affecting the levels of surface CD3, TCR alphabeta, CD4 or CD8 . Deletion of superantigen-responsive T cells was delayed in mice with down-modulated CD2 following the administration of staphylococcal enterotoxin B (SEB) . This was paralleled by diminished apoptosis of SEB-responsive cells . The findings suggest a model whereby the level of CD2 expression influences the intensity of TCR signaling and the ability to undergo apoptosis.

Ann Thorac Surg, 1998 Feb, 65(2), 359 - 64
Operation for infective endocarditis: results after implantation of mechanical valves; Bauernschmitt R et al.; BACKGROUND: Operation for acute endocarditis during the active phase violates a basic surgical rule not to implant a foreign body into an infective process, resulting in a high operative mortality and the risk of early recurrent endocarditis . Several investigators analyzing risk factors for perioperative mortality and morbidity presented strategies for more favorable outcomes, but most studies suffer from the drawback of heterogeneous populations observed over a long period of time . METHODS: We present a prospective study on 138 patients operated on from March 1988 to March 1996 . Patients were only included if the activity of the infection was proved by positive culture of the valve leaflets or by histologic staining . During the observation period, indication for operation, surgical approach, and postoperative antibiotic therapy were standardized as much as possible . After radical debridement of all parts of infected tissue, valve replacement was carried out with mechanical prostheses . RESULTS: The early mortality was 11.5% overall . High New York Heart Association functional classification, advanced age, and staphylococcal disease were significant risk factors for early mortality . The site of infection, multiple valve involvement, and prosthetic valve endocarditis did not affect the outcome . Early recurrent endocarditis was recorded in only 3 patients of the entire series . CONCLUSIONS: In case of acute infective endocarditis, valve replacement with mechanical prostheses is a safe procedure, if radical operation and aggressive postoperative antibiotic therapy are performed . For further improvements of the results, earlier operation is advisable in patients with rapidly progressive cardiac deterioration and in most cases of staphylococcal endocarditis.

Biochemistry, 1998 Feb 10, 37(6), 1623 - 31
Urea denaturation of staphylococcal nuclease monitored by Fourier transform infrared spectroscopy; From NB et al.; Fourier transform infrared (FTIR) amide I spectroscopy has not been widely used as a method to study protein folding . Some thorough studies of thermal unfolding have been carried out; however, protein unfolding in the presence of the widely used denaturants guanidine hydrochloride and urea has only recently been reported . Guanidine hydrochloride and urea both absorb strongly in the amide I region, as does H2O . Here, we have used deuterated 13C-urea as the chemical denaturant and monitored the unfolding transition with deuterium-exchanged staphylococcal nuclease (SNase) in D2O . These conditions circumvent all subtraction difficulties as the absorption bands of D2O and denaturant are shifted out of the amide I' region {Fabian, H., and Manstch, H . H . (1995) Biochemistry 34, 13651-13655} . A very reproducible unfolding transition is obtained for SNase . 13C-Urea-induced unfolding of SNase was found not only to be comparable to previous FTIR thermal unfolding data but also to have a denatured-state spectrum similar to those of other thermally denatured proteins . The unfolding is approximately two-state . The infrared spectra in the denatured state show evidence of some residual beta-sheet structure as well as other band components not attributable to random structure.

Vet Immunol Immunopathol, 1997 Nov, 59(3-4), 205 - 12
Staphylococcal protein A binding to canine IgG and IgM; Scott MA et al.; Staphylococcal protein A (SpA) binds with high affinity to immunoglobulins from many species, making it a useful reagent for immunoassays and immunoglobulin purification procedures . However, its use is limited by poor reactivity with some immunoglobulin subclasses including human IgG3 and murine IgG1 . Reports of SpA's reactivity with canine immunoglobulins have been inconsistent . Because the most recent reports indicated a much greater reactivity of purified SpA with canine immunoglobulins than was suggested by our observations, we quantitatively reevaluated the binding of canine IgG and IgM to Cowan I strain SpA under the conditions of use in our laboratory . IgG and IgM were purified from pooled normal canine plasma by affinity chromatography with heavy chain specific polyclonal anti-IgG and anti-IgM antibodies . The purified IgG and IgM were assessed for SpA reactivity by affinity chromatography using a SpA-agarose column . The relative proportions of total chromatographed IgG or IgM in the flow-through (SpA-nonbindable) and eluate (SpA-bindable) fractions were determined by absorbance at 280 nm . The IgG and IgM in each immunoglobulin fraction were also nonspecifically adsorbed to microtitration plates and tested for reactivity with 125I-SpA using a solid phase immunoradiometric assay (IRMA) . Approximately 18% of the affinity purified canine IgG and 33% of the affinity purified canine IgM did not bind to the SpA affinity column and were also unreactive with 125I-SpA using the IRMA . A second approach using a different polyclonal antibody to canine IgG yielded similar results: about 21% of the purified IgG was unreactive with SpA . Incomplete reactivity of SpA with canine IgG and IgM limits the usefulness of SpA in canine immunologic procedures.

Biosens Bioelectron, 1997 Dec 30, 12(12), 1219 - 25
Development of a new kind of dual modulated QCM biosensor; Zhang C et al.; To distinguish the mass loading effect from the total frequency change is a problem in the application of a quartz crystal microbalance (QCM) biosensor in the liquid phase . Based on the characteristic damping theory, this paper proposes a new method of dual modulation to solve this problem . Using polyethyleneimine to immobilize anti-SE (staphylococcin enterotoxin) antibody (C2 type), a dual modulated QCM SE biosensor was developed and the experiment proved that it has little cross-reaction with B-type SE . The measuring curve of the sensor was also determined through experiment.

Clin Exp Immunol, 1998 Jan, 111(1), 12 - 9
Superantigen activation of CD4+ and CD8+T cells from HIV-infected subjects: role of costimulatory molecules and antigen-presenting cells (APC)
Vingerhoets J, Dohlsten M, Penne G, Colebunders R, Sansom D, Bosmans E, Kestens L, Vanham G.
T cell receptor (TCR) triggering via superantigens induces decreased proliferative responses and increased apoptosis in T cells from HIV-infected patients compared with controls . Our aim was to delineate the role of intrinsic T cell defects, of APC dysfunction and of cytokines and costimulatory signal dysregulation in the deficient responses of CD4+ and CD8+ T cells from HIV+ subjects to the superantigen Staphylococcus enterotoxin A (SEA) . Proliferation and IL-2R alpha up-regulation on SEA-stimulated CD4+ and CD8+ T cells in whole blood were reduced in HIV+ subjects with CD4 counts < 500, compared with controls . Neither addition of IL-2, IL-12 or phorbol myristate acetate (PMA) nor neutralization of endogenous IL-10, tumour necrosis factor-alpha (TNF-alpha), TNF-beta or transforming growth factor-beta (TGF-beta) could restore the decreased activation by SEA . Possible intrinsic T cell defects were studied by presenting SEA on HLA-DR-transfected Chinese hamster ovary (CHO) cells, co-expressing LFA3 and/or CD80, to purified T cells . In this system CD8+ T cells from most HIV+ patients were hyporesponsive with regard to IL-2 production, IL-2R alpha up-regulation and proliferation, whereas clearly reduced responses were only shown in CD4+ T cells from AIDS patients . Similarly, apoptosis was increased in CD8+ T cells from all patients, but only in CD4+ T cells from AIDS patients . During HIV infection, the responses to TCR triggering through SEA are deficient in both T cell subsets . The intrinsic defect appears earlier during disease progression in purified CD8+ T than in CD4+ T cells, it occurs in conjunction with both CD2 and CD28 costimulation, and it is correlated with increased levels of apoptosis.

J Hand Surg {Am}, 1997 Nov, 22(6), 1091 - 2
Acute hematogenous staphylococcal infections involving upper-extremity musculotendinous compartments: report of 2 cases; Gabuzda G et al.; Two unusual cases involving 2 patients with 3 hematogenously spread acute staphylococcal infections that occurred in musculotendinous compartments and required surgical debridement are reported . Infections that persist despite intravenous administration of antibiotics may indicate rare closed-space infections.

J Immunol, 1998 Feb 15, 160(4), 1841 - 9
Superantigen-activated T cells redirected by a bispecific antibody inhibit vesicular stomatitis virus replication in vitro and in vivo; Fernandez-Sesma A et al.; A bispecific Ab (BsAb) that binds the TCR on T cells and the G protein of the vesicular stomatitis virus (VSV) can redirect staphylococcal enterotoxin B (SEB)-activated T cells to kill VSV-infected cells and to inhibit VSV replication in vitro . Inhibition of virus replication in our system is dependent upon the specificity of the Ab for the viral protein . IFN-gamma does not play a very important role in this phenomenon, which is mainly mediated by the release of Pfp from CD8+ T cells . We have used a Stat1 knockout mouse model in which VSV infection is lethal . Infusion of staphylococcal enterotoxin-activated B T cells and bispecific Ab significantly slowed virus progression and prolonged the survival of VSV-infected Stat1 knockout mice in vivo.

Pediatr Infect Dis J, 1998 Jan, 17(1), 10 - 7
Intravenous lipid emulsions are the major determinant of coagulase-negative staphylococcal bacteremia in very low birth weight newborns; Avila-Figueroa C et al.; BACKGROUND: Intravenous lipid emulsions and the i.v . catheters through which they were administered were the major risk factors for nosocomial coagulase-negative staphylococcal (CONS) bacteremia among newborns in our neonatal intensive care units a decade ago . However, medical practice is changing, and these and other interventions may have different effects in the current setting . OBJECTIVES: We determined the independent risk factors for CONS bacteremia in current very low birth weight newborns after adjusting for severity of underlying illness . METHODS: We surveyed 590 consecutively admitted newborns with birth weights < 1500 g hospitalized in 2 neonatal intensive care units and conducted a case-control study in a sample of 74 cases of CONS bacteremia and 74 pairs of matched controls . Adjusted relative odds of bacteremia were estimated for a number of attributes and therapeutic interventions in 2 time intervals before CONS bacteremia: any time before bacteremia and the week before bacteremia . RESULTS: Using conditional logistic regression to adjust for indicators of severity of illness, two procedures were independently associated with subsequent risk of CONS bacteremia at any time during hospitalization: i.v . lipids, odds ratio (OR) = 9.4 {95% confidence interval (CI) 1.2 to 74.2}; and any surgical or percutaneously placed central venous catheter, OR = 2.0 (95% CI 1.1 to 3.9) . Considering only the week immediately preceding bacteremia, the independent risk factors were: mechanical ventilation, OR = 3.2 (95% CI 1.3 to 7.6); and short peripheral venous catheters, OR = 2.6 (95% CI 1.0 to 6.5) . CONCLUSIONS: During the last decade exposure to i.v . lipids any time during hospitalization has become an even more important risk factor for CONS bacteremia (OR = 9.4) . Of these bacteremias 85% are now attributable to lipid therapy . In contrast the relative importance of intravenous catheters as independent risk factors has declined . Mechanical ventilation in the week before bacteremia has emerged as a risk factor for bacteremia.

Microbios, 1997, 91(367), 89 - 95
Effect of a Staphylococcus epidermidis-extracted slime factor on human natural killer cell activity; Sultan N et al.; A slime factor produced by Staphylococcus epidermidis was a complex glycoconjugate extracted by the phenol extraction method . The potential stimulatory or inhibitory capacity of the phenol-extracted slime (PES) was tested on human natural killer cell cytotoxic activity . Various concentrations of the PES preparation were incubated with the effector cells 30 min before and during the assay period . The PES factor inhibited natural killer cell cytotoxic activity at a concentration of 250 micrograms/ml and at higher concentrations (p < 0.05) . The inhibition of natural killer cell cytotoxic activity may probably be related to the complex composition of the slime substance.

Scand J Immunol, 1998 Jan, 47(1), 43 - 7
Cytokine induction by Mycoplasma arthritidis-derived superantigen (MAS), but not by TSST-1 or SEC-3, is correlated to certain HLA-DR types; Alvarez-Ossorio L et al.; Superantigens bind to major histocompatibility complex (MHC) class II molecules on antigen presenting cells and T cells in a V beta-restricted manner . Both cell types are activated resulting in cytokine production . Although the MHC-II binding site for superantigens has been well described, little is known as to whether this binding complex has an influence on cytokine induction . In order to assess superantigen induced cytokine production and its correlation to HLA-DR types, the authors stimulated peripheral blood from 40 subjects with superantigens toxic shock syndrome toxin-1 (TSST-1), staphylococcal enterotoxin C-3 (SEC-3) and Mycoplasma arthritidis-derived superantigen (MAS), and measured cytokine levels thereafter . The HLA-DR type was determined in each subject . A statistical evaluation was carried out between the highest superantigen cytokine induction and the presence of certain HLA-DR types . Whereas MAS presented a statistical association between the highest cytokine production with HLA-DR4, DR7 and DR12, no such associations were observed for TSST-1 and SEC-3 . These results demonstrate that T cell stimulation, and consequently its cytokine production by MAS but not by TSST-1 and SEC-3, depends on the presenting HLA-DR type . Because the diverse HLA-DR specificities are given according to the variability of the beta chain of the HLA-DR molecule, the data suggest the participation of the human MHC-II beta chain in the MAS/MHC-II binding.

Onderstepoort J Vet Res, 1997 Sep, 64(3), 201 - 3
The binding potential of commercial antibody conjugates with sera of various small terrestrial mammals; Pretorius A et al.; Infectious diseases of wild animals are of increasing importance, both from an economic viewpoint and because several of these diseases are pathogenic to man . However, serosurveys to determine the circulation of infectious organisms in wildlife are complicated by the fact that antibodies to species-specific immunoglobulins are not available for use in serological assays such as enzyme-linked immunosorbent assays (ELISAs) or immunofluorescence assays . To determine the binding potential of four commercially available antibody conjugates with the sera of wild animals, sera from 27 species of small terrestrial mammals were allowed to react with alkaline phosphatase-labelled protein A, anti-rabbit IgG, anti-mouse IgG and anti-human IgG by by the use of an ELISA . It was found that sera from some species of the order Lagomorpha bound optimally to anti-rabbit IgG, while anti-mouse IgG could be used for most species of Rodentia . For all Carnivora, Insectivora, Macroscelidea, Hyracoidea and other Rodentia, staphylococcal protein A demonstrated optimal binding . None of the sera that was tested bound to anti-human IgG . These results demonstrate that commercial conjugates can be used in serological assays in which wild animal sera are used, and should be useful for future serosurveys to determine the circulation of infectious agents in small terrestrial mammals.

J Mol Biol, 1998 Jan 16, 275(2), 389 - 402
Structural characterization of the pressure-denatured state and unfolding/refolding kinetics of staphylococcal nuclease by synchrotron small-angle X-ray scattering and Fourier-transform infrared spectroscopy; Panick G et al.; The pressure-induced unfolding of wild-type staphylococcal nuclease (Snase WT) was studied using synchrotron X-ray small-angle scattering (SAXS) and Fourier-transform infrared (FT-IR) spectroscopy, which monitor changes in the tertiary and secondary structural properties of the protein upon pressurization . The experimental results reveal that application of high-pressure up to 3 kbar leads to an approximate twofold increase of the radius of gyration Rg of the native protein (Rg approximately 17 A) and a large broadening of the pair-distance-distribution function, indicating a transition from a globular to an ellipsoidal or extended chain structure . Analysis of the FT-IR amide I' spectral components reveals that the pressure-induced denaturation process sets in at 1.5 kbar at 25 degrees C and is accompanied by an increase in disordered and turn structures while the content of beta-sheets and alpha-helices drastically decreases . The pressure-induced denatured state above 3 kbar retains nonetheless some degree of beta-like secondary structure and the molecule cannot be described as a fully extended random coil . Temperature-induced denaturation involves a further unfolding of the protein molecule which is indicated by a larger Rg value and significantly lower fractional intensities of IR-bands associated with secondary-structure elements . In addition, we have carried out pressure-jump kinetics studies of the secondary-structural evolution and the degree of compactness in the folding/unfolding reactions of Snase . The effect of pressure on the kinetics arises from a larger positive activation volume for folding than for unfolding, and leads to a significant slowing down of the folding rate with increasing pressure . Moreover, the system becomes two-state under pressure . These properties make it ideal for probing multiple order parameters in order to compare the kinetics of changes in secondary structure by pressure-jump FT-IR and chain collapse by pressure-jump SAXS . After a pressure jump from 1 bar to 2.4 kbar at 20 degrees C, the radius of gyration increases in a first-order manner from 17 A to 22.4 A over a timescale of approximately 30 minutes . The increase in Rg value is caused by the formation of an extended (ellipsoidal) structure as indicated by the corresponding pair-distance-distribution function . Pressure-jump FT-IR studies reveal that the reversible first order changes in beta-sheet, alpha-helical and random structure occur on the same slow timescale as that observed for the scattering curves and for fluorescence . These studies indicate that the changes in secondary structure and chain compactness in the folding/unfolding reactions of Snase are probably dependent upon the same rate-limiting step as changes in tertiary structure.

Int Immunol, 1997 Dec, 9(12), 1825 - 36
Immediate responses of leukocytes, cytokines and glucocorticoid hormones in the blood circulation of monkeys following challenge with aerosolized staphylococcal enterotoxin B; Weng CF et al.; The immediate responses to aerosolized staphylococcal enterotoxin B (SEB) in respiratory toxic shock were studied in the circulation of rhesus monkeys with low antibody levels following immunization with SEB toxoid-containing microspheres . Both the surviving and dying monkeys had toxic shock syndrome 4-48 h after SEB challenge and all showed three distinctive patterns of immediate responses . The first pattern, characterized by the responses of all T cells, HLA-DRlo cells, monocytes, IL-2R+ cells, IFN-gamma, and augmented lymphocyte mitotic responses to lipopolysaccharide (LPS) and SEB in culture, was a rapid increase at 20 min followed by a quick decrease at 90 min to approximately the original levels . The second pattern, which included responses of HLA-DRhi cells, NK cells, adrenocorticotropic hormone (ACTH) and cortisol, was characterized by a moderate decrease at 20 min and a further decrease at 90 min . The third pattern, the inverse of the second pattern, including responses of polymorphonuclear leukocytes (PMN), concanavalin A (Con A) mitogenesis, IL-6 and IL-2, was a moderate increase at 20 min and a further increase at 90 min . Between the surviving and dying monkeys, the responses of T cells, HLA-DRhi cells, PMN and cortisol did not differ significantly, suggesting that they are the basic causes that initiated toxic shock . However, significant differences were seen in the responses of HLA-DRlo cells, monocytes, IL-2R+ cells and lymphocyte mitogenesis in culture at 20 min, and of Con A mitogenesis, NK cells, IL-2, IL-6 and ACTH at 90 min . These different responses are apparently the exacerbating causes of death of the monkeys . All together, the immediate responses seem to be caused by the combined effects of SEB superantigenicity, activation of NK cells and non-lymphoid cells, and depression of the neuroimmune defense system.

Cardiovasc Surg, 1997 Oct, 5(5), 486 - 9
Mechanism of late prosthetic vascular graft infection; Jones L et al.; This study was a retrospective analysis of 41 patients with late prosthetic graft infections (> 30 days after operation) from six hospitals in the south-west of England . The 41 patients had a median age of 66 years and generally accepted risk factors for infection were documented in 19 patients preoperatively . Thirteen patients had postoperative wound complications and three had early reoperation at the site of subsequent infection . The median time between index operation and symptoms of infection was 10 (range 1-224) months . Abscess (46%) was the most common presentation followed by false aneurysm (20%) and graft thrombosis (20%) . All patients had reoperations (median two per patient, range one to seven) . Seven (17%) patients died and 10 (24%) required a major amputation . Bacteria were isolated from retrieved grafts in 23/41 patients (high virulence 14, low virulence nine) and the most frequent organism was coagulase-negative Staphylococcus epidermidis (nine patients) . In the majority of cases the aetiology of the late infections in this series was consistent with bacterial implantation at the index operation, but in four cases bacteraemia or intraperitoneal spread was more likely . Dental-type antibiotic prophylaxis would not have prevented any of the infections in this study . Aggressive treatment of recognized sources of infection in patients with vascular grafts is imperative.

Cardiovasc Res, 1997 Oct, 36(1), 86 - 91
Increased Ca2+ sensitivity of contractile elements via protein kinase C in alpha-toxin permeabilized SMA from young spontaneously hypertensive rats; Sasajima H et al.; OBJECTIVE: The purpose of the present investigation was to examine the Ca2+ sensitivity of the contractile elements via protein kinase C (PKC) in superior mesenteric artery (SMA) from young (5-6 weeks old) spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto rats (WKY) . METHODS: Staphylococcal aureus alpha-toxin, which produces pores in the plasma membrane too small to allow passage of proteins such as PKC, was used to investigate the signal transduction system in vascular smooth muscle cells . We investigated the Ca2+ sensitivity of the contractile apparatus via PKC in intact and alpha-toxin skinned SMA from young SHR and WKY . RESULTS: In intact SMA, high K+ responses were not different between SHR and WKY . However, phorbol 12,13-dibutyrate (PDBu, a PKC activator) augmented high K(+)-evoked contractions and PKC inhibitors, such as 1-(5-isoquinolinesulfonyl)-2-methylpiperazine (H-7) and calphostin C, suppressed them more in SHR as compared with WKY . In alpha-toxin skinned SMA, the {Ca2+}i-force relationship curve was not significantly different between SHR and WKY . However, PDBu augmented {Ca2+}i-evoked contractions and PKC inhibitors suppressed them more in SHR than in WKY . CONCLUSION: These results suggest that the Ca2+ sensitivity of the contractile elements via PKC is significantly greater in prehypertensive SHR than in age-matched WKY . This abnormality in small muscular arteries may be involved in the pathogenesis of hypertension in SHR.

Proc Natl Acad Sci U S A, 1997 Dec 9, 94(25), 13832 - 7
Decreased ability of HIV-1 tat protein-treated accessory cells to organize cellular clusters is associated with partial activation of T cells; Wu MX et al.; It has been shown in several animal models that HIV infection of accessory cells (ACs) plays an important role in development of AIDS . Here, we report that ACs treated with HIV-1 Tat protein (Tat-ACs) have a decreased ability to organize cellular aggregates as compared with untreated ACs, resulting in incomplete activation of T cells in responses to anti-CD3 mAb or staphylococcal enterotoxin B stimulation . The T cells failed to up-regulate adhesion molecules CD11a and CD2 on the cell surface and had reduced proliferative responses, as determined by {3H}thymidine incorporation, but they obtained lymphoblast-like morphology and expressed early activation antigens on the cell surface such as Fas and CD69 and interleukin 2 receptor, at comparable levels as those T cells undergoing a maximal proliferation . These results suggest that the Tat-AC-induced defect occurs in the late, but not in the early, phases of T cell activation . Normal expression of cell surface Fas antigen accompanied by defects in late activation thus may result in the susceptibility of these T cells to apoptosis . Our studies suggest that dysfunction, hyperactivation, and susceptibility to apoptosis, as observed with T cells isolated from HIV-infected individuals, may be, at least in part, a consequence of abnormal functions of ACs.

Chem Biol, 1995 Jun, 2(6), 391 - 400
A photogenerated pore-forming protein; Chang CY et al.; BACKGROUND: The permeabilization of cells with bacterial pore-forming proteins is an important technique in cell biology that allows the exchange of small reagents into the cytoplasm of a cell . Another notable technology is the use of caged molecules whose activities are blocked by addition of photoremovable protecting groups . This allows the photogeneration of reagents on or in cells with spatial and temporal control . Here, we combine these approaches to produce a caged pore-forming protein for the controlled permeabilization of cells . RESULTS: 2-Bromo-2-(2-nitrophenyl)acetic acid (BNPA), a water-soluble cysteine-directed reagent for caging peptides and proteins with the alpha-carboxy-2-nitrobenzyl (CNB) protecting group, was synthesized . Glutathione (gamma-Glu-Cys-Gly) was released in high yield from gamma-Glu-CysCNB-Gly by irradiation at 300 nm . Based on this finding, scanning mutagenesis was used to find a single-cysteine mutant of the pore-forming protein staphylococcal alpha-hemolysin (alpha HL) suitable for caging . When alpha HL-R104C was derivatized with BNPA, pore-forming activity toward rabbit erythrocytes was lost . Near UV irradiation led to regeneration of the cysteine sulfhydryl group and the restoration of pore-forming activity . CONCLUSIONS: Caged pore-forming proteins are potentially useful for permeabilizing one cell in a collection of cells or one region of the plasma membrane of a single cell . Therefore, alpha HL-R104C-CNB and other caged proteins designed to create pores of various diameters should be useful for many purposes . For example, the ability to introduce reagents into one cell of a network or into one region of a single cell could be used in studies of neuronal modulation . Further, BNPA should be generally useful for caging cysteine-containing peptides and single-cysteine mutant proteins to study, for example, cell signaling or structural changes in proteins.

Zh Mikrobiol Epidemiol Immunobiol, 1995 Jul-Aug, (4), 75 - 8
{Reactivity to staphylococcal peptidoglycan in the neutrophil-endothelium system}; Maianskii AN et al.; The influence of S . aureus peptidoglycan on the process of adhesion in the neutrophil-endothelium system was studied . General regularities of this reaction, as well as the discrete action of peptidoglycan on each of its components were studied . The conclusion was made that S . aureus peptidoglycan was capable of enhancing the adhesiveness of both neutrophils and endotheliocytes . The adhesiveness of peptidoglycans of different staphylococcal species in adhesive reactions of neutrophils and endotheliocytes was manifested to a different degree . The possible mechanisms of the stimulating effect of peptidoglycans is discussed.

Rev Med Chir Soc Med Nat Iasi, 1996 Jan-Jun, 100(1-2), 119 - 24
{Postoperative and posttraumatic meningitis . Comments on 87 cases}; Mihalache D et al.; Iatrogenic and traumatic cerebromeningitis infections are increasing in frequency in the last two decades . Retrospective analysis of 87 patients iatrogenic and traumatic bacterial meningitis were admitted in the Clinic of Infectious Diseases from Iasi between 1, 01, 1990-31, 12, 1994 . Head trauma, lumbar punctures and iatrogenic meningitis infections were the causes cerebromeningitis infections . The diagnosis is usually difficult because of the poor specificity of the clinical signs . Predominant pathogens were gram-negative bacteria and Staphylococcus in iatrogenic meningitis and gram-positive bacteria (S . pneumoniae) in traumatic meningitis . The treatment was based on the use penicillin G + chloramphenicol, 3rd generation cephalosporins and sometimes 2nd generation quinolones . Ten of the 87 patients with iatrogenic and traumatic cerebromeningitis infections died (10.3%).

Childs Nerv Syst, 1997 Nov-Dec, 13(11-12), 584 - 7
Reducing the incidence of infection in pediatric cerebrospinal fluid shunt operations; Rotim K et al.; We reviewed our experience with shunt implantation during two time periods . From June 1985 to December 1990, 201 children with hydrocephalus underwent 382 operations . Among these children 36 (18%) developed a proven shunt infection, with an incidence rate per procedure of 9.4% . As a result of this study, a new effective protocol for shunt procedures involving modifications to the perioperative (antibiotic prophylaxis) and intraoperative management (meticulous surgical technique, complete shunt revision) of children undergoing initial shunt implantation or revision was initiated . With this new protocol 75 children underwent a total of 112 procedures between January 1991 and December 1995 . The incidence of shunt infection decreased, with a per patient rate of 8% and a per procedure rate of 5.3% . The majority of infections in our study were caused by Staphylococcus epidermidis, which was found in 22 (52.3%) patients.

Biochemistry, 1998 Jan 27, 37(4), 1067 - 75
Correlation between native-state hydrogen exchange and cooperative residue fluctuations from a simple model; Bahar I et al.; Recently, we developed a simple analytical model based on local residue packing densities and the distribution of tertiary contacts for describing the conformational fluctuations of proteins in their folded state . This so-called Gaussian network model (GNM) is applied here to the interpretation of experimental hydrogen exchange (HX) behavior of proteins in their native state or under weakly denaturing conditions . Calculations are performed for five proteins: bovine pancreatic trypsin inhibitor, cytochrome c, plastocyanin, staphylococcal nuclease, and ribonuclease H . The results are significant in two respects . First, a good agreement is reached between calculated fluctuations and experimental measurements of HX despite the simplicity of the model and within computational times 2 or 3 orders of magnitude faster than earlier, more complex simulations . Second, the success of a theory, based on the coupled conformational fluctuations of residues near the native state, to satisfactorily describe the native-state HX behavior indicates the significant contribution of local, but cooperative, fluctuations to protein conformational dynamics . The correlation between the HX data and the unfolding kinetics of individual residues further suggests that local conformational susceptibilities as revealed by the GNM approach may have implications relevant to the global dynamics of proteins.

Clin Infect Dis, 1997 Dec, 25(6), 1327 - 33
Antimicrobial therapy for chronic osteomyelitis in adults: role of the quinolones; Rissing JP; The development of antimicrobial therapy for osteomyelitis is reviewed . The disease, especially when chronic, is notoriously resistant to antibiotic therapy . The duration of disease defining chronicity has decreased considerably in the last 30 years . Successful therapy reflects increased appreciation of the combined roles of surgical debridement and prolonged antimicrobial courses . Parenteral high-dose beta-lactam agents yield clinical success for many patients with chronic osteomyelitis, particularly with prolonged administration and surgical debridement . Over the last decade, the initial success of oral quinolone therapy for gram-negative osteomyelitis was exploited further for staphylococcal diseases . Open clinical trials and comparative trials suggest success rates approximating those achieved with parenteral beta-lactams, particularly with appropriate surgery and adequate duration of therapy . The early results with quinolones and rifampin for prosthesis-related infection are encouraging . Overall, oral quinolones provide a new and frequently proportionate response to a disease that is difficult to treat.

Biochemistry, 1998 Jan 6, 37(1), 129 - 36
NMR study of the interaction between the B domain of staphylococcal protein A and the Fc portion of immunoglobulin G; Gouda H et al.; The solution structure of the B domain of staphylococcal protein A (FB) complexed with the Fc fragment of immunoglobulin G (IgG) is reported . A previous NMR analysis has shown that in solution FB is composed of a bundle of three alpha-helices, helix I, helix II, and helix III {Gouda, H., Torigoe, H., Saito, A., Sato, M., Arata, Y., and Shimada, I . (1992) Biochemistry 31, 9665-9672} . In contrast, the crystal structure of FB in the FB-Fc complex lacks helix III . Uniformly 15N- and 15N/13C-labeled FB were prepared, and the backbone 13C resonances were assigned . The spectral data obtained in the present study indicated that in solution all three helices including helix III are preserved in the FB-Fc complex . The mode of interaction of FB with the Fc fragment was discussed on the basis of the combined data of hydrogen-deuterium exchange experiments and 1H-15N correlation spectroscopy . It was concluded that a contiguous surface shaped by F14, Y15, E16, L18, and H19 in helix I, and N29, Q33, L35, and K36 in helix II is responsible for the binding.

Blood, 1998 Jan 1, 91(1), 340 - 6
The umbilical cord blood alphabeta T-cell repertoire: characteristics of a polyclonal and naive but completely formed repertoire; Garderet L et al.; Umbilical cord blood (CB) constitutes a promising alternative to bone marrow for allogeneic transplantation and is increasingly used because of the reduced severity of graft-versus-host disease after CB transplantation . We have compared the T-cell receptor beta chain (TCRB) diversity of CB lymphocytes with that of adult lymphocytes by analyzing the complementarity determining region 3 (CDR3) size heterogeneity . In marked contrast to adult samples, we observed bell-shaped profiles in all of the 22 functional beta-chain variable (BV) subfamilies that reflect the lack of prior antigenic stimulation in CB samples . However, the mean CDR3 size and BV usage were comparable between CB and adult samples . BJ2 (65%) segments were used preferentially to BJ1 (35%), especially BJ2S7, BJ2S5, BJ2S3, and BJ2S1, in both CB and in adult lymphocytes . We therefore conclude that although naive as reflected by the heterogeneity of the CDR3 size, the TCRBV repertoire appears fully constituted at birth . The ability to expand TCRB subfamilies was confirmed by stimulation with staphylococcal superantigens toxic shock syndrome toxin-1 and staphylococcal enterotoxin A . This study provides the basis for future analysis of the T-cell repertoire reconstitution following umbilical CB transplantation.

Transfus Sci, 1998 Mar, 19 Suppl, 65 - 9
Therapeutic immunoadsorption--its role in clinical practice; Braun N et al.; Extracorporeal immunoadsorption onto staphylococcal protein A replaces conventional plasmapheresis because its elimination efficiency is higher while less adverse reactions are seen . Experience in immunoadsorption is limited to specialized centers although recent technical developments make the procedure simple and safe to use . During a workshop held in Munich, September 1996, the application of immunoadsorption in various diseases was discussed . Goodpasture's syndrome, acquired autoimmune coagulopathy, HLA-sensitized transplantation and treatment-resistant life-threatening autoimmune diseases were accepted indications for immunoadsorption . Immunoadsorption in other immunoglobuline and immune complex mediated diseases is still under discussion.

Ann Vasc Surg, 1998 Jan, 12(1), 34 - 40
Role of plasma and extracellular matrix proteins in the physiopathology of foreign body infections; Francois P et al.; Foreign body implants are highly susceptible to microorganism infection . The infectious agents may be of low pathogenicity (such as S . epidermidis) or involve more virulent strains (such as S . aureus) . The common denominator for the three main elements that play a role in the physiopathology of such infections (bacteria, neutrophils, and different biomaterials) are host proteins deposited over the surface of the devices immediately after their implantation . These proteins modulate that host cells response but can also promote Staphylococcus adhesion to the biomaterial . Neutrophils and other cells such as fibroblasts adhere to several extracellular matrix proteins such as fibronectin, fibrinogen, collagen, vitronectin, via specific cell surface receptor . The evolution of the technology and the increasing numbers of long-term artificial implants require a better understanding of fundamental mechanisms of foreign body infections to reduce their incidence and optimize their treatment.

J Med Microbiol, 1998 Jan, 47(1), 63 - 70
Molecular analysis of the macrolide-lincosamide resistance gene region of a novel plasmid from Staphylococcus hyicus; Schwarz S et al.; Resistance to macrolides and lincosamides in Staphylococcus hyicus has been shown to be encoded by a 4.0-kb plasmid designated pSES21 . It differed distinctly in its restriction map from all other staphylococcal macrolide resistance plasmids reported so far . Southern blot hybridisation with gene probes specific for staphylococcal erm genes demonstrated that the macrolide resistance gene belonged to hybridisation class C . Analysis of the ermC gene revealed that the deduced amino-acid sequence of the pSES21-encoded ErmC methylase exhibited c . 93% identity with the ErmC methylase encoded by plasmid pE194 . The ermC gene of pSES21 was expressed constitutively and sequence analysis of the regulatory region showed multiple base-pair insertions and substitutions in the translational attenuator . As a consequence of these mutations, the reading frame of the small regulatory peptide was destroyed and a novel pair of inverted repeated sequences was generated . Previous studies identified sequence deletions and sequence duplications in the ermC regulatory region as the basis for constitutive ermC gene expression . The multiple point mutations shown in the pSES21-encoded ermC translational attenuator represent a novel kind of structural alteration in this regulatory region and may explain constitutive ermC gene expression by pairing of the newly generated inverted repeated segments in the presence of a functionally deleted reading frame for the small regulatory peptide.

J Steroid Biochem Mol Biol, 1997 Sep-Oct, 63(1-3), 81 - 9
Capillary gas chromatography with chemical ionization negative ion mass spectrometry in the identification of odorous steroids formed in metabolic studies of the sulphates of androsterone, DHA and 5alpha-androst-16-en-3beta-ol with human axillary bacterial isolates; Gower DB et al.; The products of metabolism of the sulphates (0.5 micromol/l) of androsterone, dehydroepiandrosterone (DHA) and 5alpha-androst-16-en-3beta-ol have been investigated after incubation with 72 h cultures of human axillary bacterial isolates for 3 days at 37 degrees C . The medium used, tryptone soya broth (TSB), contained yeast extract and Tween 80 . The isolates used were Coryneform F1 (known previously to metabolize testosterone and to be involved in under-arm odour (UAO) production, i.e . UAO +ve), Coryneform F46 (inactive in both the testosterone metabolism and UAO tests, i.e . UAO -ve) and Staphylococcus hominis/epidermidis (IIR3) . Control incubations of TSB alone, TSB plus each of the steroid sulphates and TSB plus each of the bacterial isolates were also set up . After termination of reactions and addition of internal standards, 5alpha-androstan-3beta-ol and 5alpha-androstan-3-one (50 ng each), extracted and purified metabolites were subjected to combined gas chromatography-mass spectrometry with specific ion monitoring . Steroidal ketones were derivatized as their O-pentafluorobenzyl oximes; steroidal alcohols (only androst-16-enols in this study) were derivatized as their tert-butyldimethylsilyl ethers . Analysis was achieved by negative ion chemical ionization mass spectrometry for the pentafluorobenzyl oximes at {M-20}- and electron impact positive ion mass spectrometry for the tert-butyldimethylsilyl ethers at {M-57}+ . The incubation broth contained two compounds which had gas chromatographic and mass spectrometric properties identical to those of DHA and 4-androstenedione . It was not possible, therefore, to show unequivocally that DHA sulphate (DHAS) was converted microbially into DHA, although this is implied by the finding of small quantities of testosterone and 5alpha-dihydrotestosterone in incubations with F1 . With androsterone S, no free androsterone was recorded and only very small (5 pg or less) amounts of testosterone . Two odorous steroids, androsta-4,16-dien-3-one and 5alpha-androst-2-en-17-one (Steroid I) were formed (mean quantities 40 and 45 pg, respectively) . The sulphate of 5alpha-androst-16-en-3beta-ol was metabolized with F1 into large quantities of the odorous steroids, 5alpha-androst-16-en-3-one and Steroid I . In addition, much smaller quantities of androsta-4,16-dien-3-one were formed . In contrast, incubations of DHAS with F46 resulted in no metabolites except, possibly, DHA, but the sulphate moiety of androsterone S was also cleaved to yield the free steroid together with large amounts of Steroid I . In incubations of DHAS and androsterone S with F1, no 16-unsaturated steroids were formed, although 5alpha-androst-16-en-3beta-yl S was de-sulphated and the free steroid further metabolized . No evidence was obtained for androst-16-ene metabolism in incubations with F46 . In incubations with S . hominis/epidermidis (IIR3), androsterone S was converted into androsterone and, in high yield, to Steroid I plus some 5alpha-androst-16-en-3-one . Both DHAS and androsterone S were converted into androst-16-enols . Sulphatase activity was also manifested when 5alpha-androst-16-en-3beta-yl S was utilized as substrate with IIR3, large quantities of Steroid I and 5alpha-androst-16-en-3-one being formed, together with further metabolism of androst-16-enes . In view of the fact that both DHAS and androsterone S occur in apocrine sweat, the metabolism of these endogenous substrates by human axillary bacteria to several odorous steroids may have important implications in the context of human odour formation.

Klin Monatsbl Augenheilkd, 1997 Oct, 211(4), 245 - 9
{Endophthalmitis after intraocular interventions}; Strmen P et al.; BACKGROUND: Endophthalmitis is serious complication of intraocular surgery . At the University Eye Clinic Bratislava we performed a retrospective study on the occurrence of endophthalmitis after various intraocular procedures . MATERIAL AND METHODS: Over the years 1993-1995 we performed 2374 cataract extractions, 305 anti-glaucoma operations, 106 perforating keratoplasties and 926 pars plana vitrectomies . The charts of all patients were reviewed . The occurrence of endophthalmitis, type of surgery, perioperative complications, elapsed time between surgery and start of endophthalmitis, results of preoperative conjunctival smear cultures, final anatomical and functional results were evaluated . RESULTS: The frequency of endophthalmitis after cataract extraction was 0.84%, after antiglaucoma surgery 1.31% and 0.97% after pars plana vitrectomy . No endophthalmitis was found after penetrating keratoplasties . In most cases endophthalmitis began in the first three days after surgery . Endophthalmitis occurred after uncomplicated as well as after surgical procedures with various perioperative complications . The preoperative conjunctival smear cultures were sterile in 22 patients . Staphylococcus epidermidis was the most commonly cultured germ . All patients were treated with antibiotics parenterally and locally . Pars plana vitrectomy and intravitreal application of antibiotics was performed only exceptionally . Enucleation was required in two eyes and two other eyes become phthitical . 22 eyes recovered without serious complications . Five eyes were blind and in 11 eyes the final visual acuity was better than 6/36 . CONCLUSIONS: Pre-, peri-, and postoperative measures must be improved to prevent postoperative endophthalmitis . In cases of endophthalmitis all therapeutical options have to be performed as soon as possible to save the patient's eye and sight.

Chemotherapy, 1998 Jan-Feb, 44(1), 63 - 8
Continuous infusion of vancomycin in methicillin-resistant staphylococcus infection; Di Filippo A et al.; OBJECTIVE: The aim of the study was to verify the therapeutic response of vancomycin in methicillin-resistant staphylococcus infection (MRSA/ MRCNS) administered according to two different methods (intermittent infusion vs . continuous infusion) . METHOD: Experimental plan: retrospective study; study environment: university hospital, two intensive care units . Twenty-five critically ill patients submitted to antibiotic treatment with vancomycin for infection from MRSA/MRCNS were studied . The patients, who were classified according to SAPS II scores, were divided into two groups: group A (n = 14): dose of vancomycin of 0.5 g x 4/day and group B (n = 11): dose of 2 g/day of vancomycin administered in a continuous infusion . Before the antibiotic therapy was started (T1) and prior to its end (T2), the following parameters were evaluated: degree of impairment of the main organs and systems by means of sepsis-related organ failure assessment score (SOFA) and count of the white blood cells (WBC) . The length of the hospital stay during intensive care was calculated for both groups (statistics: Student t test) . RESULTS: No significant differences were found in the SAPS II scores and in the length of the hospital stay . In a comparison of the T1 and T2 results, we noted that patients of group A had no variations in the SOFA scores (4.84 +/- 2.48 vs . 4 +/- 3.9) and in the WBC mean values (12,415 +/- 5,099 vs . 12,841 +/- 6,864 cells/mm3) . In contrast, in the patients of group B, we noted significant variations (p < 0.05) in the mean values of the SOFA scores (6.62 +/- 2.2 vs . 4.37 +/- 3.5) and in the mean values relative to the WBC count (17,242 +/- 12,842 vs . 10,757 +/- 3,610 cells/mm3) . CONCLUSIONS: In critically ill patients suffering from MRSA/MRCNS infection, vancomycin administration in continuous infusions improved organ function and leukocyte response, but did not seem to modify the overall evolution of the disease.

J Biol Chem, 1997 Dec 19, 272(51), 32190 - 7
Conformational integrity and ligand binding properties of a single chain T-cell receptor expressed in Escherichia coli; Khandekar SS et al.; We recently showed that a soluble, heterodimeric murine D10 T-cell receptor (TCR) (Valpha2Calpha, Vbeta8.2Cbeta) expressed in insect cells binds both Vbeta8.2-specific bacterial superantigen staphylococcal enterotoxin C2 (SEC2) and a soluble, heterodimeric major histocompatibility complex class II I-Ak.conalbumin peptide complex with a low micromolar affinity . To define further the structural requirements for the TCR/ligand interactions, we have produced in Escherichia coli a soluble, functional D10 single chain (sc) TCR molecule in which the Valpha and Vbeta domains are connected by a flexible peptide linker . Purified and refolded D10 scTCR bound to SEC2 and murine major histocompatibility complex class II I-Ak.conalbumin peptide complex with thermodynamic and kinetic binding constants similar to those measured for the baculovirus-derived heterodimeric D10 TCR suggesting that neither the TCR constant domains nor potential N- or O-linked carbohydrate moieties are necessary for ligand recognition and for expression and proper folding of the D10 scTCR . Purified D10 scTCR remained soluble at concentrations up to 1 mM . Circular dichroism and NMR spectroscopy indicated that D10 scTCR is stabilized predominantly by beta-sheet secondary structure, consistent with its native-like conformation . Because of its limited size, high solubility, and structural integrity, purified D10 scTCR appears to be suitable for structural studies by multidimensional NMR spectroscopy.

Am J Rhinol, 1997 Nov-Dec, 11(6), 429 - 33
Septic thrombosis of orbital vessels due to cutaneous nasal infection; Sanchez TG et al.; The authors describe two cases of cutaneous nose infection that quickly spread and extended to the orbital venous complex . At first glance, the clinical presentation could be mistaken for a complicated sinusal infection; therefore, the evaluation of the sinuses, by means of physical examination and radiological investigation, was of great concern, showing that there was no important pathology in the sinuses . The CT scan and the color Doppler imaging (orbital ultrasound with Doppler) demonstrated, throughout the development of the disease, that the superior ophthalmic vein was affected in both patients and the cavernous sinus in one of them . On physical examination, chemosis of the conjunctiva, proptosis, and edema of the eyelids were prominent . Patients improved only after appropriate intravenous antibiotic therapy against staphylococcus (clindamycin) and corticosteroids, making one conclude that treatment of this disease should be initiated as soon as possible in order to decrease morbidity and mortality.

Vet Immunol Immunopathol, 1997 Sep 19, 58(3-4), 219 - 30
An investigation of phagocytosis and intracellular killing of Staphylococcus intermedius by canine neutrophils in vitro; Shearer DH et al.; Bacterial infection (pyoderma) of the canine skin is largely caused by Staphylococcus intermedius and may be a primary, idiopathic disease or secondary to a range of other dermatological disorders . The immune response to this organism involves phagocytosis and killing by neutrophils . In this study, the effect of opsonisation of S . intermedius by clinical sera (n = 60) and sera from normal dogs (n = 20) was investigated by in vitro phagocytosis and killing assays . All sera from dogs with pyoderma were significantly more efficient at opsonisation for phagocytosis than normal dog sera (p < 0.001) . Further, fresh serum was more effective at opsonisation of S . intermedius than heat-inactivated serum in this assay (p < 0.001) . The results indicate that opsonisation is enhanced by serum from dogs with pyoderma; therefore the humoral response in clinical cases of canine pyoderma is, in terms of phagocytosis, protective . The small but significant reduction in phagocytosis of S . intermedius opsonised by heat-inactivated serum compared to fresh frozen serum suggests that complement plays a role, although minor, in opsonisation . An in vitro killing assay showed that normal canine neutrophils were able to effectively kill phagocytosed S . intermedius over a 90-min time course, but there was no significant enhancement of killing by prior incubation of the bacteria with any clinical serum as opposed to normal sera . This suggests that the humoral response to S . intermedius does not affect intracellular killing per se.

Cancer Immunol Immunother, 1997 Nov-Dec, 45(3-4), 190 - 2
Redirected cellular cytotoxicity employing bispecific antibodies and other multifunctional binding proteins; Weiner LM et al.; Commencing with the discovery and characterization of bispecific antibodies, numerous investigations have shown that such antibodies are capable of redirecting cellular cytotoxicity . Clinical trials testing diverse strategies, including those targeting CD16-expressing effector cells, have been conducted or are in progress . This manuscript reviews our clinical trials efforts with bispecific antibodies and describes our experience employing multi-functional binding proteins containing tumor-targeting antibody Fab fragments linked to bacterial superantigens, such as staphylococcal enterotoxin A.

Cancer Immunol Immunother, 1997 Nov-Dec, 45(3-4), 180 - 3
T cell activation and retargeting using staphylococcal enterotoxin B and bispecific antibody: an effective in vivo antitumor strategy; Porter LE et al.; The aim of this work was to test for cure and immunity in a micrometastatic tumor model using in vivo T cell activation with staphylococcal enterotoxin B (SEB) and retargeting with antitumor x anti-CD3 F(ab')2 bispecific antibodies (bsAb) . All studies were performed in C3H/HeN mice using syngeneic tumor cell lines . For survival studies, mice were injected intravenously on day 0 with CL62 (a p97-transfected clone of the K1735 murine melanoma tumor) . Day-3 treatments included saline (control), SEB (50 gamma g intraperitoneal) with or without bsAb (5 micrograms i.v.) . Cured mice, surviving beyond 60 days, were rechallenged with subcutaneous CL62, K1735, or a nonmelanoma control, AG104 . SEB activation studies were performed with pulmonary tumor-infiltrating lymphocytes isolated from 10-day established CL62 tumors . Maximal tumor-infiltrating lymphocyte cytotoxicity was demonstrated 24 h following SEB injection, therefore bsAb treatments were administered 24 h after SEB . When survival was examined at 60 days, there were significantly more survivors in the group receiving SEB plus bsAb (70%) compared to the group receiving SEB alone (30%), and the controls (0%) (P = 0.02 and P < 0.01, respectively) . Mice cured of CL62 using SEB alone or with bsAb demonstrated equal immunity to CL62, however, mice treated with SEB plus bsAb were more often immune to the p97-parental cell line, K1735(P = 0.001) . Ag104 consistently grew in all mice . Results of these studies demonstrate that SEB plus bsAb can be effective, not only in curing tumors but also in providing protective immunity against targeted and non-targeted tumor antigens.

J Immunol Methods, 1997 Oct 13, 208(1), 49 - 59
Simultaneous detection of DNA synthesis and cytokine production in staphylococcal enterotoxin B activated CD4+ T lymphocytes by flow cytometry; Mehta BA et al.; Assessment of T cell activation has traditionally been performed by measuring proliferation as a function of 3{H}-thymidine incorporation, or secretion of cytokines from activated peripheral blood mononuclear cells (PBMC) in culture . An alternative method for detection of proliferation at the single cell level utilizes incorporation of bromodeoxyuridine (BrdU), an analog of thymidine, into cellular DNA . After appropriate fixation and permeabilization of the cells, a monoclonal antibody (mAb) against BrdU conjugated with a fluorescent dye is employed to measure by flow cytometry the incorporated BrdU . Here, we report a flow cytometric procedure which can be used for the simultaneous detection of BrdU incorporation, activation markers such as CD69 and CD25, and intracellular cytokines in T cell subsets from activated PBMC . Our observations are consistent with the proposal that cytokine synthesis and cell proliferation occur sequentially in CD4+ T cells stimulated with the superantigen staphylococcal enterotoxin B (SEB) . The majority of cells expressing the cytokines IFN-gamma and IL-2 at 48 h appear to have undergone DNA synthesis, however all proliferating cells do not express IFN-gamma or IL-2 . The methods presented in this report offer a unique approach for studying simultaneous expression of key cellular activation events in phenotypically resolved lymphocyte populations.

Ophthal Plast Reconstr Surg, 1997 Dec, 13(4), 256 - 8
Detection of orbital implant infection with technetium 99m-labeled leukocytes; Kristinsson JK et al.; Orbital implant infection is a rare cause of anophthalmic socket pain . Because of the potential danger of infection spreading to nearby structures, it is of paramount importance to diagnose the condition as soon as possible . Scintigraphy is a method for diagnosis of graft infections by radioisotopic imaging of inflammatory sites . We report on a patient with socket pain 3 months after implantation of an acrylic implant . The socket appearance was normal and there were no signs of infection other than culture-positive socket exudation . Three consecutive computed tomography scans revealed no abnormalities . 99mTc leukocyte scintigraphy revealed white blood cell accumulation at the implantation site . The implant was removed and cultured . This produced Staphylococcus epidermidis and R . equii . A parenteral antibiotic treatment was instituted with subsequent improvement of symptoms . Four months later, after negative scintigraphy, a hydroxyapatite implant was inserted, demonstrating full vascularization on a bone scan after 2 months . Two months later, the patient developed the previous symptoms, with all of the former findings, including positive scintigraphy . The implant was removed, revealing a microabscess on the anterior aspect, producing S . epidermidis on culture . We conclude that scintigraphy using 99mTc-labeled leukocytes is a useful technique in diagnosing low-grade orbital infection.

Acta Cardiol, 1997, 52(5), 437 - 43
Successful treatment of staphylococcal pericarditis with early catheter drainage and antibiotics; Rallidis LS et al.; We describe a 26-year-old man with staphylococcal pericarditis who was treated successfully by percutaneous catheter drainage combined with antibiotics . One year later he remains in good condition without echocardiographic evidence of constriction.

Biochemistry, 1997 Dec 9, 36(49), 15134 - 9
Effect of prolyl isomerase on the folding reactions of staphylococcal nuclease; Veeraraghavan S et al.; The low-temperature fluorescence-detected refolding of staphylococcal nuclease (SNase) can be described by three slow kinetic phases . The slowest phase is absent in the P117G mutant of SNase . Peptidyl prolyl cis-trans isomerase (cyclophilin), which has been shown to catalyze the slow folding reactions of some proteins, was employed to determine which of the refolding reactions of SNase and P117G SNase involve proline isomerization . We report here that all three folding phases of the wild type and the slower phase of P117G SNase are catalyzed by prolyl isomerase, indicating that proline isomerization is involved in these fluorescence-detected phases in the refolding of SNase . Since the rates of these phases are denaturant-dependent, we conclude that the slow folding steps involve isomerization of non-native cis proline peptide bonds and are tightly coupled to denaturant-sensitive structural changes.

Bone Marrow Transplant, 1997 Nov, 20(9), 779 - 83
A single, double lumen high-flow catheter for patients undergoing peripheral blood stem cell transplantation . Experience at the National Cancer Institute in Mexico; Volkow P et al.; Peripheral blood stem cell transplantation (PBSCT) requires a high-flow catheter for adequate cell collection by apheresis and long i.v . support, this is usually achieved by multiple catheters . We analyzed our experience with Mahurkar or Permacath for apheresis and long-term i.v . support in PBSCT, cared for exclusively by an i.v . therapy team . Fifty-six catheters were used in 53 patients that completed PBSCT (28 Permacath and 28 Mahurkar) . In 10 patients (19%) the same catheter was used for multiple PBSCT . The average stay was 58.4 days (7-219), Permacath 76.8 days (14-219) and Mahurkar 42 days (7-106) . The incidence of infectious complications was 2.2 x 1000 catheter-days (1.7 Permacath and 3.0 Mahurkar); during neutropenia it was 3.7 x 1000 cathether-days . The incidence of thrombosis was 0.9 x 1000 catheter-days . There was a total of seven infectious episodes (12.7%) . Five (9%) were local and two were (3.6%) bacteremias . The microorganism most commonly isolated was Staphylococcus sp . (57%) . Four catheters (7.1%) were removed because of complications: one thrombosis and three infections . Both catheters have proven useful and safe for long-lasting vascular access in patients undergoing PBSCT . No statistical difference was found in infectious and non-infectious complications between either catheters.

Kidney Int, 1997 Dec, 52(6), 1579 - 83
Influence of endotoxin contamination on anti-GBM antibody induced glomerular injury in rats; Karkar AM et al.; It is accepted that the main determinant of glomerular injury in experimental nephrotoxic nephritis is the administered dose of anti-glomerular basement membrane (GBM) antibody . However, there are other factors that can enhance the severity of such injury including small doses of bacterial lipopolysaccharide (LPS) . In the present study, we have assessed whether preparations of anti-GBM antibody contaminated with different concentrations of endotoxin could influence the severity of glomerular injury in the heterologous phase of nephrotoxic nephritis . We have also examined the efficacy of different laboratory methods to isolate an endotoxin-free anti-GBM antibody, and to purify anti-GBM antibody preparations from endotoxin . Preparations of anti-GBM antibody (nephrotoxic globulin) isolated from nephrotoxic serum by the sodium sulphate precipitation method contained variable concentrations of endotoxin . Administration of these preparations in equal doses into clean rats, which had no established acute phase response, markedly aggravated the severity of glomerular injury . However, preparations contained less than 50 pg/ml of endotoxin appeared to have no significant effect on such injury . Furthermore, isolation of anti-GBM antibody from nephrotoxic serum by affinity chromatography, using Staphylococcus protein-A column, proved to be a reliable method not only for the isolation of an IgG (nephrotoxic antibody) free from other serum contaminants, but also for purification of endotoxin contaminated preparations of anti-GBM antibody . These observations have practical implications in studying models of nephritis as our results show that the glomerular injury, which is usually considered to be a sole function of the mass of antibody bound to GBM, is profoundly influenced by minor endotoxin contamination of the anti-GBM antibody.

MMWR Morb Mortal Wkly Rep, 1997 Dec 19, 46(50), 1189 - 91
Outbreak of staphylococcal food poisoning associated with precooked ham--Florida, 1997; Abscess on the lateral epicondyle of the humerus as a cause of lameness in a horse; Department of Large Animal Clinical Services, College of Veterinary Medicine, Oregon State University, Corvallis 97331-4803, USAAn 18-month-old 450-kg {990-lb} sexually intact male Holsteiner was evaluated for lameness of the left forelimb of 3-months' duration . We were unable to localize the site of lameness, using intra-articular and perineural anesthesia, and radiography of the shoulder and cubital joint (elbow) did not reveal radiographic abnormalities . Nuclear scintigraphy was performed . An increase in radio-isotope uptake was evident at the lateral epicondyle of the left humerus . Radiographs of the region 3 weeks later revealed a 1.5-cm focal lucency surrounded by a 1.0-cm rim of necrotic bone . The lesion was consistent with an encapsulated osteomyelitis or bone abscess . Surgical correction was performed, using general anesthesia, and involved a lateral approach to the epicondyle of he humerus . Cyst contents were removed with curettage, and the cavity was packed with cancellous bone harvested from the tuber coxae . A coagulase-negative Staphylococcus organism was cultured from the abscess . The horse was sound 3 weeks after surgery, and radiography 10 months later revealed complete ossification and adjacent sclerosis at the surgical site . Solitary cystic lesions of long bones may represent bone abscesses capable of causing lameness in horses . Nuclear scintigraphy can provide early diagnostic capability . Curettage and cancellous bone grafting are indicated for treatment of bone abscesses.

Med Dosw Mikrobiol, 1997, 49(1-2), 27 - 33
{Incidence of methicillin-resistant and methicillin-susceptible Staphylococcus strains in clinical materials}; Wiktorowicz-Belzyt E et al.; 365 S . aureus and 165 CNS were evaluated for susceptibility to methicillin . They were received from various clinic of Clinical Hospital in Bydgoszcz . The bacterial cells that grew in the presence of 25 mg methicillin/L were evaluated as methicillin-resistant . It was found that the 129 MRSA and MRCNS . Incidence depended on the kind of clinic and material . Most often they were isolated from clinic: Surgery, Neurosurgery, Intensive Care Unit, Neurology and Ortopedics . They were from: intubation tube, catheters and vascular grafts . The methicillin-resistant S . epidermidis and S . haemolyticus showed above 50% resistance to methicillin.

Br J Haematol, 1997 Nov, 99(2), 358 - 63
Cladribine (2-CDA) given as subcutaneous bolus injections is active in pretreated Waldenström's macroglobulinaemia . Swiss Group for Clinical Cancer Research (SAKK); Betticher DC et al.; Clinical trials with intravenous cladribine infusions in pretreated patients with Waldenstrom's macroglobulinaemia have shown a response rate of 40% . Our pharmacokinetic studies revealed that the bioavailability of subcutaneous cladribine is complete but that the concentration-time profile is very different from intravenous administration . We designed this phase II multi-institutional trial to study the activity and toxicity of cladribine given as s.c . bolus injections in patients with symptomatic Waldenstrom's macroglobulinaemia . Between May 1993 and October 1995, 25 patients were accrued: male/female 18/7, median age 65 years (range 44-85) . All except one patient had been pretreated with more than one regimen (median 2, range 0-10) . 18 patients had progressed under previous therapy and six were in relapse . All patients received cladribine for a total dose of 0.5 mg/kg per cycle as s.c . bolus injections divided over 5 d at > or = 4 week intervals, for a maximum of six cycles . All 25 patients were evaluable for toxicity and response . A total of 67 cycles were administered (median 3 cycles, range 1-6) . Overall response rate including disease stabilization which had been progressive under previous therapy was 68% . 10 patients (40%, 95% CI 21-61%) achieved a partial remission . Seven responders had been progressive under previous therapy . Maximum responses were reached no later than the third cycle . Median time to treatment failure and remission duration were 4.4 (range 0.5-33) and 8 months (5-29), respectively . Four patients (16%) suffered from infections W.H.O . grade > or = 2 (pneumonia grade 2, Staphylococcus septicaemia grade 3, viral encephalitis and pneumonia, both grade 4 with complete resolution) . No other severe adverse events were observed . Cladribine given as s.c . 5 d bolus injections was found to be active in pretreated Waldenstrom's macroglobulinaemia and resulted in durable remissions.

Int Ophthalmol, 1997, 21(2), 71 - 4
Heparin and heparin-surface-modification reduce Staphylococcus epidermidis adhesion to intraocular lenses; Abu el-Asrar AM et al.; Bacterial adherence to intraocular lenses (IOLs) could be the cause of endophthalmitis following cataract surgery and lens implantation . The majority of cases of postoperative endophthalmitis are caused by microflora that reside on or near the eye of the patient . Staphylococcus epidermidis commonly colonizes the eyelid margin and conjunctiva and is the most common organism causing postoperative endophthalmitis . In this study, the in vitro adherence of S . epidermidis to regular poly-methyl methacrylate (PMMA) IOLs and to heparin-surface-modified (HSM) PMMA IOLs was investigated . The effects of heparin and antibiotics in solution on the adherence of bacteria to regular PMMA IOLs were evaluated . Adhesion of bacterial cells to IOLs was determined by counting the viable cells attached to the lenses . Significantly, fewer S . epidermidis attached to HSM-PMMA IOLs and to regular PMMA IOLs treated with heparin than to PMMA IOLs (p < 0.001) . Furthermore, bacteria attached in significantly lower numbers to regular PMMA IOLs treated with heparin than to HSM-PMMA IOLs (p = 0.0031) . Antibiotics in solution had no significant effect on bacterial adherence to PMMA IOLs . These data indicate that the use of HSM-PMMA IOLs and treatment of PMMA IOLs with heparin could diminish the incidence of postoperative endophthalmitis and intraocular inflammation associated with IOL implantation.

Neurosci Lett, 1997 Oct 24, 236(1), 53 - 6
Mechanisms and sites of pyrogenic action exerted by staphylococcal enterotoxin A in rabbits; Huang WT et al.; The febrile responses induced by i.v . administrations of staphylococcal enterotoxin A (SEA) was mimicked by direct injection of SEA into the organum vasculosum laminae terminalis (OVLT) in unanesthetized rabbits . Compared with the febrile responses induced by i.v . injection of SEA, the OVLT route of injection required a much lower dose of SEA to produce a similar fever . Furthermore, the fever induced by intra-OVLT or i.v . injection of SEA was significantly attenuated by pretreatment with intra-OVLT injection of anisomycin (a protein synthesis inhibitor), indomethacin or diclofenac (inhibitors of cyclo-oxygenase (COX)), and aminoguanidine or dexamethasone (inhibitors of inducible nitric oxide synthase (iNOS)) . These results suggest that COX or iNOS pathway in the OVLT mediate the SEA-induced fever in rabbits.

Microbiol Immunol, 1997, 41(10), 799 - 803
Protease-induced multicell formation in Staphylococcus haemolyticus; Yabu K et al.; Multicellular cells were efficiently induced in Staphylococcus haemolyticus by the addition of protease to exponentially growing cultures at 30 C . Electron microscopy revealed the formation of tetrad-shaped multicells that were septated but not separated from each other . Incubation of the multicells with extract from the cells grown without protease resulted in a fourfold increase in the number of colony-forming units as compared with the untreated control . An electrophoretic analysis showed that protease caused a loss of cell wall-lytic activity of the cell, which possibly led to the formation of multicells through cessation of cross wall separation.

Antonie Van Leeuwenhoek, 1997 Oct, 72(3), 245 - 9
Staphylococcal serine proteinase increases intracellular free calcium concentration in human polymorphonuclear leukocytes; Nowak D et al.; Staphylococcal serine proteinase (SSP) can influence various functions of human polymorphonuclear leukocytes (PMNL) including chemotaxis and phagocytosis . Since the rise in intracellular free calcium concentration is an important step in signal transduction leading to phagocyte activation, we tested the ability of SSP to increase the intracellular free calcium concentration in human PMNL using the fluorescent calcium indicator Fura-2AM . PMNL isolated from healthy donors responded to SSP in the concentration range of 10 to 100 micrograms/ml . The highest Ca2+ rise (104 +/- 47 nM) was observed for 10 micrograms/ml SSP . It was mainly dependent (81 +/- 11%) on extracellular calcium influx, however, SSP mobilized 68 +/- 7% of Ca2+ from intracellular calcium stores . Boiling of SSP or preincubation with phenylmethylsulphonylfluoride (an serine proteinase inhibitor) did not change its ability to increase intracellular free calcium concentration in PMNL . It suggests that active center of SSP is not responsible for Ca2+ mobilization . Finally, PMNL responded to each of three consecutive stimulations with SSP independently of the presence of high or low extracellular Ca2+ concentration . This may be an additional mechanism responsible for activation of human PMNL and degradation of alveolar walls during the staphylococcal infection in the lower airways.

Cell Immunol, 1997 Nov 1, 181(2), 153 - 62
Influence of retinoic acid on the differentiation pathway of T cells in the thymus; Yagi J et al.; This study investigated the ability of retinoic acid (RA) to influence T cell differentiation . All-trans-RA had marked effects on T cell differentiation in murine fetal thymic organ cultures (FTOCs) . The time course of the effect of all-trans-RA in FTOC of day 14 C57BL/6 embryos revealed a twofold increase in the frequency of CD4 single-positive (SP) cells and a high level of CD3-bearing cells (CD3high cells) at a later stage of T cell development . At an earlier stage, all-trans-RA induced a twofold increase in the frequency of CD4 SP cells, but significantly suppressed the upregulation of CD3 and TCR . Reverse transcription-PCR using RA receptor (RAR) subtype-specific primers showed that RAR alpha but not beta and gamma is expressed during T cell development in the thymus and that its expression was associated with the generation of CD4/CD8 double-positive (DP) cells . In FTOC of day 16 BALB/c embryos, the level of V beta 3high cells was greatly reduced (1.4% of the CD3high cells) in response to the mouse mammary tumor virus-6-encoded superantigen, but V beta 3-bearing cells were rescued from the deletion in the presence of all-trans-RA (5.6% of the CD3high cells) . Further, the inhibitory effect of all-trans-RA on thymocyte deletion was observed when the deletion was induced by a low concentration of staphylococcal enterotoxin B in FTOC . Taken together, these data suggest that RA increases the frequency of mature and self-reactive T cells in the thymus, possibly by inhibiting the process of negative selection at the DP stage of T cell differentiation.

Biochemistry, 1997 Nov 25, 36(47), 14539 - 50
Dissecting the catalytic mechanism of staphylococcal lipases using carbamate substrates: chain length selectivity, interfacial activation, and cofactor dependence; Simons JW et al.; p-Nitrophenyl N-alkylcarbamates with different alkyl chains were used as substrates to determine separately the carbamylation and decarbamylation rates of the lipases from Staphylococcus hyicus and S . aureus . Both enzymes are reversibly inhibited by these compounds due to a rapid carbamylation of their active site serines followed by a slow decarbamylation . The carbamylation reaction is strongly pH-dependent and the pH profile suggests that an unprotonated histidine is required for this reaction . In contrast, the decarbamylation is pH-independent suggesting the presence of a hydrogen bond between the active site histidine and the carbamyl moiety . S . hyicus lipase preferably reacts with medium to long chain carbamates with an optimum for eight carbon atoms . In contrast, S . aureus lipase is highly specific for short chain carbamates . These results are in agreement with the respective substrate preferences of both lipases toward natural lipids . The decarbamylation rates of both enzymes hardly depend on the alkyl chain length, and from this it is concluded that chain length selectivity is expressed in the first step of catalysis . Both the carbamylation and decarbamylation reaction rates of S . hyicus lipase are enhanced in the presence of micelles, the activation effect being most pronounced in the first step . For the S . aureus lipase only a small influence of interfaces on both reaction steps was observed . These results are discussed in view of a possible role of a lid covering the active site . Kinetic experiments in the presence and absence of calcium strongly suggest that calcium ions are important for the structural stabilization of the unmodified as well as of the carbamylated enzymes . This structural function of calcium was supported by urea unfolding experiments, from which it appeared that for both enzymes the free energy for unfolding is significantly lower in the absence of calcium . In conclusion our results show that kinetic differences between both lipases reside in the acylation step, and that calcium is important for the structural stabilization of the unmodified, and moreover, the acylated enzymes.

Exp Clin Immunogenet, 1997, 14(2), 141 - 8
Antibody to human MHC class I inhibits SIVsmmPBj1.9-induced proliferation of pigtailed macaque lymphocytes; Baskar PV et al.; Previously we have shown that the simian immunodeficiency virus SIVsmmPBj1.9, a molecular clone of SIVsmmPBj14, induces proliferation of human peripheral blood mononuclear cells (PBMC) . We have extended this observation to show that SIVsmmPBj1.9 induces proliferation of PBMC from pigtailed macaques . This proliferative response was markedly inhibited by mAbs against human class I MHC, class II MHC and CD4 antigens, and partially inhibited by mAbs against integrin beta 2 subunit (CD18) and LFA-1 (CD11a) . However, these antibodies differed in their ability to inhibit in vitro viral infectivity of PBMC . While anti-CD4, MHC class II, and LFA-1 strongly inhibited viral infectivity, antibodies to MHC class I demonstrated little effect on viral infectivity . A control antibody (PLM2) against porcine CD18 inhibited neither virus-induced proliferation nor viral infectivity . Based on these results, we suggest that SIVsmmPBj1.9-induced proliferation requires the participation of class I MHC, class II MHC and CD4 molecules . In addition, the observation that anti-class I MHC Ab inhibited proliferation of macaque PBMC induced by mitogen (PHA) and bacterial superantigens, such as Staphylococcus enterotoxin A and toxin shock syndrome toxin-1, suggests that SIVsmmPBj1.9 also contains a viral superantigen similar to that previously demonstrated in SIVsmmPBj14.

Infect Immun, 1997 Dec, 65(12), 5118 - 24
A mutation of F47 to A in staphylococcus enterotoxin A activates the T-cell receptor Vbeta repertoire in vivo; Rosendahl A et al.; The bacterial superantigen staphylococcal enterotoxin A (SEA) binds with high affinity to major histocompatibility complex (MHC) class II molecules and subsequently activates T cells bearing particular T-cell receptor (TCR) Vbeta chains . Structural and mutational studies have defined two distinct MHC class II binding sites located in the N-terminal and C-terminal domains of SEA . The N-terminal F47 amino acid is critically involved in a low-affinity interaction to the MHC class II alpha-chain, while the C-terminal residues H187, H225, and D227 coordinate a Zn2+ ion and bind with moderate affinity to the beta-chain . In order to analyze whether the SEA-MHC class II alpha-chain interaction plays a role in dictating the in vivo repertoire of T-cell subsets, we studied distinct Vbeta populations after stimulation with wild-type SEA {SEA(wt)} and SEA with an F47A mutation {SEA(F47A)} . Injections of SEA(wt) in C57BL/6 mice induced cytokine release in serum, strong cytotoxic T-lymphocyte activity, expansion of T-cell subsets, and modulated expression of the T-cell activation antigens CD25, CD11a, CD44, CD62L, and CD69 . SEA-reactive TCR Vbeta3+ and Vbeta11+ T cells were activated, while TCR Vbeta8+ T cells remained unaffected . The SEA(F47A) mutant protein induced a weaker T-cell response and failed to induce substantial interleukin-6 production compared to SEA(wt) . Notably, SEA(F47A) failed to activate TCR Vbeta11+ T cells, whereas in vivo expansion and modulation of T-cell activation markers on TCR Vbeta3+ T cells were similar to those for SEA(wt) . A similar response to SEA(F47A) was seen among CD4+ and CD8+ T cells . Activation of TCR Vbeta3+ and TCR Vbeta11+ T-cell hybridomas confirmed that SEA(F47A) activates TCR Vbeta3+ but not TCR Vbeta11+ T cells . The data support the view that the SEA-N-terminal MHC class II alpha-chain interaction defines a topology that is required for engagement of certain TCR Vbeta chains in vivo.

J Bacteriol, 1997 Dec, 179(23), 7573 - 6
Specificities of FemA and FemB for different glycine residues: FemB cannot substitute for FemA in staphylococcal peptidoglycan pentaglycine side chain formation; Ehlert K et al.; The femAB operon codes for two nearly identical approximately 50-kDa proteins involved in the formation of the staphylococcal pentaglycine interpeptide bridge . Sequencing and analysis of the femA region of mutants isolated by chemical mutagenesis and selection for lysostaphin resistance revealed point mutations leading to the expression of truncated FemA proteins . These femA mutants, although still producing an intact FemB, exhibited a phenotype identical as that described for femAB double mutants . Thus, FemA seems to be essential for the addition of glycine residues 2 and 3 only, whereas FemB is involved in the attachment of exclusively glycine residues 4 and 5 . Although FemB has 39% identity with FemA, it cannot substitute for FemA . The FemA and FemB proteins seem to be highly specific in regard to the position of the glycine residues that they attach.

Scand J Immunol, 1997 Nov, 46(5), 469 - 78
Increased mortality and impaired clonal deletion after staphylococcal enterotoxin B injection in old mice: relation to cytokines and nitric oxide production; Kuschnaroff LM et al.; In the present study peripheral T cell tolerance and the occurrence of shock were evaluated in young and old mice after injection of Staphylococcal enterotoxin B (SEB) . In young mice SEB immunization leads to tolerance based on deletion and anergy of SEB-reactive V beta 8+ T cells . With aging, mice developed resistance to SEB-induced deletion of V beta 8+ T cells as well as a high sensitivity to toxic shock . Compared to young mice, older mice injected with SEB showed increased serum levels of interferon-gamma (IFN-gamma), interleukin-2 (IL-2) and IL-4 . These results were confirmed by reverse transcription-polymerase chain reaction (RT-PCR), as splenic mRNA levels taken 2 h after SEB injection showed higher values of IL-2, IL-4, and IFN-gamma in old mice . In contrast, mRNA levels for FasL and tumour necrosis factor-alpha (TNF-alpha) were lower . No difference in IL-10 mRNA was found . Compared to young mice, old mice showed a high, but statistically not significantly different (P = 0.20), production of nitric oxide (NO) . Blocking of IFN-gamma with antibodies or reducing IFN-gamma by depletion of natural killer (NK) cells resulted, respectively, in a complete or partial protection against mortality in aged mice . Suppressing the NO production by the NO synthase inhibitor N-nitro-L-arginine methylester (L-NAME) increased the mortality in both young and old mice, and abrogated clonal deletion in the surviving young mice . In conclusion, in young mice NO production is a key factor in the protection against mortality and the development of clonal deletion after SEB injection . The higher mortality seen in older mice is mainly related to the elevated production of IFN-gamma and occurs despite a sufficient production of NO . The decreased clonal deletion of old mice may be related to their decreased expression of Fas ligand or TNF-alpha after SEB injection.

Scand J Immunol, 1997 Nov, 46(5), 459 - 68
Effect of treatments with cyclosporin A and anti-interferon-gamma antibodies on the mechanisms of immune tolerance in staphylococcal enterotoxin B primed mice; Kuschnaroff LM et al.; The authors were interested to investigate the effect of Cyclosporin A (CsA), known to block interleukin-2 (IL-2) production, or of anti-interferon-gamma antibodies (anti-IFN-gamma Abs) in a model of T cell tolerance induced by the injection of the superantigen Staphylococcal Enterotoxin B (SEB) in BALB/c mice . After SEB immunization, tolerance was mainly achieved through deletion and anergy of SEB-reactive V beta 8+ T cells . Association of CsA treatment with SEB led to a greater decrease of the percentage of V beta 8+ CD4+ lymphocytes in the spleen and an abolition of clonal energy . In contrast, treatment of SEB primed mice with anti-IFN-gamma Abs resulted in an increased percentage of V beta 8+ CD4+ cells without affecting the induction of clonal anergy . The authors found that 1-2 h after SEB priming, splenic mRNA levels of IFN-gamma and IL-4 were decreased by either CsA and anti-IFN-gamma Abs, whereas FasL, Bcl-2, p . 53, and c-myc levels were not influenced by either treatment . However, SEB-induced IL-2 and IL-10 mRNA expression was suppressed only by CsA, whereas tumour necrosis factor-alpha (TNF-alpha) was decreased only by anti-IFN-gamma Abs . To investigate whether the effect of CsA on the tolerance mechanisms was related to suppression of IL-2, CsA was administered together with recombinant IL-2 . Whereas anergy was not influenced, the decreased percentage of V beta 8+ CD4+ cells seen in CsA-treated animals in the second week after SEB injection was partially corrected by the administration of IL-2 . Experiments involving bromodeoxiuridine incorporation revealed that the latter effect of IL-2 was mainly due to a correction of the defective proliferation of V beta 8+ T cells after SEB injection in CsA-treated mice . These results suggest that the effect of CsA and anti-IFN-gamma Abs on tolerance mechanisms are in part explained by their action on cytokines.

Hua Xi Yi Ke Da Xue Xue Bao, 1996 Jun, 27(2), 182 - 4
{Improvement in model of mesangial proliferative glomerulonephritis of rat}; Liu Z et al.; To establish a model of mesangial proliferative glomerulonephritis (MsPGN), we gave Sprague-Dawley rats staphylococcal enterotoxin B (SEB) intravenously twice in different times, bovine serum albumine (BSA) through digestive tract every other day, and Freud's adjuvant subcutaneously twice in different times, respectively . The results showed that measangial cells and matrix in glomeruli of rats began to proliferate at the end of 6th week and this change became obvious at the end of 8th week . The duration of experiment was shorter than that using only SEB, and the change of pathomorphology was stable . The experiment confirms the thesis that mucosal immunization of gastroenteric tract plays an important role in the development of MsPGN.

J Biomol NMR, 1997 Sep, 10(2), 143 - 64
Solution structures of staphylococcal nuclease from multidimensional, multinuclear NMR: nuclease-H124L and its ternary complex with Ca2+ and thymidine-3',5'-bisphosphate; Wang J et al.; The solution structures of staphylococcal nuclease (nuclease) H124L and its ternary complex, (nuclease-H124L).pdTp.Ca2+, were determined by ab initio dynamic simulated annealing using 1925 NOE, 119 phi, 20 chi 1 and 112 hydrogen bond constraints for the free protein, and 2003 NOE, 118 phi, 20 chi 1 and 114 hydrogen bond constraints for the ternary complex . In both cases, the final structures display only small deviations from idealized covalent geometry . In structured regions, the overall root-mean-square deviations from mean atomic coordinates are 0.46 (+/- 0.05) A and 0.41 (+/- 0.05) A for the backbone heavy atoms of nuclease and its ternary complex, respectively . The backbone conformations of residues in the loop formed by Arg81-Gly86, which is adjacent to the active site, are more precisely defined in the ternary complex than in unligated nuclease . Also, the protein side chains that show NOEs and evidence for hydrogen bonds to pdTp (Arg35, Lys84, Tyr85, Arg87, Tyr113, and Tyr115) are better defined in the ternary complex . As has been observed previously in the X-ray structures of nuclease-WT, the binding of pdTp causes the backbone of Tyr113 to change from an extended to a left-handed alpha-helical conformation . The NMR structures reported here were compared with available X-ray structures: nuclease-H124L {Truckses et al . (1996) Protein Sci., 5, 1907-1916} and the ternary complex of wild-type staphylococcal nuclease {Loll and Lattman (1989) Proteins Struct . Funct . Genet., 5, 183-201} . Overall, the solution structures of nuclease-H124L are consistent with these crystal structures, but small differences were observed between the structures in the solution and crystal environments . These included differences in the conformations of certain side chains, a reduction in the extent of helix 1 in solution, and many fewer hydrogen bonds involving side chains in solution.

J Mol Biol, 1997 Oct 10, 272(5), 790 - 804
Contributions to protein entropy and heat capacity from bond vector motions measured by NMR spin relaxation; Yang D et al.; The backbone dynamics of both folded and unfolded states of staphylococcal nuclease (SNase) and the N-terminal SH3 domain from drk (drkN SH3) are studied at two different temperatures . A simple method for obtaining order parameters, describing the amplitudes of motion of bond vectors, from NMR relaxation measurements of both folded and unfolded proteins is presented and the data obtained for 15N-NH bond vectors in both the SNase and drkN SH3 systems analyzed with this approach . Using a recently developed theory relating the amplitude of bond vector motions to conformational entropy, the entropy change between the folded and unfolded forms of SNase is calculated on a per residue basis . It is noteworthy that the region of the molecule with the smallest entropy change includes those residues showing native-like structure in the unfolded form of the molecule, as established by NOE-based experiments . Order parameters of backbone 15N-NH bond vectors show significantly larger changes with temperature in the unfolded states of both proteins relative to the corresponding folded forms . The differential temperature dependence is interpreted in terms of differences in the heat capacities of folded and unfolded polypeptide chains . The contribution to the heat capacity of the unfolded chain from rapid 15N-NH bond vector motions is calculated and compared with estimates of the heat capacity of the backbone unit, -CHCONH-, obtained from calorimetric data . Methyl dynamics measured at 14 and 30 degrees C establish that the amplitudes of side-chain motions in the folded SH3 domain are more sensitive to changes in temperature than the backbone dynamics, suggesting that over this temperature range side-chain ps to ns time-scale motions contribute more to the heat capacity than backbone motions for this protein.

Biochem Biophys Res Commun, 1997 Oct 29, 239(3), 782 - 8
Fas/Fas ligand interaction regulates cytotoxicity of CD4+ T cells against staphylococcal enterotoxin B-pulsed endothelial cells; Urayama S et al.; Infiltration of activated CD4+ T cells and apoptosis of endothelial cells are present in the synovium of rheumatoid arthritis (RA) . Using staphylococcal enterotoxin B (SEB) as an antigen, we examined the possible role of antigen (Ag)-dependent activation of CD4+ T cells by endothelial cells, in inducing endothelial cell apoptosis . The human endothelial cell line, EA.hy926 cells, was cultured with or without interferon-gamma (IFN-gamma) and further incubated with CD4+ T cells in the presence or absence of SEB . After this cocultivation, the cytotoxicity and Fas ligand (FasL) expression of CD4+ T cells were examined . A small percentage of EA.hy926 cells expressed HLA-DR and -DQ, and this expression was significantly augmented after IFN-gamma stimulation . Anti-Fas IgM-induced apoptosis was exhibited by both unstimulated and IFN-gamma-stimulated EA.hy926 cells . Cytotoxicity of CD4+ T cells toward SEB-pulsed unstimulated EA.hy926 cells was detected . Furthermore, when CD4+ T cells were incubated with IFN-gamma-stimulated, SEB-pulsed EA.hy926 cells with augmented HLA-DR and -DQ expression, this cytotoxicity was more significant . The addition of anti-HLA-DR and -DQ monoclonal antibodies (mAbs) or human Fas chimeric protein (hFas-Fc) reduced the cytotoxicity . FasL expression was induced in CD4+ T cells cocultured with SEB-pulsed EA.hy926 cells, especially when the EA.hy926 cells were IFN-gamma-stimulated . Furthermore, the addition of mAbs against CD54 and CD58 inhibited both the cytotoxicity and FasL expression of CD4+ T cells induced by SEB-pulsed EA.hy926 cells, indicating the importance of costimulatory molecules on EA.hy926 cells in activating CD4+ T cells . Our results suggest that CD4+ T cells are activated by endothelial cells in an Ag-dependent manner and subsequently express FasL, which induces Fas-mediated apoptosis of endothelial cells . This phenomenon may counteract the growth of RA synovium by inhibiting the proliferation of endothelial cells.

J Immunol, 1997 Nov 15, 159(10), 4737 - 44
Impaired peripheral deletion of activated T cells in mice lacking the common cytokine receptor gamma-chain: defective Fas ligand expression in gamma-chain-deficient mice; Nakajima H et al.; Mice lacking the common cytokine receptor gamma-chain (gamma c) exhibit severely compromised lymphoid development . T cells that develop in these mice exhibit decreased Bcl-2 levels and accelerated apoptosis; nevertheless, these mice exhibit an age-dependent accumulation of activated CD4+ T cells . To investigate the basis for this accumulation, we analyzed both thymic and peripheral deletion in these mice . Gamma c-deficient mice had increased numbers of V beta 11+ T cells, consistent with a possible defect in Mtv-9-induced deletion; however, the deletion of V beta 5+ T cells by Mtv-9 and that of V beta 6+ T cells by Mls-1a were normal . Moreover, antigenic peptide could induce wild-type levels of deletion of CD4+ CD8+ thymocytes in TCR-transgenic gamma c-deficient mice . In contrast to this relatively normal deletion of thymocytes, bacterial superantigen (staphylococcal enterotoxin B)-induced elimination of peripheral T cells was greatly impaired, suggesting that defective peripheral deletion contributed to the accumulation of activated T cells . Interestingly, despite CD4+ T cell accumulation, these cells exhibited increased sensitivity to Fas-mediated death in vitro . Correction of the defect in Bcl-2 expression by mating to Bcl-2 transgenic mice augmented the splenic T cell accumulation and substantially enhanced the survival of gamma c-deficient T cells; however, these cells still exhibited significant Fas-mediated death, indicating that the increased Fas-mediated death was not simply due to diminished Bcl-2 expression . Moreover, these T cells exhibit decreased expression of Fas ligand, suggesting that Fas-bearing cells cannot be effectively eliminated in vivo in gamma c-deficient mice . Thus, gamma c-dependent signals play a role in peripheral T cell deletion, presumably by inducing Fas ligand on activated T cells.

Biosens Bioelectron, 1997, 12(7), 661 - 7
Detection of staphylococcal enterotoxin B employing a piezoelectric crystal immunosensor; Harteveld JL et al.; This paper reports on a study concerning the development of a biochemical sensor for Staphylococcal Enterotoxin B (SEB), a substance within the bracket of biological warfare . A 20 MHz piezoelectric quartz crystal sensor device was employed in a flow injection system . The assay for SEB is based on a competition scheme using polyclonal antibodies (anti-SEB) . Three parameters, i.e . the flow rate in the flow cell, the incubation time and the anti-SEB concentration, were optimized . A detection limit of 0.1 microgram/ml was obtained, whereas at concentrations of SEB of 10 micrograms/ml or higher the sensor response was completely inhibited . The results were compared with a competition enzyme linked immunosorbent assay.

Am J Med Sci, 1997 Nov, 314(5), 354 - 6
Emphysematous pyelonephritis in a nonfunctioning renal allograft of a patient undergoing hemodialysis; Goral S et al.; A hemodialysis patient with insulin-dependent diabetes mellitus and a non-functioning renal allograft in whom fever, low blood pressure, and confusion developed is reported . She underwent extensive evaluation and allograft nephrectomy for emphysematous pyelonephritis that was diagnosed by the presence of air in the collecting system of the transplanted kidney during computerized tomography of the abdomen . In nine patients with emphysematous pyelonephritis in renal allografts reported previously, this is the first instance of emphysematous pyelonephritis in a renal allograft with coagulase-negative staphylococcus.

Ann Vasc Surg, 1997 Nov, 11(6), 634 - 6
Mycotic aneurysm of the extracranial carotid artery: an uncommon complication of bacterial endocarditis; Hubaut JJ et al.; This report describes a case involving mycotic aneurysm of the extracranial internal carotid artery occurring as a complication of staphylococcal endocarditis in a patient with systemic lupus erythematosus . Three main points are emphasized: (1) this complication occurred in an immunodepressed patient; (2) surgical treatment consisted of aneurysmorraphy using absorbable suture; (3) the outcome was successful with a follow-up of 24 months.

Biosci Biotechnol Biochem, 1997 Oct, 61(10), 1786 - 8
Identification of the minimum segment essential for the H gamma II-specific function of staphylococcal gamma-hemolysin; Nariya H et al.; Staphylococcal gamma-hemolysin consists of H gamma I (or LukF) of 34 kDa and H gamma II of 32 kDa, which cooperatively lyse human erythrocytes . Our previous data showed that the N-terminal 57-residue segment of H gamma II is the essential region for the H gamma II function {H . Nariya and Y . Kamio, Biosci . Biotech . Biochem., 59, 1603-1604 (1995)} . To identify the minimum amino acid residues in the 57-residue segment responsible for the specific hemolytic activity, a series of mutant genes were constructed and expressed in Escherichia coli . The mutant proteins were purified and assayed for their hemolytic activity . The results indicate that the 5-residue segment (K23R24L25A26I27) of H gamma II is the minimum region essential for the H gamma II function.

Spine, 1997 Oct 15, 22(20), 2444 - 50; discussion 2450-1
Delayed infection after elective spinal instrumentation and fusion . A retrospective analysis of eight cases; Viola RW et al.; STUDY DESIGN: A retrospective analysis of eight cases of delayed spinal infection after elective posterior or combined anterior and posterior spinal instrumentation and fusion . OBJECTIVES: These cases are reviewed to identify risk factors for delayed spinal infection after elective instrumentation and to describe the treatment of this complication . SUMMARY OF BACKGROUND DATA: Delayed spinal infection after elective spinal instrumentation and fusion is uncommon . This diagnosis is frequently difficult . METHODS: Five cases seen in the senior author's practice and three referral cases are reviewed . RESULTS: Of these eight cases, the organisms were Staphylococcus epidermidis in six cases, Propionibacterium acnes in one cases, and in the final patient, all intraoperative cultures were negative . Clinical presentations were variable; however, all patients reported back pain . Seven patients had elevated erythrocyte sedimentation rates, averaging 57 mm/hour . Only two had elevated white blood cell counts . No distant foci of infection were identified in any patient . Five-patients were found to have at least one pseudarthrosis . All patients were treated with debridement, instrumentation removal, and primary wound closure over drains followed by a minimum 6-week course of culture-directed postoperative antibiotics . At an average follow-up of 18 months, no patient has evidence of infection . CONCLUSIONS: The diagnosis of delayed infection after elective spinal instrumentation and fusion requires a high index of suspicion . These infections may have been caused by intraoperative inoculation . All patients were successfully treated with debridement, instrumentation removal, and culture-directed postoperative antibiotics.

J Pharmacol Exp Ther, 1997 Nov, 283(2), 426 - 33
Atrial natriuretic peptide inhibits evoked catecholamine release by altering sensitivity to calcium; Kanwal S et al.; Natriuretic peptides are cyclized peptides produced by cardiovascular and neural tissues . These peptides inhibit various secretory responses such as the release of renin, aldosterone and autonomic neurotransmitters . This report tests the hypothesis that atrial natriuretic peptide reduces dopamine efflux from an adrenergic cell line, rat pheochromocytoma cells, by suppressing intracellular calcium concentrations . The L-type calcium channel inhibitor, nifedipine, markedly suppressed dopamine release from depolarized PC12 cells, suggesting that calcium entering through this channel was the predominant stimulus for dopamine efflux . Atrial natriuretic peptide maximally reduced depolarization-evoked dopamine release 20 +/- 3% at a concentration of 100 nM and this effect was abolished by nifedipine, but not by pretreatment with the N-type calcium channel inhibitor, omega-conotoxin, or an inhibitor of calcium-induced calcium release, ryanodine . In cells loaded with Fura-2, atrial natriuretic peptide both augmented depolarization-induced increases of intracellular free calcium concentrations and accelerated the depolarization-induced quenching of the Fura-2 signal by manganese, findings consistent with enhanced conductivity of calcium channels . Dopamine efflux induced by either the calcium ionophore, A23187, or staphylococcal alpha toxin was attenuated by atrial natriuretic peptide . Additionally, a natriuretic peptide interacting solely with the natriuretic peptide C receptor in these cells, C-type natriuretic peptide, also suppressed calcium-induced dopamine efflux in permeabilized cells . These data are consistent with natriuretic peptides attenuating catecholamine exocytosis in response to calcium but inconsistent with the neuromodulatory effect resulting from a reduction in intracellular calcium concentrations within pheochromocytoma cells.

Eur J Ophthalmol, 1997 Jul-Sep, 7(3), 241 - 4
Influence of optic and haptic materials on the adherence of Staphylococcus epidermidis to intraocular lenses: a pilot study; Prosdocimo G et al.; To evaluate in vitro the adherence of Staphylococcus epidermidis to intraocular lenses (IOL) of different optic and haptic materials and design, we used a quantitative cultural method . Polymethylmethacrylate (PMMA), PMMA-prolene, polyHEMA, silicone and surface-modified PMMA (wet and dry) implants were tested . Adherence differed significantly in the various groups, with the best performance by all-PMMA IOL.

Int Orthop, 1997, 21(4), 243 - 8
Subacute osteomyelitis presenting as a bone tumour . A review of 21 cases; Cottias P et al.; Twenty-one patients with subacute osteomyelitis who were initially considered to have bone tumours were reviewed, with an average follow up of 3 years . The clinical symptoms were not specific and laboratory investigations were normal . The radiographic findings were limited osteolysis surrounded by bone sclerosis in 14 cases, osteolysis without definite borders in 6, and onion-layer periosteal bone formation in one . The preoperative diagnoses included osteoid osteoma, osteosarcoma, chondroblastoma, Ewing's sarcoma, giant cell tumour, fibrosarcoma, eosinophilic granuloma, and bone tumour of unknown aetiology . The definitive diagnosis was made by surgical biopsy, histology and cultures which grew staphylococcus in 9 cases . The gross specimens all showed lymphocytes, plasma cells and granulation tissue with osteogenesis . All the patients recovered completely; 17 were treated with antibiotics and immobilisation, and 4 did not need an antibiotic . There was no recurrence of infection after curettage and excision of the infected tissues.

J Biol Chem, 1997 Oct 31, 272(44), 27796 - 803
Destabilization of peptide binding and interdomain communication by an E543K mutation in the bovine 70-kDa heat shock cognate protein, a molecular chaperone; Ha JH et al.; We have compared 70-kDa heat shock cognate protein (Hsc70) isolated from bovine brain with recombinant wild type protein and mutant E543K protein (previously studied as wild type in our laboratory) . Wild type bovine and recombinant protein differ by posttranslational modification of lysine 561 but interact similarly with a short peptide (fluorescein-labeled FYQLALT) and with denatured staphylococcal nuclease-(Delta135-149) . Mutation E543K results in 4 . 5-fold faster release of peptide and lower stability of complexes with staphylococcal nuclease-(Delta135-149) . ATP hydrolysis rates of the wild type proteins are enhanced 6-10-fold by the addition of peptide . The E543K mutant has a peptide-stimulated hydrolytic rate similar to that of wild type protein but a higher unstimulated rate, yielding a mere 2-fold enhancement . All three versions of Hsc70 possess similar ATP-dependent conformational shifts, and all show potassium ion dependence . These data support the following model: (i) in the presence of K+, Mg2+, and ATP, the peptide binding domain inhibits the ATPase; (ii) binding of peptide relieves this inhibition; and (iii) the E543K mutation significantly attenuates the inhibition by the peptide binding domain and destabilizes Hsc70-peptide complexes.

J Pediatr, 1997 Sep, 131(3), 466 - 9
Staphylococcal necrotizing fasciitis in the mammary region in childhood: a report of five cases; Bodemer C et al.; OBJECTIVE: Necrotizing fasciitis is a highly lethal soft tissue infection rarely reported in childhood . The initiating site is usually a local trauma or a surgical wound . We observed five cases of necrotizing fasciitis the initiating site for which was the mammary region and discuss their management . STUDY DESIGN: We describe these five patients and review the clinical characteristics of their presentation . RESULTS: Staphylococcal necrotizing fasciitis was observed in the mammary region in all five cases . Four children were newborn infants with a mammitis preceding the onset of necrotizing fasciitis . Surgical debridement was done only after the fourth day from onset of illness . All children were discharged in good condition after 1 month . Two have been followed until puberty, with destruction of the mammary gland in one case and good development in the other one . CONCLUSION: Mammitis may be the initiating event for necrotizing fasciitis in neonates . Necrotizing fasciitis is a life-threatening disease; patients require early intensive care, parenteral antibiotic therapy, and surgical debridement . In a few instances surgery can be carefully delayed until the necrotic area is more delineated if the condition is diagnosed early during disease evolution and appropriate treatment is instituted in intensive care units.

FEBS Lett, 1997 Sep 22, 415(1), 96 - 100
Identification of the minimum segment in which the threonine246 residue is a potential phosphorylated site by protein kinase A for the LukS-specific function of staphylococcal leukocidin; Nariya H et al.; Staphylococcal leukocidin and gamma-hemolysin consist of LukF and LukS for leukocidin and LukF and Hlg2 for gamma-hemolysin . In this report, we identify the minimum segment responsible for the LukS-specific function of leukocidin . After chemical analysis and homology study of the amino acid sequence of the C-terminal region between LukS and Hlg2, we found a unique 5-residue sequence I242K243R244S245T246 in LukS in which the 4-residue KRST is identical with that of the phosphorylated segment of a protein phosphorylated by protein kinase A . To elucidate whether the 5-residue segment is essential for the LukS function, we created plasmids containing a series of mutant genes corresponding to the 5-residue sequence and expressed them in Escherichia coli . The mutant proteins were purified and assayed for their leukocytolytic activity with LukF . The mutant MLS-TS, in which the T246 in the 5-residue sequence was replaced by S, showed leukocidin activity 10 times higher than that of the intact LukS . However, neither mutant MLS-TY nor MLS-TA, in which T246 was replaced by Y or A, respectively, showed leukocidin activity . The 5-residue segment was found to be deleted in Hlg2 . The mutant of Hlg2, in which the 5-residue segment was inserted at the position that the segment is deleted, showed leukocidin activity . The boiled LukS, MLS-TS, and MHS-Z were strongly phosphorylated with {gamma-32P}ATP in the presence of protein kinase A in a cell-free system . Thus, we conclude that the 5-residue segment 1242K243R244S245T246 is the pivotal segment of LukS responsible for the LukS function of staphylococcal leukocidin.

Int J Immunopharmacol, 1997 Mar, 19(3), 167 - 79
Spontaneous programmed cell death (PCD) process of lymphocytes of FIV-infected cats: pharmacological modulation in vitro; Guiot AL et al.; We previously reported that unstimulated lymphocytes in culture from FIV-infected cats undergo spontaneous apoptosis in vitro as indicated by internucleosomal DNA fragmentation and hypodiploid DNA content of nuclei . Unlike what is reported in HIV-infected individuals, we observed that cell death of cat lymphocytes was inhibited by activation . Spontaneous apoptosis was reduced by the addition of cat serum, interleukins {interleukin (IL)1, Il2, IL6 and interferon-gamma (IFN gamma)} and after activation by phorbol ester {phorbol myristyl acetate (PMA)}, superantigens {staphylococcal enterotoxin B (SEB), staphylococcal enterotoxin A (SEA)}, and to a lesser extent by mitogens such as Concanavalin A and pokeweed mitogen, IN contrast, apoptosis of lymphocytes from FIV-infected, but not from control cats was increased in the presence of calcium ionophore (ionomycin) . In this study, we studied the spontaneous programmed cell death (PCD)-inducing pathways, and the mechanisms of action of PMA, SEB and SEA . Spontaneous lymphocyte apoptosis of FIV-infected cats was inhibited by cycloheximide, ZnSO4 and N-acetyl-cystein . The preventive effect of SEB and SEA was inhibited by actinomycin, but not by inhibitors of kinases . Calyculin, an inhibitor of phosphatase, had no effect either on spontaneous apoptosis, or on the action of PMA, SEB and SEA . Ionomycin-induced apoptosis was found sensitive to PMA and cytokines . In FIV-infected cats, these data suggest that the mature lymphocytes appear programmed to die by apoptosis, unless rescued by specific agents, such as protein kinase C activators or growth factors, and that spontaneous PCD seems to be dependent of de nove protein synthesis (see effect of cycloheximide) . The effects of PMA, SEB and SEA are probably mediated by de novo proteins which for PMA, undergo a phosphorylation involving serine-threonine and/or tyrosine groups . Our data suggest a clear difference between lymphocytes from FIV-infected cats and lymphocytes from HIV-infected humans, with regard to their metabolic regulations.

Anaesthesiol Reanim, 1997, 22(3), 69 - 79
{Epiduritis after long-term pain therapy with an epidural catheter--review of the literature with a current case report}; Michel H et al.; Patients suffering from vascular disease are often a challenge for the acute pain service . Ischaemia, impaired wound healing, stump and phantom limb pain often require a complex analgesic regimen . Invasive measures such as spinal or epidural catheters can be very helpful but carry the risk of infection, as shown by this case report . A 53-year-old woman with a ten-year history of diabetes developed arterial vascular disease . Her right lower leg had been amputated two years previously . She was now admitted with necroses of the left forefoot . A bypass operation was performed under general anaesthesia . Because of intractable ischaemic pain, she was provided with an epidural catheter by the acute pain service . The bypass occluded, however, and a few days later her left lower leg also had to be amputated, this operation being performed under epidural anaesthesia with bupivacaine . The catheter was subsequently used for postoperative pain control and as a means to prevent phantom limb pain . When signs of superficial catheter infection were noticed days later, the catheter was immediately removed . Intractable pain then developed in the left leg which could not be sufficiently controlled with opioids and NSAIDs, and so a second epidural catheter was inserted one segment rostrally . Several days later the infected vascular prosthesis had to be removed followed by amputation of the thigh, this operation also being performed in epidural anaesthesia . Eleven days after insertion of the first epidural catheter, the patient complained of low back pain and headache . Examination by a neurologist revealed no signs of intraspinal infection . The second epidural catheter dislocated at this point in time and it was decided to introduce a third one, this being the only means to treat the otherwise intractable stump pain . Ten days later meningism, Kernig's sign and leucocytosis developed . NMR tomography detected intraspinal fluid in the epidural space at the dorsal border of the spinal canal . A hemilaminectomy was performed . The spinal epidural space showed signs of inflammation of the adipose tissue, but no pus . A little necrotic material and residues of an old haematoma were removed and the epidural space was lavaged . Specimens taken from the epidural material revealed colonisation with staphylococcus epidermidis, which was sensitive to the broad spectrum antibiotics formerly given to the patient to treat the infection in the left stump . By the next day, all signs of epiduritis had disappeared and the patient recovered completely.

J Vet Med Sci, 1997 Sep, 59(9), 775 - 83
Phylogenetic analyses of Staphylococcus based on the 16S rDNA sequence and assignment of clinical isolates from animals; Takahashi T et al.; The nucleotide sequences of the 16S rDNA in 17 strains of 16 taxa of the genus Staphylococcus were determined . The sequences were compared phylogenetically together with the gene sequences of 10 (including 7 other species) Staphylococcus species retrieved from the DNA Data Bank of Japan . Although the primary and secondary structures of most of Staphylococcus species were very similar (homology values 96.4% or more) except for S . caseolyticus MAFF 911387T (homology values 95.4% or less), the 23 staphylococcal species were divided into 10 groups based on similarity, evolutionary distance and phylogenetic tree analysis . Nucleotide stretches in several variable domains in the 16S rDNA sequences appeared to be specific for the bacterial groups or the species . By comparing such characteristics in the sequence and phylogenies of 5 staphylococcal clinical isolates from bovine mastitis, canine and feline pyoderma, and feline urogenital syndrome with the information obtained in this study, the species level of each organism was identified.

Biochemistry, 1997 Oct 28, 36(43), 13298 - 304
Staphylococcal alpha-toxin: formation of the heptameric pore is partially cooperative and proceeds through multiple intermediate stages; Valeva A et al.; Staphylococcal alpha-toxin is a 293 residue polypeptide that assembles into pore-forming heptamers, residues 118-140, thereby inserting to form an amphipathic beta-barrel in the lipid bilayer . Fluorometric analyses were here conducted using cysteine-substitution mutants site-specifically-labeled at positions 35 or 130 with the environmentally-sensitive fluorophore acrylodan . In conjunction with functional assays, three conformational states of the heptamer were defined, which may represent transitional configurations of the toxin molecule along its way to membrane insertion and pore formation . The first was the freshly assembled, SDS-sensitive heptamer alpha7*a, where a minor alteration in the environment of H35 with no change in the environment of the membrane-inserting stem domain was observed . In transition stage alpha7*b, the stem domain moved from a hydrophilic to a more hydrophobic environment, due to protein-protein interaction . Transition to alpha7*c involved a cooperative effect, in which residue 35 was forced by a neighboring molecule into a markedly hydrophobic environment . At this stage, the heptamers acquired SDS stability . The final pore conformation alpha7 resulted when the stem domain inserted into the lipid bilayer, an event that was driven by H35 within the respective protomer . A model thus evolved in which cooperative forces first lever H35 into a position that subsequently drives the pore-forming sequence within each respective protomer into the membrane . In accord with this model, when hybrid heptamers were formed between a functionally defective H35 substitution mutant and active toxin, only the latter inserted their pore-forming domain into the membrane . In a satisfying functional correlate, pores of reduced size were then generated.

Zh Mikrobiol Epidemiol Immunobiol, 1997 Jul-Aug, (4), 81 - 4
{The role of Staphylococcus epidermidis persistence factors in the infectious process}; Chernova OL et al.; The species composition of microflora was determined and the etiological role of S . epidermidis was shown in some purulent inflammatory diseases (maxillary sinusitis, urethritis) . The role of the complex of S . epidermidis persistence factors in the chronization of the infectious process and the formation of the resident carrier state was established.

Inflamm Res, 1997 Sep, 46(9), 361 - 5
The pharmacological profile of mouse hind paw inflammation induced by staphylococcal enterotoxin type A; Desouza IA et al.; OBJECTIVE AND DESIGN: The present paper examines the possible pharmacological mediators involved in mouse hind paw inflammation induced by staphylococcal enterotoxin type A (SEA) . MATERIALS AND METHODS: The edema and the Evans blue exudation were measured in male Swiss mice (20-25 g) using methods described by Levy and Griswold, respectively . RESULTS: SEA (32 micrograms/paw) produced a biphasic, long-lasting, dose- and time-dependent edematogenic response . The acute phase edema was pronounced while the chronic edema was of a low intensity . Exudate was the principal component of the edema . The edematogenic effect of SEA appears to involve cyclooxygenase products and was dose-dependently reduced by pretreating the mice with dexamethasone, indomethacin, BW755C, WEB2086, capsaicin, diphenhydramine or cimetidine . CONCLUSIONS: These results demonstrate that SEA-induced mouse hind paw inflammation is a useful model for studying SEA-mediated enterotoxemia and may be sufficiently sensitive to differentiate between the effects of SEA and those of SE type B (SEB).

Rev Prat, 1997 Sep 1, 47(13), 1447 - 51
{Diagnosis of neonatal eruptions}; Wallach D; The aetiological diagnosis of a skin rash in a neonate requires an accurate clinical analysis . Smear examination of blister fluid is sometimes necessary . Macular-papular eruptions include erythema toxicum and generalized infections, viral as well as bacterial . Pustular eruptions include benign transient dermatoses and infections . Bullous eruptions include staphylococcal toxic syndromes and hereditary epidermolysis bullosa . So, whatever the clinical appearance of the rash, the main problem is to document an infection and to implement early treatment.

Reg Anesth, 1997 Sep-Oct, 22(5), 428 - 31
Culture of bacteria from lumbar and caudal epidural catheters used for postoperative analgesia in children; McNeely JK et al.; BACKGROUND AND OBJECTIVES: Continuous epidural analgesia has been used with increasing frequency to provide postoperative pain relief for children . Epidural space infection is a potential complication of epidural catheter placement . This study investigated the incidence of bacterial colonization on lumbar and caudal epidural catheter tips in postoperative pediatric patients . METHODS: In this prospective study, lumbar and caudal epidural catheters were placed in the operating room with aseptic technique . Dilute local anesthetic and/or opioid infusions were used for postoperative analgesia . On discontinuation of the epidural infusion, the skin site was decontaminated with 70% alcohol and then cultured . The distal catheter tip and hub were cultured . Semiquantitative and qualitative aerobic cultures were performed . RESULTS: Data from 91 epidural catheters were available (45 caudal versus 46 lumbar) . Of the 45 caudal catheter tips 9 (20%) were colonized, compared with 2 of the 46 (4%) lumbar catheter tips (P < .02) . Staphylococcus epidermidis was the predominant skin and catheter tip organism isolated in both groups . Four of nine caudal catheter tips grew gram-negative bacteria . Statistical analyses did not show that time, skin site inflammation, or dressing condition were independent predictors of catheter tip colonization . No patient developed a clinical epidural infection during the study period . CONCLUSIONS: The results of this study suggest that the risk of clinical epidural infection associated with caudal or lumbar postoperative catheters is low . However, the incidence of epidural catheter tip colonization is increased with the caudal route of insertion, and the bacteria differ from those cultured from the lumbar insertion site.

Vet Immunol Immunopathol, 1997 Sep, 58(2), 107 - 20
Aspects of the humoral immune response to Staphylococcus intermedius in dogs with superficial pyoderma, deep pyoderma and anal furunculosis; Shearer DH et al.; Bacterial infection (pyoderma) of the canine skin is largely caused by Staphylococcus intermedius and may be a superficial or deep infection . Pyoderma may be a primary, idiopathic disease or secondary to a range of other dermatological disorders . In this study, the serum concentrations of IgG, IgA, antistaphylococcal IgG and antistaphylococcal IgA were measured by ELISA in normal dogs (n = 22), dogs with idiopathic deep pyoderma (n = 22), atopic dermatitis and superficial pyoderma (n = 24), atopic dermatitis without pyoderma (n = 25), flea bite dermatitis with superficial pyoderma (n = 8), pustular demodicosis (n = 8) and German shepherd dogs with anal furunculosis (n = 28) . The serum IgG was significantly increased in dogs with atopy and superficial pyoderma (p < 0.001), and lower than normal in dogs with idiopathic deep pyoderma (p < 0.015) . The concentration of serum IgA was significantly lower than normal in dogs with atopy uncomplicated by pyoderma (p < 0.015) . The concentration of antistaphylococcal IgG in all clinical sera was significantly elevated (p < 0.001) when compared to normal dogs but concentrations of antistaphylococcal IgA were no greater than in normal dogs . Western blotting analysis for determination of the specificity of serum IgG antistaphylococcal antibody revealed that there were nine major epitopes . Discriminant analysis demonstrated that particular combinations of these epitopes were recognised more frequently by sera from dogs in different clinical groups.

Can Vet J, 1997 Oct, 38(10), 632 - 6
A benefit to cost analysis of the effect of premilking teat hygiene on somatic cell count and intramammary infections in a commercial dairy herd; Ruegg PL et al.; A field trial was conducted to determine the effect of premilking teat disinfection (predipping) on several measures of mastitis in a commercial dairy farm where the predominant organisms isolated from intramammary infections were coagulase negative Staphylococcus spp . Cows were randomly assigned to a treated (predipped with 0.5% iodine germicide plus "good udder preparation") or a control group ("good udder preparation" alone) . Sterile composite milk samples were collected at the initiation of the trial and on an approximately bimonthly basis throughout the duration of the trial . There was no difference in the prevalence of isolation of coagulase-negative Staphylococcus spp . from composite milk samples obtained during the 6 herd cultures . The incidence rate for clinical mastitis in the control group was 1.38 cases per 1000 cow days . The incidence rate for clinical mastitis in the treatment group was 1.06 cases per 1000 cow days . The ratio of these 2 was 1.3, suggesting a higher rate in the control group, but the ratio was not statistically significant (P = 0.34) . Logistic regression analysis indicated that the effect of treatment group was not significant, although the coefficient suggested that predipping reduced the risk of clinical mastitis . The benefit to cost ratio of 0.37 indicated that the benefit of reduced incidence of clinical cases of mastitis would not have justified the added expense required to predip the herd.

Arq Neuropsiquiatr, 1997 Mar, 55(1), 75 - 81
{Infections of cerebrospinal fluid shunts in children . Review of 100 infections in 87 children}; Martins R et al.; An analysis of 100 infections in 87 children treated with shunts in the period of 1982 to 1995 is reported . The clinical presentation has been more frequently secondary to inflammatory signals . Staphylococcus were the most frequently microorganisms found . Infection by Gram negative agents was more aggressive and directly related with failure of therapy . Treatment included since only systemic antibiotics until withdrawal of shunt with use of systemic and intrathecal antibiotics . The best therapeutic results were obtained with withdrawal of shunt system and replacement by external shunt system associated to systemic antibiotics . In our experience this management must be accepted for treatment of this severe complication.

Adv Exp Med Biol, 1997, 418, 805 - 7
Erythrogenic toxin-induced arteritis in a rabbit ear model . Comparison with Arthus reaction angiitis; Abe Y et al.; We have developed a local type experimental model of angiitisin rabbits . Repeated intracutaneous injections of erythrogenic toxins types A and C (ETA, ETC) along the intermediate auricular artery of the rabbit ear produced subacute type arteritis, characteristic of lymphocyticinfiltration, simulating Kawasaki disease angiitis . Staphylococcal enterotoxin B (SEB) and toxic shock syndrome toxin 1 (TSST) also induced similar lesions, indicating superantigens as inflammogens in vivo . Conversely, the Arthus reaction included acute type angiitis when tested similarly in rabbits immunized to human serum albumin . The ear artery was infiltrated by heterophil leukocytes, often together with venules and capillaries affected, resembling periarteritis nodosa and leukoclastic vasculitis in human disease.

J Bone Joint Surg Br, 1997 Sep, 79(5), 816 - 9
The prevention of prosthetic infection using a cross-linked albumin coating in a rabbit model; An YH et al.; We evaluated the effects of a serum protein coating on prosthetic infection in 29 adult male rabbits divided into three groups: control, albumin-coated and uncoated . We used 34 grit-blasted, commercially pure titanium implants . Eleven were coated with cross-linked albumin . All the implants were exposed to a suspension of Staphylococcus epidermidis before implantation . Our findings showed that albumin-coated implants had a much lower infection rate (27%) than the uncoated implants (62%) . This may be a useful method of reducing the infection of prostheses.

Curr Eye Res, 1997 Oct, 16(10), 1036 - 43
Immunopathologic features of Staphylococcus epidermidis-induced endophthalmitis in the rat; Ravindranath RM et al.; PURPOSE: To investigate the clinical, histopathologic and immunologic responses to Staphylococcus epidermidis endophthalmitis in a rat model . METHODS: Experimental rats received an intravitreal injection of viable S . epidermidis (7000 organisms), while control rats received sterile saline . The clinical scores, cellular infiltrate in vitreous, and levels of serum and vitreous IgM, IgG and IgA to glycerol teichoic acid (GTA), the major antigenic determinant of S . epidermidis cell wall, were all measured from day 1 to day 30 after injection . RESULTS: The ocular inflammation was largely resolved by day 14 . The red reflex was abolished in 50% of rats between days 3 and 10 . The bacteria were cleared from the vitreous by day 7 . In vitreous, the neutrophils peaked at day 1 and decreased by day 7, and plasma cells were seen between days 1 and 3 . Presence of B cells (CD45+/CD3-) was confirmed by flow cytometric analysis of pooled vitreous humor . IgM and IgG but not IgA antibodies to GTA were found in vitreous of injected eyes . The peak of anti-GTA IgM was observed in vitreous of S . epidermidis-infected rats on day 1 and declined by day 7 . In contrast to vitreous antibodies, serum anti-GTA IgM antibodies were significantly elevated throughout the course of S . epidermidis endophthalmitis . A weak IgG but no IgA response were observed in serum . Anti-GTA antibodies were also found in low level in normal sera but not in normal vitreous . CONCLUSIONS: The vitreous antibodies may be involved in neutrophil-mediated opsonophagocytosis leading to 'spontaneous sterility' of the bacteria, and may play a role in the immunopathogenesis of staphylococcal endophthalmitis in the rat.

Adv Exp Med Biol, 1997, 421, 247 - 51
Expression of several matrix metalloproteinase genes in human monocytic cells; Machein U et al.; Matrix metalloproteinases (MMP) are a family of structurally related endopeptidases that resorb macromolecules of the extracellular matrix (ECM) . They are involved in normal tissue remodeling and wound repair as well as in pathological processes such as the irreversible destruction of joints observed in rheumatoid arthritis (RA) . In addition, MMP catalyze the cleavage of the transmembrane form of tumor necrosis factor (TNF) . Since cells of the monocyte lineage are major producers of TNF in the rheumatoid synovium we analysed the expression of MMP genes in these cells . To examine the transcriptional activity of MMP genes in undifferentiated monocytic cell lines (MonoMac6, U937) and in nature human monocytes isolated from peripheral blood, we developed an assay that is based on reverse transcription (RT) followed by a polymerase chain reaction (PCR) . This screening procedure demonstrates that several MMP genes are transcriptionally active in the cells tested after exposure to a variety of stimuli such as phorbol ester, lipopolysaccharide (LPS) and staphylococcal enterotoxin B (SEB) . The data were confirmed by quantitative Northern blot analysis . In conclusion, cells of the monocyte lineage produce high mRNA levels of at least six members of the MMP gene family that could participate in joint destruction by resorption of the ECM and secretion of TNF.

J Heart Valve Dis, 1997 Sep, 6(5), 550 - 2
Repair of left ventriculo-atrial fistulas due to posterior mitral annular abscesses; Baumgartner F et al.; Two cases of acute staphylococcal mitral annular abscesses resulting in transannular left ventriculoatrial fistulation are reported . Mitral annular abscess debridement and repair of the annulus, obliteration of the fistulation, and preservation of the native valve were successful in both cases.

Proc Natl Acad Sci U S A, 1997 Oct 14, 94(21), 11607 - 11
Transmembrane beta-barrel of staphylococcal alpha-toxin forms in sensitive but not in resistant cells; Valeva A et al.; Staphylococcal alpha-toxin is a 293-residue, single-chain polypeptide that spontaneously assembles into a heptameric pore in target cell membranes . To identify the pore-forming domain, substitution mutants have been produced in which single cysteine residues were introduced throughout the toxin molecule . By attaching the environmentally sensitive dye acrylodan to the sulfhydryl groups, the environment of individual amino acid side chains could be probed . In liposomes, a single 23-amino acid sequence (residues 118-140) was found to move from a polar to a nonpolar environment, indicating that this sequence forms the walls of the pore . However, periodicity in side chain environmental polarity could not be detected in the liposomal system . In the present study, the fluorimetric analyses were extended to physiological target cells . With susceptible cells such as rabbit erythrocytes and human lymphocytes, the 23 central amino acids 118-140 were again found to insert into the membrane; in contrast to the previous study with liposomes, the expected periodicity was now detected . Thus, every other residue in the sequence 126-140 entered a nonpolar environment in a striking display of an amphipathic transmembrane beta-barrel . In contrast, human granulocytes were found to bind alpha-toxin to a similar extent as lymphocytes, but the heptamers forming on these cells failed to insert their pore-forming domain into the membrane . As a consequence, nonfunctional heptamers assembled and the cells remained viable . The data resolve the molecular organization of a pore-forming toxin domain in living cells and reveal that resistant cells can prevent insertion of the functional domain into the bilayer.

Infect Immun, 1997 Oct, 65(10), 4190 - 8
Allelic polymorphisms at the H-2A and HLA-DQ loci influence the response of murine lymphocytes to the Mycoplasma arthritidis superantigen MAM; Cole BC et al.; Mycoplasma arthritidis, an agent of rodent arthritis, produces a potent superantigen (SAg), MAM . Previous work established that MAM is presented to T cells by murine H-2E or the homologous human HLA-DR molecules and that lymphocytes lacking a functional H-2E molecule fail to respond to MAM . Recently, more potent and purified preparations of MAM of known protein content have become available . This enabled us to more effectively compare the response of MAM with that of other SAgs by using lymphocytes from mice whose cells express different H-2A and HLA-DQ molecules . Here we demonstrate that cells from some H-2E-negative mouse strains respond to higher concentrations of MAM . By use of inbred, congenic, and recombinant mice, we show that these differences are, in fact, exercised at the level of the major histocompatibility complex (MHC) and that allelic polymorphisms at H-2A influence reactivity to MAM . In addition, polymorphisms at HLA-DQ, the human homolog of H-2A, also influence responsiveness to MAM . Cells expressing DQw6 (HLA-DQA1*0103 and DQBI*0601 chains) gave much higher responses to MAM than did cells expressing DQw8 (DQA1*0301 and DQB1*0302 chains) . In fact, responses of lymphocytes expressing DQB1*0601 chains homozygously were as high as those observed for cells expressing a functional H-2E molecule . Murine lymphocytes responded less well to staphylococcal enterotoxin B (SEB) and SEA, but mouse cells expressing human MHC molecules gave much higher responses . The patterns of reactivity observed with cells expressing the various murine and human alleles differed for MAM, SEB, and SEA, suggesting that each of these SAgs interacts with different regions or residues on MHC molecules . It has been hypothesized that SAgs might play a role in susceptibility to autoimmune disease . Allelic polymorphisms at MHC loci might therefore influence susceptibility to autoimmune disease by affecting immunoreactivity to specific superantigens.

J Clin Microbiol, 1997 Oct, 35(10), 2537 - 41
Human isolates of Staphylococcus caprae: association with bone and joint infections; Shuttleworth R et al.; Staphylococcus caprae is a coagulase-negative, DNase-positive member of the genus Staphylococcus usually associated with goats, but since 1991 a few laboratories have reported isolating the organism from human clinical specimens . We report on the isolation of 14 strains from human specimens and note that 10 strains were obtained from patients with bone and joint infections . Nine of the 10 infections started with traumatic fractures, and the other was a case of mastoiditis . Seven of these 10 infections were in patients with orthopedic prostheses, which appears to be a risk factor . Three of the 14 strains were from transplant patients . For three of the patients, S . caprae was the only organism isolated . S . caprae may be misidentified because it is not represented in the current MicroScan or Vitek identification systems which are in use in many laboratories, but the organism can be differentiated by a few biochemical tests . S . caprae produces positive results for DNase, pyrrolidonyl aminopeptidase, and acid production from mannitol and maltose; it produces negative results for ornithine decarboxylase and tube coagulase.

Am J Orthop, 1997 Sep, 26(9), 598 - 600
Midterm results in metal-backed patella revisions for metallosis; Arredondo J et al.; Nine patients underwent isolated patellar revisions and synovectomies for metallosis secondary to the mechanical failure of metal-backed patellar components . These patients were followed for an average of 5 years (range, 2 to 8 years) . In all cases, the femoral and tibial components were left in place, and in one patient the patellar bone was deemed too thin to resurface . Two patients (22%) developed deep infection . One infection (enterococcal) required arthrodesis, and the other (staphylococcal) was successfully managed with a two-staged reimplantation . A third patient sustained an inferior patella pole fracture that was treated conservatively . The remaining six patients are functioning satisfactorily . Metallosis is a serious complication in knee arthroplasty, and these patients merit close follow-up.

Biochemistry, 1997 Oct 7, 36(40), 12167 - 74
Stability studies of amino acid substitutions at tyrosine 27 of the staphylococcal nuclease beta-barrel; Bhat MG et al.; In order to help determine the extent to which side chain interactions within the staphylococcal nuclease beta-barrel affect its global stability, a full set of point mutants was generated for residue 27 . Intrinsic tryptophan fluorescence was monitored during solvent denaturation with guanidine hydrochloride (GuHCl) and was used to calculate DeltaGH2O unfolding and m values for each mutant . In the wild type protein, residue 27 is a tyrosine which is at the first position of a type I' beta-turn, and which participates in both hydrophobic interactions and side chain to side chain hydrogen bonding . The hydrophobicity of the mutant residue was found to be the dominant factor in determining global protein stability within this series of nuclease mutants.

Biomed Environ Sci, 1997 Sep, 10(2-3), 253 - 9
Ebselen--an in vivo immune response modifier; Wendel A et al.; Numerous biochemical activities are known for the selenoorganic drug ebselen {(2-phenyl-1,2-benzoisoselenazol-3-(2H)-one} . The initial interest focussed on its GSH peroxidase-like activity and its inhibitory activity on 5-lipoxygenase, however, further activities of the drug on oxidative burst of leukocytes, nitric oxide synthases, protein kinases and on leukocyte migration were recognized . Here we report that Ebselen differentially interacts in vivo with the production and action of systemically released cytokines in various hyperinflammation models in mice . Ebselen given orally to mice protected dose-dependently from Concanavalin A-induced, T-cell dependent inflammatory liver injury . The release of the proinflammatory cytokines Tumor Necosis Factor alpha and Interferon gamma was downregulated, while the production of the anti-inflammatory cytokine Interleukin 10 was increased . Similar results were found in galactosamine-sensitized mice in an inflammatory liver model using the superantigen staphylococcal enterotoxin B as a T-cell activator . Moreover, Ebselen inhibited the release of TNF alpha initiated by endotoxin in vivo in mice . In galactosamine-pretreated mice . Ebselen also attenuated liver injury induced by recombinant Tumor Necosis Factor alpha and initiated enhanced release of Interleukin 10 . These findings expand the pharmacological knowledge on ebselen to hitherto unknown immunomodulatory properties which encourage to develop the drug for treatment of T-cell related autoimmune diseases.

Scand J Immunol, 1997 Sep, 46(3), 262 - 7
The alpha beta T cell receptor clustering upon superantigen/MHC recognition; Mazda O et al.; Antigen recognition by T cells is the key event for the antigen specific immune responses to be triggered . This recognition is initiated by the binding of the T cell receptor (TCR) to antigen peptide/major histocompatibility complex (MHC) on the surface of the antigen presenting cells . TCR on most of the T cells is a heterodimer composed of alpha and beta chains which are associated with CD3 gamma delta epsilon as well as zeta chains, the signal transmission molecules . The dynamics of this TCR complex upon antigen/MHC recognition, however, has not been well understood . In this paper the authors analyse the configuration of TCR complex on T cells from a TCR beta chain gene transgenic mouse (TGM) strain . Unlike many other TGM strains reported, a considerable proportion of T cells from this TGM expresses both transgene-encoded (V beta 3) and endogenous TCR beta chains on their surface . By immunoprecipitation and immunoblotting analysis of T cells stimulated with a superantigen, staphylococcal enterotoxin B (SEB), the authors found that V beta 3 was coprecipitated with V beta 8, demonstrating the clustering of TCR alpha beta upon superantigen/MHC recognition.

Scand J Immunol, 1997 Sep, 46(3), 230 - 4
A unique response to staphylococcal enterotoxin B by intrahepatic lymphocytes and its relevance to the induction of tolerance in the liver; Matsui S et al.; It has been reported that the intrahepatic lymphocyte (IHL) population is somewhat differently constituted from lymphocytes in other lymphoid tissues . Staphylococcal enterotoxin B (SEB) is a superantigen which can induce T-cell tolerance in mice . The authors investigated the in vitro and in vivo responses of mouse IHL to SEB . An intravenous injection of SEB did not result in the augmentation of the proliferative response of IHL, while mesenteric and splenic lymphocytes (mLNC and SPC, respectively) had augmented responses . Interleukin-2 (IL-2) mRNA was clearly detected in mLNC and SPC by reverse transcriptase-polymerase chain reaction (RT-PCR) shortly after the administration of SEB, but it was scarcely expressed in IHL . The expression of CD25 (IL-2 receptor) was also augmented in mLNC and SPC in the early period, while it was not changed in IHL . These findings suggested that the time required for tolerance induction is different locally and that the loss of augmentation of IL-2 and IL-2 receptor production by IHL may be relevant to the rapid induction of T-cell tolerance in the liver.

Biol Reprod, 1997 Oct, 57(4), 887 - 93
Bovine luteal cells elicit major histocompatibility complex class II-dependent T-cell proliferation; Petroff M et al.; Major histocompatibility complex (MHC) class II molecules are expressed in the bovine corpus luteum (CL) in a manner correlating with luteolysis . Whether bovine luteal cells can stimulate T-cell proliferation in a class II-restricted manner was investigated . Staphylococcal enterotoxin B (SEB) enhances T-cell proliferation by a mechanism requiring MHC class II molecules and was used to examine stimulation of T-cell proliferation by luteal cells . Luteal cells from midcycle or regressing CL (induced by prostaglandin F2 alpha) were cocultured with autologous T cells in the presence of no treatment, SEB (1 microgram/ml), or SEB + anti-MHC class II antibody (3 micrograms/ml); and proliferation was assessed by incorporation of tritiated thymidine . T cells proliferated in the presence of cells from regressing CL more than when in the presence of midcycle cells (118,309 +/- 20,567 vs . 75,261 +/- 12,494 cpm; p < 0.05) . Anti-MHC attenuated this response of cells from regressing CL (81,108 cpm +/- 13,249; p < 0.05) . Without SEB, T cells proliferated when cultured with cells from regressing, but not midcycle, CL (4637 +/- 816 vs . 2117 +/- 589 cpm; p < 0.03) . These results suggest that luteal cells can function as antigen-presenting cells in vitro and that prostaglandin F2 alpha may enhance their ability to present antigen.

Clin Infect Dis, 1997 Sep, 25(3), 733 - 5
Cerebrospinal fluid (CSF) pharmacokinetics of intraventricular vancomycin in patients with staphylococcal ventriculitis associated with external CSF drainage; Pfausler B et al.; We studied the efficacy and pharmacokinetics of intraventricularly administered vancomycin in three patients with shunt-associated staphylococcal ventriculitis . We instilled 10 mg of the drug intraventricularly every 24 hours . Cerebrospinal fluid (CSF) levels were measured 1 hour after instillation and then every 2 hours . Peak vancomycin levels reached a mean of 292.9 microg/mL . The mean trough levels, measured immediately before readministration of vancomycin, were 7.6 microg/mL; this level has proved to be sufficient for maintaining the necessary steady-state serum concentration of vancomycin . All three patients were cured clinically and bacteriologically, and CSF parameters returned to normal within 5-13 days . No side effects were observed . Our results suggest that intraventricularly administered vancomycin is a valuable therapeutic strategy for treating shunt-associated staphylococcal ventriculitis . In addition, we provide evidence that 10 mg of vancomycin, administered intraventricularly every 24 hours, allows maintenance of therapeutic drug levels in the CSF for at least 24 hours.

J Virol, 1997 Oct, 71(10), 7560 - 6
Abrogation of in vitro suppression of human immunodeficiency virus type 1 (HIV-1) replication mediated by CD8+ T lymphocytes of asymptomatic HIV-1 carriers by staphylococcal enterotoxin B and phorbol esters through induction of tumor necrosis factor alpha; Kubo M et al.; CD8+ T lymphocytes of asymptomatic human immunodeficiency virus type 1 (HIV-1) carriers (AC) suppress HIV-1 replication in vitro . Failure of host defense mechanisms and increased virus proliferation are associated with disease progression . The exact mechanisms inducing these changes at the advanced stage of the disease are still obscure . In this study, we searched for experimental conditions favoring the abrogation of the suppression of viral replication in peripheral blood mononuclear cells (PBMC) of AC by using various pharmacological and biological probes modifying cell activation . Among such agents, staphylococcal enterotoxin B (SEB) and phorbol 12-myristate 13-acetate (PMA) markedly increased otherwise low levels of HIV-1 replication in cultures of phytohemagglutinin-stimulated AC PBMC following in vitro HIV-1 LAI infection . A similar but less pronounced virus induction was also observed in macrophage-tropic HIV-1 . Individual pretreatment of CD4+ and CD8+ PBMC fractions with these agents caused a reduction in CD8+ cell proliferation and enhanced HIV-1 replication in CD4+ cells . SEB- and PMA-mediated augmentation of HIV-1 replication in AC PBMC was significantly blocked by neutralizing antibody to tumor necrosis factor-alpha (TNF-alpha), although recombinant TNF-alpha alone failed to reproduce the effects of SEB or PMA . Our results suggest that the induction of TNF-alpha may be one of the mechanisms that overcomes the CD8+-induced suppression of HIV-1 replication in AC and that it may induce HIV-1 replication.

Int J Food Microbiol, 1997 Jul 22, 37(2-3), 237 - 40
Method for the rapid quantitative detection of lipolytic activity among food fermenting microorganisms; Leuschner RG et al.; A standard method for the detection of free fatty acids (FFAs) in milk was modified and applied to the measurement of the lipolytic activity of microorganisms in a model system containing either homogenised pork or beef fat tissue . The increase in FFAs was measured colorimetrically using palmitic acid as a standard . Among the strains tested, two strains of Staphylococcus xylosus and one strain of Staphylococcus carnosus were found to display lipolytic activity . For all strains, a higher increase in FFA was observed in broth supplemented with pork fat than with beef fat . All three strains displayed lipolytic activity when tested on tributyrin agar plates.

Int Immunol, 1997 Sep, 9(9), 1393 - 403
Bacterial superantigens reactivate antigen-specific CD8+ memory T cells; Coppola MA et al.; Superantigens stimulate naive CD4+ and CD8+ T cells in a TCR V beta-specific manner . However, it has been reported that memory T cells are unresponsive to superantigen stimulation . In this study, we show that staphylococcal enterotoxins (SE) can activate influenza virus-specific CD8+ memory cytotoxic T cells . In vivo SEB challenge of mice that had recovered from influenza virus infection (memory mice) resulted in the generation of vigorous influenza-specific cytotoxic T lymphocyte (CTL) activity and in vitro SEA or SEB stimulation of splenic T cells from memory mice, but not naive mice, also induced influenza-specific CTL . Analysis of the mechanism of activation suggested that although there may be a component of cytokine-mediated bystander activation, the CTL activity is largely generated in response to direct TCR engagement by superantigen . Moreover, influenza-specific CTL could be generated from purified CD8+ CD62L loCD44hi (memory phenotype) T cells cultured in the presence of T cell-depleted splenic antigen-presenting cells and SE . Purified CD8+ memory T cells also secreted lymphokines and synthesized DNA in response to superantigen . These results definitively demonstrate that CD8+ memory T cells respond to SE stimulation by proliferating and developing appropriate effector function . Furthermore, the data raise the possibility that otherwise inconsequential exposure to bacterial superantigens may perturb the CD8+ T cell memory pool.

Int Immunol, 1997 Sep, 9(9), 1367 - 74
Normal thymic selection, superantigen-induced deletion and Fas-mediated apoptosis of T cells in IL-2 receptor beta chain-deficient mice; Suzuki H et al.; Mice lacking the IL-2 receptor beta chain (IL-2R beta) exhibit an autoimmune reaction characterized by generalized T cell activation, production of autoantibodies, myeloproliferation and severe anemia . T cells of IL-2R beta-/- mice were examined to elucidate the mechanism responsible for their abnormal activation and to determine how such abnormal activation might affect other cell lineages . Elevated levels of IgG, IgE and autoantibodies in IL-2R beta-/- mice were found to be associated with activated CD4+ T cells which secreted elevated levels of IL-4 . Thymocytes in IL-2R beta-/- mice showed normal negative and positive selection patterns when analyzed in transgenic mice bearing a TCR specific for HY antigen, suggesting that neither IL-2 nor IL-15 is essential for thymic selection . Peripheral T cells in IL-2R beta-deficient mice underwent normal programmed cell death in response to staphylococcal enterotoxin B superantigen, in contrast to cells from mice deficient for either IL-2 or IL-2R alpha . Activated T cells in IL-2R beta-deficient mice expressed normal levels of Fas antigen and underwent normal apoptosis in response to induction with anti-Fas mAb . Thus, the accumulation of activated T cells in IL-2R beta-/- mice does not appear to be derived from abnormalities in either thymic selection or Fas-mediated apoptosis.

AIDS Res Hum Retroviruses, 1997 Sep 20, 13(14), 1221 - 7
Activation of the HIV type 1 long terminal repeat and viral replication by dimethylsulfoxide and related solvents; Klebanoff SJ et al.; The HIV-1 long terminal repeat (LTR) introduced into the macrophage cell line THP-1 and the T lymphocyte cell line Jurkat in association with the luciferase reporter gene is activated by the polar, aprotic solvents dimethylsulfoxide (DMSO), dimethylacetamide (DMAC), and dimethylformamide (DMF) . These solvents also greatly potentiated the activation of the LTR in THP-1 cells by phorbol myristate acetate (PMA), tumor necrosis factor alpha (TNF-alpha), H202, and a Staphylococcus epidermidis product . Lipopolysaccharide (LPS) and lipoteichoic acid (LTA) at 1 microg/ml had no effect on the LTR in THP-1 cells unless the solvents were added . The aprotic solvents also greatly potentiated the activation of the LTR in Jurkat cells by PMA, TNF-alpha, and H202, whereas LPS, LTA, or the S . epidermidis product had no effect in the presence or absence of the solvents . DMSO, DMAC, and DMF also increased the production of intact virions by latently HIV-1-infected ACH-2, J1.1, U1, and OM10.1 cells under some experimental conditions . The use of the polar aprotic solvents DMSO, DMAC, and DMF, by amplification, may allow the better detection of a weak activator of the LTR and facilitate studies of the mechanism of activation.

Pacing Clin Electrophysiol, 1997 Sep, 20(9 Pt 1), 2268 - 70
Successful management of an infected implantable cardioverter defibrillator with oral antibiotics and without removal of the device; Turkisher V et al.; Infection of an implantable cardioverter defibrillator developed 2 weeks after implantation, presenting with fever, swelling, redness, and tenderness of the skin above the generator site . A cloxacillin resistant coagulase-negative staphylococcus was repeatedly cultured from the abdominal wall pocket fluid . The infection was successfully treated with a combination of two antibiotics, fusidic acid and rifampin, given orally for 3 months . Although the device was not removed, infection did not recur during a 24-month follow-up.

J Pept Res, 1997 Sep, 50(3), 199 - 209
Modulation of dimerization by residues distant from the interface in bovine neurophysin-II; Zheng C et al.; The crystal structure of bovine neurophysin-II in its liganded state (Chen et al . {1991} Proc . Natl . Acad . Sci . USA 88, 4240-4244) indicates that the 1-6 sequence has a disordered conformation, lacks noncovalent contacts to other regions of the protein and is distant from the monomer-monomer interface . Cleavage of the 1-6 sequence by Staphylococcus protease V8 yielded a protein that, for the first time, crystallized in both liganded and unliganded states . Insights into the role of the 1-6 sequence in the unliganded state were obtained by NMR and related biophysical comparisons of the native and des-1-6 proteins . NMR spectra demonstrated that the environment and/or conformation of residues in the 1-6 sequence differed in liganded and unliganded states . Additionally, the unliganded des-1-6 protein exhibited a dimerization constant four to five times that of the native protein, potentially accounting for the observation that its peptide affinity was also increased . NMR studies further indicated that the increased dimerization constant of the des-1-6 protein correlated with the presence in the native protein of two isoenergetic forms of the monomer, in contrast to only a single form in the des-1-6 protein, as evidenced by signals from an internal dimerization-sensitive alpha-proton . Thus, the 1-6 sequence reduces the dimerization constant by stabilization of an alternative monomer conformation . A second product of Staphylococcus protease V8 digestion of the native protein was identified as the des-1-6 protein with an internal clip after binding site residue Glu-47, the clip presumably breaking the short 3,10 helix that most directly connects the interface to the interface to the binding site . This product, although unable to bind peptide, retained the dimerization constant of the des-1-6 protein, suggesting a lack of importance of the helix in dimerization and contrasting with the effects of the 1-6 sequence . A model is proposed in which the 1-6 sequence stabilizes the second conformation of the unliganded monomer via interactions affecting the loop region that separates the two neurophysin domains and which has been shown to influence neurophysin self-association.

Immunology, 1997 Jul, 91(3), 479 - 85
Variable region structure and staphylococcal protein A binding specificity of a mouse monoclonal IgM anti-laminin-receptor antibody; Feijo GC et al.; Staphylococcal protein A is a cell wall-attached polypeptide that acts as a B-lymphocyte superantigen . This activation correlates with specific VH gene segment usage in the B-cell receptor . B-cell receptor assembled from members of the VH3 family in humans, or S107 family in mice, has an intrinsic affinity for protein A . Human VH3-derived antibodies bind to domain D of protein A . We have characterized a mouse IgM monoclonal antibody that binds protein A . The sequencing of the variable region suggests an almost germline-encoded VH derived from S107 family and a V kappa 8-derived VL . The binding specificity of the monoclonal antibody was tested with various recombinant constructions derived from protein A . We show that, unlike human VH3-derived antibody, mouse S107-derived immunoglobulin binds to the B domain of the bacterial superantigen.

Res Immunol, 1997 May, 148(4), 257 - 66
"PERFEXT": a direct method for quantitative assessment of cytokine production in vivo at the local level; Villavedra M et al.; A method termed "PERFEXT", based on sequential perfusion and detergent extraction of lymphoid and non-lymphoid organs, has been developed for the quantitative measurement of cytokines produced at a local level in a given tissue . In vivo treatment of mice with Staphylococcus enterotoxin B (SEB) or lipopolysaccharide (LPS) served as the model systems . Interleukin-2 (IL2) and interferon-gamma (IFN gamma) levels were monitored by ELISA analysis of extracted samples . After local footpad (FP) injection with SEB, spleen and serum IL2 levels peaked at 2-4 h, while IL2 levels peaked at around 4-8 h in both FP and popliteal lymph nodes . SEB injection resulted in increased IFN gamma levels both in the FP and the draining lymph node . The detection of cytokines in the intestine allows for the application of the method at mucosal sites as well, provided enzyme inhibitors are present during the extraction procedure . After FP injection with LPS, IFN gamma production was significantly increased in the draining lymph node and was detectable in the FP, whereas IL2 was undetectable in any organ examined . IL2 and IFN gamma could also be detected at the site of elicitation of a delayed-type hypersensitivity reaction following local FP challenge . Local cytokine production correlated with the swelling response, whereas cytokine production in the spleen did not . IL2 peaked early, followed by a late increase in IFN gamma production, corresponding to the maximum swelling . This simple method should prove useful for analysing the production of other cytokines in vivo in distinct anatomical compartments.

Curr Opin Pediatr, 1997 Aug, 9(4), 396 - 405
Distinguishing vesicular and pustular disorders in the neonate; Wagner A; Vesicular and pustular disorders are common in the neonatal period . Most are benign, but several serious infectious and noninfectious diseases can present in the neonate . This review focuses on the clinical features of these disorders as well as recent advances in the field . The following diseases are discussed: erythema toxicum, transient neonatal pustular melanosis, miliaria, neonatal acne, eosinophilic folliculitis, mastocytosis, acropustulosis of infancy, incontinentia pigmenti, scabies, neonatal and congenital candidiasis, syphilis, neonatal herpes simplex infections, neonatal varicella, and staphylococcal infections.

J Mol Biol, 1997 Sep 12, 272(1), 9 - 20
Comprehensive NOE characterization of a partially folded large fragment of staphylococcal nuclease Delta131Delta, using NMR methods with improved resolution; Zhang O et al.; Comprehensive NOE results from detailed structural characterization of a 131 residue partially folded fragment of staphylococcal nuclease (Delta131Delta) made possible by NMR methods with improved resolution are presented . The resulting NOE patterns reflect sampling of both alpha and beta regions of phi, varphi conformational space, yet demonstrate significant preferences for both native-like and non-native-like turn and potentially helical conformations . Together with data from studies of the unfolded state of the drkN SH3 domain, NOE patterns observed for partially folded or unfolded proteins are summarized . It is surprising that few long-range NOEs were observed in Delta131Delta . The two longest-range NOEs are both native-like; one of these, an (i,i+5) NOE, provides evidence for a Schellman capping motif for helix termination . Many aliphatic-aliphatic and aliphatic-amide NOEs, which are not normally observed in folded proteins, were detected . We have ruled out significant contributions from spin-diffusion for a number of these NOEs and suggest that one source may be sampling of non-prolyl cis peptide bond configurations in the disordered state of Delta131Delta .

Vopr Virusol, 1997 May-Jun, 42(3), 123 - 6
{Gamma-plant as a new nontoxic interferon inducer}; Chekanovskaia LA; The interferon-inducing activity of gamma plant was studied in vivo in mice and in vitro in human whole blood cells . The maximum serum interferon (IF) production after intraperitoneal injection of gamma plant was as high as 256 IU/ml . The titers of IF produced in response to gamma plant were 16 to 32 times higher than the level of IF induced by gossipol . The capacity of human whole blood cells to produce IF under the effects of various doses of different inducers (Newcastle disease virus, staphylococcal enterotoxin A, gossipol, and gamma plant) was studied in vitro . In contrast to gossipol and staphylococcal enterotoxin A, gamma plant stimulated the cell production of IF with a high antiviral effect in culture medium . The findings permit a conclusion on a high potency of gamma plant as an IF inducer.

Przegl Lek, 1997, 54(3), 201 - 3
{Thrombus in the right ventricle in an infant--a good result of thrombolytic therapy with recombinant tissue plasminogen activator}; Aleszewicz-Baranowska J et al.; In a 2 month-old infant presenting with the symptoms of cardio-respiratory failure in the course of staphylococcal septicaemia a thrombus in the right ventricle was detected in an echocardiographic examination . Scintigraphy of the lungs revealed of microthrombosis . Thrombolytic therapy with recombinant tissue plasminogen activator (Actylise) was started on the dose of 0.5 mg/kg/hour in 7-hour constant infusion . After 5 days of treatment the thrombus was completely dissolved . No complication were observed . Indicators of serum clothing remained normal during the treatment.

Eur J Immunol, 1997 Aug, 27(8), 1933 - 41
A recombinant single-chain human class II MHC molecule (HLA-DR1) as a covalently linked heterotrimer of alpha chain, beta chain, and antigenic peptide, with immunogenicity in vitro and reduced affinity for bacterial superantigens; Zhu X et al.; Major histocompatibility complex (MHC) class II molecules bind to numerous peptides and display these on the cell surface for T cell recognition . In a given immune response, receptors on T cells recognize antigenic peptides that are a minor population of MHC class II-bound peptides . To control which peptides are presented to T cells, it may be desirable to use recombinant MHC molecules with covalently bound antigenic peptides . To study T cell responses to such homogeneous peptide-MHC complexes, we engineered an HLA-DR1 cDNA coding for influenza hemagglutinin, influenza matrix, or HIV p24 gag peptides covalently attached via a peptide spacer to the N terminus of the DR1 beta chain . Co-transfection with DR alpha cDNA into mouse L cells resulted in surface expression of HLA-DR1 molecules that reacted with monoclonal antibodies (mAb) specific for correctly folded HLA-DR epitopes . This suggested that the spacer and peptide did not alter expression or folding of the molecule . We then engineered an additional peptide spacer between the C terminus of a truncated beta chain (without transmembrane or cytoplasmic domains) and the N terminus of full-length DR alpha chain . Transfection of this cDNA into mouse L cells resulted in surface expression of the entire covalently linked heterotrimer of peptide, beta chain, and alpha chain with the expected molecular mass of approximately 66 kDa . These single-chain HLA-DR1 molecules reacted with mAb specific for correctly folded HLA-DR epitopes, and identified one mAb with {MHC + peptide} specificity . Affinity-purified soluble secreted single-chain molecules with truncated alpha chain moved in electrophoresis as compact class II MHC dimers . Cell surface two-chain or single-chain HLA-DR1 molecules with a covalent HA peptide stimulated HLA-DR1-restricted HA-specific T cells . They were immunogenic in vitro for peripheral blood mononuclear cells . The two-chain and single-chain HLA-DR1 molecules with covalent HA peptide had reduced binding for the bacterial superantigens staphylococcal enterotoxin A and B and almost no binding for toxic shock syndrome toxin-1 . The unique properties of these engineered HLA-DR1 molecules may facilitate our understanding of the complex nature of antigen recognition and aid in the development of novel vaccines with reduced superantigen binding.

Proc Natl Acad Sci U S A, 1997 Sep 16, 94(19), 10161 - 6
Exploring the folding free energy surface of a three-helix bundle protein; Guo Z et al.; The multidimensional free energy surface for a small fast folding helical protein is explored based on first-principle calculations . The model represents the 46-residue segment from fragment B of staphylococcal protein A . The relationship between collapse and tertiary structure formation, and the order of collapse and secondary structure formation, are investigated . We find that the initial collapse process gives rise to a transition state with about 30% of the native tertiary structure and 50-70% of the native helix content . We also observe two distinct distributions of native helix in this collapsed state (Rg approximately 12 A), one with about 20% of the native helical hydrogen bonds, the other with near 70% . The former corresponds to a local minimum . The barrier from this metastable state to the native state is about 2 kBT . In the latter case, folding is essentially a downhill process involving topological assembly . In addition, the order of formation of secondary structure among the three helices is examined . We observe cooperative formation of the secondary structure in helix I and helix II . Secondary structure in helix III starts to form following the formation of certain secondary structure in both helix I and helix II . Comparisons of our results with those from theory and experiment are made.

Mil Med, 1997 Sep, 162(9), 612 - 5
Detection of interleukin-6 and interleukin-2 in serum of rhesus monkeys exposed to a nonlethal dose of staphylococcal enterotoxin B; Krakauer T et al.; The immune response to a nonlethal dose of aerosolized staphylococcal enterotoxin B (SEB) was studied in nonhuman primates to define the potential human host response to a nonlethal exposure of SEB on the battlefield . Serum levels of the cytokines interleukin 2 (IL-2) and interleukin 6 (IL-6) increased significantly (p < 0.01) in six juvenile rhesus monkeys 4 hours after receiving a nonlethal, inhaled dose of SEB . The mean (+/-SD) peak serum levels of IL-2 and IL-6 were 63 +/- 39 units/ml and 514 +/- 234 pg/ml, respectively, post-SEB treatment . Tumor necrosis factor, known to be associated with SEB-mediated lethal toxic shock, was undetectable in all samples . gamma-Interferon concentrations were also elevated, but not significantly {p < 0.089} . Hence, elevated levels of IL-2 and IL-6 might be used as a serological marker for a nonlethal, incapacitating exposure to SEB.

Arch Biochem Biophys, 1997 Sep 1, 345(1), 97 - 102
Inhibition of methionine adenosyltransferase by the polyamines; Geller AM et al.; The effect of the polyamines, putrescine, spermine, and spermidine, on the activity of extrahepatic methionine adenosyltransferase (MAT II) was studied . The polyamines inhibited MAT II activity at concentrations equal to or greater than 5 mm . Combinations of polyamines were more effective than individual polyamines in inhibiting MAT activity; maximum inhibition approached 80% with combinations of all three polyamines . S-Adenosylmethionine (AdoMet), Pi, and PPi, the products of the MAT reaction, are known to be synergistic inhibitors of the nonhepatic form of the enzyme . Combinations of polyamines plus Pi and/or PPi induced an additive inhibition of the enzyme . AdoMet plus polyamines also resulted in significant inhibition, but inhibition plateaued at about 80%, indicating the presence of a protective mechanism to maintain AdoMet synthesis . Extrahepatic MAT from human and rat tissues was inhibited by the polyamines, indicating that this phenomenon is not species specific . In addition, we examined the effect of polyamines on MAT activity in resting and activated human lymphocytes that were shown to differ in the relative expression of MAT II subunits . Although MAT from mitogen (phytohemagglutinin, PHA)- and superantigen (Staphylococcal enterotoxin B, SEB)-stimulated lymphocytes were similarly inhibited by 10 mM polyamines, at lower concentrations of polyamines (1-5 mM), MAT from SEB-stimulated cells appeared to be more susceptible to inhibition by the polyamines . Inasmuch as SEB is a more physiological stimulator of T cells than PHA, the data suggest a possible role of polyamines in regulating MAT activity.

J Biol Chem, 1997 Aug 29, 272(35), 22118 - 24
A 32-kDa proteolytic fragment of transcription factor Stat3 is capable of specific DNA binding; Dreier B et al.; Fragments of characteristic size retaining the ability of sequence-specific DNA binding were generated by partial proteolysis of transcription factor Stat3 with trypsin, chymotrypsin, or Staphylococcus V8 proteinase . The molecular masses of the smallest DNA-binding fragments were 75, 48, and 32 kDa after digestion with V8 proteinase, chymotrypsin, and trypsin, respectively . The fragments contained major parts of the domain controlling the sequence specificity of DNA binding (amino acids 406-514), the SH3 and SH2 domains, and the phosphorylated tyrosine residue Tyr-705, but not the C-terminal 20 amino acids . The N terminus of the 32-kDa tryptic fragment (ANCDASLIV) matched the sequence of amino acids 424-432 deduced from cDNA . The fragments were observed after proteolytic treatment of preformed complexes between DNA and native factors eluted from rat liver nuclei or recombinant, tyrosine-phosphorylated rat Stat3 from insect cells . It was possible to elute all three minimal fragments from their complexes with DNA and to obtain specific re-binding . The minimal fragments eluted from complexes with DNA still contained the phosphorylated Tyr-705 and the SH2 domain suggesting that they were probably bound to DNA as dimers . The DNA-binding domain of Stat3 identified by these experiments overlapped the domain previously identified by genetic experiments as the domain controlling the sequence specificity of DNA binding . The DNA-binding domain defined here by partial proteolysis probably represents an autonomously folding portion of Stat3.

J Immunol, 1997 Aug 15, 159(4), 1676 - 85
Augmentation of mature CD4+ T cell responses to isolated antigenic class II proteins by fibronectin and intercellular adhesion molecule-1; Brunmark A et al.; Mature CD4+ T cells can undergo stable adhesion to isolated antigenic MHC complexes, and upon TCR engagement exhibit up-regulated adhesion to the integrin ligands fibronectin (FN) and intercellular adhesion molecule-1 (ICAM-1) . Here, we have examined T cell responses to purified antigenic class II complexes, alone or coimmobilized in the presence of FN or ICAM-1 . T cell adhesion to immobilized peptide-MHC complexes alone stimulated suboptimal, but marked levels of IFN-gamma and IL-2 secretion, and this was accompanied by cell proliferation . T cell adhesion to both FN and ICAM-1 strongly augmented cytokine release and T cell proliferation . Activation of Vbeta3+ and Vbeta8+ T cell lines by isolated staphylococcal enterotoxin-MHC complexes was also examined, and surprisingly, a Vbeta8+ T cell line displayed significant cell adhesion or later response to staphylococcal enterotoxin B-MHC complexes only when Ag was coimmobilized with ICAM-1 or FN . The results demonstrate that adhesion of CD4+ T cells to ICAM-1 or FN activated by natural TCR ligands can strongly augment T cell signaling and downstream responses . Moreover, for some Ags, T cell interaction with accessory ligands may be critical in attaining a threshold level of receptor occupancy for cell activation.

Biochemistry, 1997 Aug 5, 36(31), 9518 - 22
The heptameric prepore of a staphylococcal alpha-hemolysin mutant in lipid bilayers imaged by atomic force microscopy; Fang Y et al.; We have used atomic force microscopy to study the oligomeric state of a genetically engineered mutant of staphylococcal alpha-hemolysin (alphaHL-H5) that can be arrested as a "prepore" assembly intermediate . AFM images of alphaHL-H5 on supported bilayers of a fluid-phase lipid, egg-yolk phosphatidylcholine (egg-PC), under conditions that lock alphaHL-H5 into the prepore state, clearly show a heptameric structure for many individual oligomers . The central dent of the prepore has a diameter of 3.2 +/- 0.2 nm . The distance between the centers of mass of neighboring subunits is 2.8 +/- 0.3 nm . The heptamer has an average diameter of 8.9 +/- 0.6 nm . These results support a recently proposed pathway for the assembly of alpha-hemolysin.

Forensic Sci Int, 1997 Aug 4, 88(2), 141 - 6
Bacterial toxins and sudden unexpected death in a young child; Bentley AJ et al.; The sudden unexpected death of a six year old child following an upper respiratory tract infection is reported . Laboratory investigations revealed the presence of staphylococcal toxic shock syndrome toxin-1 (TSST-1) in samples of brain tissue . The significance of this finding is discussed.

Anaesth Intensive Care, 1997 Aug, 25(4), 354 - 7
Bacteraemia following percutaneous dilational tracheostomy; Teoh N et al.; This study reports the incidence of bacteraemia following 106 consecutive bedside percutaneous tracheostomies . Post-tracheostomy blood culture results were compared with other blood cultures from the same population . The incidence of positive post-tracheostomy blood cultures was 10.4% (11/106), compared with 6.6% (7/106) for other blood cultures (odds ratio 1.64, 95% confidence interval 0.61-4.40, P = 0.46) . Staphylococcus epidermidis was the most common organism cultured, 7/106 (6.6%) of post-tracheostomy cultures, compared with 3/106 (2.8%) for other cultures (odds ratio 2.43, 95% confidence interval 0.61-9.65, P = 0.33) . The other four post-tracheostomy cultures grew an organism cultured from that patient's tracheal secretions . Seventy-four patients were receiving antibiotics at the time of tracheostomy, of these 7 (9.5%)-had positive blood cultures, a similar incidence (4 of 32, 12.5%) to those not receiving antibiotics (odds ratio 0.73, 95% confidence interval 0.20-2.70, P = 0.90) . We conclude bacteraemia is a common complication of percutaneous tracheostomy; the causative organisms come from the patients' trachea or skin.

Can J Ophthalmol, 1997 Aug, 32(5), 311 - 4
Bacterial endophthalmitis following extracapsular cataract extraction: recommendations for early detection; Lam SR et al.; OBJECTIVE: To determine the time between the onset of symptoms of endophthalmitis after cataract extraction and presentation to an ophthalmologist and to determine the spectrum of organisms responsible for postoperative endophthalmitis . DESIGN: Case series . SETTING: Tertiary care vitreoretinal service in Toronto . PATIENTS: Thirty-three patients with early (presentation within 2 weeks of surgery) endophthalmitis following extracapsular cataract extraction and intraocular lens implantation performed between January 1989 and December 1992 . OUTCOME MEASURES: Time to presentation to an ophthalmologist, duration of symptoms, culture results . RESULTS: Twenty-two patients (66.7%) were documented to experience identifiable symptoms of endophthalmitis before presentation to their ophthalmologist; the mean time of onset of symptoms was 3.6 (standard deviation {SD} 1.7) days after surgery . Of the 22 patients 16 (72.7%) became symptomatic by the fourth postoperative day, and 21 (95.4%) experienced symptoms by the fifth postoperative day . The mean delay between onset of symptoms and presentation was 1.9 (SD 1.6) days . Bacteria were identified in 27 cases (81.8%), confirmed by culture in 23 cases (69.7%) . The organisms were gram-positive in 25 (92.6%) of the 27 cases, and coagulase-negative Staphylococcus predominated . CONCLUSIONS: In our series a considerable delay existed between the development of symptoms of endophthalmitis following extracapsular cataract extraction and clinical diagnosis . This delay could be minimized by scheduling routine postoperative visits at 1 and 4 days following cataract surgery.

Biochem J, 1997 Aug 1, 325 ( Pt 3), 707 - 10
Perturbation of the antigen-binding site and staphylococcal protein A-binding site of IgG before significant changes in global conformation during denaturation: an equilibrium study; Wang XD et al.; Although conformational perturbation of the active sites of many enzymes has been reported to precede global molecular conformational changes {Tsou (1993) Science 262, 380-381}, little effort has been made to compare the susceptibility of the ligand-binding site of proteins and the protein molecules as a whole to perturbation by denaturants . Immunoglobulin is chosen in this study to address this problem . It is found that the variable and constant regions (Fv and Fc) of a monoclonal antibody of an IgG subclass against adenylate kinase lose their abilities to bind antigen and staphylococcal Protein A after treatment with guanidinium chloride concentrations considerably lower than those required to change the global conformation of the antibody as a whole, as detected by fluorescence and second-derivative UV absorption spectroscopy . These results indicate that both ligand-binding sites of the antibody concerned are more fragile than the molecule as a whole and that the Fv and Fc regions of the antibody molecule unfold sequentially during denaturation.

Pediatr Infect Dis J, 1997 Aug, 16(8), 773 - 9
The contribution of infectious diseases to infant mortality in Alaska; Gessner BD et al.; BACKGROUND: Based on death certificates to determine cause of death, current research suggests that infectious diseases are less important causes of infant mortality than in the past . METHODS: To determine the contribution of infectious diseases to infant mortality and the sensitivity of death certificates for identifying infectious disease causes of death, we examined information from multiple sources for a population-based sample of infant deaths that occurred in Alaska during 1992 through 1994 . RESULTS: We collected information for 181 of 272 reported infant deaths and identified 48 infants for whom an infection was a primary (n = 15), contributing (n = 12) or suspected (n = 21) cause of death (infectious disease-related infant mortality rate, 2.2/1000 live births) . Of these 48 deaths 27 were associated with a maternal peripartum infection and 15 were associated with a postneonatal respiratory tract infection . A specific organism was identified for 15 of 29 infants who died during the neonatal period and for 5 of 19 infants who died during the postneonatal period (including 2 with coagulase-negative Staphylococcus and the rest with a variety of other organisms) . Death certificates identified an infectious disease as a primary or contributing cause of death for 19 infants (sensitivity, 40%) and reported a specific organism for 4 infants . CONCLUSIONS: Infectious diseases caused or contributed to a high proportion of infant mortality in Alaska during 1992 through 1994 . Death certificates had poor sensitivity for identifying infectious disease-related infant deaths.

Clin Orthop, 1997 Aug, (341), 73 - 81
Infected total knee arthroplasty . Two-stage reimplantation with a gastrocnemius rotational flap; McPherson EJ et al.; This study reviews a consecutive series of 21 patients undergoing two-stage reimplantation total knee arthroplasty for late chronic infection . All 21 patients had late chronic infections, and 20 of 21 patients were compromised hosts . Seven different organisms were isolated at the time of prosthetic resection . Staphylococcus coagulase negative species was the most frequently isolated organism . At the time of reimplantation, a medial gastrocnemius rotational flap was rotated over the proximal tibia and knee for wound closure . The average explantation time was 25 weeks (range, 7-76 weeks), and no methylmethacrylate spacers were used . At an average 17-month followup (range, 5.1-33.1 months) all reimplanted total knee replacements remained in place with one patient having recurrent infection . At reimplantation, 11 patients had positive bacterial cultures from tissue specimens . Sixteen of the 33 (40%) positive cultures were from specimens taken from the medullary canal . At followup, the average Knee Society Score was 77.4 (range, 40-100) . The lack of a methylmethacrylate spacer and a long explantation time were considered important factors in diminishing functional performance and determining the need for a gastrocnemius flap . A medial gastrocnemius rotational flap should be considered at the time of reimplantation total knee arthroplasty if the soft tissue envelope about the knee is compromised and cannot be closed without undue tension.

Cell Immunol, 1997 Aug 1, 179(2), 153 - 64
CD4+ T cells reactivated with superantigen are both more sensitive to FasL-mediated killing and express a higher level of FasL; Wang JK et al.; Naive CD4(+) T cells proliferate strongly in response to superantigens such as staphylococcal enterotoxin B (SEB) . When these cells are rested and challenged a second time, they undergo activation-induced cell death (AICD) . Fas/FasL interactions have been shown to mediate AICD, even though the level of Fas expression in the 2 degrees SEB responder populations is no higher than in the 1 degrees cultures . To determine whether the dissimilarity between the 1 degrees and 2 degrees cultures could be attributed to differences in FasL cytotoxic activity or in the sensitivity of the Fas apoptosis signaling pathway, we compared these parameters during the 1 degrees and 2 degrees responses of lpr and gld CD4+ T cells (which do not undergo AICD due to a lack of Fas and an inactive FasL, respectively) so that each parameter could be evaluated independently . The results demonstrate that 2 degrees responders both express a higher level of functional FasL and are more sensitive to FasL-mediated killing . These findings account for the differences between the 1 degrees and 2 degrees responses of CD4+ T cells to superantigen . In addition, we found that the apparent level of FasL-mediated cytotoxic activity in the 2 degrees lpr CD4+ T cell population is much higher than that of wild-type cells, suggesting that deficient Fas expression leads to inordinately high levels of FasL expression or subsaturation of FasL binding sites.

Cell Immunol, 1997 Aug 1, 179(2), 138 - 45
Staphylococcal enterotoxin B induces arthritis in female DBA/1 mice but fails to induce activation of type II collagen-reactive lymphocytes; Omata S et al.; It has been proposed that superantigens are involved in the pathogenesis of autoimmune diseases . To test the possibility of superantigens inducing arthritis in naive mice, V beta 8-reactive superantigen staphylococcal enterotoxin B (SEB) was injected into naive mice . We used female DBA/1 mice, because they were susceptible to collagen-induced arthritis (CIA), in which the pathogenic T cells were supposed to preferentially use limited V betas of T cell receptors including V beta 8 . Mild monoarthritis developed in uninjected hindlimbs of mice administered with SEB in higher frequency (an average incidence of 24%) than the control phosphate-buffered saline-injected mice (4.2%) . Autoimmune responses in mice administered with SEB were compared with those in mice developing CIA . However, activation of type II collagen (IIC)-reactive T cells was not detected in SEB-injected mice . Production of autoantibodies, anti-IIC antibody and rheumatoid factor was also undetected . Although exact mechanisms of pathogenesis of this arthritis remain to be known, V beta 8+ T cells were activated for a long period and the unresponsiveness of V beta 8+ T cells was not detected in this strain . From these results, we discuss the pathogenesis of arthritis induced by SEB and the possibility that superantigen may play a role in the induction of autoimmune diseases.

Arch Surg, 1997 Aug, 132(8), 894 - 902
Staphylococcus epidermidis graft infection is associated with locally suppressed major histocompatibility complex class II and elevated MAC-1 expression; Henke PK et al.; OBJECTIVES: To determine the local cellular immune response in a series of human patients with Staphylococcus epidermidis prosthetic graft infection and to use a murine model to investigate the response in polytef (PTFE) and in a nonslime-producing S epidermidis variant . METHODS: Externally supported Dacron and PTFE grafts, either sterile or colonized with slime (RP-62A)- or nonslime (RP-62NA)-producing S epidermidis (10(7) colony forming units/cm2) were implanted in a dorsal subcutaneous pocket of Swiss Webster mice (Taconic, Germantown, NY) . The grafts were harvested at 7, 10, 14, and 28 days with local bacterial and leukocyte counts obtained . Perigraft and blood monocyte major histocompatibility complex class II (MHC-II) (immune antigen) and membrane attack complex type 1 (MAC-1) (glycoprotein) expression were analyzed by flow cytometry in the murine model and in 3 patients representing 4 Dacron graft infections . RESULTS: The human infected Dacron perigraft monocytes revealed a suppressed MHC-II and elevated MAC-1 expression, and early correlation with the murine model was seen . No notable perigraft monocyte MHC-II suppression occurred in the infected PTFE graft . The reciprocal relationship in Dacron between monocyte MAC-1 and MHC-II expression was exaggerated with the lack of slime production . Bacterial clearance was variable . Supranormal expression was observed at 1 month in sterile Dacron but not in PTFE grafts . CONCLUSIONS: Staphylococcus epidermidis infection is associated with local cellular immune suppression in Dacron but not PTFE grafts . Slime-producing S epidermidis induced a lesser cytotoxic-phagocytic response than the nonslime variant . The reduced immunologic response to slime-producing S epidermidis may explain, in part, its indolent nature and resistance to eradication.

Protein Sci, 1997 Aug, 6(8), 1627 - 42
Identification of cooperative folding units in a set of native proteins; Wallqvist A et al.; Cooperative unfolding penalties are calculated by statistically evaluating an ensemble of denatured states derived from native structures . The ensemble of denatured states is determined by dividing the native protein into short contiguous segments and defining all possible combinations of native, i.e., interacting, and non-native, i.e., non-interacting, segments . We use a novel knowledge-based scoring function, derived from a set of non-homologous proteins in the Protein Data Bank, to describe the interactions among residues . This procedure is used for the structural identification of cooperative folding cores for four globular proteins: bovine pancreatic trypsin inhibitor, horse heart cytochrome c, French bean plastocyanin, and staphylococcal nuclease . The theoretical folding units are shown to correspond to regions that exhibit enhanced stability against denaturation as determined from experimental hydrogen exchange protection factors . Using a sequence similarity score for related sequences, we show that, in addition to residues necessary for enzymatic function, those amino acids comprising structurally important folding cores are also preferentially conserved during evolution . This implies that the identified folding cores may be part of an array of fundamental structural folding units.

Protein Sci, 1997 Aug, 6(8), 1621 - 6
Comparison of straight chain and cyclic unnatural amino acids embedded in the core of staphylococcal nuclease; Wynn R et al.; We have determined by X-ray crystallography the structures of several variants of staphylococcal nuclease with long flexible straight chain and equivalent length cyclic unnatural amino acid side chains embedded in the protein core . The terminal atoms in the straight side chains are not well defined by the observed electron density even though they remain buried within the protein interior . We have previously observed this behavior and have suggested that it may arise from the addition of side-chain vibrational and oscillational motions with each bond as a side chain grows away from the relatively rigid protein main chain and/or the population of multiple rotamers (Wynn R, Harkins P, Richards FM . Fox RO . 1996 . Mobile unnatural amino acid side chains in the core of staphylococcal nuclease . Protein Sci 5:1026-1031) . Reduction of the number of degrees of freedom by cyclization of a side chain would be expected to constrain these motions . These side chains are in fact well defined in the structures described here . Over-packing of the protein core results in a 1.0 A shift of helix 1 away from the site of mutation . Additionally, we have determined the structure of a side chain containing a single hydrogen to fluorine atom replacement on a methyl group . A fluorine atom is intermediate in size between methyl group and a hydrogen atom . The fluorine atom is observed in a single position indicating it does not rotate like methyl hydrogen atoms . This change also causes subtle differences in the packing interactions.

Nat Biotechnol, 1997 Aug, 15(8), 772 - 7
Binding proteins selected from combinatorial libraries of an alpha-helical bacterial receptor domain; Nord K et al.; Small protein domains, capable of specific binding to different target proteins have been selected using combinatorial approaches . These binding proteins, called affibodies, were designed by randomization of 13 solvent-accessible surface residues of a stable alpha-helical bacterial receptor domain Z, derived from staphylococcal protein A . Repertoires of mutant Z domain genes were assembled and inserted into a phagemid vector adapted for monovalent phage display . Two libraries, each comprising approximately 4 x 10(7) transformants, were constructed using either an NN(G/T) or an alternative (C/A/G)NN degeneracy . Biopanning against the target proteins Taq DNA polymerase, human insulin, and a human apolipoprotein A-1 variant, showed that in all cases significant enrichments were obtained by the selection procedures . Selected clones were subsequently expressed in Escherichia coli and analyzed by SDS-PAGE, circular dichroism spectroscopy, and binding studies to their respective targets by biospecific interaction analysis . The affibodies have a secondary structure similar to the native Z domain and have micromolar dissociation constants (KD) for their respective targets.

FEMS Microbiol Lett, 1997 Aug 1, 153(1), 25 - 32
In vivo reaction of affinity-tag-labelled epidermin precursor peptide with flavoenzyme EpiD; Kupke T et al.; The Staphylococcus epidermidis genes encoding the His-tag-labelled epidermin precursor peptide EpiA and the flavoenzyme EpiD or the mutant protein EpiD-G93D, which lacks the coenzyme, were co-expressed and the proteins were synthesized in vivo in Escherichia coli . Only in the presence of EpiD was the precursor peptide converted to a reaction product with a decrease in mass of 44-46 Da . This result confirms the in vitro experiments carried out with purified EpiA and purified EpiD from Staphylococcus epidermidis {Kupke et al . (1994) J . Biol . Chem . 269, 5653-5659} . EpiD catalyzes the oxidative decarboxylation of the C-terminal cysteine residue of EpiA to a {Z}-enethiol structure . In the presence of EpiD, the amount of purified (modified) peptide EpiA was several-fold higher than in the presence of EpiD-G93D, indicating that the stabilization of EpiA against proteolysis is due to an interaction with EpiD or to the presence of the C-terminal modification . The presented experimental approach will be valuable for the analysis of enzymes that catalyze posttranslational modification reaction of peptides and proteins.

J Autoimmun, 1997 Aug, 10(4), 357 - 65
Memory T cell tolerance to superantigens is not due to increased susceptibility to apoptosis; Lee WT et al.; Naive (virgin) and memory T lymphocytes differ markedly in their response to superantigens (SAg) . When cultured with the SAg staphylococcal enterotoxin B (SEB), virgin but not memory CD4(+) T cells proliferate and secrete lymphokines . Memory cells do express increased levels of activation markers after interaction with SEB, which suggests that the cells are not ignorant of the SAg . In this report, we have considered whether SEB, rather than activating memory cells, promotes their death by apoptosis . Our results indicate that while in vivo exposure to SEB induces apoptosis, there is no greater level of cell death in the memory cell population relative to virgin cells . Further, elimination of the Fas-mediated cell death pathway does not permit memory cells to be stimulated by SEB . Memory T cells from either Fas-expressing or Fas-deficient (MRL-lpr/lpr) mice are hyporesponsive to SEB . Blockade of Fas/Fas-ligand interactions by a Fas-Fc chimeric protein does not permit BALB/c memory cells to proliferate upon culture with SEB . These results provide evidence that the failure of memory T cells to respond to SEB is not due to cell death and that inactivation (anergy) is the likely fate of these cells when they encounter SEB.

J Clin Microbiol, 1997 Aug, 35(8), 2174 - 7
Pediatric bacteremia due to Staphylococcus warneri: microbiological, epidemiological, and clinical features; Buttery JP et al.; Between 1991 and 1995, an apparent high rate of Staphylococcus warneri bacteremias at the Royal Children's Hospital, Melbourne, Victoria, Australia, raised the possibility of a virulent nosocomial strain . In a retrospective review of 30 S . warneri bacteremias in children, organisms were viable and verified in 22 episodes, 12 representing significant bacteremias . Of these 12 episodes, 2 pairs shared chromosomal DNA pulsed-field gel electrophoresis patterns in unconnected patients, dispelling concerns about a single virulent strain.

Proc Natl Acad Sci U S A, 1997 Jul 8, 94(14), 7314 - 9
In vivo zippering of inner and outer mitochondrial membranes by a stable translocation intermediate; Schulke N et al.; It was previously assumed that the import of cytoplasmically synthesized precursor proteins into mitochondria occurs through a single structure spanning both outer and inner membranes at contact sites . Based on recent findings, however, the two membranes appear to contain independent translocation elements that reversibly cooperate during protein import . This feature makes it difficult to generate a means of isolating a fully integrated and functional translocation complex . To study these independent translocases in vitro and in vivo, we have constructed a chimeric protein consisting of an N-terminal authentic mitochondrial precursor (delta1-pyrroline-5-carboxylate dehydrogenase) linked, through glutathione S-transferase, to IgG binding domains derived from staphylococcal protein A . This construct becomes trapped en route to the matrix, spanning both outer and inner membranes in such a way that the entire signal-less delta1-pyrroline-5-carboxylate dehydrogenase moiety reaches the matrix, while only the folded protein A domain remains outside . During in vivo import of this precursor, outer and inner membranes of yeast mitochondria become progressively "zippered" together, forming long stretches of close contact . Using this novel intermediate, the outer and inner mitochondrial membrane channels, which normally interact only transiently, can be tightly joined (both in vitro and in vivo), forming a stable association . This suggests a method for isolating the functional translocation complex as a single entity.

J Exp Med, 1997 Jul 7, 186(1), 71 - 81
Transferable anergy: superantigen treatment induces CD4+ T cell tolerance that is reversible and requires CD4-CD8- cells and interferon gamma; Cauley LS et al.; Bacterial superantigens induce peripheral unresponsiveness in CD4+ T cell populations that express appropriate Vbeta chains . We have used Vbeta3/Valpha11 T cell receptor transgenic (Tg) mice and the Vbeta3-specific superantigen staphylococcal enterotoxin A (SEA) to further investigate the mechanisms that contribute to such unresponsiveness . As in other models, in vivo exposure to SEA rendered the Tg CD4+ cells unresponsive to subsequent restimulation in vitro with antigen or mitogens . However, when the SEA-treated CD4+ cells were completely purified away from all other contaminating cells, they regained the ability to proliferate and secrete cytokines . Moreover, enriched CD4-CD8- cells from the SEA-treated mice suppressed the responses of fresh control CD4+ cells in mixed cultures indicating that the apparent "anergy" was both transferable and reversible . Further analysis demonstrated that interferon gamma, but not the Fas receptor, played a critical role in the suppression.

Infection, 1997 Jul-Aug, 25(4), 252 - 4
Life-threatening infection with multiresistant Staphylococcus epidermidis in a patient with end-stage renal disease: cure with chloramphenicol and quinupristin/dalfopristin (RP 59500); Mundlein E et al.; A 45-year-old man with end-stage renal disease underwent a cadaveric kidney transplantation . The allograft had to he removed 10 days after transplantation because of an acute vascular rejection . After explantation, the patient suffered from a life-threatening infection with Staphylococcus epidermidis involving lungs, eyes and liver for 11 months . Despite adequate therapy including vancomycin followed by teicoplanin, he developed spondylodiscitis requiring repeated surgical interventions . The definitive cure was achieved by a sequential therapy with chloramphenicol and quinupristin/dalfopristin.

Allergy, 1997 Jul, 52(7), 732 - 8
Neonates at risk of atopy show impaired production of interferon-gamma after stimulation with bacterial products (LPS and SEE)
Pohl D, Bockelmann C, Forster K, Rieger CH, Schauer U.
Recent studies demonstrate reduced interferon-gamma (IFN-gamma) secretion in neonates who became atopic later in life . The underlying pathomechanism is still unknown . We therefore examined the effects of bacterial products on neonatal IFN-gamma production acting through different T-cell- or antigen-presenting-cell (APC)-stimulating mechanisms: cord-blood mononuclear cells (CBMC) were incubated with lipopolysaccharide (LPS), staphylococcal enterotoxin E (SEE), or a combination of both and restimulated with PMA and ionomycin . LPS and SEE as single stimuli induced IFN-gamma production to the same extent in CBMC of neonates with high and low risk of atopy . In contrast, a combination of LPS and SEE had a multiplying effect on IFN-gamma secretion only in CBMC of neonates with low risk of atopy . Phenotype analysis revealed that only memory T cells showed impaired IFN-gamma synthesis (median 3.6% IFN-gamma-producing cells vs 14.2% in controls: P < 0.01), whereas IFN-gamma production by naive T cells did not differ in either group . Taken together, these results point to the existence of a disturbed function of costimulatory mechanisms in neonates at high risk of atopy, provoking reduced memory T-cell IFN-gamma production.

Chem Biol, 1997 Jul, 4(7), 497 - 505
Designed protein pores as components for biosensors; Braha O et al.; BACKGROUND: There is a pressing need for new sensors that can detect a variety of analytes, ranging from simple ions to complex compounds and even microorganisms . The devices should offer sensitivity, speed, reversibility and selectivity . Given these criteria, protein pores, remodeled so that their transmembrane conductances are modulated by the association of specific analytes, are excellent prospects as components of biosensors . RESULTS: Structure-based design and a separation method that employs targeted chemical modification have been used to obtain a heteromeric form of the bacterial pore-forming protein staphylococcal alpha-hemolysin, in which one of the seven subunits contains a binding site for a divalent metal ion, M(II), which serves as a prototypic analyte . The single-channel current of the heteromer in planar bilayers is modulated by nanomolar Zn(II) . Other M(II)s modulate the current and produce characteristic signatures . In addition, heteromers containing more than one mutant subunit exhibit distinct responses to M(II)s Hence, a large collection of responsive pores can be generated through subunit diversity and combinatorial assembly . CONCLUSIONS: Engineered pores have several advantages as potential sensor elements: sensitivity is in the nanomolar range; analyte binding is rapid (diffusion limited in some cases) and reversible; strictly selective binding is not required because single-channel recordings are rich in information; and for a particular analyte, the dissociation rate constant, the extent of channel block and the voltage-dependence of these parameters are distinguishing, while the frequency of partial channel block reflects the analyte concentration . A single sensor element might, therefore, be used to quantitate more than one analyte at once . The approach described here can be generalized for additional analytes.

J Hosp Infect, 1997 Jul, 36(3), 181 - 9
The potential for catheter microbial contamination from a needleless connector; Brown JD et al.; Needleless connectors have been widely introduced into clinical practice to allow the connection of syringes and luers to peripheral and central vascular catheters . The potential for microbial contamination of catheters via these devices is currently unclear . A recently introduced connector, the 'Connecta Clave', was assessed by various in-vitro methods . The 'Connecta Clave' is specifically devised to separate external components from the fluid pathway . The compression seals of 50 devices were contaminated with 1 x 10(4) cfu Staphylococcus epidermidis, disinfected with isopropanol, and fluid passed through . Only one device allowed organisms to pass through, despite this challenge, representing a contamination rate of 2% . In comparison, when 50 connectors were challenged with 20 cfu of S . epidermidis, no organisms passed through the device during use . In the clinical situation, after manipulation, < 16 cfu of skin organisms were found associated with the compression seal of the devices . It is, therefore, likely that the contamination rates in clinical practice will be extremely low . Three methods of disinfecting the compression seals and associated rims were also evaluated . A combination of alcohol chlorhexidine spray, followed by a 70% isopropanol swab, resulted in the most efficacious disinfection . The isopropanol swabs produced an adequate disinfection rate . The overall results suggest that by use of specially designed connectors, not only are needlestick injuries reduced, but the likelihood of microbial contamination of catheters via the internal route may also be diminished.

Am J Physiol, 1997 Jul, 273(1 Pt 1), L31 - 9
SEB is cytotoxic and alters EC barrier function through protein tyrosine phosphorylation in vitro; Campbell WN et al.; We studied whether Staphylococcal enterotoxin B (SEB) has direct effects on endothelial cells (EC) in the absence of effector cells or their products . Bovine or human pulmonary artery EC were grown to confluence on filters mounted in chemotaxis chambers . Barrier function was assessed by placing {14C}bovine serum albumin in the chamber and sampling the lower well for 14C activity . SEB exposures induced a significant (P < 0.001) dose- and time-dependent increase in albumin flux across both bovine and human EC monolayers . Albumin flux was temperature dependent, and cycloheximide pretreatment of the monolayers did not block the SEB-induced increase in permeability . Preincubation of SEB with trypsin or anti-SEB antibody significantly (P < 0.0001) reduced the effect, whereas pretreatment with polymyxin B did not . SEB at > or = 10 micrograms/ml significantly (P < 0.03) increased EC injury as measured by 51Cr release in a dose- and time-dependent manner . Herbimycin and genistein, inhibitors of protein tyrosine kinases, each protected against SEB-induced cytotoxicity, barrier dysfunction, and intercellular gap formation . We conclude that SEB perturbs endothelial barrier function and viability in the absence of effector cells or their mediators.

J Bone Joint Surg Br, 1997 Jul, 79(4), 567 - 9
Evaluation of sinus-track cultures in chronic bone infection; Mousa HA; Sinus-track cultures were compared prospectively with cultures from operative specimens in 55 patients with chronic bone infection . There was a total of 115 operative isolates; 102 of the sinus-track isolates were identical (88.7%), showing a specificity of 95.7% and a predictive value of 90.3% . A sinus-track specimen proved to be a reliable source for the isolation of all bacteria causing chronic bone infection except Staphylococcus epidermidis, provided that material from the depths of the sinus was aspirated by syringe from an active flowing sinus and inoculated immediately on culture media . Cotton-swab sinus specimens were unreliable for the isolation of mycobacteria, and could miss many pyogenic bacteria, or contain contaminants . Tuberculous bone infection should be suspected if there is no growth of any pyogenic bacteria or if there is growth of Staphyloccocus epidermidis alone on routine aerobic and anaerobic sinus cultures . Mycobacteria can often be identified from sinus-track culture from patients in whom operative culture, histopathology and clinical examination have failed to confirm the diagnosis of tuberculosis.

Clin Infect Dis, 1997 Jul, 25(1), 69 - 71
Polyclonal staphylococcus endocarditis; Van Wijngaerden E et al.; Coagulase-negative staphylococcus (CNS) is the most frequent cause of nosocomial bacteremia and prosthetic valve endocarditis . CNS bacteremia can be polyclonal . No data exist on the clonality of CNS causing endocarditis . We present a case of CNS aortic homograft endocarditis in which at least five different genotypes of CNS were identified in initial blood-culture isolates by genomic macrorestriction enzyme analysis and pulsed field gel electrophoresis . Since the polyclonality was accompanied by differences in antibiotic susceptibility, this observation may have important consequences for the treatment of CNS endocarditis . Because of the parallels in the pathogenesis of CNS prosthetic valve endocarditis and CNS infections of a variety of other prosthetic devices, it might also have consequences for CNS prosthetic device infections in general . We suggest that antibiotic susceptibility testing of just one blood-culture isolate may be insufficient.

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