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