Staphylococcus

J Neurochem, 1994 Oct, 63(4), 1466 - 76
Short and long form gamma 2 subunits of the GABAA/benzodiazepine receptors; Khan ZU et al.; Three novel antisera to the gamma 2 subunit of the gamma-aminobutyric acidA (GABAA) receptor/benzodiazepine receptor (GABAAR/BZDR) complex have been made . Anti-gamma 2S and anti-gamma 2L are specific antibodies to synthetic peptides that recognize the gamma 2S (short) and gamma 2L (long) forms, respectively, of the gamma 2 subunit . An antibody (anti-gamma 2IL2) to staphylococcal protein A fusion protein of the large intracellular loop (gamma 2IL) located between the putative transmembrane segments M3 and M4 of gamma 2S recognizes both gamma 2S and gamma 2L subunits . The antibodies immunoprecipitated both the solubilized and affinity-purified GABAAR/BZDR from rat and bovine brain . Immunoblots with membranes from rat brain cerebral cortex as well as with affinity-purified receptor from bovine cortex show that anti-gamma 2S and anti-gamma 2L recognize peptides of 45,000 and 47,000 M(r), respectively . Immunoprecipitation experiments indicate that gamma 2S is more prevalent in hippocampus, whereas gamma 2L is more abundant in cerebellum . Intermediate values for each form are found in the cerebral cortex . The results suggest that in the rat brain there is a considerable amount of colocalization of gamma 2S and gamma 2L in the same receptor complex . In the cerebral cortex, 15% of the BZDRs contain both gamma 2S and gamma 2L subunits and 41-48% of the gamma 2L subunit coexists with gamma 2S in the same receptor complex . In cerebellum, in 27% of the clonazepam-sensitive and 39% of the clonazepam-insensitive BZDRs the gamma 2S and gamma 2L coexist in the same receptor complex . The latter are presumably localized in granule cells and also contain alpha 6 . In addition, almost all (93%) the clonazepam-insensitive BZDRs that contain gamma 2L also contain a gamma 2S subunit in the same receptor complex . The most likely interpretation of the results is that there is an important population of granule cell receptors that contain alpha 6, gamma 2S, and gamma 2L coexisting in the same receptor complex . Nevertheless, 31% of the cerebellar receptors that contain alpha 6 subunit(s) have neither gamma 2S nor gamma 2L subunits . There are also species differences with respect to the relative abundance of gamma 2S and gamma 2L . These results might be relevant for understanding the molecular mechanisms underlying some of the GABAAR/BZDR-mediated effects of ethanol intoxication involving cerebellar granule cells.

J Leukoc Biol, 1994 Oct, 56(4), 458 - 63
Costimulatory receptors for the superantigen staphylococcal enterotoxin B on human vascular endothelial cells and T cells; Krakauer T; Cell-surface molecules on human vascular endothelial cells (ECs) and T lymphocytes that mediate staphylococcal enterotoxin B (SEB)-induced T cell proliferation and cytokine production were investigated . Expression of HLA-DR and intercellular adhesion molecule 1 (ICAM-1) on EC was induced by interferon-gamma (IFN-gamma) . IFN-gamma-treated ECs bound SEB effectively and stimulated T cells to proliferate and secrete tumor necrosis factor alpha (TNF-alpha) and IFN-gamma . SEB-induced T cell proliferation was inhibited by monoclonal antibodies to CD2, CD11a, CD28, ICAM-1, and endothelial leukocyte adhesion molecule (ELAM) . These antibodies also blocked production of the proinflammatory mediators, TNF-alpha and IFN-gamma, in SEB-stimulated T cell-EC cocultures . These results suggest that the surface molecules, CD11a:CD18/ICAM-1, CD2, CD28, and ELAM, are all important costimulatory receptors for T cell activation by superantigens with the EC as the antigen-presenting cell . Thus, like conventional antigens, multiple stimulatory signals from the interactions of these receptors are required for superantigen-induced immune responses with ECs and T cells . Reducing proinflammatory mediators such as TNF-alpha and IFN-gamma by these antibodies in SEB-induced T cell responses may be useful therapeutic strategy for circumventing SEB toxicity and pathogenesis.

Int J Radiat Biol, 1994 Oct, 66(4), 381 - 4
Metaphase yields from staphylococcal enterotoxin A stimulated peripheral blood lymphocytes of unirradiated and irradiated aged rhesus monkeys; Hill FS et al.; The mitogen phytohemagglutinin (PHA) works well in both human and cynomolgus monkey (Macaca fascicularis) lymphocyte cultures to stimulate T cell proliferation . T cells from rhesus monkeys (Macaca mulatta) are less responsive than human cells, producing few metaphases when thousands are required, e.g . in biological dosimetry studies . We show that staphylococcal enterotoxin A (SEA), one of the most potent mitogens known, at a concentration of 0.5 microgram/ml stimulated peripheral lymphocytes to grow with a mitotic index (MI) averaging 0.13 metaphases/cell in old, irradiated rhesus macaques . This was significantly greater (p < 0.001) than that produced by PHA (MI < 0.01) in lymphocytes from the same animals . Whole blood was cultured for 96, 120 and 144 h for five irradiated individuals and for two controls . All cells cultured with SEA produced a high MI with a peak response at 120 h whereas the same cultures showed low MI for each PHA stimulated culture.

J Clin Invest, 1994 Oct, 94(4), 1365 - 72
Induction of the autoantigen proliferating cell nuclear antigen in T lymphocytes by a mycobacterial antigen; Haftel HM et al.; Mycobacteria have been implicated in the pathogenesis of autoimmunity . To determine the potential effect of mycobacterial antigens on peripheral blood mononuclear cells (PBMC), we analyzed PBMC incubated with the acetone-precipitable fraction of Mycobacterium tuberculosis (APMT) for changes in cellular protein expression . Two-dimensional gel analysis showed induction of a 36-kD polypeptide identified as proliferating cell nuclear antigen (PCNA), a known autoantigen, after incubation with AP-MT . PCNA plays a role in cell proliferation and is expressed as a late growth regulated factor . However, its synthesis in response to AP-MT was induced as an early event . The early induction of PCNA was regulated at a posttranscriptional level and was restricted to T cells . Treatment of PBMC with known T cell mitogens, namely PHA, anti-CD3 antibodies, and staphylococcal superantigens failed to induce an early PCNA increase . The distinct characteristics of the AP-MT effect on PCNA expression suggest a separate mechanism of induction in response to AP-MT, compared with the late increase observed in response to mitogens . The induction of PCNA in response to mycobacterial antigens may represent a pathogenically relevant mechanism in autoimmunity.

Infect Immun, 1994 Oct, 62(10), 4626 - 31
Increased susceptibility to staphylococcal enterotoxin B intoxication in mice primed with actinomycin D; Chen JY et al.; Mice (BALB/cJ, C3H/HeN, and C3H/HeJ) primed with actinomycin D became highly susceptible to lethal intoxication with staphylococcal enterotoxin B (SEB) . The mice underwent toxicosis and toxic shock and died . Actinomycin D-primed C3H/HeN and C3H/HeJ mice showed equal sensitivity to SEB, suggesting that bacterial lipopolysaccharide derived from gram-negative bacteria in the gut may not be an important cofactor in intoxication . In a time course study of the illness, prominent pathological changes characterized by blood congestion and thickening of alveolar septa were seen in the lung, while blood congestion, inflammation, epithelial cell flattening, and villous blunting were seen in the small intestine . In lymphoid tissues, such as the spleen, congestion, inflammation, and lymphoid cell depletion were the major reactions . The pathological features of the mice had many similarities to those of rhesus monkeys intoxicated with intravenous SEB . The actinomycin D-primed C3H/HeJ mice are thus an ideal mouse model for studying SEB toxicosis and toxic shock.

Infect Immun, 1994 Oct, 62(10), 4160 - 6
Staphylococcal glycocalyx activates macrophage prostaglandin E2 and interleukin 1 production and modulates tumor necrosis factor alpha and nitric oxide production; Stout RD et al.; We have examined the effect of staphylococcal glycocalyces on the ability of murine peritoneal macrophages to produce prostaglandin E2 (PGE2) and the inflammatory cytokines interleukin 1 (IL-1) and tumor necrosis factor alpha (TNF-alpha) and to generate nitric oxide . Glycocalyx partially purified under endotoxin-free conditions from defined liquid medium cultures of Staphylococcus lugdunensis or Staphylococcus epidermidis was a strong stimulator of PGE2 and IL-1 production . The addition of 10 to 100 micrograms of glycocalyx per ml induced levels of IL-1 and PGE2 production similar to that induced by 0.1 to 1 micrograms of Escherichia coli lipopolysaccharide (LPS) per ml . In contrast, glycocalyx induced ninefold less TNF-alpha and three- to fourfold less nitrite than LPS . A modulatory effect was suggested by the observation that the amount of TNF-alpha and nitrite generated remained constant whether the macrophages were stimulated with 10 or 100 micrograms of glycocalyx per ml . A selective modulation of macrophage activation was confirmed by the demonstration that costimulation of macrophages with both glycocalyx and LPS resulted in a reduction in TNF-alpha and nitrite generation relative to stimulation with LPS alone even though costimulation had no effect on PGE2 production and increased IL-1 production . Involvement of PGE2 in this modulatory effect was suggested by the ability of indomethacin to augment glycocalyx-stimulated TNF-alpha production and to reverse the inhibitory effect of glycocalyx on LPS induction of TNF-alpha production . However, the inability of indomethacin to reverse the inhibitory effect of glycocalyx on LPS-induced nitric oxide generation suggests that the selective modulation of macrophage function by glycocalyx may be more complex than increased sensitivity to PGE2 feedback inhibition.

Eur J Immunol, 1994 Oct, 24(10), 2435 - 40
Differential expression of mRNA encoding interleukin-12 p35 and p40 subunits in situ; Bette M et al.; Interleukin-12 (IL-12) is a heterodimeric cytokine that plays an important role in the regulation of the immune response . For biological activity the expression of both subunits of IL-12, p35 and p40, is required . Moreover, in the mouse the p40 chain of IL-12 specifically inhibits the effects of the IL-12 heterodimer . In the present study we have analyzed by in situ hybridization the expression of the p35 and p40 mRNA in the spleens of BALB/c and mutant (SCID, nude, beige) mice, unstimulated and after in vivo stimulation with lipopolysaccharide (LPS) and with staphylococcal enterotoxin B (SEB) . In unstimulated spleens of BALB/c mice p35 and p40 mRNA were only detectable in a few strongly stained single cells, p35 mRNA was expressed in addition weakly in the B cell areas . After injection of LPS or SEB, p40 mRNA was strongly induced in the T cell areas all over the spleen, whereas expression of p35 mRNA and its distribution pattern did not change . Surprisingly, most of the mRNA for p35 and p40 was localized in different areas of the spleen and was apparently produced by different cells . In macrophage-depleted spleens the increased expression of p40 mRNA in response to LPS was reduced but still detectable, demonstrating that other cells besides macrophages can up-regulate IL-12 p40 mRNA . Nude mice showed a stronger expression of p35 mRNA, SCID mice lacked the weak p35 staining of the B cell areas but showed a strong basal expression of both p35 and p40 mRNA and a focal response to LPS . The pattern of IL-12 mRNA expression in beige mice was the same as in normal mice . These data demonstrate a spatial dissociation of expression of the two chains of IL-12 and are compatible with a regulatory role of the isolated IL-12 p40 chain in vivo . In addition, they indicate that the demonstration of mRNA for both chains of IL-12 in whole tissues or cell mixtures is not necessarily indicative of functional IL-12.

Eur J Immunol, 1994 Oct, 24(10), 2429 - 34
Localization of the site of the murine IgG1 molecule that is involved in binding to the murine intestinal Fc receptor; Kim JK et al.; Site-directed mutagenesis of a recombinant Fc hinge fragment has recently been used to localize the site of the murine IgG1 molecule that is involved in the control of catabolism (the "catabolic site") . In the current study, the effects of these CH2 and CH3 domain mutations (Ile 253 to Ala 253, His 310 to Ala 310, Gln 311 to Asn 311, His 433 to Ala 433 and Asn 434 to Gln 434) on intestinal transfer of Fc hinge fragments in neonatal mice have been analyzed . Studies using direct transfer and competition assays demonstrate that the mutations affect the transmission from intestinal lumen into serum in a way that correlates closely with the effects of the mutations on pharmacokinetics . Binding studies of several of the Fc hinge fragments to isolated neonatal brush borders have been used to confirm the in vivo transmission data . These analyses have resulted in the localization of the binding site for the intestinal transfer receptor, FcRn, to specific residues of the murine Fc hinge fragment . These residues are located at the CH2-CH3 domain interface and overlap with both the catabolic site and staphylococcal protein A (SpA) binding site . The pH dependence of IgG1 or Fc fragment binding to FcRn is consistent with the localization of the FcRn interaction site to a region of the Fc that encompasses two histidine residues (His 310 and His 433) . To assess whether one or two FcRn binding sites per Fc hinge are required for intestinal transfer, a hybrid Fc hinge fragment comprising a heterodimer of one Fc hinge with the wild-type IgG1 sequence and a mutant Fc hinge with a defective catabolic site (mutated at His 310, Gln 311, His 433 and Asn 434) has been analyzed in direct and competition transmission assays . The studies demonstrate that the Fc hybrid is transferred with significantly reduced efficiency compared to the wild type Fc hinge homodimer and indicate that the binding to FcRn, and possibly subsequent transfer, is enhanced by the presence of two FcRn binding sites per Fc hinge fragment.

Chest, 1994 Oct, 106(4), 1156 - 61
Invasive pulmonary aspergillosis . MRI, CT, and plain radiographic findings and their contribution for early diagnosis; Blum U et al.; A prospective study was conducted in 38 patients with nodular lesions on plain chest radiographs and the clinical suspicion of invasive pulmonary aspergillosis (IPA) to assess the diagnostic accuracy of magnetic resonance imaging (MRI) and computed tomography (CT) . For early diagnosis of IPA (clinical signs and symptoms < 10 days), CT scans with demonstration of the halo sign had a high sensitivity (16/22) and specificity (8/8) . Magnetic resonance imaging performed at the same time revealed a relatively higher sensitivity (22/22), but a very poor specificity (0/8) . Gadolinium-diethylene-triamine-pentaacetic acid (Gd-DTPA) enhanced images did not improve specificity . In the later course of infection (clinical signs and symptoms > 10 days), MRIs showed typical nodular target-like lesions with Gd-DTPA enhancement of the rim area that was not seen in the early course of the disease or in patients with Pseudomonas or staphylococcal infection . In conclusion, MRI findings are not as characteristic as the CT halo sign in diagnosing IPA in the early course of the disease, but the MRI target sign with Gd-DTPA enhancement of the rim area and the "reverse target" on T2-weighted images are strongly suggestive of IPA at a later stage of the disease.

Cell Immunol, 1994 Oct 1, 158(1), 83 - 95
Differential effect of staphylococcal enterotoxin B upon the induction of tolerance on peripheral CD4+V beta 8+ and CD8+V beta 8+ T cells; Sabapathy TK et al.; It is known that bacterial superantigens can interact with certain V beta elements of the T cell receptor to result in the activation, expansion, anergy, and/or deletion of T cells . The induction of peripheral T cell tolerance in AKR/J mice was examined in relation to the amount of staphylococcal enterotoxin B (SEB) administered and it was found that the events leading to the induction of tolerance of V beta 8+ T cells was dependent on the initial dose of superantigen employed . Following administration of a large amount (> or = 10 micrograms) of SEB into AKR/J mice, expansion of both CD4+V beta 8+ and CD8+V beta 8+ T cells was observed . This initial cell expansion was followed by the decline in the number of CD4+V beta 8+ T cells . The number of CD8+V beta 8+ T cells, however, did not decline and remained high . When a small amount (2 micrograms) of SEB was employed, it did not stimulate T cell expansion in AKR/J mice . However, when these mice were challenged with SEB, anergy was observed in the CD4+V beta 8+ T cells regardless of the initial dose of SEB . In contrast, the CD8+V beta 8+ T cells were not anergized and were able to proliferate on stimulation with a second dose of SEB . The state of anergy for the CD4+V beta 8+ T cells lasted for at least 70 days, and by 150 days the anergic state was relieved and these CD4+V beta 8+ T cells were once again able to proliferate in response to SEB . On the other hand, continuous SEB exposure resulted in the decline of both CD4+V beta 8+ and CD8+V beta 8+ T cells . Although the number of CD4+V beta 8+ and CD8+V beta 8+ T cells apparently returned to normal levels by 150 days, the state of anergy persisted, as demonstrated by the reduction of the response of these T cells following SEB stimulation in vitro . Our data suggest that the initial expansion of T cells is not an absolute prerequisite for the induction of peripheral T cell anergy . Moreover, the continuous presence of superantigen is essential for the deletion and maintenance of a state of anergy for CD8+V beta 8+ T cells.

Surg Oncol, 1994 Oct, 3(5), 279 - 85
The bispecific antibody 500A2 x 96.5 targets T-lymphocytes activated in vivo with staphylococcal enterotoxin B (SEB) against CL62 melanoma cells in vitro; Reid IM et al.; Bispecific antibodies (BAb) direct T-lymphocytes to lyse selected tumour targets, both in vitro and in vivo . Significant tumour cell lysis with BAb requires pre-expansion of T-lymphocytes, which may be achieved in vitro by the addition of anti-CD3 monoclonal antibody plus interleukin-2 (IL-2), but anti-CD3 may cause immunosuppression . We investigated an alternative agent for in vivo immunostimulation, staphyloccal enterotoxin B (SEB), which selectively activates certain T-cell subsets and may result in less immunosuppression than with anti-CD3 . We activated T-lymphocytes in vivo with SEB, expanded them in vitro with IL-2, and directed them against a tumour target with the BAb 500A2 x 96.5, specific for the murine CD3 antigen and the melanoma p97 antigen expressed by the CL62 tumour . C3H mice received SEB 50 micrograms intraperitoneally (i.p.) . After 18 h mice were sacrificed and splenocytes extracted and either passed over a nylon wool column to isolate T-lymphocytes, or cultured in vitro for 3 to 7 days with 100 U ml-1 of IL-2 . A 4-h chromium-release assay was used to assess the ability of T-lymphocytes to lyse the tumour target CL 62 in the presence or absence of the bispecific antibody 500A2 x 96.5 . The addition of BAb significantly enhanced tumour lysis by SEB activated cells after a period of in vitro culture with IL-2 . In vivo SEB results in the activation of T-lymphocytes which may be directed by bispecific antibodies to increase the lysis of selected tumour targets in vitro.

J Biochem (Tokyo), 1994 Oct, 116(4), 898 - 904
Targeting of the immunoglobulin-binding domain of protein A to the extracellular matrix using a minifibronectin expression vector; Matsuyama S et al.; A truncated form of fibronectin consisting of the N-terminal 70-kDa and C-terminal 37-kDa regions, referred to as "minifibronectin," retains the ability to assemble into the extracellular matrix, even though it lacks the central approximately 120-kDa region containing most of the type III modules (Ichihara-Tanaka, K., Titani, K., and Sekiguchi, K., FEBS Lett . 299, 155-158, 1992) . Taking advantage of the matrix assembly activity of minifibronectin, we developed a novel method to target non-matrix proteins to the extracellular matrix by inserting them between the N-terminal 70-kDa and the C-terminal 37-kDa regions of minifibronectin . Using the immunoglobulin-binding domain of Staphylococcal protein A as a model, we demonstrated that the bacterial protein expressed in mouse L cells as a chimeric protein with minifibronectin is secreted as disulfide-bonded dimers and successfully deposited onto the extracellular matrix of transfected cells . The chimeric protein retained the immunoglobulin-binding activity not only in solution but also after deposition at the matrix . This targeting strategy we developed will provide a means to manipulate the biological functions of the extracellular matrix through targeting of a wide variety of non-matrix proteins.

Bol Asoc Med P R, 1994 Oct-Dec, 86(10-12), 84 - 7
{Infections of penile prosthesis: treatment and prevention}; Olivo V et al.; To date, there are 10,000,000 men with impotence in the United States and it is estimated that at least 17,000 penile prosthesis are implanted annually . The most fearsome complication is the infection of the prosthesis which is usually caused by Staphylococcus epidermidis (in 40-80% of the cases) . In general, the incidence of infection is actually 0.8-8.3%, but it can increase to 37% in patients with tertiary implants . The initial empiric treatment is usually with vancomycin and aminoglycosides and prophylaxis is recommended with a penicillinase-resistant synthetic penicillins, first generation cephalosporins, or vancomycin in case of penicillin allergy.

Vet Immunol Immunopathol, 1994 Oct, 43(1-3), 53 - 62
Porcine peripheral blood CD4+/CD8+ dual expressing T-cells; Pescovitz MD et al.; The porcine T-cell population is unique in that there is a large percentage of CD+CD8+ dual expressing peripheral T-cells . This paper reviews the data available on these porcine T-cells and compares them to the much rarer dual expressing T-cells in humans . The percent of dual expressing cells increases with activation in in vitro culture with various antigens including pseudorabies virus . The percent of resting dual expressing cells also increases with the age of the pig . Flow-cytometric-sorted dual expressing cells responded in culture to the super antigen staphylococcal enterotoxin B . Selected CD4+CD8- cells cultured in vitro developed expression of CD8 and maintained the dual expressing phenotype for the 12 weeks of culture . Dual expressing cells freshly prepared from porcine blood did not express the IL-2 receptor as demonstrated by their failure to bind FITC-IL-2 and an anti-porcine IL-2 receptor monoclonal antibody . In response to activation with phorbol myristic acetate, CD4, but not CD8, was down regulated on the dual expressing T-cells . In summary, porcine dual expressing T-cells constitute a substantial percentage of the porcine peripheral T-cell pool . These cells appear to contain the majority of the memory T-cell with their frequency increasing with blood donor age and in vitro culture . Although the receptor specificity is not known, they have a functional receptor . Finally, the function of the two accessory molecules CD4 and CD8 in these cells is not known, but their regulation is distinct, thereby suggesting no equivalent roles in immune function.

Protein Eng, 1994 Oct, 7(10), 1209 - 20
Protein stability for single substitution mutants and the extent of local compactness in the denatured state; Miyazawa S et al.; The stability changes caused by single amino acid substitutions are studied by a simple, empirical method which takes account of the free energy change in the compact denatured state as well as in the native state . The conformational free energy is estimated from effective inter-residue contact energies, as evaluated in our previous study . When this method is applied, with a simple assumption about the compactness of the denatured state, for single amino acid replacements at Glu49 of the tryptophan synthase alpha subunit and at Ile3 of bacteriophage T4 lysozyme, the estimates of the unfolding Gibbs free energy changes correlate well with observed values, especially for hydrophobic amino acids, and it also yields the same magnitudes of energy as the observed values for both proteins . When it is also applied for amino acid replacements at various positions to estimate the average number of contacts at each position in the denatured state from the observed value of unfolding free energy change, those values for replacements with Gly and Ala at the same residue position in staphylococcal nuclease correlate well with each other . The estimated numbers of contacts indicate that the protein is not fully expanded in the denatured state and also that the compact denatured state may have a substantially native-like topology, like the molten globule state, in that there is a weak correlation between the estimated average number of contacts at each residue position in the denatured state and the number of contacts in the native structure . These results provide some further evidence that the inter-residue contact energies as applied here (i) properly reflect actual inter-residue interactions and (ii) can be considered to be a pairwise hydrophobicity scale . Also, the results indicate that characterization of the denatured state is critical to understanding the folding process.

Biochem Mol Biol Int, 1994 Oct, 34(3), 621 - 6
Expression of a biologically active human granulocyte-macrophage colony stimulating factor fusion protein in Escherichia coli; Hua Z et al.; A gene encoding human granulocyte-macrophage colony stimulating factor(GM-CSF), was cloned into plasmid pEZZ318 and fused to a DNA segment coding for the signal peptide of staphylococcal protein A and a synthetic gene coding for a protein with ability to bind immunoglobulin G(IgG) . The fusion protein was expressed in Escherichia coli and biologically actively secreted into the growth medium . Approximately all of the total activity was secreted into the culture medium, where levels of activity approached 1.96 x 10(8) units/liter . Purification of the fusion protein was performed in a single step by affinity chromatography with immobilized IgG to a specific activity of 1.2 x 10(8) units/mg.

J Cell Biochem, 1994 Oct, 56(2), 177 - 82
Triggers and switches in a self-assembling pore-forming protein; Bayley H; Protein engineering is being used to produce a collection of pore-forming proteins with applications in biotechnology . Knowledge provided by investigations of the mechanism of self-assembly of staphylococcal alpha-hemolysin has allowed the design of genetically and chemically modified variants of the protein with pore-forming activities that can be triggered or switched on-and-off by chemical, biochemical and physical inputs . Examples include alpha-hemolysins that are activated by specific proteases and alpha a-hemolysins whose activity is controlled by divalent metal ions . These proteins have potential value in drug delivery as components of immunotoxins that can be activated at the surfaces of target cells . Further applications are likely in improved encapsulation techniques for drugs, enzymes and cells.

Int Immunol, 1994 Oct, 6(10), 1555 - 60
Age-dependent changes in the response to staphylococcal enterotoxin B; Aroeira LS et al.; In the present study we investigated the response of old mice to staphylococcal enterotoxin B (SEB) immunization . Old mice were susceptible to lethal toxic shock, probably mediated by tumor necrosis factor-alpha, although lethal toxic shock was not observed in young mice . Old mice were able to produce more IL-2 and IL-4 than young mice in response to in vivo immunization with SEB . V beta 8+CD4+ T cells of old mice expanded less in vivo and were not deleted in response to SEB . However, in spite of the absence of clonal deletion, SEB was found to induce energy of SEB reactive cells in old mice, as demonstrated by reduced in vitro T cell proliferation to SEB and reduced in vitro IL-2 and IL-4 production.

Int Immunol, 1994 Oct, 6(10), 1475 - 83
Requirement of a second signal from antigen presenting cells in the clonal deletion of immature T cells; Aiba Y et al.; The role of antigen presenting cells (APC) in T cell clonal deletion was investigated by culturing murine thymic lymphocytes with the superantigen staphylococcal enterotoxin B (SEB) in the absence or presence of APC . As the APC, we used B lymphoma cell lines A20.2J and BAL17.2, both expressing MHC class II antigens at high levels . SEB reactive V beta 8+ cells were deleted only when A20.2J cells were used as APC . By using thymocytes from transgenic mice carrying a TCR beta chain transgene, it was further shown that the deletion occurred at the CD4+CD8+ stage . The other cell line, BAL17.2, failed to induce clonal deletion, although this cell line was able to stimulate the proliferative response of SEB-primed T cells . The activity of A20.2J cells to induce clonal deletion was completely abolished by fixation with paraformaldehyde, whereas the same treatment kept the ability of this cell line to induce the proliferative response of non-primed as well as SEB-primed T cells . It was further shown that the deletion was abolished by the addition of anti-MHC class II but not anti-B7 mAb in the culture . These results provided explicit evidence that a signal(s) from APC, which is distinct from that required for primary or secondary proliferative response of mature peripheral T cells, is involved in clonal deletion of thymic immature T cells.

AIDS, 1994 Oct, 8(10), 1397 - 404
Staphylococcal superantigens activate HIV-1 replication in naturally infected monocytes; Goujard C et al.; OBJECTIVES: To study the effects of microbial superantigens, Staphylococcal exotoxins (SE), on HIV replication in monocytes following binding to and signalling through major histocompatibility complex (MHC) class II molecules . METHODS: We investigated the effects of SE on HIV replication and monokine production in three different in vitro models of monocyte culture: chronically infected monocytic cell line U1, acute infection of normal monocytes by different HIV-1 strains, and naturally-infected monocytes from seropositive patients . p24 antigen, interleukin (IL)-6 and tumour necrosis factor (TNF)-alpha production was measured by specific enzyme-linked immunosorbent assay (ELISA) . RESULTS: Staphylococcal enterotoxin B and toxic shock syndrome toxin-1 (1-1000 ng/ml) are powerful inducers of HIV-1 expression in U1 cells pretreated with granulocyte macrophage colony stimulating factor . SE induce viral replication in short-term cultures (days 6-21) of monocytes infected in vitro by HIVBa-L, HIVLAI, or naturally infected in vivo . Induction of HIV expression requires direct interactions of SE with MHC class II molecules but not T-cell receptor binding and T-cell-monocyte contact . Anti-TNF-alpha and anti-IL-6 neutralizing monoclonal antibodies inhibit by over 61% SE-induced HIV replication . CONCLUSIONS: Using SE we have linked two important pathways for the regulation of HIV replication in monocytes, namely signalling through MHC class II molecules and monokine production potentially mediated by induction of the pleiotropic cellular transcription factor NF-kappa B . In HIV-infected patients bacterial infections are common and could be an important cofactor in the immunopathogenesis of AIDS by inducing HIV replication in latently infected monocytes . Their prevention might emerge as beneficial in these patients.

Br J Ophthalmol, 1994 Oct, 78(10), 772 - 4
Lack of effect of preoperative norfloxacin on bacterial contamination of anterior chamber aspirates after cataract surgery; Chitkara DK et al.; Eighty patients undergoing routine standardised extracapsular cataract surgery with lens implantation were divided randomly into two groups in a prospective double blind study comparing effects of preoperative norfloxacin eyedrops with placebo on bacterial contamination of anterior chamber aspirates after surgery . Pathogenic organisms were identified from 19 (24%) of the aspirates . The most commonly isolated organisms were coagulase negative Staphylococcus species . There was no statistical difference between the norfloxacin treated and placebo groups . This study demonstrates that routine use of topical preoperative antibiotics to eliminate the entry of bacteria into the eye during surgery is debatable.

J Microsc, 1994 Oct, 176 ( Pt 1), 8 - 16
Rapid estimation of bacterial antibiotic susceptibility with flow cytometry; Mason DJ et al.; Bacterial antibiotic susceptibility was rapidly estimated for Escherichia coli and Staphylococcus spp . by flow cytometry . This was achieved by measuring the uptake of a negatively charged membrane potential sensitive dye bis-(1,3-dibutyl-barbituric acid) trimethine oxonol and observing changes in low-angle light scatter (excitation light scattered by up to 15 degrees) . Estimations of ampicillin, gentamicin and ciprofloxacin susceptibilities were possible within 2-5 h from a plate culture, depending on the species and antibiotic used . This includes the time necessary to establish steady-state growth in liquid culture.

J Clin Epidemiol, 1994 Oct, 47(10), 1173 - 4
Are public library books contaminated by bacteria?
Brook SJ, Brook I.
The microbial flora on the surfaces of 15 books obtained from a public library and from 15 books obtained from a family household were studied . Staphylococcus epidermidis was recovered from 4 of the library books and 3 of the family household books . The number of organisms per page was between one to four . This data illustrates the safety of using library books, as they do not serve as a potential source of transmission of virulent bacteria.

Zentralbl Veterinarmed B, 1994 Oct, 41(7-8), 529 - 40
Effect of dry-cow therapy on subclinical mastitis--an evaluation of long-acting and short-acting intramammaria; Osteras O et al.; This field study was conducted to evaluate the effect of selective dry-cow therapy with long-acting and short-acting antibiotics, respectively, and also in comparison to control groups without antibiotic treatment . A total of 684 cows from 288 different herds in three Norwegian regions fulfilled the criteria of the study design . There were 104 cows in control group A (sampling only), 115 cows in control group B (placebo), 221 cows treated with long-acting intramammaria Benestermycin vet . 'Leo' for 1 day at drying off in group C, and 244 cows treated with four short-acting intramammaria Leocillin with Dihydrostreptomycin vet . 'Leo' every second day before drying off in group D . The overall effect, measured as the cow being healthy after therapy, was 14.2% in control groups and 33.7% in therapy groups 30 +/- 17 days into the next lactation . Of quarters infected with S . aureus both in late lactation (45 +/- 32 days before drying off) and at drying off, 38.4% in the control group were bacteriologically negative 30 +/- 17 days into the next lactation, compared with 49.5% in the long-acting group and 68.6% in the short-acting group . Of quarters infected with Str . dysgalactiae both in late lactation (45 +/- 32 days before drying off) and at drying off, 10 out of 27 were still infected with Str . dysgalactiae in the control group 30 +/- 17 days into next lactation, compared with 0 out of 31 in the therapy groups . Dry-cow therapy in coagulase-negative Staphylococcus spp . (CNS)-infected quarters led to a 5.2 odds ratio of being healthy quarters 30 +/- 17 days into the next lactation, compared with control groups . Despite this, the overall frequency of CNS in the material was unchanged after therapy compared with controls . Short-acting compared to long-acting preparations had a significantly better effect in preventing new infection with S . aureus or Str . dysgalactiae in untreated healthy quarters in cows with fewer than three infected quarters . This difference in preventive effect was greater in cows with one infected quarter during previous lactation (the new infection rates being 0.078 for short-acting and 0.149 for long-acting) than in those with two infected quarters (the new infection rates being 0.042 and 0.063, respectively).

Mol Pharmacol, 1994 Oct, 46(4), 618 - 26
Physicochemical and immunocytochemical analysis of the aryl hydrocarbon receptor nuclear translocator: characterization of two monoclonal antibodies to the aryl hydrocarbon receptor nuclear translocator; Hord NG et al.; The aryl hydrocarbon receptor nuclear translocator (Arnt) is a basic helix-loop-helix transcription factor that heterodimerizes with the aryl hydrocarbon receptor to mediate signal transduction pathways inducible by 2,3,7,8-tetrachlorodibenzo-p-dioxin and other planar aromatic hydrocarbons . Monoclonal antibodies (MAbs) have been raised against a carboxyl-terminal 19-amino acid peptide hapten (MAb 2B10) and against a carboxyl-terminal 378-amino acid polypeptide-staphylococcal Protein A fusion protein (MAb 4G9) of Arnt and their characterization is described . Western blot experiments show that both MAbs specifically cross-react with an approximately 85-kDa band in cytosol prepared from COS-7 cells transfected with the full length human Arnt cDNA pBMSNeo-D24-1 and in Hepa 1c1c7 cytosol but not in Arnt-deficient Hepa 1-C4 mutant cytosol . Velocity sedimentation of Hepa 1c1c7 cytosol on sucrose gradients and Superose 6 gel permeation chromatography were used to estimate the sedimentation coefficient . Stokes radius, and relative molecular mass of Arnt as approximately 3.6-4.1 S, 6.8 nm, and 101-115 kDa, respectively . These results indicate that Arnt probably exists in monomeric form in Hepa 1c1c7 cytosolic extracts . Laser scanning confocal microscopy and indirect immunofluorescence microscopy revealed Arnt to be distributed throughout the non-nucleolar portion of the nucleus of Hepa 1c1c7, VT(2) (Hepa 1-C4T mutant cell line deficient in Arnt function and stably transfected with pBMSNeo D24-1, expressing the full length human Arnt cDNA), and HeLa cells . The establishment of the nuclear localization of Arnt in human and murine cell lines shown here indicates that its nuclear localization may be conserved across species . Immunofluorescence analysis of Arnt in three cell lines using two MAbs (to distinct epitopes) provides evidence that suggests that the aryl hydrocarbon receptor heterodimerizes with Arnt in the nucleus.

J Lab Clin Med, 1994 Oct, 124(4), 537 - 45
Albumin binding surfaces for biomaterials; Keogh JR et al.; The surfaces of medical devices may promote both coagulation and infections caused by adherent microorganisms . In the case of polymeric elastomers, these iatrogenic effects are likely intermediated by absorbed host proteins that spontaneously bind to the device surface, promoting both bacterial adherence and thrombotic events . We earlier attempted to produce biomaterial surfaces that would selectively bind host albumin because albumin-coated surfaces were known to diminish both coagulation and bacterial adherence . To this end, an albumin-binding high molecular weight dextran:Cibacron blue adduct was bulk incorporated into polyetherurethane (Keogh et al., J Biomed Mater Res 1992;26:441) . The modified material bound albumin selectively and reversibly and showed evidence of enhanced biocompatibility . However, approximately 30% of the surface of this material was evidently unmodified and still capable of exerting the above adverse effects . In the present work, we have covalently surface-modified polyetherurethane with sequential additions of acrylamide, amino-propylmethacrylamide, dextran, and Cibacron blue . This derivatized polyurethane preferentially and reversibly binds albumin, even from complex mixtures of proteins such as plasma . Furthermore, this material inhibits the clotting of nonanticoagulated whole human blood (for > 16 hours at room temperature), perhaps by virtue of binding and activation of antithrombin III by the sulfonic acid residues on the surface-immobilized Cibacron blue . Finally, such surfaces, especially when bearing bound albumin, diminish the adherence of Staphylococcus epidermidis, a pathogen frequently associated with device-centered infections . We conclude that similar albumin-affinity surfaces may hold promise for the development of more biocompatible materials for implantation and blood contact applications.

J Mol Biol, 1994 Sep 30, 242(4), 527 - 46
Backbone dynamics of a highly disordered 131 residue fragment of staphylococcal nuclease; Alexandrescu AT et al.; In order to characterize the dynamic properties of the denatured state of staphylococcal nuclease, R1, R2, and NOE relaxation parameters have been measured for the backbone 15N nuclei of a 131 residue fragment that serves as a model of the denatured state under non-denaturing conditions . The relaxation data indicate a wide range of amplitudes for segmental motion and are inconsistent with a random coil conformation . An optimal value of 7.8 ns was obtained for the molecular rotational correlation time tau m based on the analysis of the 79 residues for which R1, R2, and NOE relaxation data could be obtained . This value corresponds roughly to the slowest detectable motion on the nanosecond time scale and is of a magnitude consistent with global tumbling of a large portion of the molecule . For the majority of residues, experimental data could be described most adequately in terms of a modified "model-free" formalism which includes contributions from internal motions on both an intermediate (tau e) and a fast time scale (tau f) in the context of slow overall tumbling (tau m) . The generalized order parameters S2, which gives the amplitude of motions on time scales faster than tau m, correlates with sequence hydrophobicity and suggests a relationship between chain flexibility and sequence propensity for hydrophobic collapse . The fractional populations of three alpha-helices in the protein show a stronger correlation with S2 values and hydrophobicities than with intrinsic helix propensities . These observations suggest that secondary structure may be preferentially stabilized in hydrophobic segments of the sequence.

J Immunol Methods, 1994 Sep 30, 175(1), 47 - 58
Early expression of cytokines in lymph nodes after treatment in vivo with Staphylococcus enterotoxin B; Litton MJ et al.; Excessive cytokine expression induced by superantigen may be one aspect of the pathophysiology associated with Gram positive bacteremia . We have undertaken a study of the kinetics of cytokine production in lymph nodes obtained from in vivo Staphylococcus enterotoxin B (SEB) treated animals . This study was designed to evaluate the short term cytokine profile observed using immunohistochemistry (IHC) in BALB/c mice injected intraperitoneally (i.p.) . The observed immunohistochemical kinetic profiles were corroborated using reverse transcription-polymerase chain reaction (RT-PCR) RNA analysis . We report here that TNF, IL-2, and IFN-gamma are the principal cytokines which were detected within hours of SEB administration, and that other cytokines such as IL-3, IL-4, IL-5, IL-6, IL-10, GM-CSF and M-CSF were undetectable . TNF and IL-2 appeared very early following SEB priming, and were observed by 1 h . IFN-gamma which appeared later (maximally at 14 h) was produced predominantly by CD8+ cells . In contrast, the TNF and IL-2 were produced primarily by CD4+ cells . Identical results were obtained by IHC and RT-PCR; the kinetics of mRNA expression slightly preceded the appearance of protein . The TNF and IFN-gamma staining patterns observed in lymph node sections were indicative of Golgi-localized cytokine . The IL-2 staining pattern observed in lymph node sections was distinctive, covering a significant local area of cells . This local regional concentration of IL-2, which may result from cytokine attached to extracellular binding components, may be an important aspect of the activation phase of a developing immune response . Rapid induction and excessive cytokine production elicited by superantigen in vivo, may ultimately help to explain the shock and death associated with SEB.

Biochem Biophys Res Commun, 1994 Sep 15, 203(2), 1083 - 9
Location of the binding site for the allosteric activator IMP in the COOH-terminal domain of Escherichia coli carbamyl phosphates synthetase; Bueso J et al.; Using UV-irradiation we cross-linked IMP, the allosteric activator of E . coli carbamyl phosphate synthetase (a heterodimer of 117.7 and 41.4 kDa subunits), to the large subunit of the enzyme . As in the native enzyme-IMP complex, the cross-linked complex was resistant to attack by trypsin . Thus, IMP is attached to its normal site and induces the normal conformational changes . Limited digestion of the {3H}IMP-labeled enzyme with V8 staphylococcal protease or with trypsin in the presence of SDS, and NH2-terminal sequencing, showed that {3H}IMP is cross-linked to the COOH-terminal 20 kDa domain of the large subunit, downstream of residue 912, supporting the proposal that this domain is specialized in effector binding and regulation.

J Immunol, 1994 Sep 15, 153(6), 2618 - 23
Systemic release and protective role of IL-10 in staphylococcal enterotoxin B-induced shock in mice; Florquin S et al.; Staphylococcal enterotoxin B (SEB) is a bacterial superantigen that induces the production of several pro-inflammatory cytokines, leading to a self-limited shock . In the present study, we show that SEB also triggers the systemic release of IL-10, an anti-inflammatory and immunosuppressive cytokine . Serum IL-10 was undetectable (< 1000 pg/ml) in control BALB/c mice and rose to 8500 +/- 2850 pg/ml (mean +/- SEM) 4 h after injection of 100 micrograms SEB . Cell depletion experiments and analysis of IL-10 mRNA expression indicated that CD4+ cells played a major role in SEB-induced IL-10 production . Pretreatment of mice with neutralizing anti-IL-10 mAb before SEB challenge did not modify the release of TNF but led to increased and sustained IL-2 and IFN-gamma serum levels . Furthermore, although no lethality occurred in mice injected with SEB and control mAb, injection of anti-IL-10 mAb before SEB resulted in a 30% lethality (p < 0.05) . This lethality was completely prevented by anti-IFN-gamma mAb injection, indicating that IFN-gamma plays a crucial role in the increased toxicity of SEB in anti-IL-10 mAb-injected mice . We conclude that SEB induces the production of IL-10 by CD4+ cells in vivo and that endogenous IL-10 plays an important immunoregulatory role in this model by down-regulating IL-2 and IFN-gamma production.

J Biotechnol, 1994 Sep 15, 37(1), 79 - 83
Staphylococcal protein A is a novel heterologous substrate for the HIV-1 protease; Marczinovits I et al.; Upon in vitro processing of the recombinant HIV-1/gag p24 protein, expressed in Escherichia coli as a fusion protein, by HIV-1 protease, a cleavage site within the staphylococcal protein A fusion partner was found . N-terminal sequencing of the protein A fragments showed that HIV-1 protease cleavage occurred between phenylalanine-235 and tyrosine-236 within the sequence Gln-Asn-Ala-Phe/Tyr-Glu-Ile-Leu (QNAF/YEIL) in the IgG-binding domain C of the protein A encoded by the pRIT2T fusion gene vector (Pharmacia) . Results presented here have proven that the protease-sensitive site is viable in vitro on the protein A alone and other chimeric protein, protein A/beta-galactosidase . A possible significance of this phenomenon in biotechnology work is discussed.

J Immunol, 1994 Sep 15, 153(6), 2479 - 87
Costimulation of human CD4+ T lymphocytes with B7 and lymphocyte function-associated antigen-3 results in distinct cell activation profiles; Parra E et al.; This study describes the distinct roles of B7 and LFA-3 in the regulation of T cell responses . Activation of CD4+ T cells with Chinese hamster ovary (CHO)-DR4/B7 and CHO-DR4/LFA-3 cells that present the superantigen staphylococcal enterotoxin A resulted in significant T cell proliferation and substantial production of TNF and IFN-gamma . Strong IL-2 production was recorded in B7-costimulated, but not LFA-3-costimulated, cultures . The presence of B7 induced a more vigorous and prolonged proliferative T cell response compared with LFA-3 costimulation . In contrast, LFA-3 was more efficient than B7 in mediating cell adhesion of CD4+ T cells . Costimulation with the CHO-DR4/B7/LFA-3 triple transfectant resulted in enhanced cell adhesion, proliferation, and cytokine production compared with either DR4/B7 or DR4/LFA-3 alone . Optimal production of IL-2 by naive and memory CD4+ T cells was seen only when cells were costimulated with B7, whereas IFN-gamma production was induced in memory cells by both LFA-3 and B7 . The Jurkat T cell line responded to CHO-DR4/B7/LFA-3 in a manner similar to peripheral blood CD4+ T cells . Reverse transcriptase-PCR analysis of Jurkat cells stimulated with staphylococcal enterotoxin E and the different CHO transfectants revealed that the cooperative effect of B7 and LFA-3 on IL-2 production was also seen at the mRNA level . The large amounts of IL-2 produced by B7 costimulation indicate a paracrine function of the B7/CD28 pathway, whereas the LFA-3/CD2 pathway provides strong adhesion and may facilitate autocrine T cell expansion . Combined expression of the B7 and LFA-3 molecules seems to provide an optimal Ag-presenting function that ensures strong adhesion and optimal signal transduction.

Proc Natl Acad Sci U S A, 1994 Sep 13, 91(19), 8945 - 9
Monoclonal antibody-superantigen fusion proteins: tumor-specific agents for T-cell-based tumor therapy; Dohlsten M et al.; The bacterial superantigen staphylococcal enterotoxin A (SEA) is an extremely potent activator of T lymphocytes when presented on major histocompatibility complex (MHC) class II molecules . To develop a tumor-specific superantigen for cancer therapy, we have made a recombinant fusion protein of SEA and the Fab region of the C215 monoclonal antibody specific for human colon carcinoma cells . SEA as part of a fusion protein showed a > 10-fold reduction in MHC class II binding compared to native SEA, and accordingly, the affinity of the FabC215-SEA fusion protein for the C215 tumor antigen was approximately 100-fold stronger than to MHC class II molecules . The FabC215-SEA fusion protein efficiently targeted T cells to lyse C215+ MHC class II- human colon carcinoma cells, which demonstrates functional substitution of the MHC class II-dependent presentation of SEA with tumor specificity . Treatment of mice carrying B16 melanoma cells expressing a transfected C215 antigen resulted in 85-99% inhibition of tumor growth and allowed long-term survival of animals . The therapeutic effect was dependent on antigen-specific targeting of the FabC215-SEA fusion protein, since native SEA and an antigen-irrelevant FabC242-SEA fusion protein did not influence tumor growth . The results suggest that Fab-SEA fusion proteins convey superantigenicity on tumor cells, which evokes T cells to suppress tumor growth.

Biochemistry, 1994 Sep 6, 33(35), 10842 - 50
Three-state thermodynamic analysis of the denaturation of staphylococcal nuclease mutants; Carra JH et al.; Using microcalorimetry, we found an equilibrium intermediate state during the denaturation of the wild-type and five mutant staphylococcal nuclease proteins: V66L, V66W, G88V, D77A, and E75V . The presence of two distinct heat absorption peaks allowed direct measurement of the enthalpy differences between the native, intermediate, and denatured states . Conditions of low pH and high NaCl concentration facilitated observation of the intermediate, or I-state . We propose to consider the nuclease protein as composed of two subdomains, divided along the active-site cleft . The structure of the I-state apparently consists mainly of the folded beta-barrel subdomain, as does that of a nuclease fragment protein {Shortle, D., & Abeygunawardana, C . (1993) Structure 1, 121-134} . The cooperativity of folding of the subdomains is maintained by electrostatic bonds across the active-site cleft . Removal of these bonds by the mutation D77A or E75V results in decooperation of the protein's structure and a three-state mechanism of denaturation at pH 7.0 . The origins of differences in the enthalpy change of denaturation and in the m value of guanidinium chloride-induced denaturation with mutant nucleases are discussed in terms of this three-state mechanism.

Eur J Immunol, 1994 Sep, 24(9), 2081 - 6
Induction of clonal anergy by oral administration of staphylococcal enterotoxin B; Migita K et al.; The staphylococcal enterotoxin is a major cause of food poisoning . The bacterial substance stimulates T cells expressing specific V beta T cell receptors (TcR) and is termed "the superantigen" . We have previously demonstrated that intravenous injection of staphylococcal enterotoxin B (SEB) induces functional unresponsiveness (anergy) of reactive T cells as well as a partial deletion by activation-induced programmed cell death . In the present study, we examined the effect of oral administration of SEB in mice . Our results indicate that spleen T cells from SEB-primed mice are hyporesponsive to SEB stimulation in vitro, but the response to SEA was normal . V beta 8+ T cells purified from SEB-primed mice did not respond to stimulation of TcR . This SEB-specific unresponsiveness could not be reversed by exogenous interleukin-2, but was partially reversed by phorbol 12-myristate 13-acetate . Activation of mitogen-activated protein kinase during TcR-mediated stimulation was significantly inhibited in anergic T cells . Although the mechanisms of oral tolerance are not well understood, these results show that oral administration of SEB induce clonal anergy in peripheral T cells.

J Bone Joint Surg Br, 1994 Sep, 76(5), 725 - 7
Methicillin-resistant Staphylococcus epidermidis in infection of hip arthroplasties; James PJ et al.; We investigated the incidence of cephalosporin-resistant bacteria in infected hip arthroplasties . Of 740 patients having hip replacement or related procedures performed over three years, 30 had positive bacteriological cultures from tissue removed at the time of surgery . In 18 of the 30 cultures Staphylococcus epidermidis was grown and 12 of these were methicillin-resistant . A prospective study of skin swabs taken from 100 consecutive patients at the time of admission for THR showed methicillin-resistant Staphylococcus epidermidis in 25 . This cephalosporin-resistant organism was shown to be the commonest proven cause of infection, and its presence as a skin commensal raises important questions about current antibiotic prophylaxis for joint replacement.

Infect Immun, 1994 Sep, 62(9), 3907 - 15
Binding and activation of major histocompatibility complex class II-deficient macrophages by staphylococcal exotoxins; Beharka AA et al.; Macrophages from C2D transgenic mice deficient in the expression of major histocompatibility complex (MHC) class II proteins were used to identify binding sites for superantigens distinct from the MHC class II molecule . Iodinated staphylococcal enterotoxins A and B (SEA and SEB) and exfoliative toxins A and B (ETA and ETB) bound to C2D macrophages in a concentration-dependent and competitive manner . All four toxins increased F-actin concentration within 30 s of their addition to C2D macrophages, indicating that signal transduction occurred in response to toxin in the absence of class II MHC . Furthermore, ETA, ETB, SEA, and, to a lesser extent, SEB induced C2D macrophages to produce interleukin 6 . Several molecular species on C2D macrophages with molecular masses of 140, 97, 61, 52, 43, and 37 kDa bound SEA in immunoprecipitation experiments . These data indicate the presence of novel, functionally active toxin binding sites on murine macrophages distinct from MHC class II molecules.

Infect Immun, 1994 Sep, 62(9), 3837 - 43
Serum-induced potentiation of tumor necrosis factor alpha production by human monocytes in response to staphylococcal peptidoglycan: involvement of different serum factors; Mattsson E et al.; Peptidoglycan from a Staphylococcus epidermidis strain, isolated from a patient with septicemia, was preincubated with human serum . This mixture was then investigated for its potency to induce tumor necrosis factor (TNF) secretion by human blood monocytes . TNF was measured in the supernatants by using a bioassay and/or an enzyme-linked immunosorbent assay specific for TNF alpha (TNF-alpha) . Although earlier studies indicated that staphylococcal peptidoglycan alone is a relatively poor stimulator of TNF-alpha production, the present study shows that human serum highly potentiates peptidoglycan-induced TNF-alpha release by human monocytes . In the presence of serum and in the low-dose range, peptidoglycan was almost as potent as endotoxin . At high peptidoglycan concentrations, monocytes showed an extremely high TNF-alpha response, but again only in the presence of serum . At low peptidoglycan doses, the stimulatory effect of serum was abrogated by heat treatment or depleting serum of complement components C1 and C3/C4, which suggests a role for the classical complement pathway . At high doses of peptidoglycan, the serum stimulatory effect depended mainly on immunoglobulin G.

Clin Immunol Immunopathol, 1994 Sep, 72(3), 357 - 61
Superantigens activate HIV-1 gene expression in monocytic cells; Fuleihan R et al.; Binding of superantigens to MHC class II molecules results in transduction of biochemical signals leading to cellular activation and gene expression . We demonstrate that the staphylococcal superantigens toxic shock syndrome toxin-1 (TSST-1) and staphylococcal enterotoxin A (SEA) activate HIV-1-LTR-driven transcription of chloramphenicol acetyl transferase in the human monocytic cell line THP-1 . Induction of HIV-1-LTR-driven transcription in THP-1 cells by superantigens was associated with the induction of nuclear factor-kappa B DNA-binding activity . Superantigens also increased viral protein secretion from the granulocyte-macrophage colony-stimulating factor-pretreated chronically infected human monocytic cell line U1 . Induction of HIV-1 gene expression in monocytic cells by superantigens occurred via tumor necrosis factor-alpha-dependent and -independent mechanisms . Our results suggest that superantigens and other MHC class II ligands may activate HIV-1 gene expression in monocytes/macrophages.

Diabetes Care, 1994 Sep, 17(9), 1064 - 6
Complications of the pump pocket may represent a significant cause of incidents with implanted systems for intraperitoneal insulin delivery; Renard E et al.; OBJECTIVE--To increase awareness of adverse events associated with the use of programmable implantable pumps (PIPs) . CASES--There were 7 cases of complications associated with the pump-pocket among 40 patients treated by PIP, and we searched for risk factors . RESULTS--Seven of 40 type I diabetic patients treated by PIP presented severe complications of the pump-pocket, resulting in five definitive explanations and nine other surgical interventions . The lesions included an exudative reaction in the pump-pocket and a skin retraction or atrophy, which were complicated by skin erosion in five patients . Coagulase-negative staphylococcus was identified in the pump-pocket in four patients, including three cases of skin erosion . No specific risk of local complications could be attributed to age, sex, duration of diabetes, body mass index, presence of retinopathy or peripheral neuropathy, HbA1c level since implantation, depth of implantation in the abdominal wall, or duration of experience with PIP . Usual physical activity corresponding to > 2,000 kcal energy expenditure per week, estimated by a questionnaire, appeared to be the only identified significant risk factor . CONCLUSIONS--From these results, we suggest that physical activity should be limited to moderate exercise and exclude vigorous efforts in diabetic patients treated by PIP to avoid an increased risk of complications at the implantation site.

Zhonghua Min Guo Xiao Er Ke Yi Xue Hui Za Zhi, 1994 Sep-Oct, 35(5), 423 - 8
{Coagulase-negative staphylococcal septicemia}; Huang FY et al.; In the past decade, coagulase negative Staphylococcus (CNS) has become one of the most common pathogens in nosocomial septicemia, especially in neonatal intensive care units . From January 1, 1990 to June 30, 1992, we documented 41 cases of CNS septicemia in the Department of Pediatrics at Mackay Memorial Hospital . Thirty five cases (85%) were found in infants less than 3 months of age . Nineteen patients were premature babies . Of these, 14 had a body weight less than 1,500 gms . All of the patients had intravascular catheters and 40 patients (97.5%) were receiving total parenteral nutrition . The most common causative organism isolated in this study was Staphylococcus hominis . This differs from other reports where Staphylococcus epidermidis is the most common pathogen . Eighty five percent of the organisms were resistant to methicillin and 80% of these methicillin-resistant strains were also resistant to cephalosporins . All of the organisms were sensitive to vancomycin . Because of the possibility of cross-resistance between methicillin and cephalosporin, vancomycin was chosen to treat the methicillin resistant cases . Total parenteral nutrition was discontinued before antibiotic treatment . When CNS septicemia was suspected clinically, we drew two blood cultures from different sites before beginning antibiotic therapy . This was an effort to get a definitive diagnosis . In general, the prognosis of CNS septicemia was good if early diagnosis can be made.

Br J Dermatol, 1994 Sep, 131(3), 371 - 5
Idiopathic CD4+ lymphocytopenia associated with chronic pruritic papules; Wakeel RA et al.; This is a case report and family study of a 65-year-old man with chronic prurigo lesions, in whom we demonstrated a selective deficiency of circulating T-helper/inducer lymphocytes (CD4+), in the absence of any apparent predisposing disease . He is seronegative for human immunodeficiency virus (HIV types 1 and 2) and human T-cell lymphotropic virus (HTLV-I and HTLV-II), and fulfils the criteria for the syndrome of idiopathic CD4+ T lymphocytopenia . He has an atopic diathesis, has had a severe adult chickenpox infection, chronic staphylococcal infections, tinea pedis and recalcitrant warts . He has also suffered from respiratory infections, for which no specific aetiological agent has been identified . His peripheral total lymphocyte count has been persistently abnormal since it was first measured in 1969 . He has a marked CD4+ T-cell lymphocytopenia . His son, who does not have any skin disorder, has a low CD4+ T-cell count.

Zh Mikrobiol Epidemiol Immunobiol, 1994 Sep-Oct, (5), 57 - 9
{The patterns of the corrective action of a natural immunomodulator on the secondary immunological defect caused by a corpuscular pertussis vaccine}; Semenova IB et al.; Purified staphylococcal toxoid (PST) was shown to be capable of preventing the development of secondary antigen-nonspecific immune deficiency in mice, immunized with whole-cell pertussis vaccine . The immunocorrective action of PST was manifested after its injection before (on day -1), simultaneously with and after (on day +1) the injection of whole-cell pertussis vaccine . Correction was either complete or partial, depending on the scheme of the experiment and the dose of PST.

Hum Cell, 1994 Sep, 7(3), 131 - 7
{An efficient methods for the induction of human antitumor effector CD4+ and CD8+ T cells: their application to tumor immunotherapy}; Nishimura T et al.; It is an important issues to investigate an efficient methods to induce antitumor effector T cells from peripheral blood lymphocytes of tumor patients for the development of a novel tumor immunotherapy . We established a large scale culture system of human CD4+ helper/killer T cells which have both helper and killer functions . Targeting of CD4+ helper/killer T cells to tumor using anti-CD3 x anti-c-erbB-2 mAb caused the lysis of tumor and triggering of IL-2 production . It was also demonstrated that culture of human CD4+ T cells with staphylococcal enterotoxin A (SEA) or IL-12 caused a selective induction of Th1 type of CD4+ helper/killer T cells . IL-12 also revealed a novel effect on CD8+CTL functions . Culture of CD8+ T cells with IL-12 resulted in the augmentation of IFN-gamma production and cytotoxicity . Moreover, culture of tumor-infiltrating lymphocytes with IL-12 caused a marked enhancement of CD8+CTL against autologous tumor cells . These findings suggest that IL-12 will become a useful cytokine for the tumor immunotherapy . In this paper, we will discuss the key role of CD4+ T cells for the induction of antitumor immunity in tumor-bearing host.

J Antimicrob Chemother, 1994 Sep, 34(3), 321 - 30
Disruption of the transmembrane pH gradient--a possible mechanism for the antibacterial action of azelaic acid in Propionibacterium acnes and Staphylococcus epidermidis; Bojar RA et al.; The effect of the topical acne treatment azelaic acid on the transmembrane proton gradient (delta pH) of Propionibacterium acnes and Staphylococcus epidermidis was studied in vitro at external pH values found on human skin (pH 4.0-6.0) . Bacteria were grown in defined media using continuous culture and delta pH was estimated by measuring the accumulation of {14C} benzoic by the cells using flow dialysis . In both P . acnes and S . epidermidis the addition of 30 mM azelaic acid and the membrane active inhibitors nigericin (150 microM) and CCCP (150 microM) resulted in a rapid release of {14C} label into the dialysate indicating the dissipation of delta pH between external pH values of 4.0-6.0 . The addition of 60 mM NaCl as an iso-osmotic control and 150 microM valinomycin did not induce the release of {14C} label . The addition of 30 mM azelaic acid reduced the delta pH of P . acnes by 44% at external pH 4.0 and 28% at external pH 6.0 . In S . epidermidis 30 mM azelaic acid reduced delta pH by 88% at external pH 5.0 and 20% at external pH 6.0 . Rapid loss of viability occurred in suspensions of P . acnes and S . epidermidis containing 30 mM azelaic acid at pH 4.0 with no viable cells recovered after 60 min incubation . At pH 6.0 little change in viable numbers of P . acnes and S . epidermidis were observed over a 2 h incubation period . The results indicate that the antibacterial activity of azelaic acid is associated with the perturbation of intracellular pH.

Mol Microbiol, 1994 Sep, 13(5), 897 - 909
Investigation of the role of the disulphide bond in the activity and structure of staphylococcal enterotoxin C1; Hovde CJ et al.; The goal of this study was to investigate the role of the disulphide bond of staphylococcal enterotoxin C1 (SEC1) in the structure and activity of the toxin . Mutants unable to form a disulphide bond were generated by substituting alanine or serine for cysteine at positions 93 and/or 110 . Although we did not directly investigate the residues between the disulphide linkage, tryptic lability showed that significant native structure in the cystine loop is preserved in the absence of covalent bonding between residues 93 and 110 . Since no correlation was observed between the behaviour of these mutants with regard to toxin stability, emesis and T cell proliferation we conclude that SEC1-induced emesis and T cell proliferation are dependent on separate regions of the molecule . The disulphide bond itself is not an absolute requirement for either activity . However, conformation within or adjacent to the loop is important for emesis . Although mutants with alanine substitutions were not emetic, those with serine substitutions retained this activity, suggesting that the disulphide linkage stabilizes a crucial conformation but can be replaced by residues which hydrogen bond.

Pathol Int, 1994 Sep, 44(9), 688 - 96
Lymphokine-activated killer cytotoxicity against pancreas adenocarcinoma cell lines and vascular endothelial cells; Sugiura H et al.; Eight pancreas carcinoma cell lines of duct cell origin (PCI-6, 10, 19, 24, 35, 43, 55, and 64) were established . Using one of these lines, PCI-24, human umbilical vein endothelial cells (HUVEC), and several recombinant cytokines, conditions and specificity of anti-PCI LAK induction were investigated, with the focus on a search for lymphokine-activated killer (LAK) activity that differentiates neoplastic (PCI) from non-neoplastic (HUVEC) cells . Interferon-gamma (IFN-gamma), IFN-alpha, IL-4, IL-6, and IL-7, but not tumor necrosis factor-alpha (TNF-alpha) or IL-1 beta, induced a weak LAK activity against PCI-24, whereas IL-2-induced (1000 U/mL) LAK exhibited a far more potent cytotoxicity . When these cytokines were added at the suboptimal dose IL-2 (100 U/mL), no significant augmentation in LAK activity was induced . Staphylococcal protein A (SpA) induced LAK activity as potent as that seen with IL-2 (1000 U/mL) . Both IL-2-induced and SpA-induced LAK had a potent, dose-dependent cytotoxicity against HUVEC . HUVEC inhibited both IL-2- and SpA-induced LAK cytotoxicity against PCI-24 to almost the same extent as seen with PCI-24 . Thus, two potent LAK-inducers did not generate LAK activity that differentiates neoplastic from non-neoplastic cells . Thus, in vitro cytotoxicity of LAK against non-neoplastic endothelial cells is unavoidable when handling cytokines in LAK induction.

J Neurosurg Sci, 1994 Sep, 38(3), 161 - 5
Cerebrospinal fluid shunt infections; Ersahin Y et al.; Cerebrospinal fluid (CSF) shunt infection is one of the most frequent and disabling complications . We reviewed the records of 306 patients who underwent CSF shunt surgery from 1983 through 1992 . Six hundred and twelve procedures were performed in these 306 patients . Infection occurred following 46 of the procedures for an infection rate of 7.5% per procedure . The 46 infections involved 39 patients . There were 8 recurrent infections . The infection rate per child was 12.7% . Staphylococcal species were isolated in 50% of all infections . Patients younger than 1 year old and children with multiple revisions have a greater risk of infection than those of older . Myelomeningocele and meningitis had higher infection rate among other etiologies . Patients with multiple revisions had higher infection rate than those with single revision or none . The incidence of infection was higher in cyst-peritoneal shunts than both ventriculo-atrial and ventriculo-peritoneal shunts . Mortality was high in Gram negative infections.

Biotechnol Prog, 1994 Sep-Oct, 10(5), 513 - 9
A mathematical model to predict the partitioning of peptides and peptide-modified proteins in aqueous two-phase systems; Eiteman MA et al.; A mathematical procedure was developed to predict the partition coefficients of the peptides AIIP, AWWP, AIIPAIIP and AWWPAWWP in poly(ethylene glycol) (PEG)/phosphate aqueous two-phase systems from amino acid hydrophobicities . In general, peptides containing tryptophan partition more into the PEG-enriched upper phase than analogous peptides containing isoleucine . Specifically, as the PEG concentration difference between the phases increased in a PEG/potassium phosphate aqueous two-phase system, the peptide AIIP was observed to have a partition coefficient ranging from 1.2 to 1.6, AIIPAIIP from 2.4 to 5.7, AWWP from 13.5 to 32.2, and AWWPAWWP from 43 to 170 . The model was extended to predict the partitioning of a staphylococcal protein A derivative (ZZ) modified with these four peptides . As predicted, the protein modified with isoleucine-containing peptides had lower partition coefficients than the protein modified with tryptophan-containing peptides . The partition coefficient of the ZZ protein ranged from 0.35 to 0.20, that of ZZAIIPAIIP from 0.58 to 0.48, and that of ZZAWWPAWWP from 3.5 to 5.3 in these systems . The results show that short peptide handles can significantly enhance the partitioning of proteins in aqueous two-phase systems . The relationship between the model and the surface exposure of peptide handles and the utility of the model to aid in the design of such handles to enhance purifications are also discussed.

Biotechnology (N Y), 1994 Sep, 12(9), 890 - 7
Mammalian cell expression of single-chain Fv (sFv) antibody proteins and their C-terminal fusions with interleukin-2 and other effector domains; Dorai H et al.; The production of several single-chain Fv (sFv) antibody proteins was examined by three modes of mammalian cell expression . Our primary model was the 741F8 anti-c-erbB-2 sFv, assembled as either the VH-VL or VL-VH, and expressed alone, with C-terminal cysteine for dimerization, or as fusion proteins with carboxyl-terminal effector domains, including interleukin-2, the B domain of staphylococcal protein A, the S-peptide of ribonuclease S, or hexa-histidine metal chelate peptide . Constructs were expressed and secreted transiently in 293 cells and stably in CHO or Sp2/0 cell lines, the latter yielding up to 10 mg per liter . Single-chain constructs of MOPC 315 myeloma and 26-10 monoclonal antibodies were also expressed, as were hybrids comprising unrelated VH and VL regions . Our results suggest that mammalian expression is a practical and valuable complement to the bacterial expression of single-chain antibodies.

Zhongguo Yao Li Xue Bao, 1994 Sep, 15(5), 473 - 6
{Suppressive effects of adenosine on nonspecific and humoral immunities in mice}; Feng YX et al.; Adenosine (Ade) 1.3, 13, 130 mg.kg-1 ip inhibited the ability of peripheral leukocytes and peritoneal macrophages in phagocytosing the Staphylococcus albus with {3H}TdR incorporation in mice, declined the hemolytic ability of plaque-forming cells and the production of antibody in mice immunized by sheep erythrocytes . Ade 13, 130 mg.kg-1 ip decreased the mouse serum muramidase (lysozyme) concentration . Dipyridamole (Dip) 10 mg.kg-1 ip attenuated the effects of Ade 130 mg.kg-1 on humoral immunity reaction, but the nonspecific immunity was not attenuated . These results showed that the uptake of Ade may play an important role in the effects of Ade on humoral immunity reaction . Aminophylline (Ami) 100 mg.kg-1 ip attenuated the effects of Ade 130 mg.kg-1 on hemolytic ability of plaque-forming cells and the ability of peripheral leukocytes in phagocytosing Staphylococcus albus . These results suggested that the effects of Ade on murine humoral and nonspecific immunity reaction were mediated by Ade A2 receptor (A2DR).

Arkh Patol, 1994 Sep-Oct, 56(5), 10 - 5
{The mechanisms of the pathogenicity of bacteria in different infections}; Vtiurin BV et al.; The following ultrastructural formations are found in the bacteria of various infections: fibrillar and drop-like microcapsules, an increase of nucleotide size and number, micropyles . The dynamics of staphylococcus L-form formation in sepsis as well as the phenomenon of incomplete phagocytosis and endocytobiosis were studied . The latter is observed in mixed infection: dysentery bacteria lamblia, gonococci and trichomonas . These alterations indicate increased bacterial pathogenicity and seem to reflect the evolution of the bacteria adaptive mechanisms under the conditions of antibiotic therapy.

J Exp Med, 1994 Sep 1, 180(3), 1153 - 8
The protective role of endogenously synthesized nitric oxide in staphylococcal enterotoxin B-induced shock in mice; Florquin S et al.; Nitric oxide (NO) synthesis during experimental endotoxemia has been shown to have both deleterious and beneficial effects . In the present study, we analyzed the in vivo production and the regulatory role of NO in the shock syndrome induced by staphylococcal enterotoxin B (SEB) in mice . First, we found that intraperitoneal administration of 100 micrograms SEB in BALB/c mice induced a massive synthesis of NO as indicated by high serum levels of nitrite (NO2-) and nitrate (NO3-) peaking 16 h after SEB injection . The inhibition of NO2- and NO3- release in mice injected with anti-tumor necrosis factor (TNF) and/or anti-interferon gamma (IFN-gamma) monoclonal antibody (mAb) before SEB challenge revealed that both cytokines were involved in SEB-induced NO overproduction . In vitro experiments indicated that NO synthase (NOS) inhibition by N-nitro-L-arginine methyl ester (L-NAME) enhanced IFN-gamma and TNF production by splenocytes in response to SEB . A similar effect was observed in vivo as treatment of mice with L-NAME resulted in increased IFN-gamma and TNF serum levels 24 h after SEB challenge, together with persistent expression of corresponding cytokine mRNA in spleen . The prolonged production of inflammatory cytokines in mice receiving L-NAME and SEB was associated with a 95% mortality rate within 96 h, whereas all mice survived injections of SEB or L-NAME alone . Both TNF and INF-gamma were responsible for the lethality induced by SEB in L-NAME-treated mice as shown by the protection provided by simultaneous administration of anti-IFN-gamma and anti-TNF mAbs . We conclude the SEB induces NO synthesis in vivo and that endogenous NO has protective effects in this model of T cell-dependent shock by downregulating IFN-gamma and TNF production.

J Immunol, 1994 Sep 1, 153(5), 2038 - 45
CD1+ human thymocytes proliferate in response to superantigen staphylococcal enterotoxin B1; Todd SC et al.; Exposure of human thymocytes to superantigens results in the deletion of thymocytes expressing specific TCR-V beta genes . The factors that contribute to this deletion may relate to the inherent nature of the T cell at a given stage of development . In this paper, we demonstrate that CD1+ human cortical thymocytes are capable of proliferating in response to a bacterial superantigen (staphylococcal enterotoxin B (SEB)) in the presence of autologous CD2-/low thymic APCs . Phenotypic analysis of the responding populations revealed that the majority of the CD1+ cells were CD4+CD8low or CD8+CD4low cells . The response is triggered by low concentrations of SEB, requires the participation of the TCR and IL-2R molecules, and is inhibited by cyclosporin A . Thymocytes that express specific V beta genes are expanded, which results in an engagement profile that parallels that found in PBLs . Additionally, four V beta-chains that have not been reported previously are shown to engage SEB . Once stimulated, the thymocytes failed to respond to additional SEB; however, they could be induced to proliferative with IL-2, which suggests that these expanded populations had become anergic . These data represent the first demonstration of a human cortical thymocyte subpopulation that responds to superantigen by proliferation and subsequent anergy.

Proc Natl Acad Sci U S A, 1994 Aug 30, 91(18), 8462 - 6
Binding of a soluble alpha beta T-cell receptor to superantigen/major histocompatibility complex ligands; Kappler J et al.; The genes for the alpha and beta chains of a murine T-cell receptor were truncated just prior to the portions encoding the transmembrane regions and introduced into baculovirus by recombination . Insect cells infected with the virus secreted a soluble form of the receptor that could be purified to homogeneity . This soluble receptor reacted with a set of six monoclonal antibodies originally raised to different epitopes on the natural transmembrane-region-containing receptor and bound with appropriate specificity to a cell surface complex of the human major histocompatibility complex class II molecule DR1 with the bacterial superantigen staphylococcal enterotoxin B.

Biochemistry, 1994 Aug 30, 33(34), 10457 - 62
Conformational flexibility in a staphylococcal nuclease mutant K45C from time-resolved resonance energy transfer measurements; Wu P et al.; Thermal fluctuations exist in native proteins and other macromolecules in solution . Some may play a role in ligand or receptor binding, control rates of enzymatic catalysis, or define a range of conformations a segment can adopt in solution . We apply the method of time-resolved resonance energy transfer to study the conformational flexibility of a staphylococcal nuclease mutant, K45C, where lysine 45 located at a flexible loop is replaced by a cysteine . We labeled the thiol group with DTNB (5,5'-dithiobis(2-nitrobenzoic acid)) and used the TNB group covalently attached to the protein as an energy acceptor from a single tryptophan at residue 140 as the donor . Conformational flexibility occurring on the time scale of nanoseconds or longer is dispersed as an apparent distance distribution in time-resolved resonance energy transfer measurements . Below room temperature the apparent distance distribution was fitted with a symmetric Lorentzian model with a full width at half maximum height of about 6 A, indicating substantial degrees of heterogeneity between residues 45 and 140 . At room or higher temperature where the protein is in its native state, the apparent distance distribution is asymmetric, indicating the presence of static disorders . Segments in the protein that contribute to the static disorder can be converted to mobile ones with the addition of denaturing guanidinium chloride.

FEMS Microbiol Lett, 1994 Aug 15, 121(2), 189 - 97
Evidence for importance of the Staphylococcus hyicus lipase pro-peptide in lipase secretion, stability and activity; Demleitner G et al.; To investigate the function of the pro-peptide (PP) region of the Staphylococcus hyicus exolipase, restriction sites were created in the lipase gene to facilitate the construction of deletions in this region . Lipase gene expression was carried out in Staphylococcus carnosus . In the presence of the entire PP region, the 86-kDa pro-lipase was efficiently exported, had high lipolytic activity, and hardly any degradation products were seen in Western blot analysis . In addition to the 86-kDa pro-lipase, the membrane fraction contained a 106-kDa immunoreactive form . If the PP was completely or partially deleted, signal peptide processing, lipase secretion, lipase activity and/or lipase stability were impaired . The results obtained with lipase PP deletion mutants indicate that the PP region may have two functional domains . The N-terminal region of the lipase PP appears to be more important for lipase activity and the C-terminal portion for lipase secretion and proteolytic stability . In the presence of only the C-terminal part of the PP lipase, secretion was hardly affected . However, the activity of the extracellular lipase was markedly reduced . If only a small portion of the C-terminal part of the PP was present, lipase secretion was again markedly reduced and no lipase activity was detectable . In the presence of the N-terminal half of the PP region, lipase secretion was affected to a lesser extent . However, the resulting 60-kDa form, which showed comparably good specific lipase activity, suffered severe proteolytic degradation.

Appl Environ Microbiol, 1994 Aug, 60(8), 2876 - 83
Producer immunity towards the lantibiotic Pep5: identification of the immunity gene pepI and localization and functional analysis of its gene product; Reis M et al.; The lantibiotic Pep5 is produced by Staphylococcus epidermidis 5 . Pep5 production and producer immunity are associated with the 20-kb plasmid pED503 . A 1.3-kb KpnI fragment of pED503, containing the Pep5 structural gene pepA, was subcloned into the Escherichia coli-Staphylococcus shuttle vector pCU1, and the recombinant plasmid pMR2 was transferred to the Pep5- and immunity-negative mutant S . epidermidis 5 Pep5- (devoid of pED503) . This clone did not produce active Pep5 but showed the same degree of insensitivity towards Pep5 as did the wild-type strain . Sequencing of the 1.3-kb KpnI-fragment and analysis of mutants demonstrated the involvement of two genes in Pep5 immunity, the structural gene pepA itself and pepI, a short open reading frame upstream of pepA . To identify the 69-amino-acid pepI gene product, we constructed an E . coli maltose-binding protein-PepI fusion clone . The immunity peptide PepI was detected in the soluble and membrane fractions of the wild-type strain and the immune mutants (harboring the plasmids pMR2 and pMR11) by immunoblotting with anti-maltose-binding protein-PepI antiserum . Strains harboring either pepI without pepA or pepI with incomplete pepA were not immune and did not produce PepI . Washing the membrane with salts and EDTA reduced the amount of PepI in this fraction, and treatment with Triton X-100 almost completely removed the peptide . Furthermore, PepI was hydrolyzed by proteases added to osmotically stabilized protoplasts . This suggests that PepI is loosely attached to the outside of the cytoplasmic membrane . Proline uptake and efflux experiments with immune and nonimmune strains also indicated that PepI may act at the membrane site.

Ann Thorac Surg, 1994 Aug, 58(2), 429 - 32; discussion 432-3
Infective endocarditis in patients who had replacement of the aortic root; Ralph-Edwards A et al.; In 12 patients who had had composite replacement of the aortic valve and ascending aorta, infective endocarditis developed 2 months to 17 years after operation . Six patients had mechanical valves and 6 had biological ones (four homograft and two porcine valves) . All patients needed operation because of shock, heart failure, persistent sepsis in spite of adequate antibiotic therapy, or the development of a paravalvular false aneurysm . The predominant microorganism was Staphylococcus . All 6 patients who had mechanical valves were found to have an abscess in the junction between the aortic annulus and the prosthesis; in patients who had biological valves the infection was limited to the leaflets in 3 (one homograft and two porcine valves) and leaflets and annulus abscess in 3 (three homograft valves) . Operation consisted of radical resection of tissues suspected of being infected and reconstruction of the left ventricular outflow tract and of the surrounding structures with glutaraldehyde-fixed bovine pericardium . The aortic valve and ascending aorta were replaced with a new valved conduit . An aortic homograft was used in only 1 patient . There was only one operative death due to right ventricular infarction but most patients experienced serious postoperative complications . Operative survivors were followed up from 3 to 156 months (mean, 42 months) . One patient died 35 months postoperatively due to bleeding complications of anticoagulation; 1 patient suffered a cardiac arrest at home 2 months after operation, sustained permanent cerebral damage, and died 4 months later . The remaining patients are asymptomatic from the cardiovascular viewpoint.(ABSTRACT TRUNCATED AT 250 WORDS)

Ann Thorac Surg, 1994 Aug, 58(2), 425 - 8
Surgical treatment of infected composite graft after replacement of ascending aorta; Soyer R et al.; Infection of a composite graft is a serious complication . However, reports of such cases are rare even in large series . We report our experience with 4 patients in whom infection of a composite graft developed with pseudoaneurysm formation . Two of the patients had Marfan's syndrome and were treated by Bentall procedure and 2 were treated by Cabrol technique for non-Marfan cystic medial necrosis . Staphylococcus epidermidis was detected in 2 patients and Enterococcus in 1 . Reoperation was carried out between 1 and 32 months after the first intervention . One patient died of cerebral embolism and 3 remained free of infection 11 to 82 months later . These cases and guidelines for managing abdominal and peripheral vascular prosthetic infection indicate the need for prompt reintervention when infection is suspected from chronic sepsis, septicemia, positive blood cultures, fistula, anastomotic leak, hemolysis, embolism, graft deformity, or false aneurysm . When the organism is isolated, appropriate antibiotic therapy should be administered . All prosthetic material should be removed and all adjacent infected or necrotic tissue excised . Local antiseptic irrigation may be helpful . Dead space around the prosthesis should be filled with well-vascularized transposed pedicled flaps . Antibiotic therapy should be intravenously administered for at least 6 weeks.

Epidemiol Infect, 1994 Aug, 113(1), 75 - 81
Application of pulsed-field gel electrophoresis to the epidemiological characterization of Staphylococcus intermedius implicated in a food-related outbreak; Khambaty FM et al.; An outbreak of food intoxication involving over 265 cases in western United States occurred in October 1991 . Staphylococcus intermedius was implicated as the aetiologic agent . Representative outbreak isolates (five clinical and ten from foods) produced type A enterotoxin . DNA fragments generated by four restriction endonucleases and analysed by pulsed-field gel electrophoresis (PFGE) provided definitive evidence that all isolates from nine different counties in California and Nevada were derived from a single strain . The PFGE pattern of these outbreak isolates was distinct from those of a heterogeneous collection of seven S . intermedius strains of veterinary origin and five unrelated S . aureus laboratory strains . The data show a significant PFGE pattern heterogeneity not only among members of different Staphylococcus species but also within members of the same species and even the same enterotoxin type . The results indicate that PFGE is a valuable strain-specific discriminator for the epidemiological characterization of S . intermedius . To our knowledge, this represents the first documented foodborne outbreak caused by S . intermedius . These findings suggest that the presence of S . intermedius and other species such as S . hyicus in food should be reason for concern.

Eur J Immunol, 1994 Aug, 24(8), 1893 - 902
Exogenous superantigens acutely trigger distinct levels of peripheral T cell tolerance/immunosuppression: dose-response relationship; Miethke T et al.; Ligand-specific immunosuppression requires an understanding of the parameters that control peripheral T cell tolerance . T cell receptor (TcR) transgenic mice offer a clear advantage for studying post-thymic tolerance mechanisms in vivo that are operational in a monoclonal T cell population with preselected antigen specificity . Yet it is unclear whether the rules defined in monoclonal T cells of genetically manipulated mice reflect those operative in clonally diverse peripheral T cells of normal mice . To analyze acute tolerance mechanisms in unselected peripheral T cells, we challenged normal mice with the superantigen staphylococcal enterotoxin B (SEB) and analyzed ligand-reactive V beta 8+ T cells for TcR-triggered tolerance mechanisms such as anergy, TcR down-regulation, or apoptosis . Upon challenge with graded doses of SEB (0.001-10 micrograms) V beta 8+ T cells become anergic within 6-16 h . Importantly, a dosage effect of SEB in regard to the level of anergy induced was observed . Anergy induced by low concentrations of SEB (0.001-0.1 microgram) is transient and is overcome by clonal growth, while higher concentrations of SEB (0.1-10 micrograms) cause long-lasting anergy resistant to cell cycle progression . At high SEB concentrations (1-10 mg) about 50% of the anergic V beta 8+ T cells additionally down-regulate their TcR-CD3 complex, followed by a loss of CD2, CD4, CD8 accessory molecules . In parallel, T cell phenotype-negative but genotypically V beta 8+ T cells are generated . The T cell phenotype-negative cells reacquire their V beta 8+ T cell phenotype upon culture in vitro . In vivo, a subset of V beta 8+ cells, defined by an intermediate stage of TcR down-regulation, i.e . V beta 8lowCD3+ cells, but not T cell phenotype-negative cells are selectively programmed for apoptosis, which occurs within 1 h . These data suggest that SEB triggers distinct tolerance pathways which operate in a hierarchical fashion in clonally diverse ligand-reactive T cells . Specifically, the results illustrate the power of exogenous superantigens to exploit these distinct tolerance pathways, thereby achieving distinct levels of immunosuppression.

Eur J Immunol, 1994 Aug, 24(8), 1757 - 64
New infectious mammary tumor virus superantigen with V beta-specificity identical to staphylococcal enterotoxin B (SEB); Luther S et al.; Only few infectious mouse mammary tumor viruses (MMTV) have been characterized which induce a potent superantigen response in vivo . Here we describe the characterization of an MMTV which was isolated from milk of the highly mammary tumor-prone SHN mouse strain . Exposure of newborn mice to milk-borne MMTV (SHN) results in a very slow deletion of V beta 7, 8.1, 8.2 and 8.3 expressing peripheral T cells . Subcutaneous injection of adult mice with this virus induces a rapid and strong stimulation of all four affected V beta-subsets in vivo . Besides the strong T cell effect we observed an early proliferation and activation of the local B cell pool leading to the initial secretion of IgM followed by preferential secretion of IgG2a by day 6 . Sequence comparison of the polymorphic C terminus with known open reading frames revealed high homology to the endogenous provirus Mtv-RCS . This is the first report of a virus having a complete overlap in V beta-specificity with a bacterial superantigen stimulating as many as 35% of the whole CD4+ T cell repertoire including V beta 8.2.

Arch Pediatr Adolesc Med, 1994 Aug, 148(8), 853 - 5
Escherichia coli septicemia in nonperforated appendicitis; Ruff ME et al.; OBJECTIVE: To determine the association between nonperforated appendicitis and Escherichia coli septicemia, and the frequency with which blood cultures are obtained in the clinical setting of appendicitis . DESIGN: Three case reports of E coli septicemia and nonperforated appendicitis and a retrospective survey . SETTING: Children's Medical Center, Dallas, Tex, a primary care and tertiary referral center . PATIENTS: All children admitted in a 2-year period with a diagnosis of appendicitis . INTERVENTIONS: None . RESULTS: Preoperative blood cultures were obtained in 20 (21%) of 96 patients with histologic evidence of appendicitis . Fifty percent of the patients had gross or microscopic evidence of appendiceal perforation . Twelve (25%) of the 48 patients with perforated appendicitis had blood cultures obtained before the initiation of antimicrobial therapy, and in two of these patients (17%) the results were positive . Blood cultures were drawn before antibiotic therapy in four (8%) of the 48 patients with nonperforated appendicitis, and in two of these the results were positive . The blood culture isolates (coagulase-negative Staphylococcus and E coli) were the same in both groups . CONCLUSIONS: Nonperforated appendicitis and septicemia may be more common than formerly appreciated . Only a prospective study can determine the true incidence of septicemia in children with perforated or nonperforated appendicitis.

Plast Reconstr Surg, 1994 Aug, 94(2), 300 - 5
Periprosthetic bacteria and the breast implant patient with systemic symptoms; Dowden RV; This report presents seven women with breast implants who experienced systemic symptoms which resolved rapidly after implant removal . A hypothesis is that these symptoms (which have been labeled "silicone poisoning" or "silicone adjuvant disease") may actually be caused by periprosthetic bacteria which have generally been considered innocuous, e.g., Staphylococcus epidermidis . In these cases, systemic symptoms such as malaise, fatigue, diarrhea, muscle aches, and arthralgia rapidly resolved after an antibacterial regimen plus implant removal without capsulectomy . Of cultures taken by swab in four patients, all were positive; of those taken by irrigation in three patients, one was positive . I believe that these patients' symptoms were real and offer the hypothesis that treatment of periprosthetic bacteria might explain rapid clinical improvement following explantation.

Infect Immun, 1994 Aug, 62(8), 3408 - 15
Identification of staphylococcal enterotoxin B sequences important for induction of lymphocyte proliferation by using synthetic peptide fragments of the toxin; Jett M et al.; A series of 13 synthetic peptides, approximately 30 amino acids each, which spanned the entire sequence of staphylococcal enterotoxin B (SEB) were tested to evaluate their effects on T-cell proliferation in a culture system containing elutriated human peripheral blood lymphocytes incubated with a specific ratio of mononuclear cells . Four peptide regions were found to inhibit SEB-induced proliferation; they included sequences 1 to 30 (previously thought to be involved in major histocompatibility complex class II binding), 61 to 92 (sequences which relate to the T-cell receptor site), 93 to 112 (a linear sequence corresponding to the cysteine loop), and 130 to 160 (containing a highly conserved sequence, KKKVTAQEL) . Antisera raised to this last peptide were capable of neutralizing SEB-induced proliferation . Antisera raised against the peptides which overlapped this sequence also were somewhat inhibitory . Neutralizing antisera were not produced from any other peptide sequence tested . To determine if any of these effects were nonspecific with regard to SEB-induced proliferation, the peptides were tested for inhibition of phorbol dibutyryl ester-induced proliferation, and only the sequence 93 to 112 (corresponding to the cysteinyl loop region) was consistently inhibitory (40%) . Of the regions which displayed inhibition of SEB-induced proliferation, the peptide 130 to 160 inhibited binding of 125I-SEB to lymphocytes . These data suggest that the residues containing and surrounding the sequence KKKVTAQEL may be critical in the SEB-induced proliferation and may be useful for developing neutralizing antisera to SEB.

Infect Immun, 1994 Aug, 62(8), 3396 - 407
Predictions of T-cell receptor- and major histocompatibility complex-binding sites on staphylococcal enterotoxin C1; Hoffmann ML et al.; We have focused on regions of staphylococcal enterotoxin C1 (SEC1) causing immunomodulation . N-terminal deletion mutants lacking residues 6 through 13 induced T-cell proliferation similar to that induced by native toxin . However, mutants with residues deleted between positions 19 and 33, although nonmitogenic themselves, were able to inhibit both SEC1-induced T-cell proliferation and binding of the native toxin to major histocompatibility complex (MHC) class II . Presumably, these deletions define a part of SEC1 that interacts with the T-cell receptor . Three synthetic peptides containing residues located in a region analogous to the alpha 5 groove of SEC3 had residual mitogenic activity or blocked T-cell proliferation induced by SEC1 and appear to recognize the same site as SEC1 on a receptor for the toxin, presumably MHC class II . We conclude that isolated portions of the SEC1 molecule can retain residual mitogenic activity but that the entire protein is needed to achieve maximal superantigenic stimulation . Our results, together with the results of other investigators, support a model in which SEC1 binds to an alpha helix of MHC class II through a central groove in the toxin and thereby promotes or stabilizes the interaction between antigen-presenting cells and T cells.

Cell Immunol, 1994 Aug, 157(1), 29 - 37
Macrophages are dispensable for superantigen-mediated stimulation and anergy induction of peripheral T cells in vivo; Koesling M et al.; Bacterial superantigens provoke T lymphocyte activation by cross-linking the variable part of the T cell receptor (TCR) beta-chain with MHC class II molecules on antigen-presenting cells . Although the molecular mechanisms of this interaction are well characterized, the in vivo accessory cell requirements for this stimulation of T lymphocytes by bacterial superantigens remain unknown . In the present study we have addressed the role of splenic macrophages in the activation of V beta 8+ peripheral T cells by staphylococcal enterotoxin B (SEB) in BALB/c mice . SEB-triggered clonal expansion and subsequent induction of unresponsiveness of both CD4+ and CD8+ T cells were investigated in naive animals, or in mice injected intravenously with dichloromethylene diphosphonate-containing liposomes . Such a treatment resulted in the complete and long-lasting elimination of the splenic macrophage population . Remarkably, however, this complete depletion of peripheral macrophages had only a rather minor effect on the superantigen-induced T cell response in the spleen, and macrophage-depleted animals exhibited overall the same magnitude and kinetics of SEB-mediated T cell activation and anergy-induction as their nondepleted counterparts . Our data thus exclude an essential role of peripheral macrophages or macrophage-secreted cytokines in the systemic T cell activation caused by bacterial superantigens in vivo.

Mol Microbiol, 1994 Aug, 13(4), 733 - 43
The nlpD gene is located in an operon with rpoS on the Escherichia coli chromosome and encodes a novel lipoprotein with a potential function in cell wall formation; Lange R et al.; rpoS is the structural gene for sigma s, which is a second vegetative sigma subunit of RNA polymerase in Escherichia coli and is involved in the expression of many stationary phase-induced genes . Upstream of rpoS is an open reading frame (ORF) whose function and regulation have not been studied . Strong overproduction of its gene product using the IPTG-inducible tac promoter leads to the formation of bulges at the cell septum and the cell poles, and in rapidly growing cells brings about cell lysis, indicating that the gene product has a hydrolytic function in cell wall formation or maintenance . This is corroborated by sequence homology to lysostaphin, a cell wall lytic exoenzyme synthesized by two Staphylococcus strains . Using globomycin, a specific inhibitor of signal peptidase II, we demonstrate that the product of the ORF is a novel lipoprotein (NlpD) . Two transcriptional start sites for nlpD have been localized . In contrast to rpoS, nlpD is not induced during entry into stationary phase . Growth-phase-regulated transcription of rpoS is initiated at additional sites within the nlpD ORF, but the nlpD promoters contribute substantially to the basal level of rpoS expression in exponentially growing cells, indicating that nlpD and rpoS form an operon.

Int J Pept Protein Res, 1994 Aug, 44(2), 130 - 8
Bony fish neurophysins . Identification of MSEL- and VLDV-neurophysins of the pollack (Pollachius virens); Chauvet J et al.; The two types of neurophysins known in vertebrate species, namely MSEL-neurophysin (vasopressin-like hormone-associated neurophysin) and VLDV-neurophysin (oxytocin-like hormone-associated neurophysin) have been purified from the pollack (Pollachius virens) pituitary through a combination of molecular sieving and high-pressure liquid chromatography (HPLC) . Homogeneity has been checked by gel electrophoresis and return in HPLC . The apparent molecular masses measured by SDS-electrophoresis are near 12 kDa, significantly higher than those found for their mammalian homologues (10 kDa) . The two types of neurophysins have been recognized through their N-terminal amino acid sequences . The primary structure of MSEL-neurophysin has been partially determined using automated Edman degradation applied on native and reduced-alkylated protein, as well as peptides derived by trypsin or staphylococcal proteinase hydrolyses . Comparison of pollack MSEL-neurophysin with ox, goose and frog counterparts reveals that particular positions in the polypeptide chain are subjected to substitutions and that the numbers of substitutions do not seem closely related to the paleontological times of divergence between the different vertebrate classes.

Vet Immunol Immunopathol, 1994 Aug, 42(2), 137 - 47
Antistaphylococcal antibodies in dogs with recurrent staphylococcal pyoderma; Morales CA et al.; Staphylococcus intermedius skin infection (pyoderma) may be perpetuated in some dogs by a hypersensitivity reaction to staphylococcal organisms . Dogs with idiopathic superficial or deep recurrent staphylococcal skin infections may thus have quantitative differences in serum antistaphylococcal IgE antibodies compared with healthy dogs . To test this hypothesis, antistaphylococcal IgG and IgE antibodies were measured by ELISA in groups of dogs with idiopathic recurrent pyoderma, recurrent pyoderma secondary to atopic disease, non-recurrent pyoderma, and in healthy dogs . All groups of dogs with prior staphylococcal skin infection had significantly higher mean serum antistaphylococcal IgG levels than healthy dogs (P < 0.05) . Dogs with recurrent deep pyoderma had the highest mean levels of antistaphylococcal IgG . Dogs with idiopathic recurrent superficial pyoderma and those with recurrent pyoderma secondary to atopy had significantly (P < 0.05) higher mean levels of serum antistaphylococcal IgE than other groups tested . It is concluded from these findings that S . intermedius can behave as an allergen in some dogs and elicit an IgE response . These results support the concept that bacterial hypersensitivity may be responsible for initiating or perpetuating skin lesions in these animals.

Vet Microbiol, 1994 Aug 1, 41(3), 259 - 66
Characterization of Staphylococcus intermedius from healthy dogs and cases of superficial pyoderma by DNA restriction endonuclease patterns; Hesselbarth J et al.; In order to study the possible clonal relation of Staphyloccocus (S.) intermedius from canine superficial pyoderma and from healthy carriers, isolates from pustular swabs and from vaginal, nasal and normal skin sabs were typed using macrorestriction analysis with Sma I and pulsed-field gel electrophoresis . From the size of the resulting fragments the size of the chromosome of S . intermedius could be determined to be roughly 1500 +/- 200 kb on the average . The fingerprints were very heterogeneous though characteristically distinct from patterns of (human) S . aureus as published by others . Strains from superficial pyoderma were not found to be more similar to each other than strains from healthy carriers . Therefore it was concluded that strains from this type of skin infection probably did not belong to a certain subpopulation of S . intermedius, which might have indicated a higher virulence of these strains.

Indian J Exp Biol, 1994 Aug, 32(8), 567 - 70
Effect of Staphylococcus protein-A on adrenal gland in mice; Sahu AP; Staphylococcus protein-A (SpA) was administered (ip) to Balb/c male mice for two weeks, twice a week at the dose level of 1, 6 and 12 micrograms in 200 microliters of normal saline . A significant change in the relative weights of liver, spleen, thymus, tracheobronchial lymph node, lung and testis was observed in 1 microgram SpA treatment group . In the adrenal, marked changes at the dose level of 1 microgram SpA treatment after 2 weeks were observed in the form of cellular proliferation on zona glomerulosa (ZG) and zona fasciculata (ZF) of adrenal cortex . With 6 and 12 micrograms SpA, adrenocortical masses were observed outside the capsule of adrenal gland . The enhanced effects of SpA at 4 weeks after treatment with the dose level of 6 and 12 micrograms were in the form of adrenal cell masses outside adrenal gland and histological changes in the adrenal cortex . The results suggest that long term and high doses therapy with SpA may be a risk factor to sensitive endocrine glands.

Curr Opin Pediatr, 1994 Aug, 6(4), 442 - 6
Vesiculopustular diseases of neonates and infants; Sahn EE; Vesiculopustular diseases of neonates and infants can be divided into infectious and noninfectious categories . Recent clinical and scientific literature has focused mainly on the infectious diseases and on epidermolysis bullosa . This review covers the following disorders: neonatal sepsis, staphylococcal scaled skin syndrome, neonatal herpes simplex, neonatal varicella, congenital syphilis, scabies, mastocytosis, incontinentia pigmenti, eosinophilic pustular folliculitis, and epidermolysis bullosa.

Protein Expr Purif, 1994 Aug, 5(4), 385 - 90
Antigen detection using recombinant, bifunctional single-chain Fv fusion proteins synthesised in Escherichia coli; Gandecha A et al.; A gene fusion approach has been used to produce antibody conjugates for use in immunoassays . Escherichia coli expression vectors encoding fusions between the outer membrane protein A signal peptide, an anti-phytochrome single-chain Fv protein, and either Escherichia coli alkaline phosphatase or Staphylococcal protein A downstream of the T7 O10 promoter were constructed . A crude lysate from cells expressing the single-chain Fv-alkaline phosphatase fusion protein could be used directly for the sensitive and specific staining of phytochrome on protein blots by a single-step immunoassay procedure . Following purification by immunoglobulin G affinity chromatography, the Staphylococcal protein A-single-chain Fv fusion protein was also used for selective immunostaining of phytochrome on protein blots by a two-step procedure in which a rabbit immunoglobulin-alkaline phosphatase conjugate was used to detect antigen-bound Staphylococcal protein A . Recombinant antibody conjugates of the types described here are simple and inexpensive to produce and are a realistic alternative to conventional antibody conjugates.

Int J Hematol, 1994 Aug, 60(2), 129 - 36
Infection in myelodysplastic syndromes before evolution into acute non-lymphoblastic leukemia; Oguma S et al.; The frequencies, kinds, pathogens, and risk factors of infections in the myelodysplastic syndromes (MDS) were analysed in 430 cases . The overall tendency was for one infectious episode per 1023.5 patient days . The frequency of infectious episodes was highest just after diagnosis of MDS when more than 4 episodes per 1000 patient days occurred . Thereafter, the rate declined rapidly to about 0.3 episodes per 1000 patient days within 4 years . The most frequent infection was that of the respiratory tract followed by sepsis and fever of unknown origin (FUO) . Among the types of infection resulting in death, sepsis and FUO comprised the highest proportion (40%) followed by respiratory tract infections (39%) . The most frequent pathogen observed was Staphylococcus bacteria . The significant multivariate risk factors for fatal infections were subtype, hemoglobin, dependence on red blood cell transfusion, age, and sex . A staging system was created using these five simple variables at diagnosis.

Int J Hematol, 1994 Aug, 60(2), 119 - 27
A randomised dosage study of ceftazidime with single daily tobramycin for the empirical management of febrile neutropenia in patients with hematological diseases; Gibson J et al.; A single-institution, randomised trial was conducted to compare the clinical and microbiological efficacy of two different doses of ceftazidime in combination with a single daily dose of tobramycin for the empirical management of febrile neutropenic patients with hematologic disorders (absolute neutrophil count < 1 x 10(9)/l) . Upon the development of fever or signs of sepsis, patients received either 2 g ceftazidime every 8 h plus a single daily dose tobramycin (5 mg/kg/day) (C2T, n = 73) or 1 g ceftazidime every 8 h plus a single daily dose of tobramycin (C1T, n = 77) . In addition, flucloxacillin (1-2 g every 4 h) could be added if there was clinical suspicion of staphylococcal infection . Analysis was performed for the whole group and for the subset which did not receive flucloxacillin . When evaluated at 96 h, 70% (95% CI, +/- 11%) of patients randomised to C2T and 60% (95% CI, +/- 11%) randomised to C1T had responded (chi 2 = 1.27, P = 0.26) . The response rates at 96 h for those who did not receive flucloxacillin were 77% (+/- 12%) and 74% (+/- 13%), respectively (chi 2 = 0.01, P = 0.92) . Overall, 68 (93% +/- 6%) and 72 (94% +/- 6%) patients, respectively, eventually became afebrile (chi 2 = 0.06, P = 0.81) . Renal function, as judged by serum creatinine, was unaffected by either antibiotic schedule . Within 10 days of antibiotic commencement there was one death in each arm and overall there were five deaths in each arm.(ABSTRACT TRUNCATED AT 250 WORDS)

Am J Reprod Immunol, 1994 Aug, 32(1), 15 - 25
Spontaneous abortion in immunodeficient SCID mice; Clark DA et al.; PROBLEM: Immunodeficient SCID mice on the CB-17 have been used to test the role of "rejection" in a xenogeneic blastocyst transfer model of recurrent miscarriage, but interpretation of the data requires knowing syngeneic within-species matings have a high success rate and do not require immunotrophic factors expected only in immunocompetent non-T-cell deficient mice . METHOD: Resorption rates were studied in a SCID CB-17 barrier facility that provided the mice used to test the role of immunology in the resorption model . RESULTS: Spontaneous resorption in syngeneically mated immunodeficient SCID mice on the CB-17 background occurred at an unexpectedly high rate and could not be prevented by treatment with anti-asialo GM1 antibody or GM-CSF, both of which are effective in ameliorating abortion in DBA/2J-mated CBA/J mice . Immunocompetent CB-17 +/+ mice showed an even higher rate of loss . The latter was also not affected by treatment with antiasialo GM1 antibody or by GM-CSF and was not prevented by tetracycline (which is effective in the DBA/2-CBA/J system) or progesterone treatment . Mating experiments showed a scid/+ x scid@+ cross gave the highest rate of loss, and it appeared that the presence of +/(+)-type embryos in the uterus could be augmenting abortion with selective discrimination against scid/scid embryos . High abortion rates were associated both with appearance of a coagulase-negative Staphylococcus sp . in feces and with loss of one component of the SPF flora . Decidual tissue from mated CB-17 +/+ mice showed premature release of TNF-alpha in absence of TGF-beta 2-related suppressor activity, and vascular lesions (fibrinoid necrosis), varying in extent, were associated with both scid/scid x scid/scid and +/+ x +/+ pregnancies . TNF-alpha also appeared prematurely in pregnant scid/scid mice, but the levels were lower (and areas of necrosis smaller than in +/+ x +/+ pregnancies) . Outcrossing onto a C57B1/6 background dramatically reduced the abortion rate, indicating an important genetic effect on susceptibility with heterogeneity protecting against abortion . CONCLUSIONS: SCID mice on the CB-17 background do not have a high rate of successful syngeneic pregnancies, and a TNF-alpha induced vasculopathy may be responsible . Abortion was not caused by immunodeficiency leading to loss of immunotrophism because immunocompetent non-SCID CB-17 mice had a higher rate of loss . Factors augmenting the abortion rate included the presence of embryos of the +/+ genotype in the uterus and treatment with anti-asialo GM1 antibody . Abortion rates were not reduced by treatments effective in the DBA/2-mated CBA/J mouse model but were reduced by re-establishing a new colony with defined flora (a temporary effect) and by outcrossing mice with a different (C57B1/6) background . Together, the data suggest an infectious trigger (identity uncertain) of the vasculopathy and an important genetic influence on susceptibility with heterozygosity and a SCID mouse mutation providing against abortion a degree of protection.

Photochem Photobiol, 1994 Aug, 60(2), 147 - 53
Accessory cell ability of Langerhans cells for superantigen is resistant to ultraviolet-B light; Tokura Y et al.; We examined the effects of ultraviolet-B (UVB) irradiation on the accessory cell ability of Langerhans cells (LC) to induce a T-cell response to a superantigen, staphylococcal enterotoxin B (SEB) . The ability of LC-enriched epidermal cells (LC-EC) to evoke a T-cell response to SEB was retained at the doses of UVB (up to 40 mJ/cm2) that profoundly affected the antigen-presenting function of LC-EC for a hapten, trinitrophenyl (TNP), and a protein antigen, conalbumin . Thus, the LC accessory function for superantigens is more resistant to UVB irradiation than that for ordinary antigens . This UVB resistance is presumably due to no requirement of antigen processing for superantigens as chemically fixed or chloroquine-treated LC-EC still retained their ability to induce T-cell responses to SEB . Higher doses of UVB (more than 60 mJ/cm2) reduced the accessory cell ability of LC-EC for SEB up to 50% of control . The addition of monoclonal antibodies against adhesion molecules between LC and T cells to the culture resulted in a substantial suppression of the T-cell response to SEB induced by nonirradiated LC-EC, while the UVB-irradiated LC-EC-induced T-cell response was not significantly blocked with these monoclonal antibodies . This suggested that the reduction of LC ability for superantigen by high doses of UVB is at least partly due to the impairment of adhesion molecules on LC by UVB irradiation.

J Allergy Clin Immunol, 1994 Aug, 94(2 Pt 1), 189 - 94
Vancomycin hypersensitivity: synergism with narcotics and "desensitization" by a rapid continuous intravenous protocol; Wong JT et al.; BACKGROUND: We examined the clinical spectrum of patients with persistent adverse reactions to vancomycin, assessed contributing factors, and evaluated the efficacy and safety of a rapid continuous intravenous "desensitization" protocol in these patients . METHODS: Seven patients with serious staphylococcal infections resistant to beta-lactam antibiotics whose adverse reactions to vancomycin persisted despite slowing of the vancomycin infusion and pretreatment with H1-antihistamine were studied . All seven patients underwent a rapid continuous intravenous desensitization protocol with multiple small increases in vancomycin concentration tightly regulated with a syringe pump . RESULTS: Most of the seven patients safely achieved, during the first day, a vancomycin infusion rate (VIR) sufficient, or close to sufficient, to provide the desired vancomycin dose . In three patients there appeared to be a threshold VIR beyond which adverse reactions were repeatedly elicited; these reactions abated when the VIR was slightly lowered . Narcotic administration was found to adversely affect treatment with vancomycin . After concurrent narcotic administration was discontinued in three patients, they and the other four patients successfully completed the full course of treatment with vancomycin . CONCLUSION: Patients whose adverse reactions to vancomycin did not respond to slowing of the infusion rate and additional H1-antihistamines can be safely treated with a rapid continuous intravenous desensitization protocol and discontinuance of narcotic administration.

Clin Exp Immunol, 1994 Aug, 97(2), 266 - 72
Flow cytometric analysis of the stimulatory response of T cell subsets from normal and HIV-1+ individuals to various mitogenic stimuli in vitro; Medina E et al.; A novel technique is described which allows the study of the responses of T cell subpopulations stimulated in bulk cultures without interfering with cell-cell interactions . The number and phenotype of lymphoblasts developing following stimulation with phytohaemagglutinin (PHA), anti-CD3, staphylococcal protein A (SPA) and pokeweed mitogen (PWM) was determined in HIV-1- and HIV-1+ patients using a new five-parameter flow cytometric method . We found that normal T cells responded faster to PHA than to any of the other mitogens tested . The peak of the PHA response occurred on day 3, followed by anti-CD3 and SPA on day 4 and PWM mitogen on day 5 . Although PHA and anti-CD3 stimulated up to 95% and 80% of lymphocytes, respectively, SPA and PWM stimulated only 40% and 30% of cells, respectively . A defective T cell response was observed in lymphocytes cultured from asymptomatic HIV-1+ patients compared with negative controls . This loss of response was related to a selective mortality of T cells following mitogenic stimulation, referred to as activation-associated lymphocyte death (AALD) . The results showed that stronger mitogens (PHA and anti-CD3) induced AALD in a larger proportion (50-60%) of T cells than weaker mitogens such as SPA and PWM (30-40%), and that AALD affected different lymphocyte subsets to different extents . AALD occurred more frequently in total CD8+ and CD45RO+ T cells compared with CD4+ and CD45RA+ T cells, but memory CD4+ T cells were the population most severely affected in samples from HIV-1+ donors.

J Antimicrob Chemother, 1994 Aug, 34 Suppl A, 75 - 84
Hydrocephalus shunt infections; Bayston R; Hydrocephalus is most commonly diagnosed in the first few months of life, though cases also arise in later life . Cerebrospinal fluid shunts used to control the condition are prone to colonization particularly by Staphylococcus epidermidis . The incidence is very much higher in infancy than in older age groups, and this is probably due to prolonged hospital stay as a result of the underlying pathology, combined with the propensity for a high skin bacterial density with more adherent strains, rather than to any immune immaturity . Diagnosis of shunt colonization is often very difficult and serological tests have an important role to play even in infancy . There are several pitfalls in diagnosis, particularly in the elderly . Treatment of shunt infections should include removal of the colonized shunt, though regimens to avoid this are currently being investigated . Intraventricular therapy with vancomycin along with intravenous rifampicin offers the best changes of success at the first attempt . Shunted patients who contract purulent bacterial meningitis should not have their shunts removed but should be treated in the same way as those without shunts.

FEMS Immunol Med Microbiol, 1994 Aug, 9(2), 109 - 15
The effects of extracellular slime from Staphylococcus epidermidis on phagocytic ingestion and killing; Rodgers J et al.; Extracellular slime (Ecs) from three strains of Staphylococcus epidermidis was prepared and added to fresh suspensions of polymorphonuclear neutrophils . Phagocytic ingestion and killing of opsonised and unopsonised S . epidermidis strains was assessed over time using slide preparations stained by the Gram's method and microbiological culture . Both phagocytic ingestion and killing were inhibited . Investigation as to one possible mechanism of action of Ecs on phagocytes was performed using 1 mu polystyrene spheres which were incubated overnight with Ecs . It was found that the surface tension was altered with Ecs making the beads more hydrophilic, a factor which may interfere with the phagocytic response to infection.

Biochem Mol Biol Int, 1994 Aug, 33(6), 1215 - 20
Expression of a fibrinolytically active human pro-urokinase fusion protein in Escherichia coli; Hua Z et al.; The gene encoding human pro-urokinase(pro-UK) was cloned into plasmid pEZZ318 and fused to the gene coding for the signal peptide of staphylococcal protein A and IgG bindinging domain . The fusion protein which was synthesized under the control of T7 promoter in Escherichia coli and secreted into the growth medium, was found to be fibrinolytically active . Approximately 60% of the total activity was secreted into the culture medium, where levels of activity approached 150,000 I.U./liter and about 40% of the total activity remained in the cell lysate with levels of activity around 100,000 I.U./liter . The fusion protein was purified in a single step by IgG affinity chromatography . These results demonstrate that human pro-UK can be synthesized and secreted by E . coli as a fibrinolytically active fusion protein.

J Exp Med, 1994 Aug 1, 180(2), 615 - 21
Monoclonal antibodies defining functional sites on the toxin superantigen staphylococcal enterotoxin B; Hamad AR et al.; Four monoclonal antibodies (mAbs) were produced binding to four nonoverlapping epitopes on the superantigen staphylococcal enterotoxin B (SEB) . The mAbs were tested for their ability to detect SEB bound to major histocompatibility complex (MHC) class II, to inhibit SEB binding to MHC class II, to inhibit SEB stimulation of T cell hybridomas, to bind to various nonfunctional mutants of SEB, and to capture and present SEB and its mutants to T cells in the absence of MHC class II . We concluded that two mAbs, B344 and B327, bound to epitopes not required for superantigen function, one mAb, 2B33, blocked an MHC interaction site on SEB, and the fourth mAb, B87, blocked the T cell recognition site on SEB . Moreover, two mAbs (B344 and 2B33) were capable of presenting SEB, although much less efficiently than APC, to CD4- but not CD4+ T cell hybridomas . The results confirm the functional domains on SEB originally defined by mutation and show that MHC class II is not always an essential component of the superantigen ligand.

Gene, 1994 Jul 22, 145(1), 41 - 7
Primary structure and biological features of a thermostable nuclease isolated from Staphylococcus hyicus; Chesneau O et al.; The nucH gene, encoding a thermostable nuclease (TNase), was isolated from the cellular DNA of Staphylococcus hyicus strain E80 and sequenced . NucH, the 169-amino-acid (aa) protein encoded by this gene, contains, at its N-terminus, a signal peptide which appears to be cleaved at the same site in S . hyicus and Escherichia coli, yielding a mature protein which is exported extracellularly from S . hyicus, but not from E . coli . The aa sequence of NucH is highly homologous with that of the TNase from S . intermedius strain LRA076, whereas significant similarities are observed with the TNase from S . aureus, as well as with three other bacterial proteins of which only one has been shown to exhibit DNase activity . As seen in a multiple sequence alignment, the invariant residues are mostly located in the regions involved in the biological activity of the S . aureus TNase . The ability of crude cell extracts of E . coli strains carrying nucH to degrade various forms of nucleic acids with or without Ca2+ supplementation was studied . Under our experimental conditions, the enzyme encoded by nucH was active at 37 degrees C on both DNA and RNA, had the potential to act as an endonuclease, and functioned in the presence of Ca2+ . Moreover, activity was retained after heating at 100 degrees C, suggesting that the enzyme could undergo reversible unfolding.

Biochem J, 1994 Jul 15, 301 ( Pt 2), 523 - 9
Dynamics of Ca2+ and guanosine 5'-{gamma-thio}triphosphate action on insulin secretion from alpha-toxin-permeabilized HIT-T15 cells; Jonas JC et al.; The time course of Ca2+ and GTP-analogue effects on insulin secretion was investigated in HIT-T15 cells permeabilized with Staphylococcus alpha-toxin . These cells responded to Ca2+ in the range 0.1-10 microM and could be used in a dynamic perifusion system because of the minimal run-down of the secretory response . High Ca2+ (10 microM) elicited a monophasic ATP-dependent stimulation of insulin secretion that reached a peak within 5 min (approximately 20-fold increase) and rapidly decreased during the subsequent 15 min to a plateau remaining above basal rates (0.1 microM Ca2+) . The decrease in Ca(2+)-induced insulin secretion with time could not be attributed to decreased capacity to respond to Ca2+ after prolonged perfusion at low Ca2+ (run-down), nor to depletion of a particular secretory-granule pool . It was rather due to desensitization of the secretory machinery to Ca2+ that was not reversed by selective inhibition of the Ca2+/calmodulin-dependent kinase II with KN-62 . However, an intermediate Ca2+ concentration (2 microM) increased insulin secretion to stable level without causing any desensitization . Imposed oscillations of Ca2+ (0.1-10 microM) produced phasic oscillations of insulin secretion, but did not prevent desensitization to Ca2+ . Poorly hydrolysable GTP analogues increased insulin secretion at low Ca2+, whereas they strongly inhibited Ca(2+)-induced insulin secretion . By contrast, GTP did not affect basal secretion, and slightly increased Ca(2+)-evoked secretion . These results indicate the following . (1) Oscillations of insulin secretion are tightly coupled to cytosolic Ca2+ oscillations . (2) Oscillations of Ca2+ do not prevent high-Ca(2+)-induced desensitization to Ca2+; this result does not support the idea of a greater efficiency of oscillations compared with sustained Ca2+ rises in triggering exocytosis . (3) Activation of G-proteins modulates exocytosis in a bimodal manner.

Biochemistry, 1994 Jul 5, 33(26), 8017 - 28
NMR docking of a substrate into the X-ray structure of the Asp-21-->Glu mutant of staphylococcal nuclease; Weber DJ et al.; To understand the structural basis of the 1500-fold decrease in catalytic activity of the D21E mutant of staphylococcal nuclease in which an aspartate ligand of the essential Ca2+ has been enlarged to glutamate, the conformation of the enzyme-bound substrate dTdA has been determined by NMR methods and has been docked into the X-ray structure of the D21E mutant (Libson, A . M., Gittis, A.G., & Lattman, E . E . Biochemistry, preceding paper in this issue) based on distances from the bound metal ion to dTdA and on intermolecular nuclear Overhauser effects from assigned aromatic proton resonances of Tyr-85, Tyr-113, and Tyr-115 to proton resonances of dTdA, using energy minimization to relieve small overlaps . Like the wild-type enzyme, the D21E mutant forms binary E-M and E-S and ternary E-M-S complexes with Ca2+, Mn2+, Co2+, and La3+ . D21E enhances the paramagnetic effects of Co2+ on 1/T1 and 1/T2 of the phosphorus and on 1/T1 of four proton resonances of dTdA, and these effects are abolished by the binding of the competitive inhibitor 3',5'-pdTp . From the paramagnetic effects of enzyme-bound Co2+ on 1/T1 of phosphorus and protons, with the use of a correlation time of 1.1 ps based on 1/T1 values at 250 and 600 MHz, five metal-nucleus distances and 11 lower limit metal-nucleus distances have been calculated . The Co2+ to 31P distance of 4.1 +/- 0.9 A agrees with that found on the wild-type enzyme (Weber, D . J., Mullen, G . P., & Mildvan, A . S . (1991) Biochemistry 30, 7425-7437) and indicates at least 18% inner sphere phosphate coordination . Fourteen interproton distances and 109 lower limit interproton distances in dTdA in the ternary D21E-La(3+)-dTdA complex were determined by NOESY spectra at 50-, 100-, and 200-ms mixing times . Both the metal-nucleus and interproton distances were necessary to compute a narrow range of conformations for enzyme-bound dTdA . As on the wild-type enzyme, the conformation of dTdA on the D21E mutant is highly extended, with high-anti C-2' endo conformations for the individual nucleosides . However, significant conformational differences are found in the torsional angles chi of dA (delta chi = 49 +/- 3 degrees), in gamma of dT (delta gamma = 108 +/- 30 degrees) and in zeta of dT (delta zeta = 124 +/- 38 degrees).(ABSTRACT TRUNCATED AT 400 WORDS)

Biochemistry, 1994 Jul 5, 33(26), 8007 - 16
Crystal structures of the binary Ca2+ and pdTp complexes and the ternary complex of the Asp21-->Glu mutant of staphylococcal nuclease . Implications for catalysis and ligand binding; Libson AM et al.; The crystal structure of the Asp21-->Glu mutant (D21E) of staphylococcal nuclease (SNase) has been determined in three different complex forms . The structure of the D21E ternary complex in which D21E is bound to both Ca2+ and the transition-state analogue, thymidine 3',5'-diphosphate (pdTp), was determined to 1.95-A resolution . The structures of both binary complexes, D21E bound either to Ca2+ or pdTp, were determined to 2.15- and 2.05-A resolution, respectively . In the ternary structure, we find a 1.5-A movement of the Ca2+ in the active site, evidence of bidentate coordination of Ca2+ by Glu21 and inner-sphere coordination of the Ca2+ by Glu43 . Comparison of the D21E binary structures with the ternary model shows large movements of active site side chains expected to play a direct role in catalysis . Glu43 moves in the binary nucleotide complex, whereas Arg35 is oriented differently in the binary metal complex . From these changes, we seek to explain the basis for the 1500-fold decrease in Vmax of D21E relative to wild-type SNase (WT) . Furthermore, we describe direct structural evidence which explains the cooperativity of Ca2+ and pdTp binding in the ternary complex relative to that of the binary complexes.

J Urol, 1994 Jul, 152(1), 213 - 6
Protamine sulfate and vancomycin are synergistic against Staphylococcus epidermidis prosthesis infection in vivo; Teichman JM et al.; We have previously demonstrated that the quaternary amine, protamine sulfate (PS), is bactericidal against Staphylococcus epidermidis . In an attempt to decrease genitourinary prosthesis infection rates, we examined the ability of PS as a wound irrigant to inhibit Staphylococcus epidermidis viability . Eighty-seven Sprague-Dawley rats were studied by implanting a sterile silicone pellet in their dorsum . The pellet was inoculated with Staphylococcus epidermidis and the rats were divided into four groups based on the wound irrigant employed after inoculation: (1) control (sterile water) (2) vancomycin; (3) PS; (4) vancomycin + PS . All rats received perioperative and daily intramuscular vancomycin, and the pellets were explanted on postoperative day 28 and cultured . The infection rates were: (1) control 77%, (2) vancomycin 50%, (3) protamine sulfate 67%, and (4) protamine sulfate and vancomycin 19% . The differences between (2) vancomycin versus (4) vancomycin + PS and (3) PS versus (4) vancomycin + PS were significant (p = 0.05 and p < 0.005) . The data suggest that PS potentiates vancomycin as a wound irrigant in prosthesis implantation.

Nippon Kyobu Geka Gakkai Zasshi, 1994 Jul, 42(7), 1101 - 4
{Successful surgical treatment in a case of complete rupture of the posterior papillary muscle of the mitral valve caused by infective endocarditis}; Terai H et al.; Forty three year-old male who had cough and easy fatigability since three weeks prior to admission was diagnosed to have acute pulmonary edema with severe mitral regurgitation caused by active infective endocarditis . Transesophageal echocardiograms under the endotracheal intubation for controlled respiration suggested rupture of the posterior papillary muscle of the mitral valve and the emergency surgical treatment was performed . Intraoperatively the total rupture of the posterior papillary muscle was confirmed and mitral valve replacement was carried out with a SJM prosthetic valve . Histological examination of the ruptured papillary muscle revealed hemorrhage, muscle necrosis and small cell infiltration suggesting the presence of active inflammation with bacteria on it . Staphylococcus epidermidis was demonstrated by the bacteriological studies of the ruptured papillary muscle.

J Dermatol, 1994 Jul, 21(7), 453 - 60
Comparative study of staphylococcal flora on the skin surface of atopic dermatitis patients and healthy subjects; Ogawa T et al.; Bacterial samples were obtained from the facial and forearm skin of atopic dermatitis (AD) patients and healthy subjects in humid summer weather . Staphylococcus (Staph) species bacteria were selectively sampled by a plate-contact technique using mannitol salt medium-coated film . Staph species were isolated from the film after the number of colonies was counted, and the species were determined by the API-Staph system and coagulase test . Large numbers of S . aureus were isolated in the samples from the facial skin of patients with severe dermatitis, as previously reported . The number of S . aureus colonies in the samples from the forearm skin of the patients was significantly greater than from the healthy subjects; the mean and standard deviation of colony forming units per 10 cm2, expressed as log10 value, were 1.64 +/- 0.96 for patients and 0.02 +/- 0.07 for healthy subjects (p < 0.01) . One hundred and seventeen strains were isolated, and their sensitivities to ten antibiotics were tested . Ten of the strains were resistant to some antibiotics; all of these were isolated from patients . Penicillin type antibiotics had weak effects, but cephem type had strong effects . The number of S . aureus colonies isolated in this study was compared with the number isolated during the winter, in our previous study, in order to identify seasonal variations . The only significant seasonal difference was on the forearm skin of the AD patients, which had significantly more S . aureus colonies (p < 0.01) in this study . There were no significant seasonal differences in S . aureus numbers on the facial skin of patients or healthy subjects.

J Biomol NMR, 1994 Jul, 4(4), 543 - 51
Estimates of phi and psi torsion angles in proteins from one-, two- and three-bond nuclear spin-spin couplings: application to staphylococcal nuclease; Edison AS et al.; Calculated coupling constants (3JHNH alpha, 1JC alpha H alpha, 2JC'H alpha, 1JC alpha N and 2JC alpha N) from our accompanying paper {Edison, A.S . et al . (1994) J . Biomol . NMR, 4, 519-542} have been used to generate error surfaces that can provide estimates of the phi and psi angles in proteins . We have used experimental coupling data {3JHNH alpha: Kay, L.E . et al . (1989) J . Am . Chem . Soc., 111, 5488-5490; 1JC alpha H alpha: Vuister, G . W . et al . (1993) J . Biomol . NMR, 3, 67-80; 2JC'H alpha: Vuister, G.W . and Bax, A . (1992) J . Biomol . NMR, 2, 401-405; 1JC alpha N and 2JC alpha N: Delaglio, F . et al . (1991) J . Biomol . NMR, 1, 439-446} to create error surfaces for selected residues of the protein staphylococcal nuclease . The residues were chosen to include all those with five experimental couplings, as well as some with four experimental couplings, to demonstrate the relative importance of 3JHNH alpha and 1JC alpha H alpha . For most of the cases, we obtained good agreement between the X-ray structure {Loll, P.J . and Lattman, E.E . (1989) Protein Struct . Funct . Genet., 5, 183-201} and the NMR data.

J Biomol NMR, 1994 Jul, 4(4), 519 - 42
Calculations of one-, two- and three-bond nuclear spin-spin couplings in a model peptide and correlations with experimental data; Edison AS et al.; We present ab initio calculations of the Fermi contact term and experimental correlations of six coupling constants, 3JHNH alpha, 1JC alpha H alpha, 2JC'H alpha, 1JC alpha N, 2JC alpha N and 1JC'N, in a peptide as functions of the backbone dihedral angles, phi and psi . Given estimates of experimental uncertainties, we find semiquantitative experimental correlations for 3JHNH alpha, 1JC alpha N and 2JC alpha N, qualitative correlations for 1JC alpha H alpha and 2JC'H alpha, but no experimental correlations of practical utility for 1JC'N, owing to its complex dependence on at least four dihedral angles . Errors in the estimation of dihedral angles from X-ray crystallographic data for proteins, which result from uncertainties in atom-to-atom distances, place substantial limitations on the quantitative reliability of coupling constant calculations fitted to such data . In the accompanying paper {Edison, A.S . et al., J . Biomol . NMR, 4, 543-551} we apply the results of the coupling constant calculations presented here to the estimation of phi and psi angles in staphylococcal nuclease from experimental coupling constants.

Int J Syst Bacteriol, 1994 Jul, 44(3), 404 - 9
Comparison of the SmaI-digested chromosomes of Staphylococcus epidermidis and the closely related species Staphylococcus capitis and Staphylococcus caprae; George CG et al.; Pulsed-field gel electrophoresis was used to examine the chromosomal polymorphisms existing within and between four closely related members of the Staphylococcus epidermidis species group, S . epidermidis, Staphylococcus caprae, Staphylococcus capitis subsp . capitis, and S . capitis subsp . ureolyticus . SmaI was chosen as the restriction endonuclease for this study because it generated only a few well-separated chromosomal fragments . Each of the species and subspecies showed distinct SmaI digest patterns . The strains examined in this study were collected over a 20-year period from various geographical locations . The results indicate that DNA fragment patterns are unique to each species and subspecies and represent a reasonably stable component in the chromosome structure . S . caprae and S . capitis demonstrated considerable conservation in chromosome structure as indicated by the large numbers of conserved SmaI digest fragments . The polymorphisms found within each species appear to be linked to the species' character variability . The genome size of each Staphylococcus strain was extrapolated from the SmaI digest fragment pattern obtained by pulsed-field gel electrophoresis . The average genome size for S . epidermidis is 2,364 +/- 119 kb; for S . caprae strains from humans it is 2,600 +/- 157 kb and for S . caprae strains from goats it is 2,493 +/- 15 kb; for S . capitis subsp . capitis it is 2,456 +/- 71 kb; and for S . capitis subsp . ureolyticus it is 2,276 +/- 90 kb.

J Orthop Res, 1994 Jul, 12(4), 526 - 31
Infection at the site of implanted materials with and without preadhered bacteria; Chang CC et al.; A study was undertaken to examine the role of bacterial adherence in the development of infection at the site of an implant . The amount of in vitro adherence of Staphylococcus epidermidis was greatest for stainless steel, followed by polymethylmethacrylate and commercially pure titanium, and was least for polymethylmethacrylate with gentamicin . These materials then were preincubated with S . epidermidis and implanted . The number of organisms that were isolated and the rate of infection followed the same pattern as that in the in vitro studies . Materials that were not preincubated with bacteria also were implanted and bacteria were injected into the site . The number of organisms isolated from the site and the rate of infection were lower than those for the preincubated materials, but the trend was the same as in both the in vitro and the in vivo studies . The rates of infection and colonization correlated with the propensity for the organisms to adhere to a given material . Materials colonized with S . epidermidis at the time of implantation caused a high rate of infection . The ability of organisms to adhere to a material in vitro is correlated with their propensity to cause biomaterial-based infection.

Eur J Immunol, 1994 Jul, 24(7), 1604 - 11
Flexibility of the T cell receptor repertoire; Liang HE et al.; Alternative T cell receptor (TcR) gene usage between mice of different Mls alleles has been demonstrated in a number of T cell responses . A clear illustration of a flexible TcR V beta usage in the same strain of mice remains to be established . Using a model system in which I-Ek-restricted T cells recognizing lambda repressor cI protein (cI) 12-26 and pigeon cytochrome c (pcc) 81-104 predominantly use V beta 3 in B10.A and B10.BR mice, and V beta 1 in Mls-2a-bearing A/J and C3H mice, we have first demonstrated that the hierarchy of TcR V beta usage can not be inferred from one strain of mice to the other . The presumed flexibility of V beta 3 to V beta 1 did not exist in B10.BR mice in the given responses . Instead, a switch of dominant TcR from V beta 1/V beta 3 to V beta 8 was identified in C3H and B10.BR mice . In contrast, there was an absolute rigidity in TcR repertoire usage in some mouse strains such as A/J . The lack of flexibility was not due to slow generating kinetics of replacing T cells; since A/J mice treated with staphylococcal enterotoxin A from birth on still responded poorly to cI 12-26 and pcc 81-104 . Therefore, whether TcR V beta usage in a T cell response would be flexible or rigid is highly dependent on each strain of mice . However, even the plasticity seen in B10.BR mice is very limited and further tolerance of the V beta 8+ population results in non-responsiveness toward the given antigens.

Cell Immunol, 1994 Jul, 156(2), 310 - 21
Differential expression of activation markers during tolerance induction by superantigens in T-cell receptor (beta-chain) transgenic mice; Perkins DL et al.; To investigate the process of tolerance induction we have developed an in vivo model using TCR beta-chain transgenic mice tolerized with the superantigen staphylococcal enterotoxin B . We have previously demonstrated that tolerized peripheral T cells were anergic when stimulated in vitro with immunogenic peptides, superantigens, mitogens, and immobilized anti-TCR mAb . However, the development of anergy is preceded by an induction phase which produces expansion followed by contraction of the peripheral T cell population presumably due to proliferation and programmed cell death, respectively . The current experiments focus on the induction phase of tolerance . A kinetic functional analysis showed that the inhibition of proliferation was apparent 2-3 days post-tolerization . Interestingly, the inhibition of proliferation correlated with the loss of IL-2R alpha expression, which occurred 2 days post-tolerization following an initial increase in IL-2R alpha expression . In addition, the expression of multiple activation markers including CD44, Ly-6A/E, and very early activation marker H1.2F3 is induced, whereas the expression of CD45RB is decreased during tolerance induction . Elevated expression of Ly-6A/E persists up to 28 days post-tolerization; however, altered expression of the other markers does not persist and near baseline levels of the other markers are noted 7 to 28 days post-tolerization . These results show that tolerance induction is an active process which has functional and phenotypic similarities to antigen-specific immunity . However, tolerance induction in our system differs from immunity in terms of the early loss of IL-2R alpha expression, the persistent increased expression of Ly-6A/E, and the lack of development of CD45RBlo memory-type T cells.

J Bacteriol, 1994 Jul, 176(14), 4204 - 9
Glycerol monolaurate inhibits the production of beta-lactamase, toxic shock toxin-1, and other staphylococcal exoproteins by interfering with signal transduction; Projan SJ et al.; Glycerol monolaurate (GML) is a naturally occurring surfactant that is used widely as an emulsifier in the food and cosmetics industries and is generally regarded as lacking in important biological activities . The recent observation that it inhibits the production of staphylococcal toxic shock toxin-1 (P . M . Schlievert, J . R . Deringer, M . H . Kim, S . J . Projan, and R . P . Novick, Antimicrob . Agents Chemother . 36:626-631, 1992) is therefore rather surprising and raises the interesting question of how such a compound might interact with cells . In this report, we show that GML inhibits the synthesis of most staphylococcal toxins and other exoproteins and that it does so at the level of transcription . We find that GML blocks the induction but not the constitutive synthesis of beta-lactamase, suggesting that it acts by interfering with signal transduction.

Am J Gastroenterol, 1994 Jul, 89(7), 1090 - 5
Liver abscess in Crohn's disease; Vakil N et al.; Liver abscess is a rare but serious complication of Crohn's disease . Intra-abdominal abscesses, fistulous disease, and metronidazole or steroid therapy have all been reported to be important predisposing factors in the pathogenesis of the disease, and the mortality has been reported to be high . We report six patients who developed a liver abscess as a complication of Crohn's disease . Three patients presented with a liver abscess as the first manifestation of Crohn's disease and two others had quiescent disease at presentation . The diagnosis was delayed by 1-8 wk after the onset of fever because of the paucity of signs indicating a hepatic infection . None of the patients had intra-abdominal abscesses, active fistulas, or metronidazole therapy before the onset of symptoms . The only predisposing conditions identified were two minor skin infections in patients developing staphylococcal liver abscesses . Nonoperative catheter drainage was successful in four of the six patients . One patient required surgical placement of drains, and the patient with the longest delay before diagnosis required hepatic lobectomy because of extensive necrosis . Shaking chills, fever with leukocytosis, and an elevated alkaline phosphatase are suggestive of a liver abscess and should prompt an ultrasound examination . Catheter drainage with antibiotic therapy is effective if the liver abscess is diagnosed before extensive necrosis has occurred . Minor skin infections may predispose to staphylococcal liver abscess in some cases.

J Infect Dis, 1994 Jul, 170(1), 100 - 9
Antibiotic treatment of experimental endocarditis due to methicillin-resistant Staphylococcus epidermidis; Entenza JM et al.; The natural history and treatment of experimental endocarditis due to heterogeneous and homogeneous methicillin-resistant Staphylococcus epidermidis was investigated . Amoxicillin/clavulanate or vancomycin were administered for 3 days via a computerized pump to mimic human drug kinetics in animals . After challenge with the minimum inoculum producing 90% of infections (ID90), bacteria in the vegetations grew logarithmically for 16 h . Then, bacterial densities stabilized (at approximately 10(8) cfu/g) and growth rates sharply declined . Both regimens cured > or = 60% of endocarditis (due to heterogeneous or homogeneous bacteria) when started 12-16 h after infection, although the bacterial densities in the vegetations had increased by 20 times in between . In contrast, treatment started after 24 h failed in most animals, while bacterial densities had not increased any more . Thus, while both regimens were equivalent, the therapeutic outcome was best predicted by growth rates in the vegetations, not by bacterial densities . These observations highlight the importance of phenotypic tolerance developing in vivo.

Bioorg Khim, 1994 Jul, 20(7), 759 - 71
{Synthesis and cloning of genes for the antisense peptides for human calcitonin and miniproinsulin}; Efimov VA et al.; Because of the potential significance of the 'molecular recognition' theory for studies in molecular biology and biotechnology, the theory is worth being examined using methods of chemical-enzymatic gene synthesis, recombinant DNA construction and microbiological peptide synthesis . We therefore undertook the synthesis of human Va18-calcitonin, miniproinsulin, and the corresponding antisense peptides as model compounds . In designing an experimental system the idea was to combine sense and antisense polypeptides into a single chain and to examine their intramolecular interaction . In this paper the chemical-enzymatic synthesis, cloning and expression of the genes for calcitonin, miniproinsulin, the corresponding antisense peptides and their combinations are described . The recombinant DNAs obtained were able to direct in vivo expression of the target polypeptides as hybrid proteins with the IgG-binding domain of the staphylococcal A protein in bacterial cells.

New Microbiol, 1994 Jul, 17(3), 211 - 6
Effect of subinhibitory concentrations of lomefloxacin on bacterial adherence; Ravizzola G et al.; We studied the effect of subinhibitory concentrations of lomefloxacin on bacterial adherence, assessing the presence of fimbriae in E . coli and the adherence to uroepithelial cells of Staphylococcus saprophyticus . The degree of inhibition of fimbriae in E . coli in the log-phase is directly proportional to subminimal inhibiting concentrations (sub-MIC) . On the contrary, there is no effect when the bacteria are in a stationary phase . In the case of Staphylococcus saprophyticus the adherence to epithelial cells was observed at concentrations of 1/2 and 1/4 the MIC.

Gen Pharmacol, 1994 Jul, 25(4), 729 - 37
Fc epsilon RI-stimulated Ca(2+)-dependent secretion from rat basophilic leukemia (RBL-2H3) cells permeabilized with Staphylococcal alpha-toxin: Fc epsilon RI-operated signals are not mimicked by the actions of GTP gamma S; Oishi K et al.; 1 . RBL-2H3 cells permeabilized with alpha-toxin responded to dinitrophenol (30-40 mol/mol)-conjugated human serum albumin, as antigen, to secrete {14C}serotonin in the micromolar range of free Ca2+ . 2 . Calcium ion alone did not cause substantial secretion . 3 . Guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S) (100 microM) in combination with Ca2+ produced only negligible {14C}serotonin secretion . 4 . GTP gamma S, in the presence of cytochalasin D, caused optimal secretion of {14C}serotonin in a Ca(2+)-dependent manner.

Int Immunol, 1994 Jul, 6(7), 983 - 9
Mechanism underlying superantigen-induced clonal deletion of mature T lymphocytes; Biasi G et al.; We observed that peripheral T cells activated in vivo or in vitro by superantigens are susceptible to cell death when their antigen receptor is cross-linked with the appropriate anti-alpha beta TCR mAb . TCR ligation by mAbs specifically drove the T cell clonal deletion in both CD4+ and CD8+ cell subsets . An IL-2/IL-2R interaction seems to be a critical step in predisposing superantigen activated cells to death; in fact, in vivo IL-2R blockade reversed T cell deletion in superantigen plus anti-alpha beta TCR mAb treated mice . TCR ligation by mAbs also produced cell death of the relevant targets in in vitro IL-2 activated T cells . Surprisingly, no T cell deletion was demonstrable in IL-2 activated cells following staphylococcal enterotoxin B--TCR interaction, ruling out the possibility that superantigen in itself can induce cell death . Thus, while superantigen activation opens the cell death program, a subsequent TCR--antigen (self) interaction appears necessary to produce clonal deletion in mature T lymphocytes.

Br J Ophthalmol, 1994 Jul, 78(7), 546 - 8
Ocular penetration of topical ciprofloxacin and norfloxacin drops and their effect upon eyelid flora; Leeming JP et al.; A double blind, prospective study was undertaken to compare aqueous humour penetration of topical 0.3% norfloxacin and 0.3% ciprofloxacin and their effect upon normal eyelid flora in 39 patients undergoing cataract surgery . Lid swabs were taken before and after six 1 hourly applications of single drops of ciprofloxacin or norfloxacin given before surgery . Aqueous humour was aspirated at surgery and antibiotic concentration assayed using high performance liquid chromatography . The mean aqueous humour concentrations were: ciprofloxacin 220 ng ml-1, norfloxacin 140 ng ml-1 . Although this difference was not statistically significant (p = 0.112) the trend demonstrated may be relevant clinically, especially considering the greater activity of ciprofloxacin . Both coagulase negative staphylococcal (p = 0.004) and total bacterial (p = 0.019) lid counts dropped sixfold after ciprofloxacin treatment but the smaller reductions noted after norfloxacin application did not achieve statistical significance (p > 0.1) . The reduction of external eye flora experienced with ciprofloxacin suggests that this may be a useful presurgical prophylactic agent.

Rev Hosp Clin Fac Med Sao Paulo, 1994 Jul-Aug, 49(4), 168 - 72
{Hospital bacteremia at the "Instituto do Coração do Hospital das Clínicas da FMUSP": a four-year retrospective study}; Camargo LF et al.; We conducted a retrospective study to establish mortality rates and prevalence of nosocomial bacteremias at our institute . We found 1.21 nosocomial bacteremias per 100 hospital discharges with an overall Mortality rate of 29.5% . Primary bacteremias increased during the four-year-study-period from 31 to 41% . Staphylococcus, both coagulase-positive and coagulase-negative, was the bacteria most frequently isolated . An abrupt increase in the isolation of P.aeruginosa occurred in 1992 . We concluded that a blood-culture surveillance program is required for determining an endemic rate.

Int J Syst Bacteriol, 1994 Jul, 44(3), 454 - 60
Identification of the Staphylococcus sciuri species group with EcoRI fragments containing rRNA sequences and description of Staphylococcus vitulus sp . nov; Webster JA et al.; Strains of a new species, Staphylococcus vitulus, were isolated from food and a variety of mammals . This species was recognized on the basis of the results of an analysis of genomic EcoRI fragments containing portions of the rRNA operons . The patterns of hybridized fragments obtained from strains belonging to the new taxon were sorted into a distinguishable cluster and were distinct from the Staphylococcus lentus and Staphylococcus sciuri patterns . The results of DNA-DNA hybridization reactions demonstrated that strains in this cluster were more closely related to S . lentus and S . sciuri than to other Staphylococcus species and yet were significantly different . While these strains had some of the phenotypic characteristics of the S . sciuri species group, the newly recognized taxon could be distinguished by its very small colonies on P agar, absence of alkaline phosphatase activity, and lack of acid production from L-arabinose, maltose, N-acetylglucosamine, D-mannose, and raffinose . The type strain of the new species is strain DD 756 (= ATCC 51145).

J Immunol, 1994 Jul 1, 153(1), 117 - 27
Human T cell-dependent B cell differentiation induced by staphylococcal superantigens; Stohl W et al.; Microbial superantigens (SAgs), by virtue of their binding to TCR V beta elements on T cells and to class II MHC molecules on accessory cells (AC), trigger T cell activation . Although anti-CD3 mAb (which also trigger T cell activation via surface CD3/TCR) can readily induce T cell-dependent B cell differentiation in unmanipulated PBMC cultures, induction of Ig production in SAg-stimulated cultures has usually required special manipulation of the T cells, such as irradiating them or treating them with mitomycin C . We now demonstrate that eight different staphylococcal SAgs, typically at concentrations 10- to 100-fold lower than those required for proliferation, can each trigger unmanipulated peripheral blood and tonsil T cells to drive polyclonal B cell differentiation . Such SAg-induced T cell-dependent generation of Ig-secreting cells (IgSC) requires T cells and B cells only and occurs in the absence of monocytes as long as there are adequate numbers of B cells to serve as (DR+) AC . Physical contact among T cells, responder B cells, and AC (when different from the responder B cells) is required . The fusion protein CTLA4Ig inhibits SAg-induced IgSC generation in a dose-dependent fashion, whereas a control fusion protein has no such effect . In contrast, CTLA4Ig has, at best, only modest effects on SAg-induced T cell proliferation, indicating that CD28 (CTLA4)/B7 (B7-like) interactions play a more prominent role in SAg-induced IgSC generation than in SAg-induced T cell proliferation . These results establish SAg-induced T cell-dependent B cell differentiation as a useful model for T cell/B cell interactions, inasmuch as no other cell types are necessary for successful B cell differentiation; these results also demonstrate the importance of CD28 (CTLA4)/B7 (B7-like)-dependent mechanisms in this process.

Biochem Biophys Res Commun, 1994 Jun 30, 201(3), 1242 - 7
Staphylococcal protein A containing phospholipid monolayers on aqueous and solid surfaces; Lu B et al.; The staphylococcal protein A containing monolayer is characterized on both aqueous and solid surfaces . The density of SpA in dipalmitoylphosphatidic acid (DPPA) monolayers is determined from ultraviolet spectroscopy . The influence of calcium on the SpA concentration in the monolayer is also investigated . The spectroscopic results indicate that the incorporation of SpA in the monolayer is influenced by the calcium concentration in the subphase, and more than 90% of adsorbed protein is excluded from the monolayer when calcium concentration is in the order of millimoles . The IgG binding activity of the SpA in the reconstituted membrane is influenced by the surface pressure at which the membrane is formed.

Gene, 1994 Jun 24, 144(1), 69 - 73
Vectors to facilitate the creation of translational fusions to the maltose-binding protein of Escherichia coli; Aitken R et al.; A set of vectors has been constructed to facilitate the fusion of heterologous sequences to the C terminus of the maltose-binding protein (MBP) of Escherichia coli . The plasmids carry a cloning region comprising two blunt cloning sites, a BamHI site and multiple stop codons, and this has been placed in each reading frame so that translational fusions to MBP can be generated and manipulated with ease . To demonstrate the utility of this system, recombinant proteins have been engineered in which staphylococcal enterotoxin A has been fused to MBP.

Biochem Biophys Res Commun, 1994 Jun 15, 201(2), 855 - 60
Signalling via MHC class II molecules selectively induces IL-1 beta over IL-1 receptor antagonist gene expression; al-Daccak R et al.; Activation of human monocytes or human monocytic cell lines by several types of stimuli coordinately induces IL-1 beta and its antagonist (IL-1Ra) gene expression; alterations in their balance seem to mediate the inflammatory response . Using the human monocytic cell line THP-1, we report that superantigens, such as staphylococcal enterotoxin A (SEA) and Mycoplasma arthritidis -derived superantigen (MAM) induce an increase in the level of IL-1 beta mRNA without any detectable effect on IL-Ra mRNA . Unlike MAM-induced IL-1 beta mRNA, SEA-induced IL-1 beta mRNA was adequately translated into protein . Superantigen-induced gene expression is mediated by signalling, via their receptors, the MHC class II molecules . Thus, it appears that this mode of signalling selectively induces the proinflammatory cytokine IL-1 beta gene expression which, by itself, can have major importance in disease pathology especially in autoimmune diseases.

Biochem Biophys Res Commun, 1994 Jun 15, 201(2), 596 - 602
Multiple binding sites on the superantigen, staphylococcal enterotoxin B, imparts versatility in binding to MHC class II molecules; Soos JM et al.; To determine MHC class II molecule binding regions of staphylococcal enterotoxin B (SEB), we employed a structurally based approach in which eight overlapping peptides of the entire SEB molecule were synthesized to encompass discrete secondary structures based on the SEB crystalline structure . SEB peptides encompassing amino acid residues 1-33, 31-64 and 179-212 successfully competed with {125I}SEB for binding to DR1 transfected L cells . In contrast, SEB peptides encompassing amino acid residues 1-33, 124-154, 150-183 and 179-212 successfully competed with {125I}SEB for binding to Raji cells (HLA-DR3, DRw10, DQw1 and DQw2) . In addition, the SEB peptide (124-154) inhibited the mitogenic function of SEB . Thus, we have identified multiple regions, including the C-terminus, of SEB that are involved in binding to MHC class II and have shown that these interactions are complex and dependent on the haplotype of the MHC class II molecule.

Circulation, 1994 Jun, 89(6), 2684 - 7
Usefulness of transesophageal echocardiography for diagnosis of infected transvenous permanent pacemakers; Vilacosta I et al.; BACKGROUND: Transesophageal echocardiography is superior to transthoracic echocardiography in detecting left-sided valvular vegetations . There are no data on the value of transesophageal echocardiography in the diagnosis of infected transvenous permanent pacemakers . METHODS AND RESULTS: Transthoracic and transesophageal echocardiography was performed in 10 patients for whom there was clinical suspicion of infected permanent transvenous pacemakers . Transthoracic echocardiography detected pacemaker lead vegetations in 2 patients, whereas transesophageal echocardiography visualized pacemaker lead vegetations in 7 patients . Surgical confirmation was obtained in 6 of these 7 patients . Most patients had more than one pacemaker electrode in place . Local complications at the generator pocket were present in 6 patients . Staphylococcus was the predominant causative organism . CONCLUSIONS: Transesophageal echocardiography is superior to transthoracic echocardiography in the detection of pacemaker lead vegetations.

J Bacteriol, 1994 Jun, 176(11), 3218 - 23
Biochemical properties of a novel metalloprotease from Staphylococcus hyicus subsp . hyicus involved in extracellular lipase processing; Ayora S et al.; Two extracellular proteases from Staphylococcus hyicus subsp . hyicus, ShpI and ShpII, have been characterized . ShpI is a neutral metalloprotease with broad substrate specificity; the gene has been cloned and sequenced . ShpII, characterized here, is mainly produced in the late logarithmic growth phase in contrast to ShpI, which is mainly produced in the late stationary growth phase . ShpII was purified from culture medium of S . hyicus by ammonium sulfate precipitation and DEAE-Sepharose chromatography . The molecular mass, estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, was 34 kDa . The temperature optimum of ShpII was 55 degrees C, and the pH optimum was 7.4 . ShpII, a neutral metalloprotease, was strongly inhibited by zinc and calcium chelators . The amino-terminal sequence of the active enzyme was similar to the corresponding region of a Staphylococcus epidermidis metalloprotease . The substrate specificity of ShpII was similar to that of thermolysin-like proteases, with the exception that ShpII also recognized aromatic amino acids . We demonstrated in vitro that ShpII, but not ShpI, cleaved the 86-kDa S . hyicus subsp . hyicus prolipase between Thr-245 and Val-246 to generate the mature 46-kDa lipase . Results of additional in vivo experiments supported the model that ShpII is necessary for the extracellular processing and maturation of S . hyicus subsp . hyicus lipase.

Infect Immun, 1994 Jun, 62(6), 2249 - 56
Histidine residues near the N terminus of staphylococcal alpha-toxin as reporters of regions that are critical for oligomerization and pore formation; Jursch R et al.; Chemical modification of histidine residues in staphylococcal alpha-toxin leads to loss of functional activity . Site-directed mutants of the toxin in which each of the four histidine residues was replaced by several amino acids were therefore produced . The mutant proteins were purified and characterized . Exchange of H-259 or H-144 was sometimes tolerated without reduction in hemolytic activity . These histidine residues are thus not essential for toxin function . Exchange of H-35 and H-48, however, had marked effects . H-35 mutant toxins bound with high affinity to rabbit erythrocytes but displayed faulty oligomerization and were unable to form pores . H-48 mutant toxins also had severely impaired hemolytic activity due probably to faulty hexamerization . We interpret these results to indicate that the N-terminal domain of alpha-toxin in the region of H-35 and H-48 is involved in protomer-protomer interactions that underlie the hexamerization and pore-forming process.

Antimicrob Agents Chemother, 1994 Jun, 38(6), 1251 - 5
Lack of mecA transcription in slime-negative phase variants of methicillin-resistant Staphylococcus epidermidis; Mempel M et al.; Five phase variants (PV1 to PV5) of the well-characterized, slime-producing, methicillin-resistant, pathogenic strain of Staphylococcus epidermidis sensu strictu RP62A (ATCC 35984) were isolated by the Congo red agar method . In comparison with the parent strain, the phase variants showed a different colonial morphology on Congo red agar, a strongly reduced adherence capacity, and decreased levels of resistance to methicillin, oxacillin, and penicillin . All phase variants yielded biochemical reaction patterns and profiles in pulsed-field gel electrophoresis identical to those of parent strain RP62A, indicating a common origin . All phase variants proved to have the capacity to shift back to the original phenotype of parent strain RP62A . A search for the resistance mechanisms of strain RP62A revealed beta-lactamase production and the presence of mecA in PV1 to PV5 as well as parent strain RP62A . In Northern blots of total staphylococcal RNA, the phase variants showed no detectable mecA-specific transcription product, whereas parent strain RP62A revealed a strong signal, indicating that mecA transcription is not the mechanism responsible for the decreased methicillin resistance phenotype of phase variants PV1 to PV5.

Int Immunol, 1994 Jun, 6(6), 925 - 30
Exposure of monocytes to heat shock does not increase class II expression but modulates antigen-dependent T cell responses; Mariethoz E et al.; Expression of heat shock (HS) proteins (HSP) increases after exposure to elevated temperatures or other types of injury, such as oxidative injury . Because of their function as 'molecular chaperones', HSP are suggested to participate in antigen processing and presentation . We have previously reported that HS modulates antigen presentation in a human EBV-transformed B cell line . Here we investigated the effects of HS on MHC class II expression and on antigen processing and presentation by human monocytes . Monocytes were isolated from peripheral blood of normal human volunteers, purified by adherence, then exposed to temperatures ranging from 37 to 45 degrees C for 20 min, allowed to recover for 2 h at 37 degrees C and used for immunofluorescence or as antigen presenting cells in autologous and heterologous lymphocyte proliferation assays . No increase in class II expression was detected as assessed by flow cytometry . Monocytes (3 x 10(4)) and lymphocytes (1 x 10(5)) were co-cultured for 5 days in the presence of several antigens {diphtheria toxoid, tetanus toxoid or purified peptide derivative (PPD)} and labeled with 1 microCi {3H}thymidine for 16 h . Pre-exposure to HS (44 degrees C) significantly (P < 0.001) increased T cell responses to diphtheria toxoid, whereas the effect on the responses to other antigens (tetanus toxoid or PPD) were not significant . HS did not increase heterologous T cell responses nor T cell proliferation induced by the non-processed superantigens such as staphylococcal enterotoxin B . The effect of HS was inhibited by actinomycin B and thus appeared dependent upon HSP synthesis . HSP-mediated increases in antigen processing may potentiate the ongoing immune response at inflammatory sites.

Protein Sci, 1994 Jun, 3(6), 952 - 9
Thermodynamics of staphylococcal nuclease denaturation . II . The A-state; Carra JH et al.; Staphylococcal nuclease, at low pH and in the presence of high salt concentrations, has previously been proposed to exist in a partially folded or molten globule form called the "A-state" (Fink et al., 1993, Protein Sci 2:1155-1160) . We have found that the A-state of nuclease at pH 2.1 in the presence of moderate to high salt concentrations and at low temperature exists in a substantially folded form structurally more similar to a native state . The A-state has the far-UV circular dichroism spectra characteristic of the native protein, which indicates that it has a large degree of secondary structure . Upon heating, the A-state denatures with a sigmoidal change in far-UV ellipticity and an observable peak in a differential scanning calorimeter trace, indicating that it is thermodynamically distinct from the denatured state . Three different mutations in a residue normally buried in the protein's core stabilize or destabilize the A-state in the same way as they affect the denaturation of the native state . The A-state must, therefore, contain at least some tertiary packing of side chains . Unlike the native state, which shows cold denaturation at low temperatures, the A-state is most stable at temperatures below 0 degrees C.

Protein Sci, 1994 Jun, 3(6), 944 - 51
Thermodynamics of staphylococcal nuclease denaturation . I . The acid-denatured state; Carra JH et al.; Using high-sensitivity differential scanning calorimetry, we reexamined the thermodynamics of denaturation of staphylococcal nuclease . The denaturational changes in enthalpy and heat capacity were found to be functions of both temperature and pH . The denatured state of staphylococcal nuclease at pH 8.0 and high temperature has a heat capacity consistent with a fully unfolded protein completely exposed to solvent . At lower pH values, however, the heat capacity of the denatured state is lower, resulting in a lower delta Cp and delta H for the denaturation reaction . The acid-denatured protein can thus be distinguished from a completely unfolded protein by a defined difference in enthalpy and heat capacity . Comparison of circular dichroism spectra suggests that the low heat capacity of the acid-denatured protein does not result from residual helical secondary structure . The enthalpy and heat capacity changes of denaturation of a less stable mutant nuclease support the observed dependence of delta H on pH.

Mol Immunol, 1994 Jun, 31(9), 675 - 81
Bacterial superantigen signaling via HLA class II on human B lymphocytes; Mooney NA et al.; Staphylococcus enterotoxins and toxic shock syndrome toxin-1 (TSST-1) are members of the family of staphylococcal exoproteins (SE) which binds specifically to HLA class II molecules and certain V beta T cell receptor phenotypes . These bacterial products have been termed "superantigens" due to their capacity to stimulate a greater proportion of T lymphocytes than peptide antigens without a requirement for antigen processing . The SE stimulate monocytes to secrete IL-1 and TNF-alpha and affect B lymphocyte proliferation in response to anti-human IgM and Ig production by PBMC . The current study concerns the transmission of signals in human B lymphocytes following fixation of TSST-1 . Activation of both PLC and PKC are observed while intracellular calcium levels remain unchanged . Levels of HLA class II mRNA were increased suggesting that a pathway leading to activation was triggered . This study therefore identifies some of the second messengers involved after SE fixation on HLA class II molecules and suggests that the signals transmitted via class II antigens as well as those via the TCR may have a role in the physiological responses to bacterial superantigens.

Transfusion, 1994 Jun, 34(6), 536 - 8
Resolution of refractory, classic thrombotic thrombocytopenic purpura after staphylococcal protein A immunoadsorption; Drew MJ; BACKGROUND: Multiple therapeutic interventions are available for treatment of thrombotic thrombocytopenic purpura . Resolution of thrombotic thrombocytopenic purpura may require use of several of these interventions . CASE REPORT: A patient presenting with classic (non-cancer chemotherapy-associated) thrombotic thrombocytopenic purpura had an initial response to intensive, daily plasma exchange with fresh-frozen plasma and cryosupernatant . Multiple attempts over a period of 2 months to decrease the frequency of plasma exchange resulted in exacerbations of disease activity, indicated by increased schistocytosis, decreased hematocrit, increased serum lactate dehydrogenase, and decreased platelet counts . After a total of 39 plasma exchanges, the patient was begun on immunoadsorption therapy utilizing a staphylococcal protein A immunoadsorption treatment column . After six 2000-mL protein A immunoadsorption treatments, the patient's platelet count, lactate dehydrogenase, and peripheral smear normalized, and they have remained normal over nearly 4 months of follow-up . CONCLUSION: Treatment by protein A immunoadsorption may be of benefit in patients with classic thrombotic thrombocytopenic purpura who are not achieving a sustained remission with conventional plasma exchange therapy.

Biomaterials, 1994 Jun, 15(8), 628 - 34
Site-specific adhesion of Staphylococcus epidermidis (RP12) in Ti-Al-V metal systems; Gabriel BL et al.; Staphylococcus epidermidis (RP12) adhesion patterns were studied on the following titanium (Ti)-aluminium (Al)-vanadium (V) metal systems: (i) microfabricated samples consisting of Ti, Al and V islands deposited onto Ti or V substrata, (ii) pure Ti, Al and V metals, and (iii) medical grade Ti6Al4-V alloy . All of these surfaces were covered with their respective oxides formed upon exposure of the metals to air . Quantitative analysis of the number of cells bound per unit area indicates that S . epidermidis (RP12) exhibits greatest adhesion to pure V surfaces . When exposed to surfaces having controlled spatial variations in chemical composition on the 10 microns scale (microfabricated samples), the bacteria preferentially populate V islands versus Ti or Al substrata . In the case of the biphasic Ti6Al4V alloy, the bacteria tend to adhere to V-rich, mixed phase regions and phase boundaries . These findings demonstrate that enhanced and preferential adhesion of S . epidermidis (RP12) occurs on V surfaces in Ti-Al-V metal systems and suggest that bacterial interactions are influenced by surface oxide composition.

Immunology, 1994 Jun, 82(2), 207 - 10
Chlorpromazine specifically inhibits peripheral and brain TNF production, and up-regulates IL-10 production, in mice; Mengozzi M et al.; We have previously shown that chlorpromazine (CPZ) inhibits tumour necrosis factor (TNF) production and protects against endotoxic shock in mice . In this paper we investigated the effect of pretreatment with CPZ, 4 mg/kg i.p . 30 min before, compared with dexamethasone (DEX; 3 mg/kg) on the induction of other endotoxin (lipopolysaccharide; LPS)-induced cytokines in the serum of mice, i.e . interleukin-1 alpha (IL-1 alpha), IL-6 and IL-10, and TNF . We also studied the effect of CPZ on serum and spleen-associated TNF . Both DEX and CPZ inhibited TNF production, whereas induction of IL-1 and IL-6 was inhibited by DEX but not by CPZ . DEX did not affect IL-10, while CPZ potentiated its induction . CPZ also inhibited spleen-associated TNF induction in LPS-treated mice, suggesting an effect on the synthesis of TNF . CPZ inhibited TNF induction by Gram-positive bacteria (heat-killed Staphylococcus epidermidis) and by anti-CD3 monoclonal antibodies . Intraperitoneal administration of CPZ also inhibited the induction of brain-associated TNF induced by intra-cerebroventricular injection of LPS . Therefore, CPZ is a more specific inhibitor of TNF production than DEX; in particular, CPZ increased the induction of IL-10, which is a 'protective' cytokine known to inhibit LPS toxicity and TNF production . CPZ inhibited TNF production in vivo, irrespective of the TNF stimulus used to induce TNF . Finally, CPZ did not induce the 'rebound' effect of DEX that, when given 24 hr before LPS, potentiates TNF production, but it did inhibit TNF production after 24 hr.

J Exp Med, 1994 Jun 1, 179(6), 1885 - 93
Antigen-specific activation, tolerization, and reactivation of the interleukin 4 pathway in vivo; Rocken M et al.; The outcome of immune responses critically depends on the pattern of lymphokines secreted by CD4+ T cells . CD4+ T cells may differentiate into interleukin 2 (IL-2) and interferon gamma secreting T helper 1 (Th1)-like cells or IL-4/IL-5/IL-10 secreting Th2-like cells . However, the mechanisms that regulate production of IL-4 or other T cell lymphokines in vivo remain unknown . We use the superantigen, Staphylococcus enterotoxin A (SEA), as a model antigen to characterize the signals that regulate the production of IL-4 in vivo . Induction of IL-4 in normal CD4+ T cells required stimulation with both antigen and IL-4 . SEA-specific CD4+ T cells produced large amounts of IL-4 when restimulated within 10 d after in vivo priming . Repetitive application of both signals was required to prevent downregulation of IL-4 production . Although controversy exists regarding the susceptibility of Th2-like cells to tolerogenic signals, high doses of superantigen readily abolished the capacity to produce IL-4 in both naive T cells and in T cells already primed for IL-4 production . Infection with the nematode, Nippostrongylus brasiliensis, reversed the established T cell tolerance, whereas the signals which induced IL-4 production in normal T cells, antigen and IL-4, were not capable of reversing superantigen-specific tolerance in vivo . The major parameter that correlated with the capacity of parasitic infection to break tolerance was magnitude of the lymphoproliferation seen during the course of the infection . The capacity to activate or tolerize the IL-4 pathway in an antigen-specific fashion should prove useful in the design of antigen-specific therapies for autoimmune and allergic diseases.

Cell Immunol, 1994 Jun, 156(1), 220 - 9
Costimulation of superantigen-activated T lymphocytes by autologous dendritic cells is dependent on B7; Nestle FO et al.; Highly purified populations of dendritic cells (DCs) can be isolated from various tissues such as skin and blood . These sites are likely to encounter secreted toxins of bacteria such as superantigens . In vivo, DCs express the cell surface molecule B7, a counterreceptor for CD28 which provides costimulation to resting T cells . Highly purified preparations of DCs obtained from the epidermis and dermis of normal skin as well as blood were used to study the role of B7 in superantigen presentation to autologous T cells, as well as in alloantigen responses . We compared these results to those obtained with nondendritic antigen presenting cells (APCs) such as mononuclear cells derived from the Ficoll-Hypaque interface (PBMCs) . All DC populations strongly express B7, and in a purely autologous system staphylococcal enterotoxin B (SEB)-mediated T cell proliferation was inhibited by 55-85% using a soluble chimeric fusion protein (i.e., CTLA4Ig), a potent inhibitor of CD28:B7 interaction . In contrast, while T cells also proliferated vigorously when stimulated by SEB in the presence of autologous PBMC (which only weakly express B7), costimulation was not inhibited by CTLA4Ig . In allogeneic responses, DCs were also more potent stimulators compared to PBMC, but both types of APC:T cell reactions were almost completely inhibited by CTLA4Ig (> 90%) . For both SEB-mediated reactions and alloantigen reactions, the relative importance of LFA-1 and HLA-DR was similar between DCs and PBMCs . The data indicate that these DCs express B7, which can function in the SEB-driven response of autologous T cells, as well as in allogeneic T cell reactions . Overall, when comparing the relative costimulatory capabilities of different types of APCs, it appears the relatively low level of B7 expressed by PBMC functioned effectively in allogeneic reactions, whereas only the higher levels of B7 expressed by these DC populations functioned in SEB-mediated T cell responses.

J Mol Biol, 1994 May 27, 239(1), 154 - 7
Crystallization and preliminary X-ray analysis of an anti-staphylococcal nuclease-staphylococcal nuclease complex and of a second anti-staphylococcal nuclease antibody; Chang CY et al.; The Fab fragments of several monoclonal antibodies that bind Staphylococcal nuclease have been screened for crystallization conditions . Two of these, N10 and N25, have been crystallized in forms suitable for X-ray structural analysis . The anti-Staphylococcal nuclease antibody complex N10 Fab-nuclease crystallizes with symmetry consistent with space group C2 and cell parameters of a = 234.7 A; b = 43.5 A; c = 74.4 A; beta = 106.4 degrees . A second anti-Staphylococcal nuclease antibody, N25, although crystallized starting with the Fab-nuclease complex, apparently crystallizes as uncomplexed N25 Fab with symmetry consistent with space group P3(1)21 (or its enantiomorph P3(2)21) and cell parameters of a = b = 80.9 A; c = 138.4 A.

Biochemistry, 1994 May 24, 33(20), 6158 - 66
Modeling compact denatured states of proteins; Lattman EE et al.; We propose a model for the conformations of compact denatured states of globular proteins: that they are broad ensembles of chain backbone conformations that involve common localized hydrophobic clustering and helical contacts, depending on the amino acid sequence . We construct representative ensembles for chain lengths up to 136 monomers on three-dimensional cubic lattices using the "hydrophobic zippers" method (Fiebig & Dill, 1993) . We find that model conformations with radii of gyration about 20% larger than native conformations commonly have bimodal distributions of P(r), of the pairwise interatomic distances, r, and Kratky plots in agreement with recent small-angle X-ray scattering (Sosnick & Trewhella, 1992; Flanagan et al., 1992; Kataoka et al., 1993; Flanagan et al., 1993) experiments on three different proteins . We also find that the lattice model of the Shortle 1-136 fragment of staphylococcal nuclease does not appear capable of forming a single hydrophobic core by hydrophobic zippering, consistent with experiments.

Biochem Biophys Res Commun, 1994 May 16, 200(3), 1463 - 9
Characterization of modified staphylococcal protein A containing phospholipid monolayer on both solution and slide surfaces; Lu B et al.; A method is described for incorporation of water-soluble protein Staphylococcal protein A (SpA) into phospholipid monolayer using covalent protein-lipid conjugates in detergent solution . The amphiphilic conjugates have solubility properties very similar to intergral membrane proteins . When the conjugates are applied into dipalmitoyl-phosphatidic acid monolayer, a protein containing monolayer is formed on subphase surface . The monolayer is transferred to pre-coated substrate surface to form an artificial membrane . Results show that unmodified SpA is readily ejected from the monolayer when compressing the monolayer but modified SpA incorporates into the monolayer stably . The incorporation of the protein is proportional to the lipid coupling degree . When the protein is excessively modified, the IgG binding activity of the SpA in the membrane is lost significantly.

Cancer Res, 1994 May 15, 54(10), 2738 - 43
Antitumor x anti-CD3 bifunctional antibodies redirect T-cells activated in vivo with staphylococcal enterotoxin B to neutralize pulmonary metastases; Penna C et al.; T-cell antitumor activities are limited by the requirement of two specific major histocompatibility complex restricted steps, T-helper cell activation and cytotoxic T-lymphocyte targeting . The aim of this study was to investigate whether bypassing these major histocompatibility complex restricted steps using nonspecific in vivo activation of T-cells with staphylococcal enterotoxin-B (SE-B) and retargeting with antitumor x anti-CD3 bifunctional antibody (BFA) could provide an effective antitumor response . C3H/HeN mice were injected i.v . with CL-62 melanoma cells, which express the human melanoma antigen p97, and were treated 10 min later with SE-B and/or anti-CD3 (500A2) x anti-p97 (96.5) BFA . Pulmonary metastases were counted 14 days following injections . SE-B alone induced a dose-dependent activation of T-cells as measured by increased interleukin-2 receptor expression and enhanced proliferative responses . SE-B doses greater than 10 micrograms significantly reduced the number of pulmonary metastases versus control (P < 0.01) . Combined treatment with SE-B (50 micrograms) and BFA (5 to 50 micrograms) significantly decreased pulmonary metastases compared to treatment with SE-B alone (P < 0.05) . Similar reductions in metastases were observed with the F(ab')2 BFA but not with the unconjugated antitumor component of the BFA . Combined treatments with SE-B plus BFA accomplished better tumor neutralization than adoptively transferred in vitro activated splenocytes (4 x 10(7)) retargeted with BFA (5-100 micrograms; P < 0.05) . These studies demonstrate that T-cells can be activated in vivo by SE-B and retargeted with small doses of BFA . In this immunocompetent syngeneic pulmonary metastasis model, SE-B plus BFA provided a dramatic antitumor response.

Biochemistry, 1994 May 10, 33(18), 5510 - 7
A thermodynamic scale for the beta-sheet forming tendencies of the amino acids; Smith CK et al.; The results of a study to measure the beta-sheet forming propensities of the 20 naturally occurring amino acids are presented . The protein host for these studies is the 56 amino acid B1 domain of staphylococcal IgG binding protein G {Fahnestock, S.R., Alexander, P., Nagle, J., & Filpula, D . (1986) J . Bacteriol . 167, 870-880} . This protein was selected because it exhibits a reversible two-state thermal denaturation transition and its structure is known at high resolution . A suitable guest position in the protein was identified, and its neighboring environment was modified to minimize the potential for artifactual interactions . All 20 amino acids were individually substituted at the guest site, and their effect on the protein's thermal stability was determined . NMR was used to verify the structural integrity of several of the proteins with different amino acid substitutions at the guest site . The results of these studies provide a thermodynamic scale for the relative beta-sheet forming propensities of the amino acids that shows a clear correlation with the beta-sheet preferences derived from statistical surveys of proteins of known structure.

J Biol Chem, 1994 May 6, 269(18), 13107 - 14
hsp70-protein complexes . Complex stability and conformation of bound substrate protein; Palleros DR et al.; The presence of bound substrate protein increases the thermostability of hsp70 molecular chaperones (heat shock proteins of molecular mass 70 kDa) . Complexes between hsp70 and unfolded substrate proteins were isolated by size-exclusion high performance liquid chromatography . The isolated complexes were observed to dissociate at a significant rate even in the absence of ATP . The presence of ADP caused a substantial increase in the stability of the complex . Both ADP and inorganic phosphate were found to inhibit the ATP-induced dissociation of complex . ADP was also observed to increase both the rate of complex formation and its stability as a function of temperature, suggesting an important regulatory role for nucleotides during heat shock . Circular dichroism and fluorescence studies of the complex between DnaK and a thermally unstable mutant of staphylococcal nuclease indicate that the bound substrate protein is significantly unfolded . A model for hsp70 cycle of complex formation and dissociation, which accounts for the regulatory role of nucleotides, is proposed.

Biochemistry, 1994 May 3, 33(17), 5021 - 30
Engineering alternative beta-turn types in staphylococcal nuclease; Hynes TR et al.; We have refined the crystal structures of three point mutants of staphylococcal nuclease designed to favor alternative beta-turn types . Single amino acid substitutions were made in a type VIa beta-turn (residues 115-118; Tyr-Lys-Pro-Asn) containing a cis Lys 116-Pro 117 peptide bond . The mutations result in two new backbone conformations, a type I beta-turn for P117T and a type I' beta-turn for P117G and P117A . The P117G and P117A structures exhibit a dramatic difference in backbone conformation in the region of the mutation compared to the nuclease A structure such that the side chain of Lys 116 is reoriented to point into the nucleotide binding pocket . The distinct conformation observed for the nuclease A, P117G, and P117T beta-turn sequences agrees with correlations between beta-turn type and sequence identified from protein crystal structures . The P117A turn conformation provides an exception to these correlations . The results demonstrate that single residue changes can significantly alter backbone conformation, illustrating the process by which diversity in the structure of the protein surface can evolve on a conserved structural core, and suggest protein engineering applications in which the positioning as well as the identify of side chains can be modified to design new enzyme functions . Nuclease variants at the type VIa beta-turn site also allow the relationship between the amino acid sequence and beta-turn conformation to be examined in the context of an identical protein fold in crystallographic detail.

Plast Reconstr Surg, 1994 May, 93(6), 1277 - 81
Occult chin abscess associated with an autologous chin implant; Caputy GG et al.; The case of an occult abscess associated with an autologous nasal hump cartilage chin graft is presented . The graft was placed 30 years prior to secondary aesthetic surgery performed on December 22, 1992 . In the course of the removal of the autologous tissue, an abscess pocket was discovered which subsequently cultured positive for Staphylococcus epidermidis . The area was thoroughly debrided and irrigated, and a preoperatively custom-fabricated silicone elastomer implant was placed . A 2-week course of appropriate antibiotic therapy was administered following receipt of the culture and sensitivity data . The patient suffered no postoperative complications . The bacteriology of implant colonization and infection in the light of host defenses would predict such a course, although, if known preoperatively, the more prudent course of abstinence from alloplastic implantation would likely be dictated in the aesthetic surgery patient.

J Infect Dis, 1994 May, 169(5), 1042 - 9
Transposon mutants of Staphylococcus epidermidis deficient in elaboration of capsular polysaccharide/adhesin and slime are avirulent in a rabbit model of endocarditis; Shiro H et al.; Virulence comparisons were made in a rabbit model of endocarditis between wild-type and transposon mutants of Staphylococcus epidermidis deficient in elaboration of the capsular polysaccharide/adhesin (PS/A) and slime . The parental phenotype grew from 36 (61%) of 59 cultures of blood . The PS/A-negative phenotype grew in 1 (1%) of 98 cultures of blood (P < .001) . No animals infected with PS/A-negative strains developed endocarditis compared with 75% of rabbits infected with PS/A-positive strains . PS/A-producing strains survived better than did PS/A-deficient strains in intact, absorbed rabbit or human serum plus human leukocytes . There was also greater deposition of C3 onto the PS/A-deficient strains than with the PS/A-producing isogenic strains . PS/A functions as an antiphagocytic bacterial capsule preventing C3 deposition and phagocytosis; loss of this structure increases the strain's susceptibility to opsonic killing and decreases its virulence.

J Antimicrob Chemother, 1994 May, 33(5), 969 - 78
Prevention of bacterial colonization of intravenous catheters by antiseptic impregnation of polyurethane polymers; Bach A et al.; The use of intravascular catheters is associated with infectious complications . Using plastic materials with antibacterial activity may reduce catheter-related bacterial colonization . A novel intravascular catheter impregnated with the antiseptics silver-sulphadiazine and chlorhexidine was tested in an in-vivo model using implantation of catheters into the internal jugular veins of rats . The rate and magnitude of bacterial colonization in groups with implantation of silver-sulphadiazine and chlorhexidine bonded (SSC) and control (C) catheters were assessed 3 and 7 days after intravenous implantation, and local challenge at the exit site by 10(7) cfu Staphylococcus epidermidis ATCC 35984 . Significant reductions in the culture positivity of catheters were observed in the test compared with control groups . After 3 and 7 days, the magnitude of bacterial colonization of implanted catheter segments was significantly lower compared with control catheters (P < 0.01) . These findings indicate that antiseptic-bonded catheters substantially reduce the incidence and magnitude of catheter-related bacterial colonization, and may subsequently reduce catheter-related infection.

Immunol Lett, 1994 May, 40(2), 147 - 55
B- and T-cell responses in congenic mice to repeat sequences of the malaria antigen Pf332: effects of the number of repeats; Ahlborg N et al.; The Plasmodium falciparum antigen Pf332 comprises degenerated 11-amino-acid repeats with regularly spaced pairs of glutamic acid . Epitopes formed by such repeats are recognized by polyclonal and monoclonal antibodies that interfere with the life cycle of the blood stages of the malaria parasite . In order to study the immunogenicity of one such Pf332 repeat sequence (SVTEEIAEEDK), fusion proteins containing ZZ (two IgG binding domains of staphylococcal protein A) and dimers, trimers or tetramers of the malarial sequence were injected into mice . To analyse possible major histocompatibility complex class II restrictions of the immune response, mice of different H-2 haplotypes were used . A significant antibody response was elicited by administration of all the three fusion proteins in mice expressing the I-Ak allele (B10.BR, B10.A(2R) and B10.A(4R)) whereas B10 and C57BL/6 (H-2b) mice were low responders . In comparison, B10.D2 (H-2d) mice were low responders to fusion proteins with 2 or 3 repeats but responded well to the protein containing 4 repeats . Lymph node cells from B10.BR (H-2k) mice, primed in vivo with ZZ-fusion proteins containing either 2 or 4 repeats, proliferated in vitro in response to repeat sequences fused to ZZ or to an unrelated fusion partner, as well as to a synthetic peptide containing less than two repeats . In contrast, a response of lymph node cells from B10.D2 (H-2d) mice was only obtained when a fusion protein containing 4 repeats was used both for in vivo priming and in vitro restimulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Protein Eng, 1994 May, 7(5), 705 - 13
Gene synthesis and functional expression of a protein exhibiting monodomain IgG Fc binding; Trumble WR et al.; A gene encoding a bacterial IgG Fc binding domain was designed and synthesized . The synthetic DNA fragment was cloned 3' to an inducible trpE promoter such that expression of the gene in Escherichia coli produced abundant Fc binding protein fused to the first seven amino acids of the trpE protein . The recombinant protein contained a single Fc binding domain and demonstrated efficient binding to human IgG in Western blot analysis . This protein degraded rapidly following cell lysis in the absence of protease inhibitors, but could be effectively protected by the addition of protease inhibitor . After purification of the protein by IgG affinity chromatography, IgG Fc binding ability was retained for at least 24 h at either 23 or 37 degrees C and on heating for 15 min at temperatures up to 65 degrees C . No immunoprecipitation was observed in interactions between the monodomain Fc binding protein and IgG molecules . Unlike staphylococcal protein A, no detectable binding of the monodomain IgG Fc binding protein was observed to either IgM or IgA . Truncated proteins, expressed from a series of 3' deletions of the synthetic gene, were used to estimate the minimum portion of a monodomain Fc binding protein that retained Fc binding ability.

Eur J Clin Microbiol Infect Dis, 1994 May, 13(5), 417 - 20
Role of serological tests in the diagnosis of immune complex disease in infection of ventriculoatrial shunts for hydrocephalus; Bayston R et al.; Seven cases of ventriculoatrial shunt infection with immune complex disease are reported in order to demonstrate the usefulness of measurement of levels of specific antibody to Staphylococcus epidermidis in diagnosis . Blood and cerebrospinal fluid cultures gave misleading results, and there was initial doubt about the diagnosis in all seven cases . All showed grossly elevated titres of antibody to Staphylococcus epidermidis, with raised serum C-reactive protein levels and depressed complement levels . Measurement of antibody to Staphylococcus epidermidis enables the diagnosis of chronic ventriculoatrial shunt infection to be made rapidly and reliably.

APMIS, 1994 May, 102(5), 347 - 55
Long-term cyclosporin A nephrotoxicity in the rat . Evaluation of a morphological scoring system and of co-treatment with isradipine; Starklint H et al.; Cyclosporin A (CyA) nephrotoxicity was examined in Spraque-Dawley rats given CyA (12.5 (n = 45) or 25 (n = 45) mg/kg/day perorally for 16 weeks . Control rats (n = 45) received CyA vehicle . All rats were given either isradipine (ISRA) 1 or 5 mg/kg/day orally, or isradipine vehicle . Fifteen rats died from interstitial pneumonia caused by Staphylococcus xylosus . A predefined morphological CyA nephrotoxicity scoring system, based on semiquantitative scores for basophilic tubules and for interstitial fibrosis, performed on hematoxylin-eosin-stained tissue, yielded mean scores for basophilic tubules of 0.2 (range 0-1) in controls, 1.4 (range 0-3) in rats given CyA 12.5 mg/kg/day (p < 0.001), and 1.7 (range 0-3) in CyA 25 mg/kg/day rats (p < 0.001 as compared to controls) . Rats given CyA were grouped according to their score for interstitial fibrosis: 0.2 (range 0-1) in CyA 12.5 mg/kg/day and 1.7 (range 0-3) in CyA 25 mg/kg/day rats (p < 0.001) . When scores for basophilic tubules and interstitial fibrosis were pooled, none of the control rats had a score above 1, while 47% of the low-dose and 95% of the high-dose rats scored above 1 . Thus, this CyA nephrotoxicity scoring system provided an easy, efficacious, and reproducible identification of rats with morphological CyA nephrotoxicity, and may be of clinical interest in the assessment of CyA nephrotoxicity . Kidney tissue from rats not treated with isradipine was further investigated with periodic acid-Schiff (PAS) with and without diastase treatment, and with Sirius Red . The latter confirmed the increase in connective tissue following tubular atrophy in CyA-treated rats . PAS reaction disclosed diastase-resistant positivity in the glomerular arterioles (score in controls: mean 0.4, range 0-1, in CyA 12.5 mg/kg/day mean 2.2, range 1-3, p < 0.001 as compared to controls; in CyA 25 mg/kg/day mean 1.1, range 0-2, p < 0.005 as compared to controls, p < 0.05 as compared to CyA 12.5 mg/kg/day) . Furthermore, the straight part of the distal tubules of rats given the highest CyA dose contained considerable amounts of glycogen . The significance of this finding is unknown . Renal functional studies confirmed previous results since CyA decreased inulin clearance (Cin) from 1.2 +/- 0.5 to 0.8 +/- 0.3 ml/min/g kidney weight (kW) (p < 0.05), and lithium clearance (CLi) was reduced from 263 +/- 113 to 119 +/- 61 microliters/min/gKW (p < 0.001) . Isradipine had no significant effect.

FEMS Microbiol Rev, 1994 May, 14(1), 53 - 6
The use of fed batch cultivation for achieving high cell densities in the production of a recombinant protein in Escherichia coli; Strandberg L et al.; The production of the fusion protein staphylococcal protein A/E . coli beta-galactosidase in Escherichia coli was studied in batch and fed batch cultivations . Batch cultivation of a recombinant E . coli strain yielded a final cell dry weight of 16.4 g l-1 with a final intracellular product concentration of recombinant protein corresponding to approximately 38% of the cell dry weight . Fed batch cultivation made it possible to increase the final cell dry weight to 77.0 g l-1 . The intracellular product concentration (25%) was lower as compared to batch cultivation resulting in a total concentration of recombinant protein of 19.2 g l-1.

Immunology, 1994 May, 82(1), 117 - 25
Superantigens anergize cytokine production but not cytotoxicity in vivo; Sundstedt A et al.; We have investigated the effect of the superantigen staphylococcal enterotoxin A (SEA) on the balance between T-cell response and non-responsiveness in T-cell receptor (TcR) V beta 3 transgenic mice . One injection of SEA resulted in a substantial activation of TcR V beta 3+ cells, whereas T cells from mice injected with repeated doses of SEA displayed a diminished response to a subsequent in vitro challenge . The reduced responsiveness became apparent when SEA was injected multiple times with short intervals . Proliferation and cytokine production in anergized T cells were severely reduced when stimulated with SEA in vitro, whereas cytotoxic T lymphocyte (CTL) activity remained unaffected . The dichotomy between these functions was examined in vitro with respect to different T-cell subsets . The total number of CD4+ T cells was reduced in the hyporesponsive spleens, compatible with cell deletion . The remaining CD4+ TcR V beta 3+ T cells showed anergy of all tested functions and did not respond to exogenous interleukin-2 (IL-2) . In contrast, there was an expansion of CD8+ TcR V beta 3+ T cells with an intact cytotoxic activity . The in vitro proliferation and production of cytokines in the CD8+ compartment was impaired, but could be partially restored in the presence of exogenously added IL-2 . Analysis of the cytokine response to SEA in vivo showed that IL-2 and tumour necrosis factor (TNF) were mainly produced by CD4+ T cells, while interferon-gamma (IFN-gamma) was predominantly released by CD8+ T cells . Induction of anergy resulted in a reduction of IL-2 and TNF mRNA levels, frequencies of producing cells as well as serum protein content . In contrast, there was only a moderate influence on the IFN-gamma level in vivo . The results suggest that SEA-induced hyporesponsiveness involves CD4+ cell deletion and a failure to produce cytokines in the remaining CD4+ T-cell compartment, while IFN-gamma production and cytotoxicity in the CD8+ T-cell compartment stay relatively intact.

Rev Inst Med Trop Sao Paulo, 1994 May-Jun, 36(3), 283 - 5
"In vivo" leukocyte chemotaxis in experimental mice Schistosoma mansoni infection; Teixeira KM et al.; The "in vivo" chemotaxis was studied in C57Bl/10 mice 10, 30, 50 and 60 days after a Schistosoma mansoni infection in comparison to a control group (uninfected mice) . Staphylococcal protein A was injected into a connective tissue air pouch of control and experimental mice and the leukocyte chemotaxis was counted . A decrease in polymorphonuclear (PMN) leukocyte response was found in infected mice in comparison to the control group (p < 0.05) . The 10 day infected mice showed a decreased PMN leukocyte response respecting the control group (p < 0.05) and this finding became more evident 30 and 50 days post-infection . Although the PMN leukocyte response of 60 day infected mice increased in comparison to 50 day infected animals, it was still significantly lower the control response . The mononuclear leukocyte response was not significantly different between infected or uninfected mice.

Vestn Otorinolaringol, 1994 May-Jun, (3), 18 - 20
{Sorption methods in the treatment of nose and paranasal sinus diseases}; Tsetsarskii BM et al.; The responses to sorption treatment have been analysed for 373 patients suffering from staphylococcal, allergic, purulent, polypous-purulent rhinosinusitis . For enterosorption a coal spherical phi EH sorbent was used . Application sorption was performed with fabric sorbent AYT-M2, nonfabric sorbent AHM-pi, dust phi EH sorbent . Gelevin was tried for tampon-free postoperative management after dissection of the paranasal sinuses . Sorbents application advances the efficacy of nasal and paranasal disease treatment.

Pathol Biol (Paris), 1994 May, 42(5), 525 - 9
{Continuous infusion of vancomycin during the neonatal period}; Borderon JC et al.; Twenty-five infants with suspected or confirmed coagulase negative staphylococcal infection were studied . Continuous administration of vancomycin was used because it is usual with infusions prepared daily for catheterized patients, and because continuous infusions are well tolerated and achieve better penetration in tissues and CSF . Vancomycin acts as a time-dependent antibiotic . The aim was to obtain a level of 20-25 mg/l . in serum . Fifteen newborns term 27-35 weeks (m = 30.3) aged 7-30 days (m = 16.1) received 10 to 45 mg/kg/day of vancomycin and were monitored for 2 to 12 days . The sample for assay was taken in a peripheral vein, and the results were the same during the infusion or 15 minutes after its end . The daily dose of vancomycin necessary varied from 25 to 40 mg/kg for newborns with serum creatinine < 70 mmol/l and 10 to 30 mg/kg with serum creatinine > or = 90 mmol/l . Except for a newborn with multiorgan failure, serum creatinine rapidly decreased . Four newborns term 38-40 weeks (m = 39.5) aged 2-12 days (m = 8.3) received 20 to 45 mg/kg/day of vancomycin and were monitored for 2 to 12 days . The daily dose necessary varied from 30 to 40 mg/kg/day with important individual variations, and 20 mg/kg/day in a newborn with a high level of creatinine . In 6 infants aged 2-22 months receiving 22-45 mg/kg/day of vancomycin, a mean daily dosage of 40-45 mg/kg was adequate, with important individual variations.(ABSTRACT TRUNCATED AT 250 WORDS)

Rev Rhum Ed Fr, 1994 May, 61(5), 349 - 52
{Inferior glenohumeral subluxation: an indirect sign of sepsis of the shoulder}; Thomas E et al.; Drooping shoulder is a roentgenographic abnormality consisting in a discontinuity in the scapulo-humeral arch due to inferior subdislocation of the humeral head . Drooping shoulder is commonly seen in patients with injuries to the shoulder or neurological loss . Two cases of drooping shoulder due to staphylococcal arthritis (after local corticosteroid injections) are reported herein . The mechanism of the downward humeral displacement in septic arthritis is discussed . The other causes of drooping shoulder are reviewed . Precautions needed to avoid increasing the displacement during rehabilitation therapy are outlined.

Lett Appl Microbiol, 1994 May, 18(5), 281 - 4
Structure and putative origin of a plasmid from Staphylococcus hyicus that mediates chloramphenicol and streptomycin resistance; Schwarz S et al.; The structure and functional organization of Staphylococcus hyicus plasmid pSCGp3EB that mediates chloramphenicol and streptomycin resistance (CmrSmr) is described and compared with another CmrSmr plasmid, pSCS12, from Staphylococcus sciuri . Both plasmids appeared to be formed by co-integrate formation between plasmids that very closely resemble the chloramphenicol resistance (Cmr) plasmid pC221 and the streptomycin resistance (Smr) plasmid pS194 . In addition to the established recombination site B (RSB) in pC221 and pS194, another area suitable for recombination immediately downstream of the cat gene in pC221 and upstream of the str gene in pS194 has been identified . Co-integration at these sites would lead to the structures we have observed in the wild-type CmrSmr plasmids pSCGp3EB and pSCS12.

Vestn Khir Im I I Grek, 1994 May-Jun, 152(5-6), 12 - 6
{Endotoxinemia as a factor in the pathogenesis of thrombocytopathies in patients with acute infectious destruction of the lungs}; Shalaev SA et al.; Laboratory studies of the influence of the staphylococcal endotoxin on functional properties of thrombocytes and clinical observations of patients with a pronounced syndrome of endotoxicosis have shown an important role of endotoxinemia in pathogenesis of the arising thrombocytopathies . Possibilities of the medicamentous protection of blood plates from the damaging action of endotoxinemia are considered.

Scand J Immunol, 1994 May, 39(5), 419 - 25
Activation signal induces the expression of B cell-specific CD45R epitope (6B2) on murine T cells; Watanabe Y et al.; T lymphocytes express multiple forms of the leukocyte-common antigen CD45, transcribed by alternative usage of leukocyte-common antigen exon 4-6 . The various isoforms of CD45R expressed differentially on T cells are involved in different stages of development and activation . The monoclonal antibody (MoAb) RA3-6B2 is established as a B cell-type isoform (B220)-specific marker . However, it reacts with certain activated T cells although the relationship between 6B2 expression and T-cell activation is unclear . We have examined the 6B2 expression on activated T cells and found that concanavalin A, anti-CD3 antibody and staphylococcal enterotoxin B (SEB) induced 6B2 expression on T cells . The expression was found on both CD4+ and CD8+ T cells and also was induced by SEB in vivo predominantly on CD8+ T cells . The 6B2+ T cells are IL-2R+ and blasted cells according to flow cytometry analysis . Therefore, the 6B2+ T cells are supposed to be in an activated stage . Enzymatic analysis demonstrated that trypsin treatment decreased the 6B2 expression, whereas neuraminidase increased the intensity on activated T cells . Neither endo-D or endo-H have any effect on the expression and there are no differences, in the results of immunoprecipitation and RT-PCR analysis, between control T cells and activated T cells . Taken together, the 6B2 epitope is presumed to be the product of CD45R modification and is expressed on activated T cells . These results illustrate a novel classification of a T-cell subpopulation bearing a 6B2 epitope.

Eur J Immunol, 1994 May, 24(5), 1019 - 25
T cells of staphylococcal enterotoxin B-tolerized autoimmune MRL-lpr/lpr mice require co-stimulation through the B7-CD28/CTLA-4 pathway for activation and can be reanergized in vivo by stimulation of the T cell receptor in the absence of this co-stimulatory signal; Zhou T et al.; The CD28/CTLA-4 receptors on T cells interact with the B7 molecule on antigen-presenting cells (APC) to produce a co-stimulatory signal that determines the outcome of activation . The role of this co-stimulatory signal in T cell activation and loss of tolerance in autoimmune MRL-lpr/lpr mice has not been investigated previously . The present study examines the contribution of the CD28/CTLA-4 co-stimulatory pathway to the loss of T cell tolerance in V beta 8 transgenic MRL-lpr/lpr and (-)+/+ mice in which neonatal tolerance has been induced by the superantigen staphylococcal enterotoxin B (SEB) . An artificial APC transfected with the murine B7 gene, and a CTLA-4-Ig fusion protein were used to analyze the significance of the CD28/CTLA-4 pathway in vitro . The CTLA-4-Ig fusion protein was also used to inhibit the pathway in vivo . Our results demonstrate that CD28 and CTLA-4 mRNA was overexpressed in the lymph nodes of lpr/lpr mice (MRL, C57BL/6, C3H and AKR), but not in +/+ mice of the same background strain . Lymph node T cells and thymocytes from SEB neonatally tolerized MRL-lpr/lpr mice that had undergone tolerance loss, proliferated when cultured with SEB and B7+ fibroblasts in vitro, but did not proliferate when the SEB was presented in the context of B7- fibroblasts . This in vitro tolerance loss could be prevented by blocking of B7 signaling by CTLA-4-Ig . This loss of tolerance did not occur in lymph node T cells from thymectomized MRL-lpr/lpr mice . SEB challenge of tolerized MRL-lpr/lpr mice in vivo led to weight loss, increased serum cytokine levels and depletion of V beta 8+ T cells . These effects were blocked by blocking of the co-stimulatory pathway by treatment with the CTLA-4-Ig fusion protein prior to and during challenge with SEB . T cells from thymus and lymph nodes of these mice did not proliferate later in response to stimulation in vitro with SEB even in the presence of B7+ APC . Nonresponsiveness was not due to deletion of V beta 8+ CD28+ T cells, as the number of these cells was increased after treatment with SEB and the CTLA-4-Ig fusion protein.(ABSTRACT TRUNCATED AT 250 WORDS)

Biochem Biophys Res Commun, 1994 Apr 29, 200(2), 1059 - 65
Identification of an HIV-1 Nef peptide that binds to HLA class II antigens; Torres BA et al.; Overlapping peptides corresponding to the entire Nef (HIVBRU) sequence were tested for their relative abilities to block binding of staphylococcal enterotoxins (SEs) to Raji cells . An internal sequence, Nef(123-160), blocked binding of two highly homologous SEs, SEA and SEE, while it was less effective against SEB and SEC1 . Nef(123-160) bound directly class II DR antigens, as assessed by specific antibodies, and its binding was significantly inhibited by SEE, and also by SEA to a lesser degree . Purified Nef inhibited specific binding of SEA to Raji cells in a dose-dependent manner . Nef induced IL 2 production and proliferation by human mononuclear cells . Definitive expansion of specific V beta T cell populations was not observed, possibly due to lesser mitogenic activity of Nef relative to SEs . Thus, Nef may have mitogenic activity similar to that of superantigens.

J Biotechnol, 1994 Apr 15, 33(3), 293 - 306
Expression of human parathyroid hormone in mammalian cells, Escherichia coli and Saccharomyces cerevisiae; Rokkones E et al.; The entire human parathyroid hormone (hPTH) cDNA gene with its natural signal and pro-region is expressed in transfected mouse mammary tumor cells (C127I cells) and Chinese hamster lung cells (DON cells) under control of the murine metallothioneine-1 promoter in a vector in which replication functions are provided by the entire genome of bovine papilloma virus type I (BPV-1) . Authentic hPTH is efficiently produced by the non-endocrine cells and secreted to the growth medium without any abberant processing . Immunoblots from SDS-PAGE gels of concentrated growth medium reveal one band corresponding to intact, undegraded hPTH . Purification by reversed-phase HPLC results in a peptide with an amino acid content and N-terminal sequence identical to hPTH . For comparison, hPTH cDNA with deleted prepro-region is also expressed as secretory proteins in Escherichia coli and in Saccharomyces cerevisiae . In E . coli the vector construct is based on the staphylococcal protein A promoter employing protein A signal sequence . In S . cerevisiae a mating factor alpha expression system containing the factor alpha-signal sequence is employed . The results show that intact hPTH is secreted in addition to proteolytically cleaved fragments in both microorganisms . Thus, the signal sequences promote efficient secretion, and correct N-terminal processing of hPTH in both mammalian, bacterial and yeast cells . However, the folding characteristics of hPTH make it susceptible to internal proteolytical cleavage which appears to be species specific in yeast and E . coli.

Biochemistry, 1994 Apr 12, 33(14), 4207 - 11
Enhanced in vitro refolding of insulin-like growth factor I using a solubilizing fusion partner; Samuelsson E et al.; We have previously shown that human insulin-like growth factor I (IGF-I), fused to ZZ (two domains derived from staphylococcal protein A), can be refolded at relatively high concentrations, without the use of solubilizing agents {Samuelsson, E., Wadensten, H., Hartmanis, M., Moks, T., & Uhlen, M . (1991) Bio/Technology 9, 363-366} . Here we have studied this phenomenon in detail by characterizing the in vitro refolding of IGF-I, fused to one or two solubilizing Z domains and without a solubilizing fusion partner . The characterization included solubility studies of the reduced proteins and an evaluation of the aggregation occurring during the refolding process . The results suggest that the applied fusion protein strategy can be used to obtain a cis-acting chaperone-like effect during refolding in vitro . Fusion to one or two Z domains resulted in more than a 100-fold increase in the solubility of reduced IGF-I . In addition, the Z or ZZ fusion partners decrease multimerization of the IGF-I moieties during the renaturation . The fusion protein strategy may be an option to overcome the obstacles of insolubility and aggregation, frequently encountered when designing in vitro refolding processes.

Proteins, 1994 Apr, 18(4), 353 - 66
Monte Carlo simulations of protein folding . II . Application to protein A, ROP, and crambin; Kolinski A et al.; The hierarchy of lattice Monte Carlo models described in the accompanying paper (Kolinski, A., Skolnick, J . Monte Carlo simulations of protein folding . I . Lattice model and interaction scheme . Proteins 18:338-352, 1994) is applied to the simulation of protein folding and the prediction of 3-dimensional structure . Using sequence information alone, three proteins have been successfully folded: the B domain of staphylococcal protein A, a 120 residue, monomeric version of ROP dimer, and crambin . Starting from a random expanded conformation, the model proteins fold along relatively well-defined folding pathways . These involve a collection of early intermediates, which are followed by the final (and rate-determining) transition from compact intermediates closely resembling the molten globule state to the native-like state . The predicted structures are rather unique, with native-like packing of the side chains . The accuracy of the predicted native conformations is better than those obtained in previous folding simulations . The best (but by no means atypical) folds of protein A have a coordinate rms of 2.25 A from the native C alpha trace, and the best coordinate rms from crambin is 3.18 A . For ROP monomer, the lowest coordinate rms from equivalent C alpha s of ROP dimer is 3.65 A . Thus, for two simple helical proteins and a small alpha/beta protein, the ability to predict protein structure from sequence has been demonstrated.

J Dairy Sci, 1994 Apr, 77(4), 958 - 69
Coagulase-positive Staphylococcus intramammary infections in primiparous dairy cows; Roberson JR et al.; Objectives were to determine the prevalence of coagulase-positive staphylococcal IMI in primiparous cows at first parturition, to contrast the differences in coagulase-positive staphylococcal IMI in primiparous cows at parturition in herds with high and low prevalences of coagulase-positive staphylococcal IMI in the lactating herd, and to determine the percentage of primiparous cows having persistent coagulase-positive staphylococcal IMI . Milk samples were collected aseptically from cows at the start and end of the study, at dry-off, and at parturition . Herds (n = 18) were split evenly into two categories: high (> 10%) or low (< 5%) prevalence of coagulase-positive staphylococcal IMI . At the start, the mean prevalence of coagulase-positive staphylococcal IMI in high prevalence herds was 30%, ranging from 13 to 65%, and in low prevalence herds was 2%, ranging from 0 to 5% . Overall the prevalence of coagulase-positive staphylococcal IMI in primiparous cows at parturition was 8.1% (67 of 828), ranging from 0 to 27% . Although primiparous cows from high prevalence herds had a higher prevalence of coagulase-positive staphylococcal IMI (9.2%; 40 of 436) at parturition than did primiparous cows from low herds (6.9%; 27 of 392), the difference was not significant . Of primiparous cows with coagulase-positive staphylococcal IMI at parturition, 43% had coagulase-positive staphylococcal IMI at least 2 mo after parturition . Primiparous cows with coagulase-positive staphylococcal IMI at parturition may represent significant reservoirs of infection to uninfected herdmates.

FEMS Microbiol Lett, 1994 Apr 1, 117(2), 231 - 5
Plasmid-encoded catalase of Staphylococcus simulans biovar staphylolyticus; Fondren LM et al.; Staphylococcus simulans biovar staphylolyticus contains five plasmids designated pACK1 through pACK5 . Non-denaturing electrophoretic analysis of an extract prepared from wild-type cells revealed three bands of catalase activity, whereas an extract of cells cured of pACK1 produced only two catalase bands . Cloning and Southern hybridization analysis showed that there is a catalase structural gene on pACK1 . The plasmid-specified catalase was the major activity produced under both aerobic and anaerobic conditions of growth.

Eur J Surg Oncol, 1994 Apr, 20(2), 122 - 9
Catheter-related complications in 52 patients treated with continuous infusion of low dose recombinant interleukin-2 via an implanted central venous catheter; Vlasveld LT et al.; In this study we evaluated the catheter-related complications in 52 patients with advanced melanoma, renal cell cancer or non-Hodgkin's lymphoma treated with continuous infusion of low-dose recombinant interleukin-2 by central venous access (CVA) of the port-a-cath type . We noted a high incidence (55.5%) of catheter infection, defined as positive blood cultures drawn from the CVA in symptomatic or asymptomatic patients . Six infections were noted before rIL-2 treatment was started . Twelve of the 30 documented infections were symptomatic (fever and/or chills), with only four documented bacteraemias . The most frequently cultured microorganism was Staphylococcus epidermidis (73%) . Treatment initially consisted of systemic antibiotics via the CVA, but as experience increased, the mostly asymptomatic CVA infections were not treated . In 30% of the documented CVA infections a thrombus at the tip of the catheter was found by radiological contrast examination . Local thrombosis can be effectively treated with constant infusion of low dose streptokinase via the CVA.

Nippon Yakurigaku Zasshi, 1994 Apr, 103(4), 175 - 86
{A mechanism underlying histamine-induced contraction in isolated rabbit lingual artery}; Suyama N et al.; To understand the contractile response to histamine, drug-induced tension developments were measured in intact and staphylococcal alpha-toxin-treated permeabilized smooth muscle preparations of rabbit lingual artery . Histamine produced an endothelium-independent contraction; the observed effect was antagonized by diphenhydramine and was attenuated by nifedipine . Histamine produced only transient contraction in the Ca(2+)-free bathing media . In permeabilized preparations, histamine-induced contraction was abolished by ryanodine (in the presence of caffeine) or prior repeated application of caffeine; however, contraction produced by IP3 was still observed under the same experimental condition as that of the histamine study . Histamine- or caffeine-induced contraction was abolished in the permeabilized preparations by prior repeated application of IP3; caffeine or IP3 produced contraction after repeated application of histamine . Ryanodine in the presence of histamine was ineffective on caffeine-induced contraction, suggesting that histamine by itself may not have the ability to act on caffeine-sensitive Ca2+ release channels . Neomycin and H-7 completely abolished the histamine-induced contraction . Hence, it is suggested that histamine can contract the lingual artery via H1-receptor-coupled phospholipase C activation, and the contraction consists of a phasic response evoked by Ca2+ release from the IP3- and caffeine-sensitive Ca2+ storage site and the tonic response generated by the voltage-dependent influx of Ca2+ . It is also postulated that IP3 produced by histamine may be able to produce Ca2+ release from only a part of the intracellular Ca2+ stores in the smooth muscle of rabbit lingual artery.

Cancer Immunol Immunother, 1994 Apr, 38(4), 265 - 71
Combined activation of murine lymphocytes with staphylococcal enterotoxin and interleukin-2 results in additive cytotoxic activity; Belfrage H et al.; This report demonstrates that in vitro activation of murine spleen cells with interleukin-2 (IL-2) or the bacterial superantigen staphylococcal enterotoxin A (SEA) results in different patterns of activation and function of cytotoxic cells . Lymphokine-activated killer activity and antibody-dependent cellular cytotoxicity (ADCC) are mainly mediated by IL-2 activated natural killer (NK) cells . SEA is the most powerful T cell mitogen known so far and retargets cytotoxic T lymphocytes (CTL) to tumors expressing major histocompatibility complex (MHC) class II in staphylococcal-enterotoxin-dependent cellular cytotoxicity (SDCC) . Culture of mouse spleen cells with SEA led to expansion and activation of T cells, which demonstrated strong SDCC activity and some NK-like cytotoxicity after 5 days in culture . Cell sorting revealed that both CD8+ and CD4+ T cells mediated SDCC but the former were more effective . Phenotypic analysis showed that SEA preferentially stimulated and expanded T cells expressing T cell receptor V beta 11, in particular CD8+ T cells . Combined activation with SEA and IL-2 resulted in simultaneous induction of T and NK cell cytotoxicity . Moreover, IL-2 had additive effects on SEA-induced SDCC . Combined treatment with SEA and IL-2 might therefore be an approach to induce maximal cytotoxicity against tumors and to recruit both T and NK cells in tumor therapy.

J Vasc Surg, 1994 Apr, 19(4), 739 - 41
Treatment of vascular graft infection by in situ replacement with a rifampin-bonded gelatin-sealed Dacron graft; Goeau-Brissonniere O et al.; PURPOSE: The purpose of this study was to treat an established prosthetic vascular graft infection by in situ replacement with a rifampin-bonded gelatin-sealed Dacron graft in an animal model . METHODS: The infrarenal aorta of 18 dogs was replaced with a gelatin-sealed graft contaminated in vitro by soaking it in a solution with Staphylococcus epidermidis . One week later, animals were randomized into three groups . In group I (control, (n = 6), the dogs did not undergo repeat operations . The dogs in groups II and III underwent repeat operation . In these animals the infected grafts were removed for bacteriologic analysis and replaced in situ with one of two types of grafts: group II (n = 6) received an untreated, gelatin-sealed graft; group III (n = 6) received a rifampin-bonded, gelatin-sealed graft . Antibiotic bonding was obtained by soaking grafts for 15 minutes in a 60 mg/ml saline solution of rifampin at 37 degrees C . All 18 dogs received no systemic adjunct antibiotic therapy . Control grafts and replacement grafts were removed 4 weeks after the initial implantation for bacteriologic analysis . When harvested, all the grafts were cut into two fragments, and quantitative bacterial cultures were obtained from all the fragments . Results were expressed as colony-forming units (CFU)/cm2 of graft material . RESULTS: All 18 initially implanted grafts and all the untreated replacement grafts were grossly infected at the time of removal, whereas all the rifampin-bonded replacement grafts had normal incorporation . None of the rifampin-bonded grafts grew bacteria, whereas all the initially implanted and all the untreated replacement grafts were infected (p < 0.01) . Bacterial counts from the infected fragments were similar in control grafts (2.6 +/- 1.9 x 10(6) CFU/cm2), in initially implanted grafts of groups II (9 +/- 1.1 x 10(5) CFU/cm2) and III (1.3 +/- 1.5 x 10(6) CFU/cm2), and in untreated replacement grafts of group II (1.7 +/- 2.5 x 10(6) CFU/cm2) . Blood culture results and culture results of liver, spleen, kidney, and lung specimens at the time of sacrifice were negative . CONCLUSION: This study demonstrates that rifampin-bonded gelatin-sealed Dacron grafts are resistant to infection when used for in situ replacement of an infected graft in the dog.

Chest, 1994 Apr, 105(4), 1147 - 50
Significance of iatrogenic pneumothoraces; Despars JA et al.; The purpose of this study was to review the cases of iatrogenic pneumothorax that occurred between October 1983 and December 1988 at the Veterans Administration Medical Center, Long Beach, Calif, to determine the treatment and complications . During this time period, 106 patients were identified with iatrogenic pneumothorax, and the charts of 98 were available for review . There were 90 cases of spontaneous pneumothorax at this institution during the same time period . The most common cause of iatrogenic pneumothorax was transthoracic needle aspiration (35), followed by thoracentesis (30), subclavian venipuncture (23), and positive pressure ventilation (7) . In 11 cases, the cause was due to miscellaneous triggers . The majority of the patients (65 of 98) were treated with chest tubes . The chest tubes were in place 4.7 +/- 3.9 days . Nine of the patients required a second chest tube . Aspiration of the pneumothorax only was attempted in five patients, and all patients subsequently received a chest tube . Two patients died from iatrogenic pneumothorax . One patient receiving positive pressure ventilation developed an unrecognized tension pneumothorax . The other patient developed a pneumothorax after thoracentesis and was treated with a chest tube, which led to a staphylococcal empyema and death . From this study, we conclude that the incidence of iatrogenic pneumothorax exceeds that of spontaneous pneumothorax and that there is substantial morbidity and some mortality from iatrogenic pneumothorax.

Br J Anaesth, 1994 Apr, 72(4), 415 - 7
Do infusions of midazolam and propofol pose an infection risk to critically ill patients?
Farrington M, McGinnes J, Matthews I, Park GR.
In order to investigate bacterial contamination of i.v . anaesthetic agents, given by infusion to critically ill patients, we have cultured residual infusion fluid from infusion syringes, 50 containing midazolam and 50 propofol . The infusions had been prepared with routine aseptic precautions and had been running for between 0.75 and 21.25 h . Only scanty growths of Staphylococcus epidermidis were isolated from seven syringes (four midazolam and three propofol) . Small volume samples were more likely to produce bacterial growth than large volume specimens . Midazolam infusions made up in 5% glucose were more likely to be contaminated than those made up in 0.9% saline . Antibacterial activity was detected in 18 midazolam and one propofol filtrate . Midazolam infusions inhibited the growth of all seven of the S . epidermidis isolates, whereas propofol supported similar rates of multiplication to that obtained with control broth medium . The results of this study imply that contamination of the infusions probably occurred after they were disconnected from the patient . Despite the ability of propofol to support microbial multiplication, we have no evidence to suggest that this is clinically significant when infusions are prepared with conventional aseptic precautions.

Neonatal Netw, 1994 Apr, 13(3), 33 - 9
Vancomycin dosing in neonatal patients: the controversy continues; Wandstrat TL et al.; During the past decade, an increasing incidence of staphylococcus organisms resistant to penicillinase-resistant penicillins has necessitated the use of vancomycin . This increased utilization has revitalized research concerning efficacious vancomycin dosing regimens for premature neonates, infants, and children . Vancomycin dosing in neonates is variable because this patient population has decreased renal clearance and a larger volume of distribution than infants, children, or adults . The observation of the variability in vancomycin clearance and volume of distribution in infants with the same postconceptional age (PCA) but different gestational age (GA) suggests that the rates of maturation both extrauterine and intrauterine for disposition mechanisms of vancomycin are similar when PCAs are equal . Conditions such as patent ductus arteriosus, respiratory distress syndrome, sepsis, and asphyxia may further complicate the renal maturation process . Few investigations suggest vancomycin dosing regimens . Most of these studies propose dosing regimens based on retrospective analysis of vancomycin pharmacokinetics obtained from regimens based on physician discretion . To ensure efficacious and rational vancomycin dosing for premature neonates and infants, regimens should consider PCA as well as body weight.

Scand J Immunol, 1994 Apr, 39(4), 387 - 94
Interactions between staphylococcal superantigens and human T-cell clones are predominantly but not exclusively governed by their T-cell receptor V beta usage; Lundin KE et al.; Staphylococcal exotoxins (SE) are potent mitogens for human and murine T cells . Extensive studies in mice have demonstrated strict correlations between T-cell responses to individual SE and TCR V beta expression . Studies examining the TCR V beta expression of SE-activated human peripheral blood T cells also suggest close correlations, whereas the data reported using human T-cell clones (TCC) are conflicting . We have determined the cDNA TCRB sequences of 52 different human TCC, expressing 35 different T-cell receptor V beta (TCRBV)-encoded sequences . The TCC were tested in proliferative assays using nine different SE . Most of these TCC express V beta s which have not been tested previously in studies examining interaction between TCC and SE . The SE stimulated a variable fraction (1/48-31/52) of the TCC . The ability of a given SE to stimulate TCC in many cases correlated with V beta expression, but several exceptions were found . With one possible exception, comparisons between deduced amino-acid sequences within the 'fourth hypervariable region' of the TCR beta chain and SE responsiveness did not reveal potential SE binding motifs . We conclude that the reactivity of T cells towards SE is governed mainly by their TCR V beta expression . However, the authors' results also suggest that the interaction between SE and human T cells may involve elements unidentified as yet which are in addition to the beta chain of the TCR.

J Exp Med, 1994 Apr 1, 179(4), 1087 - 97
The V beta complementarity determining region 1 of a major histocompatibility complex (MHC) class I-restricted T cell receptor is involved in the recognition of peptide/MHC I and superantigen/MHC II complex; Bellio M et al.; We investigated the role of the complementarity determining region 1 (CDR1) of T cell receptor (TCR) beta chain both in antigen/major histocompatibility complex I (MHC I) and in superantigen (SAg)/MHC II complex recognition . Residues 26 to 31 of the V beta 10 domain of a TCR derived from an H-2Kd-restricted cytotoxic clone were individually changed to alanine, using site-directed mutagenesis, and the mutated TCR beta chains were transfected along with the wild-type TCR alpha chain into a TCR alpha-beta-T hydridoma . These mutations affected antigen/H-2Kd complex recognition, although to a different extent, as estimated by interleukin 2 production . Certain mutations also affected differently the recognition of two Staphylococcal toxins, exfoliative toxin and Staphylococcal enterotoxin C2, presented by HLA-DR1 . Whereas mutation of residues D30 or T31 affect the recognition of both toxins, residues T26, L27, and H29 are critical for the recognition of only one of the SAgs . These observations demonstrate the participation of the CDR1 region in the recognition of peptide/MHC class I as well as SAg/MHC II complexes.

J Leukoc Biol, 1994 Apr, 55(4), 523 - 9
Differential RNA regulation by staphylococcal enterotoxins A and B in murine macrophages; Chapes SK et al.; Staphylococcal enterotoxin A (SEA) is significantly better than enterotoxin B (SEB) in activating tumor necrosis factor (TNF) secretion by B6MP102 cells . Both toxins bound to B6MP102 cells; however, SEB competed less effectively with SEA than SEA competed with SEB . This suggested that receptors unique to SEA were present on B6MP102 cells . Signal transduction occurred in response to both toxins . Within 30 s after addition, SEA and SEB significantly increased the F-actin concentration in B6MP102 cells . However, only SEA induced increased TNF mRNA levels . B6MP102 cells incubated with interferon-gamma and SEB secreted TNF . However, enhanced mRNA expression was delayed and the concentration of TNF secreted was less than that of B6MP102 cells stimulated with SEA . Although these data suggest that receptors unique to SEA are present on B6MP102 cells, they also indicate that staphylococcal enterotoxins differentially regulate TNF at the RNA level, perhaps because of differences in binding to the plasma membrane.

J Immunol, 1994 Apr 1, 152(7), 3578 - 85
Involvement of the cystine transport system xc- in the macrophage-induced glutamate-dependent cytotoxicity to neurons; Piani D et al.; Macrophages have been found to release glutamate and thereby induce neuronal cell death by excitotoxicity, a mechanism that seems to be operative in various neurologic diseases . In this study, it is shown that the presence of both cystine and glutamine in the culture medium is indispensable for brain macrophages to release glutamate and to cause neuronal cell death . Furthermore, release of glutamate requires protein synthesis since cycloheximide prevented accumulation of the neurotoxic molecule in supernatants of microglial cell cultures . Aminoadipate, which was shown to inhibit the uptake of cystine by system xc- in fibroblasts, efficiently reduced the release of glutamate . The requirement of glutamine and cystine for the release of glutamate by microglial cells as well as the inhibitory effect observed with aminoadipate shows the transport system xc- to be essential for the release of the excitotoxin glutamate by microglial cells . Phagocytosis of zymosan particles and stimulation with different bacterial components, such as LPS, protein A, tuberculin, and Staphylococcus enterotoxin A increased glutamate release two- to threefold above basal values . In addition, the effect of bacterial components was mimicked by TNF-alpha, but not by IL-1 and IL-6 . Cytokines known to deactivate macrophages, such as TGF-beta, IL-4, and IL-10, did not affect the transport system xc- in microglial cells . However, dexamethasone suppressed the glutamate release up to 50%.

J Immunol, 1994 Apr 1, 152(7), 3522 - 9
Cooperation between staphylococcal enterotoxin B and low dose melphalan in the cure of mice bearing a large MOPC-315 tumor and extensive metastases; Rubin M et al.; We have recently demonstrated the importance of V beta 8+/CD8+ cells for the curative effectiveness of a suboptimal low dose (0.75 mg/kg) of melphalan (L-phenylalanine mustard; L-PAM) for mice bearing a large s.c . MOPC-315 tumor and extensive metastases . Here we show that staphylococcal enterotoxin B (SEB), which is known to selectively stimulate T cells expressing members of the TCR-V beta 8 gene family, substantially improved the curative effectiveness of the suboptimal dose of L-PAM for mice bearing a large MOPC-315 tumor . Moreover, treatment of mice with mAb F23.1 (anti-V beta 8) abrogated the in vivo therapeutic effect of SEB for low dose L-PAM-treated MOPC-315 tumor bearers (L-PAM TuB mice) . Analysis of the effect of SEB on the tumor-infiltrating lymphocytes (TILs) demonstrated that the SEB-mediated therapeutic effect was associated with a significant increase in: 1) the percentage of V beta 8+ cells among the CD8+ T cells that accumulated in the s.c . tumor nodules, 2) the total number of V beta 8+/CD8+ cells present per tumor on day 4 after low dose L-PAM therapy, and 3) the ability of the TILs to lyse MOPC-315 tumor cells in vitro in a short term assay . Furthermore, treatment of mice with mAb F23.1 abolished the ability of SEB to render the TIL population of L-PAM TuB mice more cytotoxic in vitro for MOPC-315 tumor cells . Thus, the SEB-mediated improvement in the therapeutic outcome of low dose L-PAM therapy for mice bearing a large MOPC-315 tumor may be due in part to SEB-mediated increase in the contribution of the V beta 8+ T cells to tumor eradication through enhancement in the magnitude of the anti-MOPC-315 lytic activity exhibited by the TIL population.

J Immunol, 1994 Apr 1, 152(7), 3324 - 32
Gene transfer studies of T cell receptor-gamma delta recognition . Specificity for staphylococcal enterotoxin A is conveyed by V gamma 9 alone; Loh EY et al.; gamma delta T cells bearing the V gamma 9 gene segment have been shown to recognize staphylococcal enterotoxin A (SEA) and a range of other Ags including mycobacterial Ags . We have established an experimental system to analyze the recognition properties of human TCR-gamma delta on a molecular level by transferring the receptor from its original T cell into a Jurkat T cell host that does not express an endogenous TCR . Three groups of transfectants that express the same delta-chain, V delta 1, but different gamma-chains (V gamma 9-J2-C gamma 2, V gamma 3-J2-C gamma 2, and V gamma 9-JP-C gamma 1) together with the endogenous CD3 were obtained . The transfectant T cells each expressing different gamma delta receptors all produced IL-2 after stimulation with plastic bound anti-CD3 Ab, but only those expressing V gamma 9 responded to stimulation with SEA in the presence of an autologous lymphoblastoid B cell line . In addition, transfectants that expressed V delta 2 combined with V gamma 9 could also respond to SEA . These results indicate that the V gamma 9 portion of the receptor, independent of the J region and C region or the delta-chain, is responsible for recognizing SEA.

Cell Immunol, 1994 Apr 1, 154(1), 440 - 52
Early activation and cell trafficking induced by staphylococcal enterotoxin B: effects of high- versus low-dose challenge on induction of anergy; Bell SJ et al.; The in vivo challenge with exogenous superantigen, staphylococcal enterotoxin B (SEB), selectively induces vigorous polyclonal proliferation of T cells bearing the V beta 8+ TcR domain, whereafter the responsive cells become anergic . We used kinetic analyses to compare the effects of primary (1 degree) and secondary (2 degrees) challenge with a high and a low dose of SEB and the conventional antigen, sperm whale myoglobin, to determine the differential effects of in vivo challenge with a superantigen compared with a conventional antigen . We demonstrate that SEB induces very early activation-associated intralymphatic proliferation and trafficking of more T cells than can be accounted for by V beta 8+ T cells alone . Overall, this study indicates that challenge with SEB causes an apparent loss of CD4+ T-helper cell function and provides an essential foundation for the understanding of the mechanisms of peripheral tolerance induction and T cell "memory."

J Chemother, 1994 Apr, 6(2), 107 - 10
Differential inhibition by clindamycin on slime formation, adherence to teflon catheters and hemolysin production by Staphylococcus epidermidis; Shibl AM et al.; Slime formation was detected in Staphylococcus epidermidis strains isolated from either infected patients or healthy individuals . Cells of S . epidermidis, that either formed slime or not, adhered to teflon catheters . There was no correlation between adherence of bacteria to teflon catheters and slime formation . Clindamycin at subinhibitory concentration significantly inhibited slime formation without inhibiting bacterial growth . Adherence of S . epidermidis to teflon catheters was affected by the presence of clindamycin whether slime was produced or not . Clindamycin at subinhibitory concentrations markedly inhibited hemolysin production by S . epidermidis without appreciably altering the cell density, and cells grown in the presence of the drug showed very low hemolytic activity upon disruption . These results suggest that clindamycin at low concentration alters S . epidermidis virulence properties, apart from inhibiting growth.

Pol Arch Med Wewn, 1994 Apr, 91(4), 288 - 93
{Diagnostic difficulties with a tumor in deep tissues of the buttock}; Urbaniak-Kujda D et al.; Diagnostic difficulties in a case of a large tumor localized in the deep tissue of buttock in a 29-year old woman is described . The patient was very severely ill presenting with neurological symptoms and immobilization of the left coxal joint, what strongly suggested a neoplasm and this was supported by the CT scan . USG examination allowed diagnosis of a deep hematogenic staphylococcal abscess of the buttock, probably due to extensive oral sepsis and staphylococcal septicaemia . Systemic antibiotic therapy as well as evacuation of pus caused a complete recovery of the patient.

Biol Chem Hoppe Seyler, 1994 Apr, 375(4), 271 - 80
Production of the immunoglobulin variable domain REIv via a fusion protein synthesized and secreted by Staphylococcus carnosus; Pschorr J et al.; REIv--the variable domain of an immunoglobulin x light chain--was produced by heterologous gene expression in a Gram-positive bacterium, purified to homogeneity and characterized . A host/vector combination based on secretion of Staphylococcus hyicus lipase by Staphylococcus carnosus was exploited . A gene encoding a fusion protein, composed of an aminoterminal portion of the pre-pro-peptide of S . hyicus lipase, a hexahistidine affinity tag, followed by the recognition sequence of IgA protease and REIv was constructed . Expression of the fusion gene in S . carnosus causes selective secretion and accumulation of a soluble fusion protein in the culture medium (5-10 mg/l), which can be purified from the supernatant by immobilized metal ion affinity chromatography (IMAC) . REIv is released from the fusion protein with an additional threonine and proline residue at the aminoterminus (REIvTP) by site-specific cleavage with IgA protease and can be separated from the hexahistidine-tagged fusion partner and the protease by a second passage through an IMAC gel matrix . Like authentic REIv, the isolated protein (> 1 mg/l culture medium) migrates as a dimer in gel filtration chromatography and undergoes cooperative, reversible unfolding in urea . The isolated immunoglobulin REIvTP and authentic REIv have indistinguishable free energies of unfolding (approx . 26 kJ/mol, 6.3 kcal/mol).

J Antimicrob Chemother, 1994 Apr, 33(4), 747 - 55
In-vitro synergy between aminoglycosides deployed against Staphylococcus spp . harbouring a 6'-aminoglycoside acetyltransferase, 2"-aminoglycoside phosphotransferase enzyme; Culebras E et al.; In-vitro synergistic effects between commercial aminoglycosides are described for gentamicin resistant Staphylococcus strains harbouring 6'-aminoglycoside acetyltransferase activity . Seventy eight strains were studied using the double-disc test and synergy was observed with combinations in which at least one of the components has a garosamine-like 6'-aminosugar . These results were confirmed in chequerboard titrations (sigma FIC < or = 0.5) carried out on Staphylococcus epidermidis strains RYC13036 and RYC4904 . Additionally, a clear reduction in tobramycin acetylating activity was observed in the presence of gentamicin or netilmicin in crude extracts of these strains . These experiments suggest that the observed synergy is due to inhibition of the aminoglycoside-modifying enzyme by aminoglycosides with a garosamine like 6'-aminosugar component.

Anal Biochem, 1994 Apr, 218(1), 26 - 33
Reversible immunoprecipitation using histidine- or glutathione S-transferase-tagged staphylococcal protein A; Poon RY et al.; We have constructed, expressed, and purified hexahistidine- and glutathione S-transferase (GST)-tagged Staphylococcal protein A . The histidine-tagged protein A bound efficiently to iminodiacetic acid (IDA)-Sepharose loaded with Zn2+, and the GST-protein A was efficiently retained by glutathione-Sepharose . Both recombinant forms of protein A can be used in the normal way to harvest immune complexes with IgG . Both forms of protein A can be released from the Sepharose matrix by mild procedures . The his6-protein A:antibody:antigen complexes can be released from the matrix with EDTA, and immunoprecipitates bound to GST-protein A can be released either by elution with glutathione or by digestion with thrombin . We tested this method with immunoprecipitates of the p40MO15 protein kinase, and found that they retained their ability to phosphorylate p33cdk2 after elution from the affinity matrices.

CLAO J, 1994 Apr, 20(2), 128 - 30
Clinical characteristics of corneal foreign bodies and their associated culture results; DeBroff BM et al.; A prospective study was performed to investigate the clinical characteristics of corneal foreign bodies and to determine if these characteristics were related to the culture results of the foreign body . The clinical characteristics included the type of foreign body, the mechanism of injury, the time present in the cornea, and the presence of a rust ring or an infiltrate . Sixty-three foreign bodies were removed from corneas and cultured for bacteria, and 14.3% cultured positive for bacteria . The major pathogen was coagulase-negative Staphylococcus . The majority of foreign bodies were metallic, and mechanisms of injury were similar for both culture-positive and culture-negative groups . There was no significant correlation between the mean time present in the cornea and the culture results . The presence of a rust ring or an infiltrate was not found to be significant when predicting culture results . No clinical features readily distinguished culture-negative foreign bodies from culture-positive foreign bodies.

Protein Eng, 1994 Apr, 7(4), 579 - 83
Inactivation of Staphylococcus hyicus lipase by hexadecylsulfonyl fluoride: evidence for an active site serine; Leuveling Tjeenk M et al.; The Staphylococcus hyicus lipase is an acyl hydrolase with broad substrate specificity including neutral glycerides and phospholipids . To obtain further insight into the mechanism of action of this enzyme, we tested several sulfonyl fluorides as active site-directed inhibitors . The enzyme is resistant to the well-known serine protease/esterase inhibitor phenylmethanesulfonyl fluoride (PMSF), but is rapidly inactivated by hexadecylsulfonyl fluoride . The kinetics of inactivation were studied in Triton X-100 micelles . Inactivation is fast and the rate of inactivation is constant over the pH range where this lipase is active . Metal ions like Ca2+ and Sr2+ do not appreciably influence the rate of inactivation, although the enzymatic activity is significantly increased, suggesting a structural role for these ions . The S . hyicus lipase contains a consensus sequence G-H/Y-S-X-G . Substitution by site-directed mutagenesis of this serine (Ser369) by a cysteine resulted in a mutant with only 0.2% residual activity . The activity of this mutant could not be inhibited with water-soluble sulfhydryl reagents either in the presence or absence of Triton X-100 micelles . In the presence of Triton X-100 micelles, inactivation of the mutant occurred with 4-nitrophenylhexadecyl disulfide (t1/2 = 125 min) while the wild-type enzyme does not react at all . We conclude that Ser369 is the active site residue and that in water this residue is inaccessible . Only after interfacial activation Ser369 (or Cys369) becomes exposed and reacts with irreversible inhibitors.

Clin Nephrol, 1994 Apr, 41(4), 233 - 6
An adult patient with hyperimmunoglobulemia E (Job's) syndrome, end-stage renal disease and repeated episodes of peritonitis; Khan GA et al.; We report the first case of an adult patient with hyperimmunoglobulemia E (Job's) syndrome, end-stage renal disease and repeated episodes of Staphylococcus peritonitis . A 42-year-old black female with end-stage renal disease was treated with hemodialysis, and later switched to peritoneal dialysis because of severe vascular access problems . On CAPD, she had numerous (> 15) episodes of staphylococcal peritonitis . Past history included chronic atopic eczema and repeated sinopulmonary infections since childhood . She was found to have serum IgE levels greater than 10,000 units/ml . This combination of repeated infections, chronic skin condition and phenomenal elevation of IgE levels establish the diagnosis of hyperimmunoglobulemia E (Job's) syndrome . The cause of repeated episodes of Staphylococcal peritonitis are presumed to be related to impaired host defense mechanisms known to be present in patients with this disorder . As nephrologists, we should be aware that patients with impaired host defense mechanisms are highly susceptible to peritonitis in the presence of an indwelling peritoneal catheter.

Int J Biochem, 1994 Apr, 26(4), 529 - 38
Isolation and partial amino acid sequence of a 78 kDa porcine gastrin-binding protein; Baldwin GS et al.; 1 . A 78 kDa protein (p78) has been partially purified from washed membranes isolated from the corpus of porcine gastric mucosa . The purification was monitored by covalent cross-linking of iodinated {Nle15}-gastrin . 2 . A single N-terminal sequence extending for 33 amino acids was obtained from the p78 preparation . Partial sequences totalling 192 amino acids were also obtained from 14 tryptic and 3 Staphylococcal V8 peptides . 3 . 10 peptides plus the N-terminal sequence were derived from a previously unsequenced protein which was distantly related to the product of the E . coli fadB gene (Baldwin G . S . (1993) Comp . Biochem . Physiol . 104B, 55-61) . The remaining 7 peptides were derived from the beta-subunit of the gastric H+/K(+)-ATPase . 4 . The gastrin-binding activity remained in association with p78, and could be separated from the beta-subunit of the gastric H+/K(+)-ATPase, during chromatography on tomato lectin-Sepharose . 5 . We conclude that p78 binds gastrin, and is a novel member of the enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase family of enzymes.

AIDS, 1994 Apr, 8(4), 443 - 9
Lack of evidence for a superantigen in lymphocytes from HIV-discordant monozygotic twins; Nisini R et al.; OBJECTIVE: An HIV-associated superantigen (SAg) has been hypothesized . Here we test whether an SAg is functionally detectable in peripheral blood mononuclear cells (PBMC) from monozygotic twins discordant for HIV infection . DESIGN AND METHODS: The V beta selective T-cell depletion found in minor lymphocyte stimulation (Mls)-positive mice is caused by an SAg encoded by the mouse mammary tumour virus . Mls is a locus whose gene product stimulates a mixed lymphocyte reaction (MLR) in mice strains identical at the major histocompatibility complex locus . If an SAg is present in PBMC and/or sorted CD4+ cells from one HIV-infected monozygotic twin, it would stimulate PBMC from the corresponding healthy monozygotic twin in an MLR . In addition, if an SAg causes V beta-selective T-cell depletion in AIDS patients, a differential proliferation to a panel of staphylococcal enterotoxins (SE) of T lymphocytes from healthy and HIV-infected monozygotic twins should become measurable . RESULTS: No positive MLR or significant differences in the SE-driven proliferation between the healthy and the HIV-infected twins were observed . CONCLUSIONS: Our results suggest that PBMC from the two HIV-infected twins do not express a functionally detectable SAg.

Eur J Haematol, 1994 Apr, 52(4), 236 - 9
Salvage therapy with low-dose cytosine arabinoside in refractory or relapsed acute non-lymphocytic leukaemia: a report on 25 patients; Jensen MK et al.; Thirteen patients with de novo acute non-lymphocytic leukaemia (ANLL) refractory to standard chemotherapy for remission induction and 12 patients with ANLL in relapse were treated with low-dose cytosine arabinoside (LD ara-C) 10 mg/m2 subcutaneously every 12 hours for 21 days . Five of 13 patients (38%) and 6 of 12 patients (50%), respectively, obtained a complete remission (CR) . Of these, 7 patients subsequently relapsed after 2-76 months, while 4 patients remain in CR after 7-131 months . Compared to standard intensive regimens treatment with LD ara-C was rather non-toxic, requiring platelet transfusions and antibiotics in only 6 and 13 cases, respectively . Three patients (12%) died during induction therapy with LD ara-C; 2 had a cerebral haemorrhage and 1 developed anuria following a staphylococcal septicaemia . In conclusion, therapy with LD ara-C may be preferable to more intensive and toxic regimens in the treatment of patients with relapsed or refractory ANLL.

Protein Sci, 1994 Apr, 3(4), 549 - 56
The importance of anchorage in determining a strained protein loop conformation; Hodel A et al.; We examine the role of the conformational restriction imposed by constrained ends of a protein loop on the determination of a strained loop conformation . The Lys 116-Pro 117 peptide bond of staphylococcal nuclease A exists in equilibrium between the cis and trans isomers . The folded protein favors the strained cis isomer with an occupancy of 90% . This peptide bond is contained in a solvent-exposed, flexible loop of residues 112-117 whose ends are anchored by Val 111 and Asn 118 . Asn 118 is constrained by 2 side-chain hydrogen bonds . We investigate the importance of this constraint by replacing Asn 118 with aspartate, alanine, and glycine . We found that removing 1 or more of the hydrogen bonds observed in Asn 118 stabilizes the trans configuration over the cis configuration . By protonating the Asp 118 side chain of N118D through decreased pH, the hydrogen bonding character of Asp 118 approached that of Asn 118 in nuclease A, and the cis configuration was stabilized relative to the trans configuration . These data suggest that the rigid anchoring of the loop end is important in establishing the strained cis conformation . The segment of residues 112-117 in nuclease A provides a promising model system for study of the basic principles that determine polypeptide conformations . Such studies could be useful in the rational design or redesign of protein molecules.

Rev Rhum Ed Fr, 1994 Apr, 61(4), 260 - 5
{Diagnosis of arthritis in black Africa . Apropos of 473 cases in Congo}; Bileckot R et al.; A cross-sectional study of arthritis was conducted in the Rheumatology Department of the Brazzaville Teaching Hospital, Congo . A total of 473 patients with arthritis seen between 1989 and 1991 were subjected to the limited tests available . Gout was the leading diagnosis (n = 83) . Septic arthritis (n = 82) and infectious discitis (n = 55) were the most common reasons for admission . Tests often failed to identify the causative organism; Staphylococcus was the most commonly recovered organism . Tuberculous discitis was less common than discitis due to pyogenic bacteria . HIV-related arthritis (n = 57) usually manifested as severe, febrile, asymmetrical, nonerosive, polyarthritis . Cases of rheumatoid arthritis (n = 29) fit the classical description of the disease . In 83 patients with monoarthritis, oligoarthritis, or polyarthritis, no etiology could be identified.

Immunology, 1994 Apr, 81(4), 513 - 20
Differentiation of V beta 8.2+ CD4+ T cells induced by a superantigen: roles of antigen-presenting cells and cytokines; McKnight AJ et al.; The activation and subsequent differentiation of naive CD4+ T cells into functionally distinct effector cells is a vital step in the generation of an effective immune response to protein antigens . To analyse the development of effector T cells following the activation of resting, naive CD4+ T cells, we have utilized a transgenic mouse model in which the majority of T cells express a common T-cell receptor V beta molecule . The resting T cells were purified and stimulated in vitro with staphylococcal enterotoxin B, in the presence of accessory cells expressing class II major histocompatibility complex (MHC) molecules . We found that the cells which developed from these primary cultures were capable of producing varying levels of interleukin-2 (IL-2), IL-4 and interferon-gamma (IFN-gamma) following restimulation with anti-V beta 8 antibody, irrespective of whether B cells or macrophages/dendritic cells were the accessory cells in the primary cultures . The addition of IL-4 during primary stimulation enhanced the differentiation of IL-4-producing cells and suppressed the expansion of IFN-gamma-producing cells, especially when B cells were the antigen-presenting cells (APC) . Neutralization of endogenously produced IL-1, even in the presence of exogenous IL-4, did not inhibit the differentiation of IL-4-producing T cells . Strikingly, IL-10 completely suppressed the development of effector T cells when adherent macrophages/dendritic cells were utilized as accessory cells in the primary cultures, but had minimal effect in the presence of B cells . IFN-gamma suppressed the generation of IL-4-producing cells, presumably by inhibiting their expansion following primary activation . Finally, in vitro-generated IL-4-producing T cells were the most potent helpers for B lymphocytes . Thus, exogenous cytokines alter the patterns of T-cell differentiation in vitro, and the effects of cytokines vary depending on the types of accessory cells present during initial T-cell activation.

J Exp Med, 1994 Apr 1, 179(4), 1285 - 95
CD4pos, NK1.1pos T cells promptly produce interleukin 4 in response to in vivo challenge with anti-CD3; Yoshimoto T et al.; Injection of anti-CD3 antibodies causes prompt expression of interleukin (IL)-4, IL-2, and interferon gamma (IFN-gamma) mRNA among spleen cells . The optimal dose of anti-CD3 for such induction was 1.33 microgram/animal; lymphokine mRNA was first observed at 30 min, peaked at 90 min, and was undetectable (for IL-4) or had declined markedly by 4 h . Cells harvested from spleens of mice injected with anti-CD3 90 min earlier secreted IL-4, IL-2, and IFN-gamma without further stimulation . By contrast, in vitro stimulation with anti-CD3 of spleen cell suspensions or splenic fragments from noninjected donors failed to cause prompt production of IL-4 and, even after 24 h of stimulation, the amount of IL-4 produced in such cells was substantially less than that secreted within 1 h by spleen cell suspensions or splenic fragments from mice injected with anti-CD3 90 min earlier . Production of IL-4 by spleen cells from anti-CD3-injected mice was not inhibited by pretreatment with anti-IL-4 antibody or with IFN-gamma or tumor growth factor beta nor enhanced by treatment with IL-4 . By contrast, CTLA-4 immunoglobulin (Ig) treatment clearly diminished IL-4 production in response to in vivo anti-CD3, indicating that cellular interactions involving CD28 (or related molecules) were important in stimulation . Cell sorting analysis indicated that the cells that produced IL-4 in response to in vivo injection of anti-CD3 were highly enriched in CD4pos cells with the phenotype leukocyte cell adhesion molecule-1 (LECAM-1)dull, CD44bright, CD45RBdull, NK1.1pos . Indeed, the small population of CD4pos, NK1.1pos cells had the great majority of the IL-4-producing activity of this population . Injection with Staphylococcal enterotoxin B also caused prompt induction of IL-4 mRNA; the cells that were principally responsible for production also had the phenotype of CD4pos, NK1.1pos . These results suggest that possibility that this rare population of T cells may be capable of secreting IL-4 at the outset of immune responses and thus may act to regulate the pattern of priming of naive T cells, by providing a source of IL-4 to favor the development of T cell helper 2-like IL-4-producing cells.

J Biol Chem, 1994 Apr 1, 269(13), 9783 - 9
The role of the hinge region of the beta 2-subunit in beta-replacement specificity of tryptophan synthase from Escherichia coli . Analysis of proteolytically modified beta species cleaved by endoproteinase Glu-C; Linkens HJ et al.; The beta 2-subunit of tryptophan synthase from Escherichia coli was proteolyzed between Glu-296 and Ser-297 (Glu-nicked beta 2), using staphylococcal protease V-8 . Recombining Glu-nicked beta 2 with native alpha-subunits yields an enzymatically active and stable complex (alpha 2.Glu-nicked beta 2) with Kd = 3 microM . Comparing the properties of alpha 2.Glu-nicked beta 2 with those of isolated native beta 2-subunits revealed that the corresponding beta-chains are almost identical with respect to enzymatic specificities as well as to some spectral characteristics . The ratio of beta-replacement to beta-elimination activity is approximately 0.6 for both species . In the presence of L-serine, alpha 2.Glu-nicked beta 2 as well as native beta 2 exhibits "aqua" fluorescence emission at 500 nm, and no "amber" absorption at 468 nm can be detected with both species . However, when ammonium ions are added, the alpha 2.Glu-nicked beta 2 complex dramatically alters its properties . Similar to the native alpha 2 beta 2 enzyme, the ratio of beta-replacement/beta-elimination activity increases by a factor of about 10, the aqua fluorescence disappears, and the amber complex becomes detectable . We conclude that the continuity of the hinge region in the tryptophan synthase beta 2-subunit plays a crucial role in the alpha-mediated structural alteration creating beta-replacement specificity . The addition of NH4+ ions to the alpha 2.Glu-nicked beta 2 complex partially restores the structure-function relationships as described for native alpha 2 beta 2 tryptophan synthase.

J Infect Dis, 1994 Apr, 169(4), 730 - 8
Expression of costimulatory molecule CD28 on T cells in human immunodeficiency virus type 1 infection: functional and clinical correlations; Brinchmann JE et al.; To study the functional integrity of T cells from human immunodeficiency virus type 1 (HIV-1)-infected persons, CD4+ and CD8+ cells were examined for proliferation and secretion of interleukin-2 (IL-2) in response to staphylococcal superantigens and antibodies to CD3 and the alpha beta T cell receptor . A functional defect within CD8+ but not within CD4+ cells from HIV-1-infected persons was observed . Within CD8+ cells, proliferation and secretion of IL-2 was restricted to cells expressing the costimulatory molecule CD28 . Such cells were proportionally reduced in HIV-1-infected persons . In patients with advanced immunodeficiency, however, evidence of functional derangement was found also within the CD28+ CD8+ cells . In a cross-sectional study of 73 HIV-infected persons and 15 controls, a significant correlation was observed between the number of CD28+ CD8+ cells and the presence of HIV-related disease . Our results suggest that regulation of expression of CD28 may play an important role in the immunopathogenesis of AIDS.

Immunology, 1994 Apr, 81(4), 584 - 91
Human eosinophils as antigen-presenting cells: relative efficiency for superantigen- and antigen-induced CD4+ T-cell proliferation; Mawhorter SD et al.; Human eosinophils become hypodense and express class II major histocompatibility (MHC) molecules when activated by granulocyte-macrophage colony-stimulating factor (GM-CSF) in vitro or in vivo in pathological conditions such as allergic disorders . In this study, we examined the capacity of class II MHC-expressing eosinophils to serve as antigen-presenting cells (APC) for resting and activated CD4+ T cells . Eosinophils were isolated from healthy donors and incubated in conditioned medium (CM) containing GM-CSF for 2-4 days, after which 15-92% of the cells expressed class II MHC (HLA-DR) . Preincubated eosinophils induced resting T cells to proliferate in response to the staphylococcal superantigens, Staphylococcus enterotoxins A, B and E . Furthermore, superantigen-induced T-cell proliferation correlated with the proportion of eosinophils expressing class II MHC molecules . When eosinophils and macrophages were compared for their ability to act as accessory cells for superantigen-induced T-cell proliferation, macrophages were more efficient than eosinophils . Eosinophils were not effective APC for microbial antigens (Ag), which required processing . Proliferative responses to purified protein derivative, tetanus toxoid, or Brugia malayi antigen were observed in only three of nine studies . The three positive studies included activated CD4+ T cells, whereas no responses were observed with resting CD4+ T cells . Macrophages and mononuclear cells were effective APC for these Ag for both resting and activated CD4+ T cells . These data indicate that although class II MHC-expressing eosinophils can serve as APC, they are relatively inefficient for the activation of CD4+ T cells by Ag, which require processing.

FEMS Microbiol Lett, 1994 Mar 15, 117(1), 113 - 9
The Staphylococcus carnosus secE gene: cloning, nucleotide sequence, and functional characterization in Escherichia coli secE mutant strains; Meens J et al.; A DNA fragment containing the genes secE, nusG and rplK of Staphylococcus carnsosus was cloned using the Escherichia coli rplK gene as a probe . The S . carnosus secE homologue encodes a protein of 65 amino acid residues which is homologous to the carboxyl-terminal region of the E . coli SecE protein . The S . carnosus SecE polypeptide which, in contrast to the E . coli SecE protein, contains only one putative transmembrane segment, could fully replace the E . coli SecE protein in two different secE mutants . These results strongly suggest that the identified secE gene encodes an important component of the S . carnosus protein export apparatus.

Eur J Biochem, 1994 Mar 15, 220(3), 893 - 900
Translocation and processing of various human parathyroid hormone peptides in Escherichia coli are differentially affected by protein-A-signal-sequence mutations; Kareem BN et al.; Two staphylococcal protein-A signal sequences were constructed and tested for function in Escherichia coli, after being linked to human parathyroid hormone (hPTH) cDNAS representing the intact form (1-84 amino acids) and two N-terminal (1-37 and 1-7 amino acids) peptides . One signal sequence was identical to the wild type, and the other signal contained a deletion of 12 bp at the 3' end . The truncated hPTH cDNAs were fused at their 3' ends to IgG-binding domains (ZZ) derived from protein A in order to facilitate purification and characterization . The expression plasmid pSPTH, containing the wild-type signal sequence, secreted efficiently the intact recombinant hPTH (1-84) into the medium . Plasmids containing the truncated hPTH genes after the wild-type signal, gave rise to hPTH-ZZ hybrid proteins which were correctly processed at the N-terminal, but the major fractions appeared in the periplasmic compartment . In contrast, the plasmid pS'PTH which harboured the 4-amino-acid signal deletion did not promote a uniform secretion of intact hPTH (1-84) to the medium, but released a non-processed form both into the periplasmic compartment and to the medium . The related plasmids pS'PTH37ZZ and pS'PTH7ZZ with the mutated signal sequence gave rise to small or trace amounts of unprocessed forms of fusion proteins in the medium and periplasm, thus the secretion competence was markedly reduced . Thus, for correct N-terminal processing, we conclude that the amino acid sequence in the signal adjacent to the expressed protein, is a key determinant . However, release into the medium or periplasmic space appeared to be dependent also on protein folding, irrespective of signal-sequence cleavage . Furthermore, we observed that the peptides with the wild-type signal sequence and correct N-terminal processing, were the only forms that showed internal cleavage of hPTH . Uncleaved signals may contribute to folding characteristics of the ensuing protein and e.g., prevent internal proteolysis.

Proc Natl Acad Sci U S A, 1994 Mar 15, 91(6), 2300 - 4
Antagonism of superantigen-stimulated helper T-cell clones and hybridomas by altered peptide ligand; Evavold BD et al.; T-cell activation by an immunogenic peptide can be antagonized by nonstimulatory analogs of that peptide . We investigated this T-cell receptor antagonism by using staphylococcal enterotoxin superantigen to stimulate hemoglobin-specific helper T (Th) cells because its activation pathway may differ from that of conventional antigen . Interestingly, superantigen activation of these Th cells was antagonized by hemoglobin peptide analogs even though agonist (superantigen) and antagonist (analog peptide) bind at different sites on the major histocompatibility complex-encoded molecule and the T-cell receptor . The antagonism appeared to be a fundamental block in T-cell activation, as phosphoinositol generation, cytokine production, and proliferation were reduced in Th1 clones, and, similarly, proliferative and cytokine responses were inhibited in Th2 cells . Even T-cell hybridoma activation (cytokine production and apoptosis) was inhibited by peptide antagonists . Furthermore, analog peptides that functioned as partial agonists for these Th cells also antagonized superantigen-induced proliferation and thus were a subset of the peptide antagonists . In summary, our results demonstrate that analogs of immunogenic peptide are potent antagonists for Th cell responses induced by superantigen as well as immunogenic peptide.

Proc Natl Acad Sci U S A, 1994 Mar 15, 91(6), 2086 - 90
Stimulation with specific antigen can block superantigen-mediated deletion of T cells in vivo; McCormack JE et al.; The T-cell response to pigeon cytochrome c peptide, residues 88-104 (pcytC), in B10.BR mice is mediated largely by cells bearing both V beta 3 and V alpha 11 variable regions of the T-cell antigen receptor . These cells are, therefore, reactive with the superantigen staphylococcal enterotoxin A (SEA) . Recent reports have shown that in vivo exposure to superantigen can lead to deletion of superantigen-reactive T cells from the pool of mature T cells in the periphery . Here we show that upon cotreatment of animals with both SEA and pcytC, bulk deletion of the population of SEA-reactive cells is maintained, while the subpopulation of SEA-reactive T cells that also responds to pcytC is not deleted but instead proliferates in response to pcytC . These results are discussed with regard to mechanisms regulating the balance between T-cell tolerance and T-cell activation in vivo.

J Immunol Methods, 1994 Mar 10, 169(2), 205 - 11
Recombinant technology in the preparation of immunogen and enzymatic tracer . Application to the development of an enzyme immunoassay for rat prolactin; Ezan E et al.; A competitive enzyme immunoassay for rat prolactin using an immunogen and a tracer obtained by recombinant DNA technology is described . Polyclonal antibodies were raised in rabbits immunized with a purified chimeric protein consisting of rat prolactin fused with two synthetic immunoglobulin G binding domains derived from staphylococcal Protein A . The enzymatic tracer was obtained using an expression system which permits insertion of rPRL DNA sequence in the alkaline phosphatase gene . Antibodies and tracer were used to develop a solid-phase competitive immunoassay for the measurement of rat prolactin in plasma with a minimal detectable concentration of 0.5 ng/ml . The mean intra- and inter-assay coefficients of variation were 7.8 and 13.2%, respectively . Rat plasma concentrations measured with this assay correlated well with those obtained with a conventional enzyme immunoassay (r = 0.993, slope = 1.037, n = 24).

J Biol Chem, 1994 Mar 4, 269(9), 6485 - 90
1,25-dihydroxyvitamin D3 dissociates production of interstitial collagenase and 92-kDa gelatinase in human mononuclear phagocytes; Lacraz S et al.; To study the effect of mononuclear cell differentiation on metalloproteinase production, the human monocytic cell lines U937 and THP-1 were exposed to two well known differentiating agents, the phorbol esters (phorbol myristate acetate (PMA)) and 1,25-dihydroxyvitamin D3 (1,25-(OH)2D3) . With U937 cells, PMA-induced differentiation increased the production of both interstitial collagenase and 92-kDa gelatinase, whereas exposure to 1,25-(OH)2D3 induced full interstitial collagenase expression in the absence of any detectable 92-kDa gelatinase production . In fact, when U937 cells were differentiated with PMA and then exposed to vitamin D3, the hormone actually suppressed phorbol-induced 92-kDa gelatinase biosynthesis . With THP-1 cells, PMA also induced the production of 92-kDa gelatinase fully, but unlike U937 cells, the combination of PMA and 1,25-(OH)2D3 was required for substantial interstitial collagenase biosynthesis . As with U937 cells, the addition of 1,25-(OH)2D3 to PMA-differentiated THP-1 cells caused a dose-dependent inhibition of 92-kDa gelatinase production . Northern hybridizations demonstrated that both phorbol esters and vitamin D3 act on monocytic cell lines at a pretranslational level . To determine whether metalloproteinase biosynthesis in normal differentiated mononuclear phagocytes was also modified by 1,25-(OH)2D3, human blood monocytes and alveolar macrophages were exposed to this hormone . In both cell types, basal and Staphylococcal-stimulated 92-kDa gelatinase production was markedly inhibited by 1,25-(OH)2D3 . In contrast, interstitial collagenase production was completely unaffected by the hormone . In summary, the two major metalloproteinases produced by monocytic cells are regulated via distinct molecular pathways by the action of PMA and 1,25-(OH)2D3 . Furthermore, vitamin D3 completely dissociates the production of 92-kDa gelatinase and interstitial collagenase in human mononuclear phagocytes.

J Cataract Refract Surg, 1994 Mar, 20(2), 158 - 61
In vitro adhesion of Staphylococcus epidermidis on heparin-surface-modified intraocular lenses; Arciola CR et al.; This quantitative study assessed the in vitro adhesion of Staphylococcus epidermidis on two types of intraocular lenses: conventional poly(methyl methacrylate) (PMMA) and heparin-surface-modified PMMA . Level adhesion was measured by microbiological turbidimetry . We also measured modifications in the bacterium's structural fatty acids after adhesion using gas chromatography.

Ophthalmic Surg, 1994 Mar, 25(3), 157 - 61
Surgical wound defects associated with endophthalmitis; Maxwell DP Jr et al.; Twenty-five consecutive cases of culture-proven postsurgical endophthalmitis were evaluated . Patients underwent wound revision and pars plana vitrectomy with intravitreal antibiotic and steroid infusion (gentamicin 8 micrograms/cc, clindamycin 9 micrograms/cc, dexamethasone 8 micrograms/cc) and injection (gentamicin 100 micrograms plus clindamycin 200 micrograms {and amphotericin 5 micrograms in one case} and dexamethasone 800 to 1000 micrograms) . Twenty cases demonstrated wound defects (eg, wound gape/malapposition, abscess/tissue necrosis, suture dehiscence, leak, vitreous wick) . Culture-proven isolates included both gram negative and positive bacteria and fungi . Visual acuity improved in 18 of the 20 (90%) gram positive cases . Ten of the 17 (59%) patients in the Staphylococcus epidermidis subgroup achieved a visual acuity of 20/50 or better . Surgical wound defects are frequently associated with culture-proven endophthalmitis . When vitrectomy is included as part of the treatment regimen, we recommend meticulous inspection and closure of any defective surgical wounds associated with endophalmitis.

J Clin Microbiol, 1994 Mar, 32(3), 811 - 8
Detection of bacteremia by Difco ESP blood culture system; Morello JA et al.; In a multicenter study, the Difco ESP blood culture system (Difco Laboratories, Detroit, Mich.) was compared with the BACTEC NR660 system (Becton Dickinson Diagnostic Instrument Systems, Sparks, Md.) . The ESP system monitors each blood culture bottle every 12 to 24 min to detect changes in oxygen consumption and gas production by microbes . Equal volumes of blood were inoculated into aerobic ESP-80A and BACTEC 6A, 16A, or PEDS Plus broths and anaerobic ESP-80N and BACTEC 7A or 17A broths and were incubated for up to 7 days . ESP bottles contain supplemented tryptic soy broth without antimicrobial agent-adsorbing resins . From 7,532 aerobic compliant sets, the ESP system detected 356 clinically significant positive cultures and the BACTEC NR660 system detected 329 . From 6,007 anaerobic cultures, the ESP system detected 234 clinically significant positive cultures and the BACTEC NR660 system detected 198 . In aerobic broths, 292 organisms were isolated from both systems and 78 organisms were isolated from the ESP system alone, whereas 54 organisms were isolated from the BACTEC NR660 system alone (P < 0.05) . Among individual organisms, pneumococci were isolated significantly more often in ESP aerobic broths . In anaerobic broths, 180 organisms were isolated from both systems and 68 organisms were isolated from the ESP system alone, whereas 35 organisms were isolated from the BACTEC NR660 system alone (P < 0.05) . Aerobic gram-positive organisms as a group and Candida spp . were isolated significantly more often in ESP anaerobic broths . Both systems detected 207 clinically significant bacteremic episodes and the ESP system alone detected 63, whereas the BACTEC NR660 system alone detected 32 (P < 0.05) . Significantly more episodes of bacteremia caused by Staphylococcus epidermidis and anaerobes were detected by the ESP system . The differences in the numbers of organisms detected >6h earlier in ESP broths compared with BACTNEC NR660 broths were significant, as were earlier times to detection . Although the total number of organisms detected was not significantly different, the ESP system alone detected more organisms in a shorter time than did the BACTEC NR660 system alone . The continuous monitoring capability of the ESP system makes it an attractive alternative to the BACTEC NR660 system.

J Clin Microbiol, 1994 Mar, 32(3), 793 - 5
Antimicrobial susceptibility of Staphylococcus hyicus isolated from exudative epidermitis in pigs; Wegener HC et al.; Exudative epidermitis or greasy pig syndrome is caused by the coagulase-variable staphylococcal species Staphylococcus hyicus . Treatment of this disease is problematic because of the limited number of antimicrobial agents available for this purpose . Thirteen antimicrobial agents were evaluated for their activities against 100 S . hyicus strains isolated from pigs with exudative epidermitis . Novobiocin was the most active compound tested, with an MIC for 90% of the strains tested (MIC90) of < or = 0.06 microgram/ml . Enrofloxacin, ampicillin, and ceftiofur were the next most active compounds, with MIC90s of 0.25, 0.5, and 1.0 microgram/ml, respectively . However, 41.4% of the 99 strains tested were positive for beta-lactamase production . The MIC90s of erythromycin, tetracycline, and streptomycin were > 32.0 micrograms/ml . Initial testing with sulfadiazine-trimethoprim yielded an MIC90 of > 64.0 micrograms/ml, but subsequent testing with thymidine phosphorylase-supplemented medium yielded an MIC90 of 0.06 microgram/ml . Both lincomycin and spectinomycin were relatively inactive against the S . hyicus strains tested, with MIC90s of > 64.0 and > 128.0 micrograms/ml, respectively . However, the combination of the two compounds at ratios of 1:2 (lincomycin to spectinomycin) and 1:8 were more active, with MIC90s of 16.0 and 4.0 micrograms/ml, respectively . These results indicate that novobiocin and sulfadiazine-trimethoprim were the most active compounds tested against the S . hyicus strains isolated from pigs with exudative epidermitis . Furthermore, the combination of lincomycin and spectinomycin was more active than the individual compounds against the strains tested.

J Clin Microbiol, 1994 Mar, 32(3), 649 - 53
Evaluation of Staf-Sistem 18-R for identification of staphylococcal clinical isolates to the species level; Piccolomini R et al.; The accuracy and efficiency of Staf-Sistem 18-R (Liofilchem s.r.l., Roseto degli Abruzzi, Teramo, Italy) were compared with those of conventional biochemical methods to identify 523 strains belonging to 16 different human Staphylococcus species . Overall, 491 strains (93.9%) were correctly identified (percentage of identification, > or = 90.0), with 28 (5.4%) requiring supplementary tests for complete identification . For 14 isolates (2.8%), the strains did not correspond to any key in the codebook and could not be identified by the manufacturer's computer service . Only 18 isolates (3.4%) were misidentified . The system is simple to use, is easy to handle, gives highly reproducible results, and is inexpensive . With the inclusion of more discriminating tests and adjustment in supplementary code numbers for some species, such as Staphylococcus lugdunensis and Staphylococcus schleiferi, Staf-Sistem 18-R is a suitable alternative for identification of human coagulase-positive and coagulase-negative Staphylococcus species in microbiological laboratories.

J Cardiovasc Electrophysiol, 1994 Mar, 5(3), 219 - 31
Ablation of left free-wall accessory pathways using radiofrequency energy at the atrial insertion site: transseptal versus transaortic approach; Deshpande SS et al.; INTRODUCTION: Transcatheter ablation of the left free-wall atrioventricular accessory pathways (AP) by delivery of radiofrequency current at the ventricular insertion site has been shown to be effective . The efficacy of such a technique targeting the atrial insertion site of the AP was evaluated . METHODS AND RESULTS: One hundred consecutive patients with left free-wall APs and symptomatic supraventricular tachyarrhythmias were included . APs were manifest in 55 patients and concealed in 45 . There were 55 men and 45 women with a mean age of 35 years . A total of 107 left free-wall APs were identified in these patients . In these 100 patients, successful ablation was accomplished in all by using a transseptal (45 patients) or transaortic (54 patients) technique . In one patient, ablation was accomplished from within the coronary sinus . Seven patients required a repeat ablative procedure, which was performed successfully . During 107 ablative procedures, six were associated with nonfatal complications including pericardial effusion (hemopericardium) in two patients, mild mitral regurgitation in two patients, swelling of the left arm in one patient, and staphylococcal bacteremia in one patient . Eighty-two (82%) patients underwent a repeat electrophysiologic study 6 to 8 weeks after successful ablation and were found to have no functioning AP or inducible supraventricular tachycardia . During a mean follow-up of 20 +/- 8 months, none of the 100 patients had a recurrence of tachyarrhythmias . CONCLUSION: These data indicate that the atrial insertion site of the AP can be successfully ablated in the majority of patients with left free-wall APs by using either a transseptal or transaortic approach . Furthermore, both techniques are associated with minimal morbidity and no mortality.

Ann Pharmacother, 1994 Mar, 28(3), 333 - 5
Otic administration of amphotericin B 0.25% in sterile water; Kintzel PE et al.; OBJECTIVE: To report otic administration of parenteral amphotericin B 0.25% in sterile water . CASE SUMMARY: A 44-year-old HIV+ man was diagnosed with otitis externa . The patient's past medical history was remarkable for positive Coccidioides immitis serology for more than five months, essential hypertension, and Barrett's esophagitis . Culture results from an ear swab revealed 4+ Aspergillus fumigatus and 3+ Staphylococcus, coagulase negative . Antiinfective therapy for the otitis externa included oral and topical antibacterial and antifungal medications . Amphotericin B 0.25% in sterile water was prepared by the pharmacy for topical otic administration . The otic amphotericin B was dispensed with instructions to refrigerate and assigned a one-week expiration date . The prescription called for instillation of 1-2 drops in each ear three times a day . The patient's signs and symptoms of otitis externa resolved during several weeks of antiinfective therapy . Topical administration of amphotericin B 0.25% in sterile water was not associated with any local adverse effects in this patient . DISCUSSION: The rationale for use of the parenteral amphotericin B formulation to prepare an otic dosage form, and the rationale for the specific concentration and expiration date chosen are discussed . CONCLUSIONS: This patient tolerated topical otic administration of amphotericin B 0.25% in sterile water when administered three times daily.

Pediatr Res, 1994 Mar, 35(3), 293 - 8
Proliferative and cytokine responses by human newborn T cells stimulated with staphylococcal enterotoxin B; Hayward A et al.; Staphylococcal enterotoxins are potentially valuable tools for investigating the development of T-cell responses because in experimental animals they can elicit either T-cell activation and proliferation or tolerance . Previous studies indicate that human T cells bearing the CD45RA phenotype (which account for the majority of newborn T cells) respond poorly to stimulation by staphylococcal enterotoxin B (SEB) compared with mature T cells from adult blood . The present studies show that the mean frequency of newborn T cells that proliferated in limiting dilution cultures stimulated by SEB was 1:3135, with a 1SD range of 3153-4191 compared with a mean of 1:493 and range of 120-1737 for adult T cells . Neither indomethacin nor the nitric oxide synthesis inhibitor, n-arginine methyl ester, increased SEB responses by newborn cells, arguing against down-regulation of the newborn response by prostaglandin or nitric oxide . Naive (CD45RA+) T cells from adult blood had a responder cell frequency to SEB similar to that of the newborn cells . IL-2 production by newborn cells was delayed compared with adult cells but was equivalent after 3 d of culture . Production of gamma-interferon and IL-4 was greater by adult than newborn cells . Our results indicate that a subset of CD45RA+ cells that is activated by SEB can mature to make IL-4 or gamma-interferon after 3-5 d . The limiting dilution assay results provide a quantitative basis for proliferation by naive T cells against which responses by T cells from healthy and premature newborns can be compared.

J Natl Med Assoc, 1994 Mar, 86(3), 203 - 8
Inappropriate breast secretions of possible bacterial etiology in the parous nonpuerperal female; Freeman JJ et al.; This article presents two cases of spontaneous green breast secretions of parous nonpuerperal patients . To understand the nature of these secretions, bacterial evaluations and subsequent treatment were undertaken . Case 1 culture and sensitivity studies from breast secretions were commenced within 24 hours yielding an isolate identified as Staphylococcus epidermidis, with sensitivity to cephalothin, erythromycin, and tetracycline but resistant to penicillin . Cephalothin, 500 mg four times a day for 10 days, followed by erythromycin 100 mg twice a day for 10 days and doxycycline 100 mg twice a day for 10 days, did not alter the breast secretions . Four weeks later, ciprofloxacin HCI 500 mg twice a day for 6 weeks caused a 50% decrement in breast secretion at 4 weeks but increased clinical depression . At 6 weeks, no evidence of breast secretions persisted . Mental depression decreased within 2 weeks postciprofloxacin treatment . In Case 2, a total of 35 minutes elapsed between sample collection and initiation of culture and sensitivity studies . Moraxella osloensis was identified and found sensitive to ampicillin and tetracycline but resistant to trimethoprim . Ampicillin 500 mg four times a day for 10 days and doxycycline 100 mg twice a day by mouth for 10 days were administered at 2-week intervals with no effect on breast discharge . After 4 weeks of treatment failure, ciprofloxacin HCI 500 mg twice a day for 6 weeks caused a 50% decrease in discharge at 2 weeks and total elimination at 6 weeks . Lethargy during treatment ceased with termination of therapy . These results support the importance of bacterial evaluation of breast secretions with subsequent antibiotic therapy.(ABSTRACT TRUNCATED AT 250 WORDS)

J Infect Dis, 1994 Mar, 169(3), 526 - 31
Endemic nosocomial transmission of Staphylococcus epidermidis bacteremia isolates in a neonatal intensive care unit over 10 years; Huebner J et al.; To assess long-term nosocomial transmission, trends in antibiotic resistance, and expression of potential virulence factors, 86 randomly selected Staphylococcus epidermidis bloodstream isolates obtained from 80 patients in a neonatal intensive care unit (NICU) over a 10-year period were studied . Pulsed-field gel electrophoresis (PFGE) analysis of SmaI-digested whole chromosomal DNA revealed distinctive banding patterns that persisted in the NICU over long periods . Pattern A included 22 isolates (26%) obtained during 1983-1990, and pattern B included 24 isolates (28%) from 1983 to 1991 . All 10 isolates examined in 1984 fell into one of these two patterns . Isolates with either pattern expressed polysaccharide/adhesin (PSA) and slime; 90% and 87% were resistant to oxacillin and gentamicin, respectively, with no trends over time . These findings suggest that distinct clones of S . epidermidis can become endemic in NICUs over periods as long as a decade and that nosocomial transmission plays an important role in neonatal S . epidermidis bacteremia.

Support Care Cancer, 1994 Mar, 2(2), 94 - 104
Infections in cancer patients: some controversial issues; Schimpff SC et al.; Despite more than two decades of clinical research into the management of infections in the neutropenic cancer patient, many patients still develop serious morbidity from infection and all too many still die . A number of controversies surround (a) the use of combination versus monotherapy for initial empiric administration; (b) the use of vancomycin as part of the initial regimen; (c) the origin of Staphylococcus epidermidis infections (i.e., mostly from vascular catheters or mostly from the alimentary canal); (d) the use of acyclovir for herpes simplex prophylaxis during remission induction for acute leukemia patients not undergoing bone marrow transplantation; (e) the use of alimentary canal microbial suppression or reverse isolation in a room with laminar air flow, or both, as infection prevention techniques . Current recommendations and observations include the following . (a) Monotherapy with ceftazidime or imipenem is effective and appropriate for patients with moderate granulocytopenia at limited risk for infection with a resistant organism . Combination therapy is recommended for patients with profound, persistent granulocytopenia who are at high risk for gram-negative bacteremia; such bacteremic patients have a better prognosis with combined-modality therapy . (b) Vancomycin need not be included in the initial regimen although some centers may choose to do so because of the high prevalence of gram-positive bacteremias . (c) Despite the ubiquitous presence of indwelling vascular catheters, most S . epidermidis infections among neutropenic patients originate from along the alimentary canal . (d) Herpes simplex infection is much more common following standard remission induction chemotherapy than previously recognized . Acyclovir will reduce these infections and concurrently probably reduce the likelihood of resultant bacterial/fungal co-infections and superinfections . (e) Selective microbial suppression is appropriate for patients expected to experience prolonged (more than 2 weeks) or profound (below 100 granulocytes/microliters) granulocytopenia . Agents chosen should suppress aerobic but not anaerobic flora (maintain colonization resistance) and need to have an effect on both the oral cavity and esophagus as well as the intestines.

Br J Dermatol, 1994 Mar, 130(3), 329 - 36
Inhibition of erythromycin-resistant propionibacteria on the skin of acne patients by topical erythromycin with and without zinc; Bojar RA et al.; Propionibacteria resistant to high concentrations of erythromycin {minimal inhibitory concentration (MIC) > or = 0.5 mg/ml} are now commonly isolated from the skin of antibiotic-treated acne patients . This double-blind study was carried out to assess the ability of 4% w/v erythromycin with and without 1.2% w/v zinc acetate to reduce the numbers of erythromycin-resistant propionibacteria in vivo, and also to monitor the acquisition of resistant strains de novo during therapy . Under laboratory conditions, erythromycin-resistant propionibacteria were shown to be as sensitive to zinc acetate as fully sensitive strains . In vivo, the erythromycin/zinc complex and erythromycin alone produced highly significant reductions in total propionibacteria (P < 0.001) and in the number of erythromycin-resistant strains (P < 0.001 at 8 weeks) . After 12 weeks, resistant propionibacteria were reacquired, or acquired de novo, by three patients treated with erythromycin alone and four patients treated with the erythromycin/zinc complex . In contrast, changes in numbers of Micrococcaceae were slight and, after 12 weeks, erythromycin-resistant strains were predominant in both treatment groups . In vitro MIC determinations suggested that this finding might be explained by the exceptionally high degree of erythromycin resistance displayed by some staphylococcal strains (MIC > 4 mg/ml) and by the relative insensitivity of all staphylococcal strains to zinc acetate . Erythromycin with and without zinc was clinically effective, and both preparations produced significant reductions in acne grade, and inflamed and non-inflamed lesion counts (P < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

J Immunol, 1994 Mar 1, 152(5), 2385 - 92
The role of T cell activation in anti-CD3 x antitumor bispecific antibody therapy; Weiner GJ et al.; Anti-CD3 x antitumor bispecific Ab can retarget T cell mediated lysis in an MHC-independent fashion and prevent tumor growth in animal models . Two bispecific Ab preparations that differ in the presence or absence of Fc were compared in the 38C13 immunocompetent murine lymphoma model to evaluate how functional Fc and T cell activation impact on response to bispecific Ab therapy . Bispecific (bs) IgG contained functional Fc and was purified from hybrid-hybridoma Ab product . Bsf(ab')2 lacked functional Fc, and was genetically constructed using the leucine zipper technique . In vitro, bsF(ab')2 induced tumor cell lysis by activated T cells more effectively than bsIgG . However, bsF(ab')2 failed to induce T cell activation in the absence of tumor cells, and did so more slowly than bsIgG when tumor cells were present . In vivo, bsIgG induced nonspecific T cell activation whereas bsF(ab')2 did not . In therapy experiments, bsIgG inhibited tumor growth in mice although a single dose of bsF(ab')2 had minimal antitumor effect . BsF(ab')2 was capable of preventing tumor growth and improving survival when mice were also treated with T cell activators (IL-2 or staphylococcal enterotoxin B), or given repeated bsF(ab')2 doses . We conclude that therapeutic response to bispecific Ab was not dependent on functional Fc, but did require T cell activation . The use of bifunctional constructs that lack functional Fc therefore allows for separate manipulation of T cell retargeting and T cell activation and deserves further evaluation as a potential immunotherapy for malignancy.

Ophthalmology, 1994 Mar, 101(3), 508 - 18
Postoperative endophthalmitis in association with diabetes mellitus; Phillips WB 2nd et al.; BACKGROUND: Endophthalmitis continues to be a potentially devasting complication of ocular surgery, despite advances in microsurgical technique and infection-preventing measures . Patients with diabetes have altered immunity at various levels and may be more susceptible to infection after ocular surgery . The authors evaluate the associations between diabetes mellitus and postoperative endophthalmitis . METHODS: The records of 162 consecutive patients treated over a 5-year period for endophthalmitis occurring within 2 weeks of ocular surgery were retrospectively reviewed . RESULTS: Twenty-one percent of this consecutive series of patients with endophthalmitis after surgery had diabetes mellitus . Both the diabetic and nondiabetic groups were similar with respect to age, type of primary surgery, duration from surgery to onset of symptoms, presenting visual acuity, and management of endophthalmitis . Seventy-nine percent of the patients with diabetes and 68% of those without diabetes had culture-proven endophthalmitis . Staphylococcus was responsible for 74% and 71% of the culture-positive cases, respectively . The patients with diabetes were more likely to have endophthalmitis secondary to a gram-negative organism (P < 0.001) than those without diabetes (18.5% versus 5.7%) . Visual outcome was worse in the diabetic group, although this may be related to preoperative visual status . CONCLUSIONS: Twenty-one percent of this consecutive series of patients with endophthalmitis after surgery had diabetes mellitus . The patients with diabetes mellitus were more likely to have endophthalmitis caused by gram-negative organisms and appear to have a poorer visual prognosis after treatment for endophthalmitis.

Invest Ophthalmol Vis Sci, 1994 Mar, 35(3), 1026 - 32
Complement system and host defense against staphylococcal endophthalmitis; Giese MJ et al.; PURPOSE . The authors studied the role of the complement system in host defense against Staphylococcus epidermidis and S . aureus endophthalmitis . METHODS . Guinea pigs in the S . epidermidis model received an intravitreal injection of 7000 viable organisms, and guinea pigs in the S . aureus model received 50 viable organisms . The experimental animals in each model were decomplemented with intraperitoneal (IP) injections of cobra venom factor, whereas the control animals received IP injections of normal saline . Mean log bacterial counts in the vitreous and mean serum complement titers were compared in the experimental and control animals in each model on days 1, 2, 3, and 7 . RESULTS . In the S . epidermidis model, mean log bacterial counts in the vitreous were significantly higher in the experimental group than the control group on days 1 and 2 (P < 0.01) and on day 3 (P < 0.05) . Mean serum complement titers were significantly lower in the experimental group at all days (P < 0.01) . In the S . aureus model, mean log bacterial counts in the vitreous were significantly higher in the experimental group than the control group on day 2 (P < 0.05) and day 3 (P < 0.01) . Mean serum complement titers were significantly lower in the experimental group on days 1, 2, and 3 (P < 0.01), but not on day 7 . CONCLUSION . These results suggest that decomplemented guinea pigs show impaired host defense to S . epidermidis and S . aureus endophthalmitis and that this defense is restored as complement levels approach normal.

Eur J Immunol, 1994 Mar, 24(3), 731 - 7
Heat-stable antigen/CD24 on mouse T lymphocytes: evidence for a costimulatory function; Hubbe M et al.; Heat-stable antigen (HSA)/mouse CD24 (formerly termed Nectadrin) is a membrane glycoprotein with an unusual structure consisting of a small protein core and extensive glycosylation . It is expressed by hematopoietic cells but not by mature T lymphocytes . HSA on accessory cells is an important costimulatory molecule required for the clonal expansion of T lymphocytes . HSA is also involved in cell-cell adhesion events and the isolated antigen has been shown to possess self-binding properties . In the present study we have re-investigated the role of HSA in T cell proliferation . We find that following stimulation of T lymphocytes with concanavalin A or of CD4+ T lymphocytes with a combination of anti-CD3/CD28 monoclonal antibodies (mAb) the HSA antigen is transiently expressed . The expression correlated with the appearance of CD25 and a CD2 activation epitope at the cell surface . Induction of HSA was also seen in vivo on V beta 8+ T lymphocytes in BALB/c mice that were injected with Staphylococcal enterotoxin B . Biosynthetic labeling and analysis of mRNA by reverse transcriptase-polymerase chain reaction showed that HSA was synthesized by activated T lymphocytes . A combination of anti-CD3 and mAb 79 to HSA was incapable of inducing proliferation of purified CD4+ T lymphocytes . However, the antibody strongly enhanced the response obtained with a combination of anti-CD3/CD28 mAb . The augmenting effect of the HSA-specific mAb was dose dependent . Since HSA is bound to the membrane via a glycosyl-phosphatidylinositol (GPI) anchor and GPI-anchored molecules have been implicated in lymphocyte activation, it is conceivable that HSA is not only a costimulatory molecule on accessory cells but is also a signaling molecule in T lymphocytes . The possibility of a homotypic HSA/HSA interaction between T lymphocytes and accessory cells is discussed.

Eur J Immunol, 1994 Mar, 24(3), 579 - 84
Absence of peripheral clonal deletion and anergy in immune responses of T cell-reconstituted athymic mice; Williams O et al.; Superantigens induce clonal deletion of reactive T cells in the thymus and clonal deletion and anergy in the periphery of euthymic mice . In this report we have assessed the ability of Staphylococcal enterotoxin B (SEB) to induce peripheral tolerance in nude mice reconstituted with normal, syngeneic T cells . Immunization of reconstituted nude mice with SEB resulted in lethal toxic shock in a large fraction of the animals . Such lethality was never observed in the normal donor mouse strain . Analysis of lymphokine production in response to SEB showed that reconstituted nude mice produced higher levels of interleukin-2 and tumor necrosis factor-alpha, but lower levels of interleukin-4, than euthymic control mice . Furthermore, SEB was unable to promote either clonal elimination or induction of anergy in the SEB-responsive peripheral T cells, despite the fact that reconstituted nude mice did produce high levels of corticosterone upon treatment with SEB . These results imply a lack of control over immune responses to superantigen in T cell-reconstituted athymic mice.

Eur J Immunol, 1994 Mar, 24(3), 542 - 8
Identifying amino acid residues that influence plasma clearance of murine IgG1 fragments by site-directed mutagenesis; Kim JK et al.; Site-directed mutagenesis has been used to change amino acid residues of a recombinant Fc-hinge fragment derived from the murine immunoglobulin (Ig)G1 molecule, and the effects of these mutations on the pharmacokinetics of the Fc-hinge fragment have been determined . Specifically, Ile-253, His-310 and Gln-311 of the CH2 domain and His-433 and Asn-434 of the CH3 domain have been changed . In the three dimensional structure of an antibody, these amino acids are in close proximity to each other at the CH2-CH3 domain interface . The mutated Fc-hinge fragments have been purified from recombinant Escherichia coli cells and their pharmacokinetic parameters determined in mice and compared with those of the wild-type Fc-hinge fragment . The results show that the site of the IgG1 molecule that controls the catabolic rate (the 'catabolic site') is located at the CH2-CH3 domain interface and overlaps with the Staphylococcal protein A binding site.

Mol Gen Genet, 1994 Mar, 242(5), 566 - 72
Binding of ArsR, the repressor of the Staphylococcus xylosus (pSX267) arsenic resistance operon to a sequence with dyad symmetry within the ars promoter; Rosenstein R et al.; arsR, the first gene of the Staphylococcus xylosus (pSX267) arsenic/antimonite resistance (ars) operon encodes a negative regulatory protein, ArsR, which mediates inducibility of the resistances by arsenic and antimony compounds . ArsR, which has no obvious DNA-binding motif in its primary structure, was purified from an ArsR-overproducing Escherichia coli strain and identified as a DNA-binding protein by its behaviour in gel mobility shift assays . ArsR had a specific affinity for a 312 bp DNA restriction fragment carrying the ars promoter; the minimum sequence complexed by ArsR was a 75 bp polymerase chain reaction (PCR) fragment, which mainly comprised the -35 and -10 regions of the promoter . The effect of inducers on the DNA-binding activity of ArsR was examined by in vitro induction assays; only arsenite inhibited DNA-binding of the repressor . DNase I footprinting revealed two protected regions within the promoter region, spanning 23 and 9 nucleotides, respectively . Furthermore, a new cleavage site for DNase I between the protected regions was made accessible by binding of the repressor . The footprints cover a region of three inverted repeats located between the -35 and -10 motifs of the ars promoter . By high resolution footprinting with the hydroxy radical, five sites of close contact between the protein and DNA were identified.

Plast Reconstr Surg, 1994 Mar, 93(3), 533 - 6
Infections requiring hospital readmission following face lift surgery: incidence, treatment, and sequelae; LeRoy JL Jr et al.; This retrospective study evaluates 11 infections requiring hospital readmission following 6166 consecutive face lifts (0.18 percent) . Seven patients had surgical drainage of an abscess, and four patients were treated for severe cellulitis, Staphylococcus was the predominant organism in all abscess cultures . Of three patients admitted 3 weeks after surgery, two cultured gram-negative organisms in addition to Staphylococcus . This suggests that all patients should be treated initially with Staphylococcus coverage . If a patient is readmitted more than 1 week postoperatively, gram-negative coverage should be considered, pending culture results . There was no consistent finding regarding past medical history, the use of perioperative antibiotics, surgical equipment used, complexity of the surgical dissection, drains, or hematoma formation . No patients demonstrated systemic signs or grossly abnormal laboratory results on readmission . Eight patients had no sequelae from the infection . Three patients developed scarring that was considered minor by the affected patients . These data support the conclusion that major infections following a face lift are rare occurrences . Standard medical and surgical care, given in a timely fashion, usually results in a satisfactory outcome with minimal morbidity.

Biologicals, 1994 Mar, 22(1), 21 - 7
Enzyme-linked immunosorbent assay assessment of bovine viral diarrhea virus antigen in inactivated vaccines using polyclonal or monoclonal antibodies; Ludemann LR et al.; An enzyme-linked immunosorbent assay (ELISA) procedure was developed to assay the cytopathic and noncytopathic bovine viral diarrhea (BVD) virus strains used in inactivated vaccines licensed by the United States Department of Agriculture . The assay uses a biotin-labeled, staphylococcal protein A purified polyclonal BVD antibody (Bab) from a calf hyperimmuned against NADL, Singer, C24v, New York-1 (NY-1) strains and a field isolate . The Bab recognized the following reference strains of BVD virus: NADL; NY-1; C24v; Singer; and a field isolate . Monoclonal antibodies (Mab) directed against gp48 and gp53 of the Singer strain of BVD could detect only the Singer and the NY-1 strains . None of the Mab tested could differentiate between cytopathic and noncytopathic BVD virus strains . In vaccines containing multiple viral and bacterial components, the Bab was specific for the BVD fraction . Two vaccines not recognized by the Bab differed from the others in the type of adjuvant . The formation of antigen-adjuvant complexes during vaccine production may inhibit the ability of Bab to detect BVD antigens in an ELISA format . This ELISA procedure enables the detection of BVD antigens and demonstrates the potential for in vitro testing of inactivated BVD vaccines in place of the currently required host animal testing.

J Antimicrob Chemother, 1994 Mar, 33(3), 431 - 41
Binding of teicoplanin and vancomycin to polymer surfaces; Wilcox MH et al.; Both teicoplanin and vancomycin were found to bind to a range of polymer surfaces . The binding of teicoplanin to specimen vessel surfaces was, on average, four times greater than that of vancomycin and was particularly marked with silconized polymers (5.2 micrograms/cm2) . Pre-exposure of a polymer surface to human body fluids caused a 60% reduction in teicoplanin binding . Reduction of the negative surface charge on a polymer surface with ferric nitrate resulted in a ten-fold increase in teicoplanin binding . The accumulation of a strain of Staphylococcus epidermidis on silicone rubber catheter segments pre-exposed to glycopeptide antibiotics was examined . In phosphate buffered saline binding of bacteria to vancomycin-treated polymer was greater than to an unexposed control surface . In contrast, in human serum both antibiotics caused reductions in adherent growth . The binding of glycopeptide antibiotics, in particular teicoplanin, to polymer surfaces may interfere with the results of in-vitro assays . However, this phenomenon may be useful in the prevention of bacterial accumulation on the surfaces of medical devices.

Nutr Hosp, 1994 Mar-Apr, 9(2), 99 - 104
{Catheter management in parenteral nutrition}; Hernandez Jaras MV et al.; Following the appearance of Staphylococcus epidermidis positive hemocultures in four patients undergoing parenteral nutrition in different services, and after microbiological controls of the mixtures prepared by the Pharmacy Service in order to discard contamination during preparation, it was decided to assess the handling o catheters and central pathways by the Hospital Nursing Staff . A survey was carried out of 34 nurses, 17 each from the morning and evening shifts, from the floors with patients undergoing parenteral nutrition, representing 13.3% of all the nurses of those floors . There were five sections in the survey, with fifty-two questions referring to the introduction of catheters, change of dressing, care of the point of insertion, uses of the administrative pathway, change and handling of the parenteral nutrition bag, intravenous administration of medicines and parenteral nutrition, and withdrawal of the catheter . According to the results, 76% of central catheters are introduced in the operating theatre: once in place, the catheter is checked by X-ray to ensure that it is in the correct position, in all cases . There were major differences in the changing of dressings . The pathway for administration of the parenteral nutrition is used for a variety of functions . Medicines are administered in "Y" with the nutrient mixture, although their stability is not known . In changing the parenteral nutrition bag and the handling of the catheter, adequate sterilization measures were not taken.(ABSTRACT TRUNCATED AT 250 WORDS)

Protein Sci, 1994 Mar, 3(3), 391 - 401
Accommodation of insertion mutations on the surface and in the interior of staphylococcal nuclease; Keefe LJ et al.; Alignment of homologous amino acid sequences reveals that insertion mutations are fairly common in evolution . Hitherto, the structural consequences of insertion mutations on the surface and in the interior of proteins of known structures have received little attention . We report here the high-resolution X-ray crystal structures of 2 site-directed insertion mutants of staphylococcal nuclease . The structure of the first insertion mutant, in which 2 glycine residues were inserted on the protein surface in the amino-terminal beta-strand, has been solved to 1.70 A resolution and refined to a crystallographic R value of 0.182 . The inserted residues are accommodated in a special 3-residue beta-bulge . A bridging water molecule in the newly created cavity satisfies the hydrogen bonding requirements of the beta-sheet by forming a bifurcated hydrogen bond to 1 beta-strand, and a single hydrogen bond to the other beta-strand . The second insertion mutant contains a single leucine residue inserted at the end of the third beta-strand . The structure was solved to 2.0 A resolution and refined to a final R value of 0.196 . The insertion is accommodated in a register shift that changes the conformation of the flexible loop portion of the molecule, relaxing and widening the omega turn . This structural alteration results in changes in position and coordination of a bound calcium ion important for catalysis . These structures illustrate important differences in how amino acid insertions are accommodated: as localized bulges, and as extensive register shifts.

J Biomol NMR, 1994 Mar, 4(2), 193 - 200
Measurement of four-bond HN-H alpha J-couplings in staphylococcal nuclease; Vuister GW et al.; Quantitative J-correlation and triple-resonance ECOSY-type experiments are used to unambiguously establish the presence of four-bond sequential HN-H alpha J-couplings in the protein staphylococcal nuclease . Substantially negative 4JH alpha HN values, ranging from -0.8 to -2.3 Hz, are observed when the psi angle is near +120 degrees, and the following phi angle near +60 degrees . For other conformations, the four-bond HN-H alpha J-couplings fall between -0.5 and +0.5 Hz.

J Dermatol, 1994 Mar, 21(3), 166 - 71
The antibiotic susceptibility of Propionibacterium acnes and Staphylococcus epidermidis isolated from acne; Nishijima S et al.; We studied the susceptibility of antimicrobial agents to Propionibacterium acnes (P . acnes) and Staphylococcus epidermidis (S . epidermidis) isolated from acne patients . We measured the minimum inhibitory concentrations (MICs) of the following five drugs: roxithromycin (RXM), erythromycin (EM), clindamycin (CLDM), minocycline (MINO) and ofloxacin (OFLX), which are frequently used to treat acne and skin infections . We found many resistant strains of S . epidermidis and some resistant strains of P . acnes . There was a correlation between the resistance of S . epidermidis and the former therapy for acne, but no distinct correlation between the resistance of P . acnes and the former therapy for acne.

J Am Soc Nephrol, 1994 Mar, 4(9), 1719 - 25
Successful use of cuffed central venous hemodialysis catheters inserted percutaneously; Swartz RD et al.; Although endogenous fistulae and grafts are preferred for permanent hemodialysis access, central venous catheters are often required for varying intervals when creating permanent access is not feasible . The prospective experience with 118 catheters in over a 3.5-yr period is reported; 93 (79%) were placed by percutaneous techniques, and 25 (21%) were placed by operative techniques . Seventy seven catheters (65%) were placed in the subclavian vein, 36 (31%) were placed in the internal jugular vein (usually right side), and 5 (4%) were placed in the femoral vein . Early postplacement complications were infrequent . Catheter function at last local follow-up ranged from several days to nearly 2 yr, averaging approximately 3 mo, even though many patients returned to their referring centers with a functioning catheter after only a short follow-up . Actuarial survival for percutaneously placed catheters was approximately 60% at 6 mo and 30% at 12 mo . Catheter failure occurred in 36% of cases, equally divided between malfunction (thrombosis refractory to fibrinolysis, extrusion, kinking, or related event) and infection with septicemia requiring removal . Such failure was not more frequent after percutaneous placement than after operative placement . Failure due to mechanical malfunction, but not that due to infection, tended to be less frequent among catheters placed in the internal jugular vein than among catheters placed in the subclavian vein . Finally, infection with septicemia involved 22% of all catheters and occurred at an average cumulated rate of approximately one infection per patient-year . Coagulase-positive staphylococcus was the most common organism isolated.(ABSTRACT TRUNCATED AT 250 WORDS)

Arch Pediatr, 1994 Mar, 1(3), 249 - 54
{Acute respiratory infections in pediatric hospital at Bobo-Dioulasso (Burkina Faso)}; Tall FR et al.; BACKGROUND--Acute lower respiratory tract infections are the primary cause of morbidity in developing countries . POPULATION AND METHODS--Six hundred and sixty seven children (379 boys, 288 girls, aged 0-30 months) admitted for lower respiratory tract infections from January 1990 to March 1991 were included in the study . Immediate medical history was collected from the mother . The weight, height, temperature and clinical manifestations, plus the results of chest X-rays, parasitologic and bacteriological examination of stools, and blood smear for malaria were recorded for each patient . Sero-immunologic study for HIV infection of 473 of the patients aged 12-36 months and their mothers also took part in a sero-immunological study for HIV infection . RESULTS--Lower respiratory tract infections were the second major cause of admission (16.8%) after malaria (28.7%) . Infections peaked between 6 and 11 months of age (29.5%) . The main diseases were pneumonia and bronchial pneumonia (54%) followed by bronchiolitis (21.7%) . Almost half the patients were admitted during the hot, dry season . Two hundred and eighty seven patients (43.4%) were referred only after they had been suffering from the disease for 3 to 9 days, partly explaining the high level of mortality . One hundred and twenty one patients (20.9%) died; the main cause of death was staphylococcal pneumonia (57.9%), followed by pneumonia and bronchial pneumonia (29.3%) . Some criteria for severity could be identified, based on pulmonary signs and symptoms and associated manifestations (dehydration, malnutrition, convulsions, anemia) . Twenty two patients were positive for HIV-1 infection . CONCLUSIONS--This study confirms that acute lower respiratory tract infections remain a public health problem . Early diagnosis and treatment are necessary to reduce mortality.

Rev Rhum Ed Fr, 1994 Mar, 61(3), 153 - 65
{Septic arthritis in rheumatoid polyarthritis . 24 cases and review of the literature}; Dubost JJ et al.; Twenty-four cases of septic arthritis in rheumatoid arthritis patients were compared with 99 cases of septic arthritis in patients without rheumatoid arthritis . In addition, 238 previously published cases of septic arthritis with rheumatoid arthritis were analyzed . Fifteen percent of our patients with septic arthritis had rheumatoid arthritis, which was typically of long duration (mean 15 years), erosive, and seropositive . Fifty-four per cent (28% in the literature) and 9% of patients with and without rheumatoid arthritis, respectively, had pyarthrosis of multiple joints . The knee represented one-third of infected joints and the elbows and wrists were more often infected in patients with than without rheumatoid arthritis . S . aureus was recovered in 80% versus only 60% of patients with and without rheumatoid arthritis, respectively . The source of sepsis was often a skin lesion, in particular at the foot, emphasizing the need for early orthopedic treatment of deformities responsible for skin lesions . Monoarticular infection was more likely to be due to an intraarticular injection . Mortality rate was 17% in patients with rheumatoid arthritis (23% in the literature) versus 7% in patients without rheumatoid arthritis . Staphylococcal infection and infection of multiple joints were associated with higher mortality rates (35% and 49%, respectively) . The mortality rate in polyarticular infections has failed to decline over the last 35 years . Initial failure to distinguish septic arthritis from an exacerbation of rheumatoid arthritis contributes to the high mortality rate . The diagnosis of septic arthritis rests on a high index of suspicion . Septic arthritis cannot be ruled out based on absence of local inflammation, fever, or hyperleukocytosis or on presence of inflammation of multiple joints . Joint fluid specimens should routinely be sent to the microbiological laboratory and should be inoculated in blood culture bottles at the least suspicion.

Immunol Lett, 1994 Mar, 39(3), 223 - 9
Activation of human CD4+45RA+ T cells using B cells as accessory cells; Kristensson K et al.; Human naive CD4+ T cells, as defined by expression of CD45RA and lack of CD45R0, can be activated in vitro using B cells as accessory cells . CD4+CD45RA+ T cells proliferate, as determined by {3H}thymidine or bromodeoxyuridine (BrdU) incorporation, after activation with the superantigen staphylococcal enterotoxin A (SEA) presented by major histocompatibility complex class II-expressing B cells . The identity of the responding cells as being CD45RA+ and not contaminating CD45R0+ T cells was determined by FACS analysis, showing that purified CD45RA-expressing T-helper cells went into S phase and progressively acquired expression of the CD45R0 isoform while simultaneously losing expression of the CD45RA isoform . Cultivation of the CD4+ T-cell subsets under limiting dilution conditions supported these findings and revealed that (i) the frequency of responding cells in the CD45RA+ population was equal to or higher than in the CD45R0+ subset and (ii) that the number of CD45R0+ cells possibly contaminating the CD45RA population was too low to be able to account for the response observed.

Biosci Biotechnol Biochem, 1994 Mar, 58(3), 602 - 5
Inactivation of gamma-hemolysin H gamma II component by addition of monosialoganglioside GM1 to human erythrocyte; Ozawa T et al.; The Staphylococcal toxins leukocidin and gamma-hemolysin consist of two protein components: F and S in leukocidin and H gamma I and H gamma II in gamma-hemolysin . The two toxins share one component (F = H gamma I) . We found that the H gamma II component was completely inactivated by the addition of monosialoganglioside GM1 at the molar ratio of 1:1 . Disialogangliosides GD1a and GD1b had little effect on the inactivation of H gamma II . The molar ratios of GD1a and GD1b to H gamma II needed for maximum inactivation were 30:1 and 100:1, respectively . Related glycolipids caused little if any inactivation . H gamma II bound to GM1 to form H gamma II-GM1 complexes . Analysis of the intrinsic aromatic amino acid fluorescence in H gamma II and H gamma II-GM1 with 280 nm as the excitation wavelength showed that GM1 in the complex reduced the fluorescence intensity of H gamma II by 12% without changing the wavelength of maximum emission (325 nm) . We concluded that GM1 is a receptor of the H gamma II component on human erythrocytes and that H gamma II takes on a different conformation when it binds to GM1.

Eur J Immunol, 1994 Mar, 24(3), 651 - 8
Engagement of MHC class II molecules by staphylococcal superantigens activates src-type protein tyrosine kinases; Morio T et al.; Staphylococcal exotoxins (SE) are superantigens that bind to monomorphic determinants on major histocompatibility complex (MHC) class II molecules and stimulate human peripheral blood T lymphocytes in a V beta-specific manner . SE also deliver activation signals via MHC class II molecules that initiate cell adhesion and cytokine gene transcription . These events are preceded by tyrosine phosphorylation and are antagonized by inhibitors of tyrosine kinases, indicating an essential role for these kinases in signaling via class II molecules . We report that stimulation of human peripheral blood monocytes with SE induced rapid and selective activation of the src-related protein tyrosine kinases (PTK) fgr and hck . SE also induced the activation of fgr and lyn in B cells . PTK activation by SE required MHC class II expression, and was greatly potentiated in the presence of T cells bearing toxin-specific V beta chains . These results indicate that in addition to their antigen and superantigen-presenting function, MHC class II molecules act as signal-transducing receptors that are coupled to src-type PTK.

Clin Diagn Lab Immunol, 1994 Mar, 1(2), 227 - 31
Primary and secondary granule release by polymorphonuclear leukocytes exposed to peritoneal dialysis effluent; Daniels I et al.; Peritoneal dialysis effluent from patients with end-stage renal failure contains a low-molecular-weight solute that inhibits the killing of phagocytosed Staphylococcus epidermidis by polymorphonuclear leukocytes (PMN) . This observation has been investigated by using luciginen-enhanced chemiluminescence to measure PMN NADPH oxidase activity, CD11b/CD18 expression and lactoferrin release to measure secondary granule discharge, and cellular levels of beta-glucuronidase (EC 3.2.1.31) to measure changes in primary granules . Peritoneal dialysis effluent had no effect on the loss of intracellular beta-glucuronidase from normal unstimulated PMN or from PMN stimulated with S . epidermidis . It did, however, cause a concentration-dependent (0 to 70%; vol/vol) increase in expression of CD11b/CD18 and NADPH oxidase activity . CD11b/CD18 expression increased over 20 min before starting to plateau . Release of lactoferrin by the same cells demonstrated a strong positive correlation with integrin expression (P < 0.001, Spearman's rank correlation coefficient) . When dialysis effluent-treated PMN were stimulated with formyl-methionylleucylphenylalanine, integrin expression, release of lactoferrin, and NADPH oxidase activity were greater than in PMN treated with formyl-methionylleucylphenylalanine alone . Under these conditions, a concentration-dependent increase in CD11b/ CD18 and lactoferrin release were observed only at a concentration between 0 and 30% (vol/vol) dialysis effluent, while a concentration-dependent increase in oxidase activity was seen at a concentration between 0 and 70% (vol/vol) . The results suggest that dialysis effluent does not affect PMN primary granule release but does cause increased release of secondary granules and an increase in NADPH oxidase activity in both unstimulated and stimulated PMN.

J Immunol, 1994 Feb 15, 152(4), 2051 - 9
Superantigen modulation of experimental allergic encephalomyelitis: activation of anergy determines outcome; Racke MK et al.; Experimental allergic encephalomyelitis (EAE) is an autoimmune disease that can be induced by the adoptive transfer of CD4, myelin basic protein (MBP)-specific T cells . Superantigens activate T cells expressing appropriate TCR V genes . In this study, MBP-specific T cells activated in vitro with a superantigen, staphylococcal enterotoxin B (SEB), could adoptively transfer a severe form of EAE in (PLxSJL)F1 mice, but did not transfer disease in PL/J or SJL/J mice . SEB treatment of donor mice anergized MBP-specific T cells using V beta 8 in (PLxSJL)F1 mice, because subsequent in vitro activation with SEB resulted in a marked decrease in proliferation to SEB and inability to transfer EAE . However, donor cells from (PLxSJL)F1 mice immunized with MBP/CFA that had been exposed to SEB in vivo before MBP stimulation in vitro still produced EAE in recipient mice . To confirm that non-V beta 8 T cells could transfer disease, donor mice were treated with antibody that eliminated V beta 8 T cells; MBP-activated T cells from these mice could still transfer EAE . Finally, EAE induced by SEB-activated T cells was substantially reduced in mice receiving anti-V beta 8 therapy in vivo . The ability of superantigens to activate encephalitogenic T cells may have relevance to human diseases such as multiple sclerosis.

J Immunol, 1994 Feb 15, 152(4), 1674 - 83
Role of DNA fragmentation in T cell activation-induced apoptosis in vitro and in vivo; Mogil RJ et al.; Apoptotic cell death, characterized by DNA fragmentation and morphologic changes, has previously been shown to occur in immature thymocytes and some T cell hybridomas after activation . Like some other forms of apoptosis, DNA fragmentation during activation-induced cell death precedes the morphologic events . For apoptosis to proceed, activation of the cells must persist at least to the time of DNA fragmentation, before which the cells can remain viable if the activation signal is removed . Aurintricarboxylic acid (ATA) blocks activation-induced apoptotic cell death in a T cell hybridoma, and kinetic studies show that this inhibition occurs at or near the time of DNA fragmentation in the cells . Taken together with the ability of ATA to inhibit DNA fragmentation in isolated nuclei exposed to Ca2+ and Mg2+, these data strongly suggest that ATA prevents apoptosis via its ability to inhibit endogenous endonuclease activity, and, conversely, that this activity is required for this form of cell death . In vivo, ATA inhibits thymocyte depletion and DNA fragmentation induced by anti-CD3 Ab . Further, specific loss of V beta 8+ thymocytes after administration of staphylococcal enterotoxin B is blocked by administration of ATA . These observations support an essential role for DNA fragmentation as an irreversible step in activation-induced apoptosis in T cell hybridomas and during T cell development . This is contrasted with heat shock-induced cell death, in which inhibition of DNA fragmentation does not prevent loss of cell viability.

J Immunol, 1994 Feb 15, 152(4), 1641 - 52
IL-2, IL-4, and IFN-gamma gene expression versus secretion in superantigen-activated T cells . Distinct requirement for costimulatory signals through adhesion molecules; Lagoo AS et al.; In the complete absence of APCs staphylococcal superantigens induced IL-2, IL-4, IL-5, IFN-gamma, and IL-2R gene transcripts in both highly purified human T cells and FACs sorted CD4+ memory (CD45RA-) T cells . Secretion of IL-2, IL-4, and IFN-gamma, as well as DNA synthesis, on the other hand, required the presence of monocytes . At cytokine gene transcript level, three patterns of expression were noted after superantigen activation of T cells in the presence vs the absence of APC . mRNA levels for IL-2 were markedly up-regulated in the presence of monocytes, IL-4 and IFN-gamma transcripts increased only modestly, and IL-5 and IL-2R mRNA levels were unaffected . Blocking mAbs against LFA-1 and LFA-3 added to staphylococcal enterotoxin B (SEB)-activated cultures of T cells and autologous monocytes, reproducibly decreased both T cell proliferation and genetic expression of IL-2, IL-4, IL-5, and IL-2R, although having little or no effect on IFN-gamma transcripts . Further, under those conditions of blocking, secretion of IL-2 and IL-4 was dramatically decreased, whereas IFN-gamma secretion remained essentially unchanged . In contrast, LFA-1 and LFA-3 mAbs completely abrogated IFN-gamma secretion from PHA-activated T cell-monocyte mixtures, although having no inhibitory effect on T cell proliferation . These results indicate a characteristic and differential involvement of adhesion molecule-mediated signals in superantigen-induced T cell proliferation, differential cytokine gene expression, and cytokine secretion.

Biochemistry, 1994 Feb 8, 33(5), 1063 - 72
Structure and dynamics of a denatured 131-residue fragment of staphylococcal nuclease: a heteronuclear NMR study; Alexandrescu AT et al.; A partially folded form of staphylococcal nuclease has been obtained by deleting residues 4-12 and 141-149 of the 149-residue wild-type protein . Sequence-specific NMR resonance assignments have been obtained for 106 of the 131 residues in this protein fragment by using multi-dimensional triple resonance NMR of samples enriched with 13C and 15N . Residues corresponding to helix 2 (residues 98-106) and helix 1 (residues 54-68) of the native state give chemical shifts and NOE effects characteristic of helical structure . These same residues, however, give coupling constants and NOE effects indicative of fast conformational averaging between helical and extended conformations . The residual helix structure observed in the nuclease fragment is thus considerably less persistent than the corresponding structure in the native state . Based on H alpha chemical shifts, we estimate the fractional population of helical conformers to be 30% for helix 2 and 10% for helix 1 . Two segments, 83-86 and 94-97, show NOE effects, coupling constants, and lowered amide temperature coefficients consistent with a native-like reverse-turn structure . The C-terminal alpha-helix as well as the fourth and fifth strands of the 5-strand beta-barrel show little evidence for ordered structure . The first three strands of the beta-sheet, part of the catalytic loop, and the first turn of helix 3 give significantly poorer NMR data than the rest of the protein, possibly as a result of exchange broadening, and could not be characterized in detail . That the most persistent elements of structure in the fragment are native-like suggests that nuclease may fold by a hierarchical mechanism.

Biochemistry, 1994 Feb 1, 33(4), 1029 - 36
Hydrogen bonding in proteins as studied by amide hydrogen D/H fractionation factors: application to staphylococcal nuclease; Loh SN et al.; The D/H fractionation factor (sigma) is the extent to which a hydrogen at a particular site becomes enriched in 2H over 1H relative to the solvent . A growing body of experimental evidence suggest that there is a correlation between the value of the fractionation factor and hydrogen-bond strength, with a lower sigma value reflecting a stronger hydrogen bond . Fractionation factors of 60% of the individual backbone amide hydrogens in the staphylococcal nuclease V8 variant (H124L) have been measured for the enzyme in the presence and absence of bound ligands (the activating ion Ca2+ and the inhibitor thymidine 3',5'-bisphosphate) . The method used employed two-dimensional 1H-15N nuclear magnetic resonance analysis of uniformly 15N-labeled protein in mixed H2O/D2O solvents . Fractionation factors of individual residues were found to range from 0.3 (T120) to 1.5 (L38) . The sigma value of 0.3 for the NH of T120, which is the lowest fractionation factor reported for any system yet studied, suggests that the hydrogen bond between T120 HN and D77 O delta 1 is unusually strong . The results of previous site-directed mutagenesis experiments {Hinck, A . P . (1993) Ph.D . Thesis, University of Wisconsin-Madison, Madison, WI} support the notion that formation of this hydrogen bond is important to maintain the stability and conformation of the native state . The sigma value averaged over all residues was approximately 0.85 for both the unligated and ligated enzymes . Residues in alpha-helices displayed a slightly lower average sigma value (0.79), whereas residues with solvent-exposed amide hydrogens exhibited a slightly higher average figure (0.98).(ABSTRACT TRUNCATED AT 250 WORDS)

Am Surg, 1994 Feb, 60(2), 138 - 42
Outcome of isolated pulmonary contusion in blunt trauma patients; Hoff SJ et al.; To determine outcome in young, healthy blunt trauma patients with isolated pulmonary contusion, and to identify factors associated with poor outcome, we reviewed 6012 consecutive adult (aged 16-49) blunt trauma admissions . Ninety-four (7.9%) presented with an isolated pulmonary contusion defined by chest radiograph and Injury Severity Score < 25; they compromise the study group . Poor outcome was defined as death, prolonged hospitalization (> 7 days), or a severe complication (pneumonia, empyema, atelectasis requiring bronchoscopy, or bronchopleural fistula) . None of the 94 study patients died . Admission chest radiograph demonstrated no contusion in 34 patients (36%) . Fifteen patients (16%) required intubation, but 13 were extubated within 48 hours . Forty-one patients (44%) required insertion of a chest tube, and 20 patients (21%) had a PaO2/FiO2 ratio of < 250 on admission . Post-injury atelectasis (n = 17), pneumothorax (n = 17), effusion (n = 8), pneumonia (n = 2), empyema (n = 1), and Staphylococcal bacteremia (n = 1) complicated hospitalizations . The following clinical factors were identified as predisposing to poor outcome by univariate analysis: 1) Pulmonary contusion on admission chest radiograph (P = 0.035); 2) Three or more rib fractures (P = 0.002); 3) chest tube insertion (P = 0.010) and drainage (P = 0.020); and 4) hypoxia on admission (PO2 < 70 torr {P = .021}, PaO2/FiO2 < 250 {P < 0.001}) . Only PaO2/FiO2 < 250 on admission was an independent predictor of poor outcome in a multivariate analysis (P = 0.040) . Our conclusion was that isolated pulmonary contusion in young, healthy patients is not associated with mortality.(ABSTRACT TRUNCATED AT 250 WORDS)

J Immunol, 1994 Feb 1, 152(3), 1277 - 88
Stimulation of tumor-draining lymph node cells with superantigenic staphylococcal toxins leads to the generation of tumor-specific effector T cells; Shu S et al.; In animal studies, lymph nodes (LN) draining progressive tumors contain immunologically sensitized but functionally deficient T cells . These preeffector cells can differentiate into mature effector cells on stimulation in vitro with anti-CD3 and IL-2 . However, anti-CD3 react with all T cells and the activated cell population expressed a broad but normal distribution of V beta phenotypes . In this study, we examined the feasibility of using bacterial superantigens to stimulate tumor-draining LN cells . Because of their TCR V beta restriction, superantigen activation may afford a means to identify T cell subsets that are important in the antitumor immune response . Stimulation of draining LN cells with staphylococcal enterotoxins A (SEA) or B (SEB) followed by culture in IL-2 resulted in selective activation and expansion of V beta 3 and V beta 11 or V beta 3 and V beta 8 T cells, respectively . However, in adoptive immunotherapy, SEB- but not SEA-activated cells mediated the regression of established pulmonary metastases . To define the relative antitumor effects of V beta 3 and V beta 8 T cells, SEB-activated cells were depleted of either V beta 3 or V beta 8 T cells with mAb and magnetic beads . The antitumor effects were demonstratably diminished after V beta 8 cell depletion but enhanced after V beta 3 cell depletion . Using antigenically distinct MCA 205 and 207 sarcomas, tumor regression mediated by the activated cells was found to be immunologically specific for the tumor that stimulated the draining LN . Furthermore, the SEB-activated cells were virtually all T cells consisting of approximately equal proportions of CD4+ and CD8+ cells and the collaboration of the two T cell subsets was required for in vivo antitumor effects . However, the helper function of CD4+ cells could be facilitated by the administration of exogenous IL-2 . Despite their in vivo antitumor reactivity, SEB-activated cells did not exhibit tumor cytotoxicity in the 4-h 51Cr release assay . However, they secreted IFN-gamma on specific stimulation with tumor cells . Taken together, these results provide for the first time clear evidence of the functional significance of superantigen interactions with immunologically committed T cells and suggest a preferential V beta use that might be associated with the T cell immune response to progressively growing tumors.

J Immunol, 1994 Feb 1, 152(3), 1022 - 31
Thymic epithelial cell lines that mediate positive selection can also induce thymocyte clonal deletion; Hugo P et al.; Negative selection of potentially autoreactive thymocytes occurs mainly in the thymus and is thought to be induced primarily by interaction with bone marrow-derived cells . However, some studies have also reported a role for radioresistant thymic cells, which are probably epithelial in origin, in the deletion of thymocytes reacting to endogenous superantigens . We have previously demonstrated that thymic epithelial cell lines could induce thymocyte-positive selection in vivo . In this study, we assessed the potential of these cells to delete thymocytes reacting to the staphylococcal enterotoxin A or B superantigens in vitro . In the presence of staphylococcal enterotoxin A or B we found that all thymic epithelial cell lines used in this study were capable of activating T cell hybrids or deleting CD4+CD8+ thymocytes expressing an appropriate TCR . The extent of superantigen-mediated thymocyte deletion mediated by thymic epithelial cell lines was comparable to that mediated by a thymic macrophage cell line . Similar results were obtained with three phenotypically distinct thymic cell lines, suggesting that the ability to induce thymocyte deletion might be a general feature of various subsets of thymic epithelium . The observations provided in this study, combined with our previous demonstration that the same thymic epithelial cell lines can participate in positive selection, suggest that a given stromal cell population might be capable of taking part both in positive and negative selection of thymocytes.

Infect Immun, 1994 Feb, 62(2), 462 - 7
Differential induction of tumor necrosis factor alpha in murine and human leukocytes by Mycoplasma arthritidis-derived superantigen; Rink L et al.; Mycoplasma arthritidis-derived superantigen (MAS) is exclusively produced by M . arthritidis, which is the only known mycoplasma to produce a superantigen . As a superantigen, MAS shows properties similar to those of the staphylococcal enterotoxins and related substances, such as binding to major histocompatibility complex (MHC) class II and V beta-specific stimulation of T cells . In this series of experiments, we demonstrate some differences between MAS and other superantigens . MAS induced the production of tumor necrosis factor alpha (TNF-alpha) mRNA in human as well as in murine leukocytes . However, only in murine leukocytes was the mRNA adequately translated into the protein . In human peripheral blood mononuclear cells, we found only small amounts of TNF, whereas in murine spleen cells we detected levels more than three times higher . The proliferative response to MAS has been shown to be restricted to I-E alpha in the murine MHC . Furthermore, TNF was induced in I-E alpha+ bone marrow-derived macrophages by MAS . In these cells, MAS rapidly induced very high levels of TNF and the amounts of mRNA detected correlated to the amount of protein produced . In comparison with other superantigens, including the staphylococcal enterotoxins, toxic shock syndrome toxin 1, and exfoliative toxin A, the failure of MAS to induce TNF-alpha in human peripheral blood mononuclear cells is specific for MAS and not common to all superantigens . The direct activation of bone marrow-derived macrophages also seems to be specific for MAS . These data suggest that the induction of TNF-alpha by MAS is dependent on the strength of binding to the MHC class II molecule.

Infect Immun, 1994 Feb, 62(2), 421 - 5
Induction of muscle-relaxing factor by staphylococcal alpha-toxin; Harshman S et al.; Brain tissue and serum from mice intracerebrally injected with 1 microgram of staphylococcal alpha-toxin contained elevated amounts of a naturally occurring brain tissue component(s) called muscle-relaxing factor (MRF) . MRF induced reversible, generalized, flaccid paralysis of mice after intracerebral but not intraperitoneal or intravenous administration . MRF (i) was soluble in Hanks balanced salt solution and in acidified (pH 2) Hanks balanced salt solution, in which it partitions into ethyl acetate, acetone, and methanol; (ii) was separated from some pigments by thin-layer chromatography on silica gel plates; (iii) did not comigrate with prostaglandin and leukotriene standards during high-pressure liquid chromatography with a mu Bondapak fatty acid column; and (iv) did not contain amino acids, exhibit absorption maxima at a wavelength range of 210 to 600 nm, or fluoresce when exposed to UV light . MRF has been detected in rabbit brain that has been stored frozen at -70 degrees C and has been enhanced in vitro in slices of both mouse and rabbit brain following incubation of the brain slices with staphylococcal alpha-toxin . Studies to identify the chemical nature of MRF and the mechanism by which, in mice, it induces reversible, flaccid paralysis of voluntary muscle are continuing.

Gastroenterology, 1994 Feb, 106(2), 450 - 8
Hepatic injury and lethal shock in galactosamine-sensitized mice induced by the superantigen staphylococcal enterotoxin B; Nagaki M et al.; BACKGROUND/AIMS: Staphylococcal enterotoxin B (SEB) acts as a superantigen binding to class II major histocompatibility complex proteins, and this complex stimulates T cells . The aim of this study was to investigate the pathogenic effects of SEB on hepatic injury and lethal shock in mice . METHODS: SEB was administered to D-galactosamine (GalN)-sensitized mice, and the degree of liver injury and levels of circulating cytokines were determined . In vitro cytokine production in response to SEB was also investigated . RESULTS: Intraperitoneal administration of SEB (50 micrograms) caused lethal shock (50% mortality) associated with massive hepatic necrosis in GalN-sensitized mice, with no mortality on injection of up to 100 micrograms SEB alone . Within 2 hours after injection of SEB, serum tumor necrosis factor alpha (TNF-alpha) levels reached a peak, followed by high levels of serum interferon-gamma (IFN-gamma) up to 10 hours after injection . Passive immunization with anti-TNF-alpha/beta-neutralizing monoclonal antibody (mAb) protected GalN-sensitized mice from the lethal effects of SEB, with less protection with anti-IFN-gamma-neutralizing mAb . SEB induced the production of TNF-alpha and IFN-gamma in a dose-dependent manner from splenic mononuclear cells in vitro . CONCLUSIONS: The results show that SEB contributes to lethal shock associated with severe hepatic injury in GalN-sensitized mice and suggest that TNF-alpha and IFN-gamma produced in response to SEB may be mediators of the lethal toxicity and hepatotoxicity of SEB.

Eur J Immunol, 1994 Feb, 24(2), 445 - 9
Differential effects of superantigen-induced "anergy" on priming and effector stages of a T cell-dependent antibody response; Lussow AR et al.; The in vitro T cell nonresponsiveness or anergy to restimulation with staphylococcal enterotoxin B (SEB) following the in vivo injection of the superantigen is well characterized . Here we use mice transgenic for a V beta 8.2+ T cell receptor (TcR) (reactive with SEB) to establish a large population of anergic T cells in vivo . As expected, peripheral T cells from the SEB injected transgenic mice failed to proliferate or produce interleukin (IL)-2 following restimulation with the superantigen in vitro . However, in this system superantigen reactivity could be restored by either addition of exogenous IL-2, or stimulation with immobilized anti-TcR antibody . To evaluate the effects of superantigen-induced anergy in vivo, SEB-injected or noninjected control transgenic mice were immunized and boosted with the T cell-dependent antigen tetanus toxin (TT) . SEB injection of the V beta 8.2+ transgenic mice 5 days prior to the TT immunization inhibited the anti-TT antibody response as measured over a 100-day period, whereas injection of a superantigen which does not interact with the V beta 8.2% TcR (such as SEA) did not . Furthermore, SEB injection of control nontransgenic mice did not interfere with the induction of a high titer anti-TT antibody response . In contrast to the inhibition seen when SEB was given prior to TT immunization, injection of transgenics with SEB either after the priming TT immunization or after the recall booster injection did not significantly influence the titers of anti-TT antibodies produced . These results demonstrate that the establishment of peripheral T cell anergy to superantigens inhibits the specific antigenic priming of helper T cells in vivo, but does not prevent primed T cells from helping B cells to mount an effective antibody response.

Eur J Immunol, 1994 Feb, 24(2), 355 - 61
A peptide binding weakly to the major histocompatibility molecule augments T cell responses; Feng MH et al.; An I-A(d)-derived peptide PB1 was found to enhance the reactivity of I-A(d)-restricted T cells . The augmentative effect was not due to the cross-reactivity of PB1 peptide with antigens . PB1 had no effect on T cells specific for I-A(b) and I-E(k), nor did PB1 increase the T cell responses to concanavalin A and staphylococcal enterotoxin B . The strict I-A(d) specificity suggests that PB1 enhances the recognition of antigen-I-A(d) complex by T cell receptor . PB1 bound to I-A(d) weakly . The augmentative effect could be found on other I-A(d)-binding peptides in appropriate conditions; however, PB1 was distinct in its prominently augmentative effect on all the I-A(d)-restricted T cells analyzed . A similar enhancing activity was demonstrated on a synthetic transferrin receptor peptide with minimum affinity for I-A(d) . The unusual enhancing activity of PB1 may thus be attributed to the low I-A(d) binding affinity . It was postulated that the binding of low-affinity PB1 would not only stabilize I-A(d) structure, but also enhance the binding of other peptides . This was supported by the increased binding of OVA 323-339 and cI 84-98 to I-A(d) in the presence of PB1 . The inclusion of PB1 in the immunization mixture also enhanced T cell responses in vivo, suggesting the possibility of using low-affinity peptide to promote specific immunity.

Clin Immunol Immunopathol, 1994 Feb, 70(2), 137 - 44
Early activation events induced by the staphylococcal superantigen toxic shock syndrome toxin-1 in human peripheral blood monocytes; Trede NS et al.; Staphylococcal exotoxins (SE) bind to MHC class II molecules and induce the production of IL-1 beta and TNF-alpha in human monocytic cells . Here we show that stimulation of peripheral blood monocytes with toxic shock syndrome toxin-1 (TSST-1) induced rapid increase in tyrosine phosphorylation of cytosolic protein substrates, accumulation of inositol phosphates, and de novo tyrosine phosphorylation of the PLC-gamma 1 isozyme . Accumulation of inositol phosphates was inhibited by preincubation of cells with inhibitors of protein tyrosine kinases (PTK) . Stimulation of monocytes with TSST-1 furthermore led to activation of protein kinase C (PKC) . PTK and PKC activation plays a role in the induction of monokine gene transcription by SE because inhibitors of PTK and PKC reduced TSST-1-stimulated IL-1 beta and TNF-alpha gene expression . We therefore propose a model in which the induction of monokine gene transcription by TSST-1 in monocytes necessitates activation of tyrosine kinase(s) and of PKC, the latter probably by way of PLC-gamma 1.

J Biochem (Tokyo), 1994 Feb, 115(2), 182 - 9
Cell-adhesive activity and receptor-binding specificity of the laminin-derived YIGSR sequence grafted onto Staphylococcal protein A; Maeda T et al.; Laminin contains multiple oligopeptide motifs to promote cell adhesion and migration . One of these motifs is YIGSR within the B1 chain . We reconstituted the cell-adhesive activity of YIGSR motif by grafting it onto a truncated form of the Staphylococcal protein A (designated tSPA) via cassette mutagenesis . When coated on a polystyrene surface, the YIGSR-grafted tSPA (YIGSR-tSPA) promoted attachment and spreading of mouse melanoma and human rhabdomyosarcoma cells, but not of hamster fibroblasts . The cell-adhesive activity of YIGSR-tSPA was abolished by amino acid substitution or scrambling of the inserted YIGSR sequence . Divalent cations Mn2+ and Mg2+, but not Ca2+, promoted the cell adhesion to YIGSR-tSPA . Interestingly, the YIGSR-tSPA-mediated cell adhesion was barely inhibited by the linear peptide CDPGYIGSR-NH2, but was strongly inhibited by the cyclic peptide CDPGYIGSRC and another peptide PEILDVPST, which is a specific inhibitor for integrin alpha 4 beta 1 . Among various anti-integrin antibodies, anti-alpha 4 and anti-beta 1 antibodies specifically inhibited the cell adhesion to YIGSR-tSPA . In support of these observations, adhesion of rhabdomyosarcoma cells to intact laminin was also partially inhibited by synthetic PEILDVPST peptide and anti-alpha 4 antibody . These results, taken together, indicate that the YIGSR motif exerts its cell-adhesive activity through interaction with integrin alpha 4 beta 1.

Semin Immunol, 1994 Feb, 6(1), 27 - 37
Defective clonal deletion and anergy induction in TCR transgenic lpr/lpr mice; Mountz JD et al.; Although autoreactive T cells are thought to play a prominent role in autoimmune disease in MRL-lpr/lpr mice, it has been difficult to directly determine if autoreactive T cells escape from the thymus and react with self-antigens in the periphery . To identify a possible defect in clonal deletion or clonal anergy induction of auto-specific T cells, we have studied C57BL/6-lpr/lpr transgenic mouse expressing TCR genes that recognize a known self-antigen, the male H-Y antigen and analyzed clonal deletion and tolerance induction after neonatal tolerance induction with the class II MHC reactive superantigen staphylococcal enterotoxin B (SEB) in V beta 8 TCR transgenic and non-transgenic MRL-lpr/lpr mice . In lpr/lpr mice, the main defect was a thymic-dependent loss of self-tolerance by auto-specific T cells and also a small defect of clonal deletion of autoreactive thymocytes . Defective expression of the Fas apoptosis antigen results from the insertion of the ETn retrotransposon . The fas defect can be partially corrected in TCR-beta TCR transgenic mice in which accelerated T cell development prevents the lymphoproliferative disease and in CD2-fas transgenic mice in which fas expression is corrected in T cells . These results suggest that expression of the TCR-beta, fas and retrovirus genes are co-regulated during early thymocyte development, most likely by common enhancer transcription factors.

Biophys J, 1994 Feb, 66(2 Pt 1), 482 - 501
The use of fluorescence methods to monitor unfolding transitions in proteins; Eftink MR; This article discusses several strategies for the use steady-state and time-resolved fluorescence methods to monitor unfolding transitions in proteins . The assumptions and limitations of several methods are discussed . Simulations are presented to show that certain fluorescence observables directly track the population of states in an unfolding transition, whereas other observables skew the transition toward the dominant fluorescing species . Several examples are given, involving the unfolding of Staphylococcal aureus nuclease A, in which thermodynamic information is obtained for the temperature and denaturant induced transitions in this protein.

Int Immunol, 1994 Feb, 6(2), 189 - 97
IL-2 promoter-driven lacZ expression as a monitoring tool for IL-2 expression in primary T cells of transgenic mice; Brombacher F et al.; A transgenic mouse system has been established to follow the pattern of IL-2 expression at the level of single T cells . This was achieved by introducing a human IL-2 promoter-driven reporter gene (Escherichia coli lacZ) into the germline of mice and monitoring its product, beta-galactosidase (beta-gal), by FACS analysis . Ex vivo experiments confirmed that the regulated expression of the transgene is comparable with that of the endogenous IL-2 gene . Transgene expression is inducible by mitogens, restricted to T cells, and diminished by immunosuppressive agents, such as cyclosporin A, at concentrations known to suppress IL-2 transcription . Depending on the mitogens used, 30-50% of peripheral T cells produced IL-2 with an asynchronous induction pattern, as measured by transgenic beta-gal activity . Both helper (CD4+CD8-) and cytotoxic T cells (CD4-CD8+) respond with comparable heterogenous expression levels but they show different frequencies of beta-gal production . Transgenic beta-gal-producing T cells were detectable as early as 2 h after mitogen stimulation . These cells represent a transitional IL-2 secreting, IL-2 receptor alpha-chain negative T cell population, which occurs in the autocrine process of T cell activation . Administration of staphylococcal enterotoxin A (SEA), a bacterial superantigen, resulted in a T cell specific (Thy-1.2) increase (2.5-fold) of reporter gene expression in vivo . In summary, we could demonstrate that IL-2 promoter-driven reporter gene expression in transgenic mice is a sensitive tool to characterize IL-2 expressing cells phenotypically.(ABSTRACT TRUNCATED AT 250 WORDS)

J Clin Microbiol, 1994 Feb, 32(2), 388 - 92
Clotting activity in Staphylococcus schleiferi subspecies from human patients; Vandenesch F et al.; Staphylococcus schleiferi subsp . schleiferi is a coagulase-negative staphylococcus, usually present as a contaminant in human specimens . A near relative, S . schleiferi subsp . coagulans, possesses coagulase activity but has not been reported from humans . We here describe three isolates of pseudocoagulase-positive S . schleiferi subsp . schleiferi and one isolate of S . schleiferi subsp . coagulans from human patients . The pseudocoagulase from the S . schleiferi subsp . schleiferi isolates differs from S . aureus staphylocoagulase by being sensitive to a combination of protease inhibitors (aprotinin, N-ethylmaleimide, and heparin) . These isolates could all easily be confused with S . aureus in a typical clinical laboratory, since they all possess a heat-stable DNase and promote clotting formation . Moreover, S . schleiferi subsp . coagulans produces protein A, and S . schleiferi subsp . schleiferi expresses a clumping factor (fibrinogen affinity factor) . Southern blot hybridization with an S . aureus coa-specific probe revealed no sequence related to the coa gene in any of the S . schleiferi isolates, and their riboprobe profiles and biochemical characteristics were typical of S . schleiferi subspecies, not of S . aureus . This study demonstrates that both subspecies of S . schleiferi can promote clotting of rabbit plasma in the standard tube test for coagulase.

Appl Environ Microbiol, 1994 Feb, 60(2), 752 - 5
Staphylococcin 1580 is identical to the lantibiotic epidermin: implications for the nature of bacteriocins from gram-positive bacteria; Sahl HG; Staphylococcin 1580 was purified to homogeneity from culture supernatants of Staphylococcus epidermidis 1580 by means of adsorption to XAD 2, cation exchange chromatography, and high-performance liquid chromatography on reversed-phase C18 . The purified active substance migrated in sodium dodecyl sulfate-polyacrylamide gel electrophoresis with an apparent M(r) of approximately 2,000 . Amino acid analysis, mass determination (2,165 Da) and N-terminal sequencing (Ile-Ala-Xaa-Lys-Phe-Ile-Xaa-Xaa-Pro-Gly-Xaa-Ala-Lys-block) demonstrated that staphylococcin 1580 is identical to epidermin, a lanthionine-containing antibiotic peptide (lantibiotic).

Appl Environ Microbiol, 1994 Feb, 60(2), 677 - 81
Evaluation of a commercial enzyme immunoassay kit (RIDASCREEN) for detection of staphylococcal enterotoxins A, B, C, D, and E in foods; Park CE et al.; The RIDASCREEN SET kit (R-Biopharm GmbH, Darmstadt, Germany), a commercial staphylococcal enterotoxin (SE) visual immunoassay kit, was evaluated for its efficacy . The kit utilizes monovalent capture antibodies against SE types A to E (SEA to SEE); therefore, it simultaneously detects and identifies the enterotoxin types . The major advantages of the kit are (i) a high degree of specificity (except for naturally occurring peroxidases, food compositions or ingredients and microbiological products due to growth of nonstaphylococcal microorganisms did not cause false-positive results; additionally, no cross-reactions among reagents of the kits were observed), (ii) excellent sensitivity (minimum detectable limits were 0.20 to 0.30 ng of SEs per ml of extracts of ham, salami, and mushroom and 0.30 to 0.35 ng of SEs per ml of cheese extracts, or 0.50 to 0.75 ng of SEs per g of foods such as noodles, ham, salami, cheese, and turkey), (iii) simplicity (the kit enabled direct assay of SEs in food extracts without the need for lengthy extraction or concentration procedures), (iv) rapidity (it took less than 3 h to complete the analysis of individual enterotoxin types SEA to SEE), and (v) its semiquantitative results (optical density values could be read against a standard curve to estimate the amount of SE in the extract) . The RIDASCREEN kit is a convenient, rapid, and reliable tool for the detection and identification of SEs in foods.

Mol Gen Genet, 1994 Feb, 242(4), 421 - 30
Genetic and biochemical properties of an extracellular neutral metalloprotease from Staphylococcus hyicus subsp . hyicus; Ayora S et al.; The gene encoding the extracellular neutral metalloprotease ShpI from Staphylococcus hyicus subsp . hyicus was cloned . DNA sequencing revealed an ORF of 1317 nucleotides encoding a 438 amino acid protein with Mr of 49,698 . When the cloned gene was expressed in Staphylococcus carnosus, a 42 kDa protease was found in the culture medium . The protease was purified from both S . carnosus (pCAshp1) and S . hyicus subsp . hyicus . The N-terminal amino acid sequences of the two proteases revealed that ShpI is organized as a pre-pro-enzyme with a proposed 26 amino acid signal peptide, a 75 amino acid hydrophilic pro-region, and a 337 amino acid extracellular mature form with a calculated Mr of 38,394 . The N-termini showed microheterogeneity in both host strains . ShpI had a maximum proteolytic activity at 55 degrees C and pH 7.4-8.5 . The protease, which had a low substrate specificity, could be inhibited by metal- and zinc-specific inhibitors, such as EDTA and 1,10-phenanthroline . Insensitivity to phosphoramidon separates ShpI from the thermolysin-like family . The conserved Zn2+ binding motif, the only homology to other proteases, and the reactivation of the apoenzyme by Zn2+, indicated that Zn2+ is the catalytic ion . Ca2+ very probably acts as a stabilizer . We also demonstrated the presence of a second extracellular protease in S . hyicus subsp . hyicus.

FASEB J, 1994 Feb, 8(2), 237 - 46
Cytolysis mediated by ionophores and pore-forming agents: role of intracellular calcium in apoptosis; Duke RC et al.; Apoptosis is a term used to describe certain forms of physiological cell death that occur during embryogenesis, differentiation, and normal cell turnover . Previous reports concerning the effects of calcium ionophores on rodent thymocytes and the pore-forming proteins perforin and staphylococcal alpha-toxin on murine tumor cells led to the suggestion that simply raising intracellular calcium causes apoptotic cell death . This hypothesis was tested using two ionophores, A23187 and valinomycin, and two pore-forming agents, melittin and staphylococcal alpha-toxin, on four murine tumor cell lines . Although treatment with these agents could raise intracellular calcium, and in some instances cause DNA fragmentation, only valinomycin caused apoptosis . In contrast to previous reports, our results suggest that raising intracellular calcium and inducing internucleosomal DNA fragmentation is not sufficient to elicit apoptotic cell death in all cell types.

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