Vet Microbiol, 1994 Dec, 42(4), 265 - 71 Evaluation of protein A and protein G as indicator system in an ELISA for detecting antibodies in mink to Pseudomonas aeruginosa; Rivera E et al.; A modified, indirect enzyme-linked immunosorbent assay (ELISA) was developed and applied in the detection of mink antibodies to Pseudomonas aeruginosa . In this assay, peroxidase conjugated protein A and protein G were evaluated as indicator systems for detecting antigen-antibody complexes . It was found that protein A has a strong affinity for mink immunoglobulins . In contrast, protein G showed no such affinity . The affinity of protein A for mink immunoglobulins was further demonstrated by immunoprecipitation assays. J Bacteriol, 1994 Dec, 176(24), 7532 - 42 The vfr gene product, required for Pseudomonas aeruginosa exotoxin A and protease production, belongs to the cyclic AMP receptor protein family; West SE et al.; The synthesis of exotoxin A (ETA) by Pseudomonas aeruginosa is a complex, regulated event . Several ETA putative regulatory mutants of P . aeruginosa PA103 have previously been characterized (S . E . H . West, S . A . Kaye, A . N . Hamood, and B . H . Iglewski, Infect . Immun . 62:897-903, 1994) . In addition to ETA production, these mutants, PA103-15, PA103-16, and PA103-19, were also deficient in the production of protease and in regA P1 promoter activity . RegA is a positive regulator of ETA transcription . We cloned a gene, designated vfr for virulence factor regulator, that restored ETA and protease production to parental levels in these mutants . In addition, transcription from the regA P1 promoter was restored . In Escherichia coli, when vfr was overexpressed from a phage T7 promoter, a protein with an apparent molecular mass of 28.5 kDa was produced . Analysis of the deduced amino acid sequence of vfr revealed that the expected protein is 67% identical and 91% similar over a 202-amino-acid overlap to the E . coli cyclic AMP receptor protein (CAP or Crp) . The cloned vfr gene complemented the beta-galactosidase- and tryptophanase-deficient phenotypes of E . coli RZ1331, a crp deletion mutant . However, the E . coli crp gene under the control of the tac promoter did not complement the ETA-deficient or protease-deficient phenotype of PA103-15 or PA103-16 . The ability of vfr to restore both ETA and protease production to these mutants suggests that vfr is a global regulator of virulence factor expression in P . aeruginosa. Biochem J, 1994 Dec 1, 304 ( Pt 2), 493 - 7 Structural heterogeneity of Pseudomonas aeruginosa bacterioferritin; Moore GR et al.; The subunit composition, amino acid sequence and haem-binding characteristics of bacterioferritin (BFR) from Pseudomonas aeruginosa have been studied . Unlike other BFRs, P . aeruginosa BFR was found to contain two subunit types, designated alpha and beta, which differed considerably in their amino acid sequences . The N-terminal 69 and 55 amino acids of the alpha and beta subunits respectively were determined . The alpha subunit differed most from other BFRs . The two subunits were present in variable proportions in different preparations . The maximum stoichiometry of haem binding was found to be sample-dependent and to be different from the previously reported one per subunit {Kadir and Moore (1990) FEBS Lett . 271, 141-143} . This previous haem-binding study was shown to have been carried out with damaged protein, which contained both normal alpha and beta subunits and shorter versions of these that appeared to have been produced by cleavage of the normal subunits . The possibility that aging processes degrade ferritins and affect their haem-binding characteristics is discussed. J Infect Dis, 1994 Dec, 170(6), 1616 - 21 Epidemiology of chronic Pseudomonas aeruginosa infections in cystic fibrosis; Romling U et al.; The epidemiology of chronic colonization of airways with Pseudomonas aeruginosa was monitored in 44 patients with cystic fibrosis (CF) by DraI/SpeI macrorestriction analyses of 489 isolates . Sequential P . aeruginosa isolates (144) that had been collected from 32 CF patients over < or = 2.5 years were investigated, and 12 patients were followed for 8 years after onset of colonization . Forty-eight different genotypes were uncovered from 481 typeable isolates . Ten genotypes were found in > 1 unrelated CF patient . The 6 most frequent clones were identified in 58% of isolates . Ten of the 12 patients monitored for 8 years were harboring their initially acquired P . aeruginosa clone at all times, with subtle shifts of fragment patterns indicating subclonal variation . During colonization, the bacteria gradually lost pyocin and phage typing responses, supporting the view that genotypically discordant P . aeruginosa strains develop a common phenotype. J Infect Dis, 1994 Dec, 170(6), 1473 - 82 Protective effects of an N-terminal fragment of bactericidal/permeability-increasing protein in rodent models of gram-negative sepsis: role of bactericidal properties; Ammons WS et al.; Effects of an N-terminal fragment of bactericidal/permeability increasing protein (rBPI21) on bacterial infections were determined . Intravenous (i.v.) rBPI21 increased survival and reduced bacteremia in rats after an iv injection of Escherichia coli O7:K1 bacteria . rBPI21 inhibited the rise in tumor necrosis factor-alpha resulting from challenge with 2 strains of E . coli . Intraperitoneal (ip) injection of rBPI21 increased survival of mice after ip injection of E . coli O7:K1 and Pseudomonas aeruginosa and reduced bacteria in peritoneal lavage fluid and blood and inhibited cytokine production in response to E . coli . rBPI21 alone did not protect mice challenged with E . coli O111:B4 but was protective and reduced bacterial counts when administered in combination with the antibiotic cefamandole . The data show that protection with rBPI21 is associated with reductions in bacterial counts and is enhanced by antibiotics . Bactericidal activity, in addition to antiendotoxin activity, is involved in the efficacy of rBPI21 in models of gram-negative infection. Infect Immun, 1994 Dec, 62(12), 5456 - 63 Pseudomonas aeruginosa selective adherence to and entry into human endothelial cells; Plotkowski MC et al.; The pathogenesis of Pseudomonas aeruginosa disseminated infections depends on bacterial interaction with blood vessels . We have hypothesized that in order to traverse the endothelial barrier, bacteria would have to adhere to and damage endothelial cells . To test this hypothesis, we studied the adherence to human endothelial cells in primary culture of the piliated P . aeruginosa strain PAK and of two isogenic nonpiliated strains: PAK/p-, which carries a mutation in the pilin structural gene, and PAK-N1, a mutant defective in the regulatory rpoN gene . PAK adhered significantly more than did the pilus-lacking strains . P . aeruginosa was also taken up by endothelial cells, as determined by quantitative bacteriologic assays and by transmission electron microscopy . This internalization of P . aeruginosa seems to be a selective process, since the piliated strain was taken up significantly more than the nonpiliated bacteria and the avirulent Escherichia coli DH5 alpha, even following bacterial centrifugation onto the cell monolayers . A significant fraction of the internalized P . aeruginosa PAK was recovered in a viable form after 6 h of residence within endothelial cells . Progressive endothelial cell damage resulted from PAK intracellular harboring, as indicated by the release of lactate dehydrogenase . An increasing concentration of PAK cells was recovered from the extracellular medium with time, suggesting that ingested bacteria were released from endothelial cells and multiplied freely . We speculate that in vivo the ability of some P . aeruginosa strains to resist intracellular residence would afford protection from host defenses and antibiotics and that the release of viable bacteria into bloodstream may represent a central feature of the pathogenesis of bacteremia in compromised patients. Infect Immun, 1994 Dec, 62(12), 5335 - 43 Pulmonary immunity to Pseudomonas aeruginosa in intestinally immunized rats roles of alveolar macrophages, tumor necrosis factor alpha, and interleukin-1 alpha; Buret A et al.; The aims of this study were to assess the role played by alveolar macrophages, tumor necrosis factor alpha (TNF-alpha), and interleukin-1 alpha (IL-1 alpha) in pulmonary immunity against Pseudomonas aeruginosa in animals that have been immunized via the gut-associated lymphoid tissue . Following intra-Peyer's patch immunization and subsequent intratracheal challenge with live bacteria, significantly enhanced bacterial clearance from the lungs correlated with an increase in bronchoalveolar neutrophils, increased recruitment and phagocytic activity of alveolar macrophages, and accelerated production of TNF-alpha in the bronchoalveolar space, while levels of IL-1 alpha remained low . Administration of recombinant TNF-alpha in physiological concentrations did not affect the proliferation of P . aeruginosa in vitro, but when given intratracheally to rats at the time of infection, recombinant TNF-alpha significantly increased bacterial clearance from the lungs . In these animals, phagocytic activity of bronchoalveolar neutrophils was enhanced, while the recruitment of alveolar macrophages and neutrophils remained unchanged . In acutely infected nonimmune animals, bronchoalveolar concentrations of soluble IL-1 alpha and TNF-alpha increased until the time of death . Levels of prostaglandin E2 and thromboxane B2 were similar in each experimental group . These results indicate that infection in immune animals enhanced both recruitment and phagocytic activity of alveolar macrophages as well as induced an accelerated production of TNF-alpha . In immune challenged animals, this cytokine enhanced the phagocytic activity of neutrophils and improved bacterial clearance from the lung . Levels of soluble IL-1 alpha and TNF-alpha in nonimmune rats increased consistently following infection until the time of death, thus implicating these cytokines in the pathogenesis of acute P . aeruginosa pneumonia. Zhonghua Liu Xing Bing Xue Za Zhi, 1994 Dec, 15(6), 357 - 9 {Assay of R-plasmid and antibiotic resistance spectrum in 286 strains of gram-negative bacteria}; Wang Q et al.; This paper reports the results of the sensitivity test to 10 antibiotics and the assay of R-plasmid on 286 strains of Gram-negative bacteria isolated from hospitals . It was found that the main prevalent resistant bacteria are Pseudomonas aeruginosa, Escherichia coli and Klebsiella pneumoniae . Their major antibiotic resistance spectrum is: Ampicillin, Carbenicillin, Chloramphemicol, Gentamicin, Furbenicillin . The results showed that 272 of 283 strains resisted to 1-9 antibiotics (95.1%) . One-hundred and seventy-eight of 286 strains resisted to more than 4 antibiotics (62.2%) . One-hundred and fifty-one of 286 strains carried R-plasmid in resistant bacteria (53.3%) . 13.3% resistance markers were transferred with the whole R-factor . But 86.7% resistance markers were transferred with only a part of R-factor. Rinsho Byori, 1994 Dec, 42(12), 1253 - 60 {Problems concerned with adult T-cell leukemia . 1) Causes of death and early diagnosis of cytomegalovirus pneumonia in adult T-cell leukemia . 2) Clinicopathological features of CD30 positive adult T-cell leukemia}; Kikuchi H et al.; 1) We studied the causes of death confirmed by autopsy or necropsy in 23 adult T-cell leukemia (ATL) patients . Of them eight showed involvement of tumor (35%), eleven infectious disease (47%) (nine cytomegalovirus (CMV) pneumonia, one varicella-zostervirus pneumonia, one pseudomonas aeruginosa pneumonia), and four others (17%) . In seven recent cases treated with a new chemotherapy regimen in combination with G-CSF administration, survival was longer than in previous cases and tumor involvement as a cause of death decreased (one case, 14%) . However, CMV pneumonia inclined to increase (six cases, 86%) . Therefore, we tried to retrospectively detect CMV DNA in the serum of ATL patients using a nested polymerase chain reaction (PCR) . CMV pneumonia was reliably diagnosed in eleven ATL patients, whereas CMV DNA was detected in all patients at the time of clinical onset of pneumonia and CMV DNA was detected only in eight patients from 7-35 days before the onset of pneumonia . These findings suggest that the nested PCR assay is a useful tool to early diagnosis CMV pneumonia in ATL patients . 2) Recently, several cases of ATL with CD30 antigen have been reported, but its clinical relevance remains unknown . Accordingly, we studied CD30 antigen expression in 36 ATL patients who had monoclonal integration of HTLV-I provirus in the tumor cells and demonstrated the immunohistochemical and clinical characteristics of these patients . CD30 antigen expression was evident in seven of these 36 patients (19.4%) . A comparison of ATL cases with and without CD30 antigen expression revealed significantly large numbers of abnormal lymphocytes in the peripheral blood and lower serum calcium levels in CD30 antigen positive ATL. Yakugaku Zasshi, 1994 Dec, 114(12), 972 - 9 {Protective effect and antibody titer of intravenous immunoglobulin (IVIG) against clinical isolates of opportunistic bacteria}; Nakae T et al.; Neonates and leukopenic, immunosuppressed patients are at high risk for severe infection of opportunistic pathogens despite of the availability of potent antimicrobial agents . In this study, antibody titers of immunoglobulin preparations (IVIG) were contrasted with the protective effect in mice against each of bacterial infection . Antibody titers were determined by ELISA . The antigens were 70-80 clinical isolates of Pseudomonas aeruginosa, Escherichia coli, Klebsiella pneumoniae and Staphylococcus aureus, respectively . Antibody titers of IVIG against these three gram-negative bacteria ranged 3200 to 102,400 . ICR mice were inoculated intraperitoneally with each of several strains against which IVIG showed various titers . IVIG showed rather high protective activities against well-reactive strains, while it showed little protective activities against poor-reactive strains . In the case of P . aeruginosa, statistical analysis of the results obtained with the antibody titer and efficacy showed a good correlation (p < 0.01) . On the other hand, IVIG showed a high and complicate antibody titer against S . aureus IVIG ranging 400,000 to 12,800,000, since apparent titers contained non-specific binding of Fc portion of IgG with protein A on the cell wall . IVIG was active in mice, where protein A was less and specific binding was stronger . Bacterial cells have various components; lipopolysaccharide, lipid A, capsule, flagella, pill, etc . that are responsive to specific antibodies . This study indicates that IVIG have such antibodies and that is associated with protective activity against bacterial infection in proportion to antibody titer. J Appl Bacteriol, 1994 Dec, 77(6), 727 - 32 Mathematical model of the interactions between Micrococcus spp . and Pseudomonas aeruginosa on agar surface; Chapuis C et al.; The interactions between six different Micrococcus species and two strains of Pseudomonas aeruginosa were studied on an agar surface . This type of interaction on solid surface could act as a model of situations occurring either in the environment, in food or in man . The hypothesis of an amensalistic relationship between a Micrococcus spp . and Pseudomonas aeruginosa, due essentially to Ps . aeruginosa bacteriolytic enzymes, is retained as the basis for a mathematical model of the variations of the colony surface of Micrococcus spp . (S) with respect to the distance (d) from Ps . aeruginosa cells . The diffusion of the bacteriolytic substance in agar explains the limitation of the growth of the Micrococcus spp . This model S = Smax (1-e-md2) is shown to be adapted to all the interactions studied. J Cardiovasc Surg (Torino), 1994 Dec, 35(6 Suppl 1), 103 - 4 Sternal wound infection: our experience with 200 cases; Shafir R et al.; More than 200 patients with sternal wound infections have been treated in the Plastic Surgery Department of our Medical Center over the years 1984-1993 . Most of these were referrals from other hospitals . In recent years, the cases have become more severe, partially due to the fact that cardiac surgeons tend to operate older and sicker patients more readily than they previously did . 80% of these were post coronary bypass surgery, and the others heart and heart-lung transplants, repair of congenital heart anomalies, valve replacements etc . Several of the cases were cardiac surgery re-do's . Risk factors for developing this complication, such as diabetes, obesity, technical errors of sternal incision, prolonged intubation, the use of aortic balloon, etc . will be discussed . Many of our earlier patients had chronic fistulae following conservative therapy with old treatment modalities . In recent years, patients are usually referred at the acute stage . Most patients undergo removal of sternum and ribs . Previously, reconstruction included mainly transfer of the rectus ahdominis muscle, whereas lately the pectoralis muscles is utilized . Omentum was used in only one case . The importance of pre-operative imaging procedures has been thoroughly studied in our series . Especially important is the definition of the extent of the infection, and localization of foreign bodies causing chronic infections, such as suture material, epicardial electrodes etc . A change in infectants has also been noticed . In the first half of the study period, Pseudomonas aeruginosa comprised at least 40%.(ABSTRACT TRUNCATED AT 250 WORDS) Curr Microbiol, 1994 Dec, 29(6), 353 - 9 Orotidine-5'-monophosphate decarboxylase from Pseudomonas aeruginosa PAO1: cloning, overexpression, and enzyme characterization; Strych U et al.; Orotidine-5'-monophosphate decarboxylase (OMPdecase) catalyzes the final step in pyrimidine biosynthesis, the conversion of orotidine-5'-monophosphate (OMP) to uridine-5'-monophosphate . The pyrF gene, encoding OMPdecase, was isolated from a chromosomal library of Pseudomonas aeruginosa PAO1 by screening for complementation of an Escherichia coli and a P . aeruginosa pyrF mutant . The nucleotide sequence of a 2510-bp chromosomal DNA fragment, complementing both strains, was determined (EMBL accession number X65613) . On this a 696-bp open reading frame capable of encoding the 24 kDa OMPdecase was identified . Despite a generally good correspondence to other OMPdecase sequences, the P . aeruginosa gene was unique in that it did not constitute part of an operon . The pyrF gene was amplified by polymerase chain reaction, overexpressed in the pT7-7/E . coli BL21(DE3) system and purified to near electrophoretic homogeneity by anion exchange chromatography . Characterization of the purified enzyme revealed the following data, a Km value for OMP of 9.91 microM and an isoelectric point of 6.65 . No major decrease in enzyme activity was observed in a pH range between 7.8 and 10.2 . Gel electrophoresis under nondenaturing conditions suggested that the native form of OMPdecase is the dimer. Immunobiology, 1994 Dec, 192(1-2), 106 - 20 Effects of interferon gamma on the antibody response to Pseudomonas aeruginosa lipopolysaccharide in mice; Taylor CE et al.; Different strains of mice were examined for the capacity to produce an Ig subclass-specific antibody response to purified Pseudomonas aeruginosa lipopolysaccharide (PALPS) . With the exception of the AKR strain, the predominant isotype for most of the strains tested was IgG3 whereas the least frequent isotype expressed was either IgG2b or IgG1 . AKR mice were unique in that the predominant isotype produced was IgG2a, rather than IgG3; however, the administration of anti-interferon gamma antibody, at the time of immunization with PALPS caused a substantial decrease in the IgG2a antibody response . Selected B10 congenic strains were used to assess the relationship between the antibody responses and the major histocompatibility complex (MHC) genes . Here, the isotype-patterns for the antibody responses were essentially the same regardless of the MHC haplotype . Interestingly, an increase in IgG2a, with a concomitant decrease in IgM and IgG1 antibody was noted when C3H mice were given interferon gamma at the time of immunization . These studies indicate that, in general, the antibody response to PALPS consists of IgG3 antibody as the predominant isotype, and that the antibody response can be modified by interferon gamma. Int J Exp Pathol, 1994 Dec, 75(6), 441 - 51 Suppression of polymorphonuclear leucocyte chemotaxis by Pseudomonas aeruginosa elastase in vitro: a study of the mechanisms and the correlation with ring abscess in pseudomonal keratitis; Ijiri Y et al.; Bacteria, or the culture supernatants of an elastase non-producing strain of Pseudomonas aeruginosa, elicited a chemotactic response from polymorphonuclear leucocytes (PMN) in vitro . The chemoattractive capacity was diminished under the presence of Boc-Phe-Leu-Phe-Leu-Phe, a receptor antagonist of N-formyl-Met-Leu-Phe (fMLP) which is a bacterial chemotactic peptide to PMN . This indicated that the chemoattractant derived from Pseudomonas aeruginosa was a fMLP-like molecule(s) . In contrast, culture supernatants of an elastase producing strain of Pseudomonas aeruginosa produced negligible chemotactic response from PMN . Indeed, an inhibitory effect of the culture supernatants or of purified Pseudomonas aeruginosa elastase (PAE) on PMN chemotaxis was observed when fMLP was used as a chemoattractant . Another fMLP-induced function of PMN, respiratory burst activation, was also diminished by pretreatment of PMN with PAE . PAE hydrolysed fMLP at the Met-Leu bond and diminished the chemoattractant capacity . In addition, a receptor analysis with fML-3H-P demonstrated a decrease in numbers of fMLP receptors on PMN without changing the dissociation constant values after the treatment of the cells with PAE . In the primary structure of the fMLP receptor previously reported, a preferential amino acid sequence for cleavage by PAE was identified in what was believed to be an extracellular portion of the receptor molecule . These results suggested that PAE could diminish PMN infiltration in response to Pseudomonas aeruginosa in vivo by cleavage of the fMLP-like pseudomonal chemotactic ligand and the receptors on PMN. J Antimicrob Chemother, 1994 Dec, 34(6), 931 - 42 Profiles of outer membrane proteins and lipopolysaccharide of Pseudomonas aeruginosa grown in the presence of sub-MICs of macrolide antibiotics and their relation to enhanced serum sensitivity; Tateda K et al.; We have previously reported that erythromycin at sub-inhibitory concentrations enhanced the serum sensitivity of some Pseudomonas aeruginosa strains . To explore the mechanism of this effect, we have now examined the influence of macrolide antibiotics on outer membrane proteins and lipopolysaccharide (LPS) of P . aeruginosa . The strains S-6 and PAO-1 of P . aeruginosa were used as susceptible and resistant strains respectively to assess enhancement of serum sensitivity by erythromycin . The strain S-6 became more serum-sensitive when grown on agar with sub-MICs of erythromycin or azithromycin, but not of josamycin, whereas no change was observed in the serum sensitivity of the strain PAO-1 after growth with any of these antibiotics . The analysis of outer membrane proteins showed that erythromycin treatment resulted in a reduction in the amount of the 38 kDa protein (OprF) and in a prominent increase of 41 kDa protein band in the strain S-6, but not in the strain PAO-1 . By an immunoblotting assay, this 41 kDa protein was shown to be highly reactive to the immune serum against untreated P . aeruginosa . LPS of the strain S-6 were examined by SDS acrylamide gel electrophoresis . The treatment with erythromycin or azithromycin, but not with josamycin, reduced the amounts of LPS species with lower molecular weights although the levels of LPS species with high molecular weights were similar to those of untreated bacteria . These results suggest that the enhanced serum sensitivity of P . aeruginosa by erythromycin is associated with changes in bacterial surface components, such as outer membrane proteins and LPS. Mol Microbiol, 1994 Dec, 14(5), 1049 - 57 PilR, a transcriptional regulator of piliation in Pseudomonas aeruginosa, binds to a cis-acting sequence upstream of the pilin gene promoter; Jin S et al.; The PilR protein of Pseudomonas aeruginosa is a transcriptional activator of the pilin gene and belongs to a two-component sensor-regulator family . PilR was overproduced by fusing pilR to the gene for the maltose-binding protein (malE), yielding a MalE-PilR hybrid protein . The plasmid with the malE-pilR fusion, when introduced into a non-piliated pilR mutant strain of P . aeruginosa, restored piliation, indicating that the hybrid protein retains PilR function in vivo . The MalE-PilR protein was purified from Escherichia coli and used in a series of DNA-binding studies . A specific pilin promoter-binding activity of MalE-PilR was observed in a gel retardation assay . Subsequent DNase I footprinting analysis revealed a 40 bp PilR-binding site located at the -120 to -80 region, relative to the transcriptional start site of the pilin gene . This PilR-binding region consists of a nine-base sequence and three consensus sequences of 5'-(N)4-6C/GTGTC-3', in a tandem array in which the first 7-9 bp are bound by the PilR on the non-coding strand, leaving the last two nucleotides (TC) unbound . On the coding strand, PilR binds to sequences complementary to the two middle consensus sequences of the non-coding strand . A sequence similar to the NifA recognition site (5'-TGT-(N)11-ACA-3') is also found within the PilR-binding region . Deletion analysis and disruption of the individual consensus PilR-binding sequences by site-directed mutagenesis revealed that all four PilR-binding sites are absolutely required for the PilS/PilR-mediated pilin gene expression.(ABSTRACT TRUNCATED AT 250 WORDS) Mol Microbiol, 1994 Dec, 14(5), 1011 - 20 The sss gene product, which affects pyoverdin production in Pseudomonas aeruginosa 7NSK2, is a site-specific recombinase; Hofte M et al.; Pyoverdin production by Pseudomonas aeruginosa strain 7NSK2 was induced by Zn(II) in the presence of iron . A mutant was isolated in which Zn(II) no longer induced pyoverdin production . The sss gene which was inactivated in this mutant was cloned and sequenced . Its protein sequence showed 50% identity to the XerC protein of Escherichia coli, which is a member of the lambda integrase family of site-specific recombinases . An open reading frame was found upstream of sss whose protein sequence showed strong identity to DapF, the diaminopimelate epimerase . In E . coli, xerC is part of a multicistronic unit that also contains dapF . The sss gene of P . aeruginosa could restore site-specific recombination at cer in an E . coli xerC mutant and the E . coli xerC gene could complement a genomic sss mutation in P . aeruginosa. Lung Cancer, 1994 Dec, 11(5-6), 345 - 52 Long-term antimicrobial prophylaxis in lung cancer surgery: correlation between microbiological findings and empyema development; Ratto GB et al.; This study was planned in order to determine the value of antimicrobial prophylaxis in preventing post-operative empyema in patients undergoing lung cancer surgery . Two-hundred consecutive subjects operated upon for lung cancer received teicoplanin and aztreonam, starting at the induction of anesthesia and lasting until removal of the pleural drains . Cultures for aerobic and anaerobic bacteria were taken from: (1) the bronchus at the time of surgical division; (2) the pleural space before closure of the chest; (3) the pleural fluid during the post-operative period; and (4) the tips of chest drains at the time of their removal . In the 200 patients receiving antibiotic prophylaxis, the number of post-operative empyemas (1%) was lower than that (7.5%) found in 53 comparable patients who were previously treated with placebo . In the 'placebo group', empyema was due to gram-positive bacteria, while in the 'prophylaxis group', it was caused by Gram-negative bacteria (Pseudomonas aeruginosa) . A significant (P < 0.05) correlation between infected bronchial secretions, pleural space contamination at surgery, contamination of chest fluid and drains during the post-operative period, and empyema development was demonstrated . In conclusion, antibiotic prophylaxis, while being effective in preventing post-operative empyema, may induce the colonization of the respiratory tract with highly resistant gram-negative bacteria. Chem Pharm Bull (Tokyo), 1994 Dec, 42(12), 2569 - 74 Pyridonecarboxylic acids as antibacterial agents . IX . Synthesis and structure-activity relationship of 3-substituted 10-(1-aminocyclopropyl)-9-fluoro-7-oxo-2,3-dihydro-7H-pyrido{1,2,3-de}- 1,4-benzoxazine-6-carboxylic acids and their 1-thio and 1-aza analogues; Todo Y et al.; A series of the title compounds listed in Chart 1 have been synthesized to study the effects of 3-alkyl substituents on the antibacterial potency and in vivo efficacy of 10-(1-aminocyclopropyl)-9-fluoro-7-oxo-2,3-dihydro-7H-pyrido{1,2,3 -de}-1,4-benzoxazine-6-carboxylic acid and its 1-thio and 1-aza variants . Compound (S)-1, which proved most active in vitro against five representative gram-positive and gram-negative organisms, was assayed in vivo using Staphylococcus aureus and Pseudomonas aeruginosa mouse infection models . It exhibited an excellent in vivo efficacy, being superior to ofloxacin and ciprofloxacin, and was then assayed for convulsion-inducing activity, mammalian cell cytotoxicity, and topoisomerase II inhibition . The biological results showed that (S)-1 displayed antibacterial and toxicological advantages over ofloxacin and ciprofloxacin . Compound (S)-1 and its methanesulfonate showed high serum concentrations after oral and intravenous administrations to mice. Eur J Clin Chem Clin Biochem, 1994 Dec, 32(12), 893 - 9 Immunoenzymometric assays for alkaline protease and exotoxin A from Pseudomonas aeruginosa: development and use in detecting exoproteins in clinical isolates from patients with cystic fibrosis; Jaffar-Bandjee MC et al.; Using immunoenzymometric assays, the production of elastase, alkaline protease and exotoxin A was determined in culture supernatants of 35 strains of Pseudomonas aeruginosa isolated from patients suffering from cystic fibrosis . The assays were simple, specific, sensitive and reproducible, and permitted the determination of low levels of exoproteins . A large strain variability of exoprotein production was found . Most of the strains secreted all three exoproteins, but six out of the 35 strains (17%) did not secrete at least one of the three (< 0.3 microgram/l) . A significant correlation was observed between elastase and exotoxin A productions (r = 0.697, p < 0.001). Pulm Pharmacol, 1994 Dec, 7(6), 357 - 66 Addition of a bacterial alginate lyase to purulent CF sputum in vitro can result in the disruption of alginate and modification of sputum viscoelasticity; Mrsny RJ et al.; Alginate is a large molecular weight exopolysaccharide present in the purulent airway secretions of cystic fibrosis (CF) patients . This polymer, produced by some of the opportunistic pathogens associated with the recurrent lung infections characteristic of CF, has been suggested to effect an increase in the viscoelastic properties of purulent CF airway secretions . We have investigated the use of an enzyme targeted at this exopolysaccharide, an alginate lyase obtained from a bacterial source, to disrupt its polymeric nature and effect a change in the rheological properties of CF sputum in vitro . Expectorated sputum samples obtained from hospitalized CF patients were found to contain 80-200 micrograms alginate per ml sputum with no measurable endogenous alginate lyase activity . Treatment with exogenous alginate lyase prepared from a mucoid strain of Pseudomonas aeruginosa resulted in the disruption of alginate and a decrease in sputum viscoelasticity in a small percentage of the samples tested . Similar treatment of these samples with recombinant human deoxyribonuclease I to cleave DNA present in purulent sputum and the use of alginate extracted from sputum as an alginate lyase assay substrate suggested that the inability of the exogenous alginate lyase to disrupt sputum alginate was not due to substrate inaccessibility or an unresponsive substrate . Concentrations of Ca2+ and Zn2+ in alginate lyase-resistant sputum samples, determined by metal ion analysis, were found to inhibit enzyme activity in studies using seaweed alginate as a substrate . High concentrations of Ca2+ and Zn2+ in sputum samples initially resistant to lyase activity could be reduced significantly in some samples by dialysis and these same samples acquired sensitivity to the lyase . Other sputum samples did not show reduced concentrations of Ca2+ and Zn2+ following dialysis and these samples remained lyase-insensitive . Together, these results suggest that bacterial alginate present within purulent CF sputum may be quite stable, that endogenous alginate lyase activities appear to be limited and that the in vitro addition of exogenous alginate lyase can lead to the disruption of alginate and a change in the viscoelastic properties of some purulent CF sputum samples. J Infect Dis, 1994 Dec, 170(6), 1524 - 31 Infection with Pseudomonas cepacia in chronic granulomatous disease: role of nonoxidative killing by neutrophils in host defense; Speert DP et al.; Pseudomonas aeruginosa and Pseudomonas cepacia are catalase-producing bacteria, but only P . cepacia causes infections in patients with chronic granulomatous disease (CGD) . The in vitro killing of P . aeruginosa and P . cepacia by polymorphonuclear leukocytes (PMNL) from patients with CGD and from healthy adults was assessed . Of 6 patients with CGD who developed severe infections with P . cepacia, 4 died . PMNL from the 2 survivors and 6 other patients with CGD killed P . aeruginosa strains efficiently and P . cepacia strains poorly . PMNL from 2 patients with autosomal recessive CGD and from 2 carriers for X-linked CGD killed P . cepacia intermediately between normal controls and patients with X-linked CGD . When superoxide anion and hydrogen peroxide were scavenged with superoxide dismutase and catalase, normal PMNL killed P . aeruginosa but not P . cepacia . Thus, P . cepacia, but not P . aeruginosa, is a pathogen in patients with CGD, because it resists neutrophil-mediated nonoxidative bactericidal effects. J Bacteriol, 1994 Dec, 176(23), 7129 - 39 Monoclonal antibodies that distinguish inner core, outer core, and lipid A regions of Pseudomonas aeruginosa lipopolysaccharide; de Kievit TR et al.; In order to examine the immunochemistry of the core-lipid A region of Pseudomonas aeruginosa lipopolysaccharide (LPS), monoclonal antibodies (MAbs) specific for this region were produced in mice . Immunogen was prepared by coating a rough mutant of P . aeruginosa with column-purified core oligosaccharide fractions in order to enhance the immune response to the LPS core-lipid A region . Fourteen hybridoma clones were isolated, characterized, and further divided into three groups on the basis of their reactivities to rough LPS antigens in both enzyme-linked immunosorbent assays and Western immunoblots . In addition, another MAb, 18-19, designated group 1, was included in this study for defining core-lipid A epitopes . MAb 18-19 recognizes the LPS core-plus-one O-repeat unit of the serologically cross-reactive P . aeruginosa O2, O5, and O16 . Group 2 MAbs are specific for the LPS outer core region and reacted with P . aeruginosa O2, O5, O7, O8, O10, O16, O18, O19, and O20, suggesting that these serotypes share a common outer core type . Group 3 MAbs recognize the inner core region and reacted with all 20 P . aeruginosa serotypes as well as with other Pseudomonas species, revealing the conserved nature of this region . Group 4 MAbs are specific for lipid A and reacted with all gram-negative organisms tested . Immunoassays using these MAbs and well-defined rough mutants, in addition to the recently determined P . aeruginosa core structures, have allowed us to precisely define immunodominant epitopes within the LPS core region. FEBS Lett, 1994 Nov 21, 355(1), 45 - 8 Hybrid proteins between Pseudomonas exotoxin A and poliovirus protease 2Apro; Novoa I et al.; Two hybrid proteins between Pseudomonas aeruginosa exotoxin A (PE) and poliovirus protease 2Apro have been generated . One hybrid protein contains the poliovirus 2Apro sequence replacing the region of PE corresponding to amino acids 413-607 . The other hybrid contains in addition the transforming growth factor sequence . The two hybrid proteins were efficiently synthesized in E . coli cells using the inducible pET vectors . Both hybrid toxins cleaved p220 (eIF-4 gamma) when the recombinant plasmids were transfected in COS cells infected with recombinant vaccinia virus bearing the T7 RNA polymerase gene. FEMS Microbiol Lett, 1994 Nov 15, 124(1), 113 - 9 Role of autolysins in the EDTA-induced lysis of Pseudomonas aeruginosa; Watt SR et al.; Treatment of Pseudomonas aeruginosa with metal ion chelators, especially ethylenediaminetetraacetic acid (EDTA), causes both release of protein-lipopolysaccharide complexes and cell death . We have examined the effect of EDTA on P . aeruginosa and found that EDTA does not induce the rapid solubilization of the peptidoglycan sacculus and complete lysis as previously thought; the decrease in optical density of cultures incubated with EDTA is primarily due to the loss of the outer membrane . Of the other potential solubilizers examined, only ethylene-bis(oxyethylenenitrilo)tetraacetic acid (EGTA) resulted in some decrease in optical density . The lytic effect of EDTA on 12 strains of P . aeruginosa was examined and was found to vary greatly between strains; the sensitivity to EDTA varies from between 96% and 10% of the decrease in optical density resulting from incubation of cells with both EDTA and lysozyme . Sensitivity to EDTA is not constant during the growth of P . aeruginosa; in the early exponential phase of growth, cells treated with EDTA exhibit a 82% decrease in optical density after 30 min while in the stationary phase the optical density decreases by only 40% . Nucleic acids were observed to leak from cells following treatment with EDTA and this was greatly facilitated by DNase and RNase . The release of genetic material was much reduced when cells were incubated at 4 degrees C, supporting an enzymatic role in cell wall solubilization . We propose that only small areas of the sacculus become hydrolysed via specific peptidoglycan hydrolases, or autolysin(s), which are activated or de-regulated by EDTA. Biochemistry, 1994 Nov 8, 33(44), 12981 - 9 Evidence for the modulation of Pseudomonas aeruginosa exotoxin A-induced pore formation by membrane surface charge density; Rasper DM et al.; The lipid requirement for the binding of wild-type Pseudomonas aeruginosa exotoxin A (ETA) to model endosomal membrane vesicles was evaluated using a fluorescence quenching technique . The binding of toxin to monodisperse model membrane vesicles (0.1 micron diameter) composed of various molar ratios of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine (POPC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylserine (POPS) prepared by an extrusion method {Hope, M . J., et al . (1986) Chem . Phys . Lipids 40 89-107} was pH-dependent, with maximal binding observed at pH 4.0 . Analysis of the binding curves indicated that the interaction of ETA with the membrane bilayer is dominated by a set of high-affinity binding sites (Kd = 2-8 microM; 60:40 (mol:mol) POPC/POPS large unilamellar vesicles (LUV)) . The binding of toxin to membrane vesicles was highly pH-dependent, but was ionic strength-independent . Toxin-induced pore formation in the lipid bilayer, as measured by the release of the fluorescent dye, calcein, from LUV was pH-dependent, with optimal dye release occurring at pH 4.0 . The rate of dye release from membrane vesicles decreased rapidly with increasing pH and approached zero at pH 6.0 and higher . The pKa for this process ranged over 4.3-4.5 . Calcein release from LUV was also sensitive to changes in the ionic strength of the assay buffer, with maximal release occurring at 50 mM NaCl . Higher ionic strength medium resulted in a dramatic reduction in the rate of dye release from vesicles, indicating that the toxin-induced pore is modulated by ionic interactions.(ABSTRACT TRUNCATED AT 250 WORDS) Plant J, 1994 Nov, 6(5), 729 - 40 A NaCl-regulated plant gene encoding a brain protein homology that activates ADP ribosyltransferase and inhibits protein kinase C; Chen Z et al.; A cDNA clone pCZ1, with a 1.1 kb insert, was isolated from a NaCl-adapted tobacco cell cDNA library that encodes an apparently full-length 29 kDa protein (251 amino acids) with a calculated pI of 5.7 . The encoded peptide had a high amino acid sequence identity with bovine 14-3-3 protein which was originally found as an abundant protein in the animal central nervous system . Recently, proteins with sequence identity to 14-3-3 protein have also been found in plants, insects and yeast, and appear to have diverse physiological functions . Similar to the bovine brain 14-3-3 protein, the recombinant pCZ1 protein stimulated ADP-ribosylation of protein substrate by ADP-ribosyltransferase from the plant and animal pathogenic bacterium Pseudomonas aeruginosa . This recombinant protein also inhibited protein kinase C activity in vitro . Southern blot analyses indicated that most likely five genes encoding 14-3-3-like proteins are present in tobacco . The pCZ1 cDNA insert hybridized to a single mRNA of 1.1 kb from cultured tobacco cells . The level of this mRNA transcript in tobacco cells was downregulated upon adaptation to NaCl but was unaffected by short-term treatment with NaCl, ABA or ethylene . In tobacco plants, expression of transcript that hybridized to pCZ1 was tissue specific, and was most abundant in roots and flower parts . Monoclonal antibody raised against GF14 protein, a maize protein with substantial sequence identity with 14-3-3 protein detected two bands on SDS-PAGE of total proteins from unadapted tobacco cells and only a single band from cells adapted to NaCl . The GF14 antibody was also used to illustrate that the G-box element of a salt-induced gene is associated with a 14-3-3-type protein. Arch Surg, 1994 Nov, 129(11), 1144 - 52 Beneficial cardiopulmonary effects of pentoxifylline in experimental sepsis are lost once septic shock is established; Ridings PC et al.; OBJECTIVE: To determine the effects of pretreatment and posttreatment with pentoxifylline in a porcine model of gram-negative sepsis . DESIGN: Nonrandomized controlled trial . STUDY SUBJECTS: Young Yorkshire swine . INTERVENTIONS: Six groups of ventilated swine were studied for 5 hours . Group 1 swine (control, n = 8) received saline solution only . Group 2 swine (sepsis, n = 8) received a 1-hour infusion of Pseudomonas aeruginosa . Groups 3, 4, and 5 swine received the P aeruginosa infusion and a 20 mg/kg bolus followed by a 6 mg/kg per hour infusion of pentoxifylline . Group 3 swine (n = 6) received pentoxifylline prior to the onset of sepsis; group 4 swine (n = 6) received pentoxifylline at 1 hour and group 5 swine (n = 4) at 2 hours after the onset of the P aeruginosa infusion . Group 6 swine (control pentoxifylline, n = 3) received pentoxifylline only . OUTCOME MEASURES: Hemodynamic variables, neutrophil counts and CD18 expression, tumor necrosis factor activity, and arterial blood gases were measured hourly . Bronchoalveolar lavage was performed at 0 and 5 hours to measure neutrophil and protein content . RESULTS: All variables remained unchanged in the control and control pentoxifylline groups . Both pretreatment and posttreatment with pentoxifylline significantly attenuated lung injury and improved arterial PaO2 . The cardiac index was significantly improved by administration of pentoxifylline in groups 3 and 4 . Administration of pentoxifylline to group 5 animals in established septic shock caused uncontrolled, fatal systemic hypotension in two of the four animals . Plasma tumor necrosis factor activity, blood polymorphonuclear leukocyte counts, and CD18 expression were unaffected by the administration of pentoxifylline . CONCLUSIONS: Pentoxifylline protects against pulmonary and systemic hemodynamic derangements in fulminant sepsis and protects against pulmonary dysfunction . Pentoxifylline has a "therapeutic window" when given in established sepsis; if administration is delayed until overt septic shock occurs, it may then have deleterious effects. Am J Physiol, 1994 Nov, 267(5 Pt 1), L551 - 6 Exoproduct secretions of Pseudomonas aeruginosa strains influence severity of alveolar epithelial injury; Kudoh I et al.; To determine whether exoenzyme S plays a role in alveolar epithelial injury, two parental strains of Pseudomonas aeruginosa, PAK and PA103, were tested that produced large quantities of exoenzyme S . Strains PAK and PA103 differ in the form of exoenzyme S they produce . Strain PAK produces a 53-kDa protein that does not possess ADP-ribosyltransferase activity and large quantities of a 49-kDa protein that expresses ADP-ribosyltransferase activity . Strain PA103 produces the 53-kDa protein and low amounts of exoenzyme S activity . A quantitative experimental protocol was used to measure the protein permeability of the alveolar epithelium and the dissemination of the bacteria to the pleural space and circulation . The results indicate that instillation of PAK and PA103 resulted in significant lung injury . Control experiments utilizing isogenic, exoenzyme S-deficient, regulatory mutants in the infection model reduced the lung injury and the dissemination of instilled bacteria . Taken together these results suggest that alveolar epithelial injury correlated with the production of the 53-kDa form of exoenzyme S or other coordinately regulated factors. J Surg Res, 1994 Nov, 57(5), 625 - 31 Monoclonal antibody to tumor necrosis factor-alpha attenuates plasma interleukin-6 levels in porcine gram-negative sepsis; Mullen PG et al.; This study examined the kinetics of IL-6 release into the systemic circulation in a porcine model of bacterial sepsis induced by infusion of live Pseudomonas aeruginosa . Three groups of animals were studied . Group I (n = 12) animals received a 1 hr infusion of live P . aeruginosa . Group II (n = 6) animals received monoclonal antibody to tumor necrosis factor-alpha (TNF-alpha) (15 mg/kg) prior to induction of sepsis . Group III (n = 7) animals received sterile saline only . TNF-alpha and interleukin-6 (IL-6) levels rose sharply, in group I following pseudomonas infusion . Following a peak at 120 min after the bacterial infusion (4.8 +/- 0.7 U/ml at 120 min vs 0.4 +/- 0.2 U/ml at 0 min), TNF-alpha levels subsequently declined prior to the end of the experiment . In contrast, IL-6 levels rose sharply, subsequent to TNF-alpha, peaked at 180 min, and remained significantly elevated throughout the study period (5.3 +/- 0.9 ng/ml vs 0.05 +/- 0.01 ng/ml, 0 min) . In animals pretreated with monoclonal antibody to TNF-alpha, no increase in TNF-alpha activity was detected at any time during the period of study . IL-6 levels in antibody-treated animals, although greatly attenuated, still rose significantly above baseline (2.02 +/- 0.8 ng/ml at 180 min vs 0.05 +/- 0.01 ng/ml at 0 min) and above levels in control animals . We conclude that although TNF-alpha plays an important role in synthesis and release of IL-6, there is a TNF-alpha-independent pathway for release of IL-6 in sepsis. J Bacteriol, 1994 Nov, 176(22), 6944 - 51 Endotoxin biosynthesis in Pseudomonas aeruginosa: enzymatic incorporation of laurate before 3-deoxy-D-manno-octulosonate; Mohan S et al.; Unlike Escherichia coli, living cells of Pseudomonas aeruginosa can complete the fatty acylation of lipid A when the biosynthesis of 3-deoxy-D-manno-octulosonate (Kdo) is inhibited (R . C . Goldman, C . C . Doran, S . K . Kadam, and J . O . Capobianco, J . Biol . Chem . 263:5217-5233, 1988) . In this study, we demonstrate the presence of a novel enzyme in extracts of P . aeruginosa that can transfer lauroyl-acyl carrier protein (ACP) to a tetraacyl disaccharide-1,4'-bis-phosphate precursor of lipid A (termed lipid IVA) that accumulates in Kdo-deficient mutants of E . coli . Comparable E . coli extracts cannot transfer laurate from lauroyl-ACP to lipid IVA, only to (Kdo)2-lipid IVA (K . A . Brozek, and C . R . H . Raetz, J . Biol . Chem . 265:15410-15417, 1990) . P . aeruginosa extracts do not utilize myristoyl- or R-3-hydroxymyristoyl-ACP instead of lauroyl-ACP to acylate lipid IVA . Laurate incorporation in P . aeruginosa extracts is dependent upon time, protein concentration, and the presence of Triton X-100 but is inhibited by lauroyl-coenzyme A . P . aeruginosa extracts transfer only one laurate to lipid IVA, whereas E . coli extracts can transfer two laurates to (Kdo)2-lipid IVA . These results demonstrate that incorporation of laurate into lipid A does not require prior attachment of Kdo in all gram-negative bacteria. J Bacteriol, 1994 Nov, 176(21), 6677 - 87 Mucoid-to-nonmucoid conversion in alginate-producing Pseudomonas aeruginosa often results from spontaneous mutations in algT, encoding a putative alternate sigma factor, and shows evidence for autoregulation; DeVries CA et al.; The mucoid phenotype is common among strains of Pseudomonas aeruginosa that cause chronic pulmonary infections in patients with cystic fibrosis and is due to overproduction of an exopolysaccharide called alginate . However, the mucoid phenotype is unstable in vitro, especially when the cells are incubated under low oxygen tension . Spontaneous conversion to the nonmucoid form is typically due to mutations (previously called algS) that are closely linked to the alginate regulatory gene algT, located at 68 min on the chromosome . Our sequence analysis of algT showed that its 22-kDa gene product shares homology with several alternate sigma factors in bacteria, suggesting that AlgT (also known as AlgU) interacts directly with RNA polymerase core to activate the promoters of alginate genes . AlgT showed striking sequence similarity (79%) to sigma E of Escherichia coli, an alternate sigma factor involved in high-temperature gene expression . Our analysis of the molecular basis for spontaneous conversion from mucoid to nonmucoid, in the cystic fibrosis isolate FRD, revealed that nonmucoid conversion was often due to one of two distinct missense mutations in algT that occurred at codons 18 and 29 . RNase protection assays showed that spontaneous nonmucoid strains with the algT18 and algT29 alleles have a four- to fivefold reduction in the accumulation of algT transcripts compared with the wild-type mucoid strain . Likewise, a plasmid-borne algT-cat transcriptional fusion was about 3-fold less active in the algT18 and algT29 backgrounds compared with the mucoid wild-type strain, and it was 20-fold less active in an algT::Tn501 background . These data indicate that algT is autoregulated . The spontaneous algT missense alleles also caused about fivefold-reduced expression of the adjacent negative regulator, algN (also known as mucB) . Transcripts of algN were essentially absent in the algT::Tn501 strain . Thus, algT regulates the algTN cluster, and the two genes may be cotranscribed . A primer extension analysis showed that algT transcription starts 54 bp upstream of the start of translation . Although the algT promoter showed little similarity to promoters recognized by the vegetative sigma factor, it was similar to the algR promoter . This finding suggests that AlgT may function as a sigma factor to activate its own promoter and those of other alginate genes . The primer extension analysis also showed that algT transcripts were readily detectable in the typical nonmucoid strain PAO1, which was in contrast to a weak signal seen in the algT18 mutant of FRD . A plasmid-borne algT gene in PAO1 resulted in both the mucoid phenotype and high levels of algT transcripts, further supporting the hypothesis that AlgT controls its own gene expression and expression of genes of the alginate regulon. Infect Immun, 1994 Nov, 62(11), 4825 - 30 Inhibition of bacterial motility with human antiflagellar monoclonal antibodies attenuates Pseudomonas aeruginosa-induced pneumonia in the immunocompetent rat; Landsperger WJ et al.; Two human monoclonal antibodies, directed against the type a and type b flagellar proteins of Pseudomonas aeruginosa, inhibited bacterial motility in vitro specifically and in a concentration-dependent manner . In order to determine if this decreased bacterial motility was associated with a decreased pathogenicity, the ability of these human antiflagellar monoclonal antibodies to attenuate P . aeruginosa-induced pneumonia in the rat was assessed . Incubation of P . aeruginosa with a 1:1 mixture of the human antiflagellar monoclonal antibodies prior to pulmonary instillation significantly (P < 0.05) ameliorated the bacterium-induced decrease in arterial blood oxygen pressure, blunted the increase in respiratory rate, and markedly reduced the area of pulmonary inflammation . Similarly, intravenous administration of the human antiflagellar monoclonal antibodies 1 h after pulmonary instillation of the bacteria also reduced the in vivo pathogenicity of P . aeruginosa . Therefore, human antiflagellar monoclonal antibodies can decrease the in vitro motility of P . aeruginosa and can reduce its in vivo pathogenicity when administered either before or after bacterial challenge. Clin Immunol Immunopathol, 1994 Nov, 73(2), 261 - 6 The effect of treatment with interleukin-1 and tumor necrosis factor on Pseudomonas aeruginosa lung infection in a granulocytopenic mouse model; Amura CR et al.; The efficacy of treatment with interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF alpha) on Pseudomonas aeruginosa pneumonia was evaluated in a granulocytopenic mouse model . Combined intravenous administration of 2000 U IL-1 beta plus 2000 U TNF alpha significantly diminished mortality from aerosol challenge with P . aeruginosa . Mice treated with IL-1 beta, TNF alpha, or both also exhibited a significant enhancement in pulmonary clearance of P . aeruginosa . Combined cytokine administration induced an increase in the pulmonary content of myeloperoxidase activity . Mature leukocytes were not detected in either circulation or bronchoalveolar lavage fluid from granulocytopenic, cytokine-treated mice . In conclusion, IL-1 beta and TNF alpha treatment exhibited a synergistic protective effect from pulmonary P . aeruginosa challenge in granulocytopenic hosts, probably due to enhancement of nonspecific antibacterial mechanisms. Rev Invest Clin, 1994 Nov-Dec, 46(6), 465 - 72 {Malignant external otitis . Experience with 12 cases}; Volkow P et al.; Malignant external otitis is a life-threatening infection occurring in aging diabetic and immunocompromised patients . The development of new antimicrobial and diagnostic aids has modified the therapy and prognosis of the disease . We describe our experience in 12 cases seen between 1982 and 1991, and review the diagnostic and therapeutic criteria during this lapse . Ten cases were males and 11 were diabetics . The most common symptoms were unilateral otalgia and otorrhea . All had edema of the external auditory channel and nine, proliferation of granulation tissue . Four had cranial nerve palsy . In ten patients Pseudomonas aeruginosa was recovered . All had axial computed tomographic scans and six sequential radionuclide scanning performed at diagnosis and follow-up . Eleven patients received combined therapy with an aminoglucoside and an anti-pseudonomas beta-lactam antibiotic; in four ambulatory treatment was continued with a quinolone . Only one patient received a quinolone as only treatment due to unavailability of other drugs in the mexican market . Presentation of granulation tissue or bone sequestrum was performed in nine patients . Only three required extensive surgical debridement procedures . We conclude that a combined antimicrobial therapy and the use of quinolones has favorably modified the prognosis and avoids extensive surgery and disminishes hospital stay. Plasmid, 1994 Nov, 32(3), 254 - 61 The surface exclusion system of RP1: investigation of the roles of trbJ and trbK in the surface exclusion, transfer, and slow-growth phenotypes; Lyras D et al.; The contiguous trbJ and trbK genes of RP1 were cloned individually to study their effects . Surface exclusion was conferred only by trbK and only when gene dosage was high or when trbJ was also present in cis or in trans . This suggests that in the low-copy-number RP1, surface exclusion is due to a two-gene interaction in which trbK is the dominant partner . Among surface exclusion genes, trbJ is novel in yielding a periplasmic product that is also essential for conjugal transfer . This cellular location and the disturbed membrane function that accompanies TrbJ-processing probably accounts for the retarded growth caused by trbJ+ clones in Pseudomonas aeruginosa strain PAO. Ophthalmic Surg, 1994 Nov-Dec, 25(10), 671 - 4 Prophylaxis of endophthalmitis; Peyman GA et al.; Prophylactic antibiotics can be administered by various routes to prevent endophthalmitis . Topical application is simple and minimally invasive, but it provides relatively poor penetration into the vitreous . Collagen shields are effective in delivering therapeutic levels of antibiotics to the cornea and aqueous but not to the vitreous cavity . Systemic administration for prophylaxis is questionable because of the poor ocular penetration of most drugs (however, some antibiotics, such as imipenem, pefloxacin, and ciprofloxacin, may provide therapeutic levels to the vitreous) . Subconjunctival injection may provide significant drug levels in the aqueous for 4 to 6 hours . However, we found subconjunctival gentamicin ineffective in experimentally induced Pseudomonas aeruginosa endophthalmitis in postoperative aphakic eyes . We recommend the use of antibiotics in infusion fluid . In cases of trauma, we inject 48 micrograms gentamicin and 54 micrograms clindamycin intracamerally or intravitreally. Eur Respir J, 1994 Nov, 7(11), 1925 - 31 Pseudomonas-induced neutrophil recruitment in the dog airway in vivo is mediated in part by IL-8 and inhibited by a leumedin; Jorens PG et al.; A bacteria-free supernatant of Pseudomonas aeruginosa induces the production of neutrophil chemotactic activity in human bronchial epithelial cells in vitro that is due to the potent chemotactic factor, interleukin-8 (IL-8) . Because P . aeruginosa supernatant itself is not chemotactic, we hypothesized that intratracheal P . aeruginosa induces the production of neutrophil chemotactic factors, including IL-8, in vivo . Because neutrophils play a key role in cystic fibrosis, inhibition of neutrophil recruitment might be therapeutic . We studied the effect of P . aeruginosa supernatant in the isolated tracheal segment of dogs in vivo, and we measured neutrophil chemotactic activity in vitro in the tracheal fluid . We also determined the local effect of intratracheal administration of leumedin NPC 15669, an inhibitor of neutrophil recruitment, on IL-8- and Pseudomonas-induced neutrophil accumulation . P . aeruginosa supernatant and IL-8 both caused time-dependent accumulation of neutrophils in the tracheal fluid . Tracheal fluid obtained after P . aeruginosa administration had neutrophil chemotactic activity in vitro that was significantly inhibited by the IL-8 antibody . Intratracheal NPC 15669 prevented both IL-8- and Pseudomonas-induced accumulation of neutrophils . We conclude that P . aeruginosa supernatant recruits neutrophils into the airway indirectly by inducing the production of chemotactic factors, including IL-8 . Our results suggest a potential therapeutic role for leumedins in chronic airway diseases. Yeast, 1994 Nov, 10(11), 1489 - 96 The sequence of a 22.4 kb DNA fragment from the left arm of yeast chromosome II reveals homologues to bacterial proline synthetase and murine alpha-adaptin, as well as a new permease and a DNA-binding protein; De Wergifosse P et al.; We report the sequencing of a 22,470 bp DNA fragment from the left arm of Saccharomyces cerevisiae chromosome II . Thirteen open reading frames longer than 300 bp provisionally called YBL0520, YBL0401 to YBL0408 and YBL0410 to YBL0413 have been detected . Five genes were previously sequenced: COR1, encoding a core protein of the mitochondrial coenzyme QH2 cytochrome c reductase complex (Tzagaloff and Crivellone, 1986), PRS3, a proteasome subunit gene (Lee et al., 1992), ERD2, coding for a protein involved in the secretory pathway (Semeza et al., 1990), URA7, which encodes a CTP synthetase (Ozier-Kalogeropoulos et al., 1991) and the gene for the ribosomal protein L16 (Pan et al., 1993) . Among the others, YBL0406 shows striking homologies to FUR4 (Jund et al., 1988) and DAL4 (Yoo et al., 1992), the uracyl and allantoin permeases; YBL0520 is a DNA-related protein, possibly involved in gene regulation; YBL0412 shares homologies with the mouse alpha-adaptins A and C; and YBL0413 is homologous to a protein of Pseudomonas aeruginosa that is likely to be involved in proline biosynthesis . YBL0401, internal to YBL0520, is probably not expressed. Vaccine, 1994 Nov, 12(14), 1288 - 94 Phase 1 trial of a 24-valent Klebsiella capsular polysaccharide vaccine and an eight-valent Pseudomonas O-polysaccharide conjugate vaccine administered simultaneously; Edelman R et al.; A Klebsiella (K) vaccine consisting of 24 capsular polysaccharide antigens and a Pseudomonas aeruginosa (P) vaccine consisting of eight O-polysaccharide antigens conjugated to P toxin A have been developed to prevent sepsis by means of active or passive immunoprophylaxis . In search for a practical immunization schedule, the two vaccines were injected in opposite arms simultaneously (20 volunteers) or 14 days apart (21 volunteers) . The vaccines were similarly well tolerated by both volunteer groups . Geometric mean antibody concentrations and mean fold antibody rises to the 33 vaccine antigens (including toxin A) were similar in the two groups at 2 months, and the decline in antibody measured at 18 months was also similar . Because the two vaccines were safe and similarly immunogenic in the two vaccine groups, they can be administered simultaneously to patients or plasma donors in a practical vaccination schedule. J Hosp Infect, 1994 Nov, 28(3), 209 - 18 Outbreak of gentamicin, ciprofloxacin-resistant Pseudomonas aeruginosa in an intensive care unit, traced to contaminated quivers; Jumaa P et al.; An outbreak of gentamicin, ciprofloxacin-resistant Pseudomonas aeruginosa in an intensive care unit, was investigated . The majority of isolates were from sputum and the organism was not isolated from any other patient in the hospital, except those admitted to the unit . A prospective study was set up, and the organism was found to be associated with contaminated quivers, used to store suction tubing between use on ventilated patients . Once the quivers were disinfected and changed between patients daily, the outbreak stopped . Suction of ventilated patients may be an important source of contamination of the respiratory tract with nosocomial pathogens . It is important that infection control teams regularly review procedures to ensure the correct practices are being followed, so that nosocomial outbreaks of infection may be prevented. Curr Eye Res, 1994 Nov, 13(11), 783 - 90 Pseudomonas aeruginosa corneal ulcer isolates distinguished using the arbitrarily primed PCR DNA fingerprinting method; Bukanov N et al.; Infection of the eye by Pseudomonas aeruginosa can result in corneal inflammation (keratitis) and ulceration, and permanent decrease in vision if not successfully treated . We tested for diversity among P . aeruginosa strains from corneal ulcers by the sensitive and efficient 'RAPD' (for 'random amplified polymorphic DNA') fingerprinting method . This method uses single oligonucleotides of arbitrarily chosen sequence as primers in low-stringency PCR amplification, and results in strain-specific arrays of DNA fragments . Tests of 20 independent P . aeruginosa corneal ulcer isolates yielded 19 different arrays of products with each of three arbitrary primers, indicating that all but two of the strains differed from one another . Additional isolates from three patients (infected eye, contact lens or eye drops) yielded fragment patterns that were identical to those of the original isolate in each case . Thus, our results demonstrate considerable diversity among P . aeruginosa corneal ulcer isolates, and suggest that just one clone may predominate in typical infections. Cornea, 1994 Nov, 13(6), 500 - 4 Comparison of topical ciprofloxacin to conventional antibiotic therapy in the treatment of experimental Pseudomonas aeruginosa keratitis; Guzek JP et al.; We evaluated the efficacy of topical ciprofloxacin (3.0 mg/ml) in the treatment of experimental Pseudomonas aeruginosa keratitis in 60 rabbits . We compared ciprofloxacin treatment with double drug therapy consisting of tobramycin (13.6 mg/ml) plus polymyxin B (25,000 U/ml) or carbenicillin (6 mg/L) . Two strains of P . aeruginosa were used . One was a strain well characterized for use in experimental Pseudomonas keratitis (ATCC organism 27853); the second was an organism from a patient with a Pseudomonas corneal ulcer . Rabbits were treated for 16 h, after which the corneas were excised, homogenized, and plated serially for residual colony-forming units . Both organisms responded significantly better to topical off-the-shelf ciprofloxacin than to therapy with two conventional antipseudomonal fortified antibiotic drugs (p < or = 0.0001). Immunology, 1994 Nov, 83(3), 362 - 9 A role for CD4+ T cells from orally immunized rats in enhanced clearance of Pseudomonas aeruginosa from the lung; Dunkley ML et al.; The role of gut-derived CD4+ T cells in clearance of Pseudomonas aeruginosa from the lung was studied by cell transfer experiments . Mesenteric lymph node cells from unimmunized rats, or rats orally immunized with either killed P . aeruginosa or Helicobacter pylori, were transferred to naive rats which were subsequently challenged intra-tracheally with live P . aeruginosa . Recipients of unseparated mesenteric lymph node cells, purifed T cells or CD4+ T cells, from P . aeruginosa-immunized donors, all exhibited enhanced bacterial clearance from the airways compared to recipients of cells from unimmunized donors . Enhanced clearance by T cells was antigen-specific as no enhanced clearance was observed by transfer of cells from donors immunized with H . pylori. Zhonghua Yi Xue Za Zhi (Taipei), 1994 Nov, 54(5), 306 - 11 Enterococcal bacteremia in a medical center; Peng MY et al.; BACKGROUND . The enterococci have become important nosocomial pathogens . They can cause multiple site infections and enterococcal bacteremia becomes more frequently associated with a high mortality rate . Previous studies of enterococcal bacteremia showed a variety of results . To establish the significance and importance of enterococci as nosocomial pathogens in this hospital, to characterize their clinical pictures and to search for the risk factors for mortality, this retrospective study was performed . METHODS . There were 208 cases of enterococcal bacteremia which occurred from 1988 to 1992 . Twenty-seven cases had no medical charts, dismissing possibility of evaluation . Finally, 181 cases of enterococcal bacteremia were analysed . RESULTS . One hundred and eighteen episodes were nosocomial infections . Polymicrobial bacteremia occurred in 68.5% of the patients and the most common co-isolate was Pseudomonas aeruginosa . Those patients (78.5%) with underlying diseases and malignancies were the most common underlying problems . The portal of entry could be found in 69.6 percent of patients, with the gastrointestinal tract the most common sources . Antimicrobial susceptibility testing showed high gentamicin resistance rate (89.5%), and ampicillin still had about 80 percent sensitivity rate . The group who received specific antibiotic therapy for enterococcus showed lower mortality (36.4% versus 47.6%) . Only one case had infective endocarditis . Forty-nine patients suffered from septic shock, the cause of 30 deaths . Totally 75 patients died during hospitalization . Besides sepsis, another major cause of death was their underlying diseases itself . CONCLUSIONS . Enterococci have no doubt become important nosocomial pathogens and enterococcal bacteremia were associated with high mortality, especially in elderly patients with underlying diseases such as malignancy or diabetes . When clinically dealing with sepsis from the gastrointestinal or biliary tract, especially when previous cephalosporins therapy showed no response, the possibility of enterococcal bacteremia should always be considered. Microbiology, 1994 Nov, 140 ( Pt 11), 2961 - 9 The effect of nutrient limitation on glycerol uptake and metabolism in continuous cultures of Pseudomonas aeruginosa; Williams SG et al.; Pseudomonas aeruginosa NM48, a non-mucoid derivative of an alginate-producing strain isolated from a cystic fibrosis patient, was grown in batch culture with glycerol, glucose or succinate as carbon source, and in continuous culture (D 0.05 h-1) under glycerol or glucose limitation . Glycerol uptake, glycerol kinase and glycerol-3-phosphate dehydrogenase were induced by glycerol, but not by glucose or succinate . Linear uptake of {14C}glycerol by washed cells (Km < or = 2 microM) was inhibited by unlabelled glycerol and glyceraldehyde, but not by cyanide or the uncoupling agent carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP), and was accompanied by substantial intracellular accumulation of glycerol-3-phosphate and/or dihydroxyacetone phosphate but not glycerol . Prolonged growth under glycerol limitation led to substantial increases in the activities and/or concentrations of the enzymes catalysing glycerol uptake and metabolism, together with a 48,000 M(r) outer-membrane protein which was also over-expressed following prolonged growth under glucose limitation . The N-terminal amino acid sequence (AEAFSPN-) and electrophoretic properties of this protein were the same as those of the previously characterized glucose porin (OprB) from P . aeruginosa, indicating that this porin is active with both glucose and glycerol . It is concluded that during growth under glycerol limitation, glycerol is transported into P . aeruginosa NM48 via OprB and a high-affinity, binding-protein-independent facilitated-diffusion system. Appl Microbiol Biotechnol, 1994 Nov, 42(2-3), 415 - 20 Degradation of chlorobenzenes in soil slurry by a specialized organism; Brunsbach FR et al.; The microbial degradation of monochloro-, 1,2-dichloro-, 1,4-dichloro-, and 1,2,4-trichlorobenzene in soil slurries was examined with single compounds as well as in mixtures . The indigenous soil populations brought about the degradation of monochlorobenzene when incubated at 27 degrees C in slurries with 29% (w/w) suspended solids . In contrast, the other chlorobenzenes persisted during an incubation period of 1 month . Supplementation with buffer, mineral salts and acetate did not significantly influence the degradation . However, inoculation with Pseudomonas aeruginosa strain RHO1, a monochloro- and 1,4-dichlorobenzene-degrading organism, to a titre of 1 x 10(5) cells/g soil, led to rapid and complete degradation of 0.8 mM growth substrate within 30 h . In addition, the strain was able to degrade 1,2-dichloro- and 1,2,4-trichlorobenzene with stoichiometric release of chloride in the presence of acetate, ethanol, monochloro- or 1,4-dichlorobenzene as growth substrates . In mixtures of chlorobenzenes the co-metabolism of 1,2-dichloro- and 1,2,4-trichlorobenzene occurred until the growth substrates monochloro- and 1,4-dichlorobenzene were degraded . The degradation was faster in the slurries of garden soil containing 8% organic carbon than in soil with the lower content of 2.6%. Zhonghua Zheng Xing Shao Shang Wai Ke Za Zhi, 1994 Nov, 10(6), 456 - 8 {Prophylactic effect of systemic ceftazidime on bacterial translocation in scalded rats}; Zong G et al.; In this study, we evaluated the prophylactic effect of Ceftazidime on bacterial translocation (BT) of Pseudomonas aeruginosa in scalded rat by qualitative and quantitative bacterial cultures of blood, organs and mesenteric lymph nodes (MLN) . Ceftazidi-me was administered systemically at 3 h, 6 h and 12 h post injury respectively . The dynamic changes in levels of Ceftazidime in blood and organ tissues were also determined . The results showed that effective levels of Ceftazidime in blood, liver and mucosa of the small intestine were rapidly reached and maintained for more than 4 h, but no drug was found in MLN . The prophylactic effect on BT was significant when systemic administration started at 3 h post injury (P < 0.001) and 6 h post injury (P < 0.05) . Although the incidence of BT was unchanged (P > 0.05) when systemic administration started at 12 h, the number of organisms present in livers and kidneys, except MLN, was dramatically reduced (P < 0.01). Pediatr Med Chir, 1994 Nov-Dec, 16(6), 551 - 4 {Pseudomonas aeruginosa antibodies in patients with cystic fibrosis: clinical implications}; Marianelli L et al.; Patients chronically colonised by P . aeruginosa develop a pronounced antibody response against P . aeruginosa that can be used to discriminate between superficial colonization and chronic infection . Anti-P . aeruginosa antibodies fail to afford protection against this pathogen; moreover high levels of antibodies are correlated with a poor prognosis . We investigated the significance of anti-P . aeruginosa antibodies (precipitins) by crossed immunoelectrophoresis (CIE) in 94 patients attending the Cystic Fibrosis Center of Florence . The highest numbers of precipitins were found in serum from patients chronically colonized in comparison to those patients who were transiently or not colonized . A negative correlation was found between the number of precipitin peaks and clinical conditions, evaluated with Schwachman score, and the number of precipitins and pulmonary functions . In summary, anti-P . aeruginosa antibodies fail to protect against P . aeruginosa bronchopulmonary infections, and are correlated to a more severe disease . Based on our experience, P . aeruginosa antibodies can be considered a reliable index of lung damage. J Antimicrob Chemother, 1994 Nov, 34(5), 697 - 705 Uptake of BRL 41897A, a C(7) alpha-formamido substituted cephalosporin, via the ferri-pyochelin transport system of Pseudomonas aeruginosa; Gensberg K et al.; Several catechol-substituted cephalosporin antibiotics are transported into Gram-negative organisms via components of the iron uptake system . We provide evidence that the C(7) alpha-formamido substituted cephalosporin BRL 41897A enters Pseudomonas aeruginosa via the high molecular weight pyochelin transport system . No MIC could be recorded when cells were grown under iron-replete conditions, but when grown in iron-depleted succinate medium strain PAO1 the MIC was 4 mg/L . A derivative of PAO1 that is deficient in the biosynthesis of pyochelin and pyoverdin, IA1, was more resistant, having MIC of 16 mg/L . This was reduced to 4 mg/L by growing the cells in the presence of pyochelin, which specifically induces the pyochelin uptake system . Growth in the presence of pyochelin also increased the rate and extent of uptake of a radiolabelled iron-antibiotic complex . Growth in the presence of 1/8 of the MIC resulted in significant reduction in expression of the ferripyochelin receptor in the outer membrane, coupled with decreased sensitivity to the agent. Arkh Patol, 1994 Nov-Dec, 56(6), 28 - 32 {The role of biologic and biochemical properties of microorganisms in the pathogenesis of experimental sepsis}; Sarkisov DS et al.; Certain biological and biochemical differences are found after biochemical and electron-microscopic examinations between different substrains of staphylococcus and Pseudomonas aeruginosa . The conclusion is made that in the development, course and form of septic process the important role is played not only by the type of infectious agent, but by its various substrains as well. Microb Pathog, 1994 Nov, 17(5), 291 - 9 Differences in eucaryotic cell binding of Pseudomonas; Cervin MA et al.; The lungs of cystic fibrosis (CF) patients are frequently chronically colonized by Pseudomonas aeruginosa . Recently there has been an increase in colonization by another pathogen Pseudomonas cepacia, which can cause a rapid decline in clinical condition or death of the patient . The nature of the factor(s) which predispose CF patients to colonization by one or both of these opportunistic pathogens is unknown . It has been suggested that the genetic defect in CF patients results in an increase in the number of epithelial cell receptors available to P . aeruginosa in the lung, thus rendering CF patients more susceptible to bacterial colonization than non-CF individuals . In this study, we have examined adherence of several strains of P . aeruginosa and P . cepacia to a variety of continuous cell lines, as well as primary cultures of CF and non-CF nasal polyp cells . The results suggested that there may be a decrease in the number of receptors available to both strains of Pseudomonas on cells of canine origin compared to human cells . Both strains appear to use pili as the primary adhesin, but there is also evidence that non-pilus adhesins contribute significantly to eucaryotic cell binding . P . cepacia exhibited microcolony formation on all cell types, which is typical of the localized adherence pattern characteristic of the enteropathogenic Escherichia coli . However, we were unable to demonstrate, with either P . cepacia or P . aeruginosa, a significant increase in adherence to CF compared to non-CF nasal polyp cultures.(ABSTRACT TRUNCATED AT 250 WORDS) Can J Microbiol, 1994 Nov, 40(11), 903 - 10 Purification and characterization of a protease from Pseudomonas pseudomallei; Sexton MM et al.; Pseudomonas pseudomallei is the causative agent of melioidosis, a glanders-like disease of humans and animals . The pathogenesis of melioidosis is not well understood, and the role of various extracellular enzymes produced by P . pseudomallei in the development of this disease is not known . The present studies were designed to purify and characterize an extracellular protease produced by P . pseudomallei isolates and to test the hypothesis that this protease may play a role in melioidosis . The protease was present in culture supernatants as an enzyme with a molecular weight of 36,000 that was optimally active at 60 degrees C and at pH 8.0 . The P . pseudomallei protease was shown to be a metalloenzyme requiring iron for maximal activity, and activity was inhibited in the presence of ethylenediaminetetraacetic acid (150 mM) . Antibodies directed against an alkaline protease produced by Pseudomonas aeruginosa cross-reacted with the P . pseudomallei protease . These data indicate that the P . pseudomallei protease belongs to the family of alkaline proteases sensitive to metal chelators . Purified P . pseudomallei protease was capable of digesting a variety of eucaryotic protein substrates including immunoglobulins . A P . pseudomallei strain deficient in protease production was shown to be less virulent than the parental strain in an animal model of lung infection . These data suggest that this protease may be a significant pathogenic determinant in infections caused by P . pseudomallei. J Med Chem, 1994 Oct 28, 37(22), 3828 - 33 Synthesis and antimicrobial activity of novel 3-{(aminopyrimidiniumyl)thio}methyl cephalosporins; Kim YZ et al.; A series of novel cephalosporin compounds which have 3-{(aminopyrimidiniumyl)thio}methyl substituents was synthesized . They show high antimicrobial activity against various bacterial species including Pseudomonas aeruginosa . Structure-activity relationships with various thiopyrimidines, thiopyrimidiniums, bicyclic thiotriazolopyrimidiniums, and bicyclic thioimidazolopyrimidiniums as 3'-substituents were also studied; cephalosporins with quarternized pyrimidinium moieties have better antimicrobial activities than neuteral pyrimidine cephalosporins, and stabilization of the positive charge on the pyrimidinium moieties is essential for better activity . According to semiempirical PM3 calculations, amino and alkylthio substituents on the pyrimidinium rings play a major role in charge stabilization and delocalization. J Mol Biol, 1994 Oct 21, 243(2), 347 - 50 Crystallization and preliminary X-ray analysis of a new crystal form of nitrite reductase from Pseudomonas aeruginosa; Tegoni M et al.; Nitrite reductase from Pseudomonas aeruginosa (EC 1.9.3.2), a redox enzyme synthesized by the bacterium grown in the presence of nitrate, is a soluble dimer of two identical subunits of 60 kDa, each containing one c and one d1 haem as prosthetic groups . A new crystal from of the Ps . aeruginosa nitrite reductase in the oxidized state, suitable for X-ray structure determination, has been obtained by vapour diffusion at 20 degrees C, in the presence of 10% polyethylene glycol 4000, 50 mM Tris-HCl (pH 8.7), 400 mM NaCl and at a protein concentration of 14 mg/ml . The crystals are dark green elongated tetragonal prisms of dimensions 1.5 mm x 0.2 mm x 0.2 mm for the largest ones . These crystals are tetragonal with space group P4(1(3))2(1)2 and cell dimensions a = b = 128.2 A, c = 172.6 A . They diffract at least up to 2.8 A . Assuming a dimer in the asymmetric unit, the VM value is 2.95 A3/Da (58% of solvent). Gene, 1994 Oct 11, 148(1), 81 - 6 Construction of improved Escherichia-Pseudomonas shuttle vectors derived from pUC18/19 and sequence of the region required for their replication in Pseudomonas aeruginosa; West SE et al.; The nucleotide sequence of the 1.9-kb PstI fragment from pRO1614, that allows stable maintenance of pMB1 (ColE1)-based cloning vectors in Pseudomonas, was determined . This fragment encodes a putative origin of replication (ori), a replication-controlling protein, and the C terminus of the Tn3 beta-lactamase-encoding gene . Improved versions of the broad-host-range plasmid vectors, pUCP18 and pUCP19, were constructed by deletion of nonessential DNA or replacement of nonessential DNA with an antibiotic-resistance cassette. Cell Immunol, 1994 Oct 1, 158(1), 71 - 82 Proliferation of immature T cells within the splenocytes of athymic mice by Pseudomonas exotoxin A; Dixon DM et al.; Pseudomonas aeruginosa exotoxin A has been shown to stimulate splenocytes from athymic nude mice . The present studies were performed to characterize the exotoxin A-responsive cells within the splenocytes of athymic nude mice . The results of these studies indicate that exotoxin A-responsive cells were represented in the nylon wool-adherent cell population and in the Ig-depleted splenocyte population (IgNA) . Flow microfluorimetry analysis indicated that exotoxin A induced the expansion of Thy1+, CD3-, CD4-, and CD8- cells which also expressed the heat-stable antigen (HSA), an antigen expressed on immature T lymphocytes . Depletion of Thy1+ cells abrogated the exotoxin A-induced proliferation; however, depletion of CD4+ and CD8+ cells did not affect the exotoxin A-induced proliferation of IgNA cells . Thus, the results indicate that exotoxin A stimulates the proliferation of immature T cells within the splenocytes of nude mice that are Thy1+, HSA+, CD3-, CD4-, CD8-. FEMS Microbiol Lett, 1994 Oct 1, 122(3), 309 - 12 Evidence for the location of OprM in the Pseudomonas aeruginosa outer membrane; Gotoh N et al.; OprM with a M(r) of 49 K is associated with the multidrug resistance of Pseudomonas aeruginosa . Detergent fractionation of bacterial cells has demonstrated that OprM is located in the outer membrane from which it sediments with the other major outer membrane proteins . In this study we have determined the location of OprM as the P . aeruginosa outer membrane . Western immunoblots of cell fractions, obtained by sucrose density gradient centrifugation of whole cell lysates, were probed with an OprM-specific murine polyclonal antiserum. FEMS Microbiol Lett, 1994 Oct 1, 122(3), 267 - 73 Isolation of OprM-deficient mutants of Pseudomonas aeruginosa by transposon insertion mutagenesis: evidence of involvement in multiple antibiotic resistance; Gotoh N et al.; Overproduction of the Pseudomonas aeruginosa outer membrane protein OprM was observed in the nalidixix acid (NalB-type) multidrug-resistant strains . To clarify the involvement of OprM in the resistance, transposon mutants were isolated from strain PAO4141 and its OprM-overexpressed mutant KG2113 and were screened for OprM production by immunoblot assays using murine polyclonal antiserum resulting from immunization with purified OprM . Two OprM-deficient mutants from PAO4141 and one from KG2113 were identified . Determination of the susceptibilities of these mutants to antimicrobial agents demonstrated that OprM was involved not only in the acquired resistance, but also in the intrinsic resistance of P . aeruginosa to quinolones, cephems, penicillins, tetracycline, and chloramphenicol. FEMS Microbiol Lett, 1994 Oct 1, 122(3), 239 - 44 Cytotoxin-converting phages, phi CTX and PS21, are R pyocin-related phages; Hayashi T et al.; phi CTX is a temperate phage of Pseudomonas aeruginosa harbouring the ctx gene that encodes cytotoxin (CTX) . We identified phi CTX as an R pyocin-related phage, by serological and molecular analysis, based on the findings that the infectivity of the phage was inhibited with the antisera directed R pyocins and R pyocin-related phages and that the phi CTX genome showed DNA homology to the genome of PS17 (a representative of the R pyocin-related phages) as well as to the pyocin R2 genes . Another new CTX-converting, R pyocin-related phage named PS21 was isolated from a CTX-producing strain of P . aeruginosa, suggesting the distribution of the ctx gene by certain members of R pyocin-related phage family. Appl Environ Microbiol, 1994 Oct, 60(10), 3739 - 45 Detection of Pseudomonas aeruginosa from clinical and environmental samples by amplification of the exotoxin A gene using PCR; Khan AA et al.; PCR was used to detect Pseudomonas aeruginosa from water samples by amplifying a 396-bp region of the exotoxin A (ETA) structural gene sequence . The identify of the amplified 396-bp fragment was confirmed by digesting it with PvuI restriction endonuclease, which produced the predicted 246- and 150-bp fragments . Specific primers amplified ETA-positive P . aeruginosa DNA, whereas other species of Pseudomonas and GC-rich bacteria did not yield any 396-bp fragment . The specificity and sensitivity of the assay were 100 and 96%, respectively, which confirms the assay's reliability for diagnostic and epidemiological studies . The assay can detect as few as 5 to 10 cells in a 10-ml water sample or 0.1 pg of P . aeruginosa DNA per reaction mixture (5 microliters) by ethidium bromide staining of an agarose gel . Ten-times-lower concentrations were detected by hybridization with a digoxigenin-labeled oligonucleotide probe internal to the PCR product . With this PCR method, ETA-positive P . aeruginosa was detected in animal cage water samples at a level of 40 cells per ml . This method is rapid and less cumbersome than other diagnostic methods for the identification of P . aeruginosa strains . The method described can be used to detect a low level of P . aeruginosa from environmental and clinical samples without the use of selective media or additional biochemical tests. Appl Environ Microbiol, 1994 Oct, 60(10), 3679 - 87 Cloning and heterologous expression of a gene encoding an alkane-induced extracellular protein involved in alkane assimilation from Pseudomonas aeruginosa; Hardegger M et al.; Pseudomonas aeruginosa PG201 produces a 16-kDa extracellular protein in media containing n-hexadecane as a carbon source but not in media containing glycerol or glucose . This protein was purified, and the N-terminal amino acid sequence was determined . The amino acid composition of the protein was found to be very similar to that of the so-called protein-like activator for n-alkane oxidation (PA) from P . aeruginosa S7B1 . This extracellular protein was previously characterized (K . Hisatsuka, T . Nakahara, Y . Minoda, and K . Yamada, Agric . Biol . Chem . 41:445-450, 1977) and found to stimulate the growth of P . aeruginosa on n-hexadecane and to possess emulsifying activity . To study the role(s) of the PA protein and to make it accessible for possible future applications, we have cloned the PA-encoding (pra) gene and determined its nucleotide sequence . This analysis revealed a protein-coding region of 162 amino acids, with the first 25 residues being reminiscent of those of a typical bacterial signal sequence . The pra gene was inactivated by insertional mutagenesis, and the resulting strain was found to lack extracellular PA protein and to be retarded in its growth in n-hexadecane-containing media . These results are consistent with the growth stimulatory role of the PA protein . The pra gene was expressed in Escherichia coli, and substantial amounts of the recombinant protein were found in the extracellular growth medium . The recombinant protein was purified by metal chelate affinity chromatography . The ability to produce secreted PA protein by E . coli provides a simple and safe means to analyze its function(s) in alkane assimilation in the future. Immun Infekt, 1994 Oct, 22(5), 177 - 80 {Extracellular enzymes of Pseudomonas aeruginosa as virulence factors}; Jaeger KE; Pseudomonas aeruginosa is an opportunistic pathogen causing a variety of diseases, especially in immunocompromised patients like those suffering from cystic fibrosis (CF) where these bacteria preferentially colonize the bronchopulmonary tract . A high intrinsic antibiotic resistance and its ability to synthesize and secrete numerous different virulence factors are regarded as biological properties contributing to the pathogenicity of P . aeruginosa . Among the virulence factors are many enzymes which have been characterized in detail with respect to their molecular properties . Environmental factors regulating the synthesis and release of extracellular enzymes have been identified as e.g . the concentration of Fe- and PO4-ions, choline, pH, and osmolarity . In addition, low molecular weight substances named autoinducers were identified as regulators which are synthesized by the bacteria . Therefore, P . aeruginosa represents an example for the remarkably complex relationship between pathogenic bacteria and their human host. J R Soc Med, 1994 Oct, 87(10), 581 - 3 Puncture wounds of the foot: can infective complications be avoided? Pennycook A, Makower R, O'Donnell AM. The bacteriological flora of the foot and shoe was studied concurrently in 200 volunteers without foot injuries, and 80 patients with puncture wounds of the foot . Seven of 28 child patients developed clinical infections, three with Pseudomonas aeruginosa . Eleven of 52 adult patients also developed infections . No patients developed infection if oral antibiotics were given within the first 24 h after injury (P < 0.05) . Oral antibiotic prophylaxis is recommended for puncture wounds of the foot. Ann Otol Rhinol Laryngol, 1994 Oct, 103(10), 771 - 4 Bacteriology of chronic suppurative otitis media; Erkan M et al.; Aspiration of exudate through an open perforation was performed in 183 patients with chronic otitis media . The pus was cultured aerobically and anaerobically . Aerobes only were isolated from 71 patients (39%); 20 patients (11%) had only anaerobes; and 91 patients (50%) had both aerobes and anaerobes . Only 1 specimen had no growth . There were 259 aerobic isolates . Pseudomonas aeruginosa was recovered from 68 patients . Other aerobes commonly recovered included Staphylococcus aureus and Klebsiella pneumoniae . There were 178 anaerobic isolates . Only anaerobic gram-positive cocci were isolated in 20 instances . Sixty-three Bacteroides isolates were recovered, including 12 Bacteroides fragilis group and 21 Bacteroides melaninogenicus. J Bacteriol, 1994 Oct, 176(19), 6023 - 9 Energy metabolism and alginate biosynthesis in Pseudomonas aeruginosa: role of the tricarboxylic acid cycle; Schlictman D et al.; Infection with mucoid, alginate-producing strains of Pseudomonas aeruginosa is the leading cause of mortality among patients with cystic fibrosis . Alginate production by P . aeruginosa is not constitutive but is triggered by stresses such as starvation . The algR2 (also termed algQ) gene has been previously identified as being necessary for mucoidy; an algR2 mutant strain is unable to produce alginate when grown at 37 degrees C . We show here that the levels of phosphorylated succinyl coenzyme A synthetase (Scs) and nucleoside diphosphate kinase (Ndk), which form a complex in P . aeruginosa, are reduced in the algR2 mutant . We were able to correlate the lower level of phosphorylated Scs with a decrease in Scs activity . Western blots (immunoblots) also showed a decreased level of Ndk in the algR2 mutant, but the presence of another kinase activity sensitive to Tween 20 provides the missing Ndk function . The effect of AlgR2 on tricarboxylic acid (TCA) cycle enzymes appears to be specific for Scs, since none of the other TCA cycle enzymes measured showed a significant decrease in activity . Furthermore, the ability of the algR2 mutant to grow on TCA cycle intermediates, but not glucose, is impaired . These data indicate that AlgR2 is responsible for maintaining proper operation of the TCA cycle and energy metabolism. J Bacteriol, 1994 Oct, 176(19), 6007 - 14 Transcriptional analysis of the Pseudomonas aeruginosa genes algR, algB, and algD reveals a hierarchy of alginate gene expression which is modulated by algT; Wozniak DJ et al.; Strains of Pseudomonas aeruginosa which colonize and infect the lungs of cystic fibrosis patients have a mucoid colony morphology due to the overproduction of the exopolysaccharide alginate . The response regulators AlgB and AlgR are required for the transcription of algD, a tightly regulated gene encoding GDP-mannose dehydrogenase, which is critical for P . aeruginosa alginate biosynthesis . Previous studies indicated that mutations in the algT gene of mucoid FRD1 P . aeruginosa result in nonmucoid derivatives . However, the specific role for algT in alginate gene regulation has not been elucidated . In this study, transcription of algB, algD, and algR was characterized by gene fusion and primer extension analysis . Expression of algR and algD was abolished in P . aeruginosa strains containing algT::Tn501 insertions because of lack of transcription initiation at the algR and algD promoters . An algR mutation was constructed in FRD1, and this resulted in the loss of alginate production and a dramatic decrease in algD transcription . RNA and gene fusion analysis revealed that algB is not required for algR expression, nor is algR necessary for transcription of algB . Thus, with the exception of a requirement for AlgT, the AlgB and AlgR pathways appear to be independent of each other . In gel band mobility shift assays, a protein(s) present in extracts from mucoid and algB and algR mutant P . aeruginosa strains formed a specific complex with algD sequences located immediately upstream of the start of transcription . No binding to these sequences was observed when extracts from algT mutant strains were examined . A model proposed suggests that a hierarchy of alginate gene expression exists in which AlgT is required for transcription of the response regulators algB and algR, which in turn are necessary for algD expression . AlgT or a protein under algT control also binds to sequences located within the algD promoter. Infect Immun, 1994 Oct, 62(10), 4572 - 9 Pili and lipopolysaccharide of Pseudomonas aeruginosa bind to the glycolipid asialo GM1; Gupta SK et al.; This study investigated which adhesins of Pseudomonas aeruginosa interact with the glycolipid asialo GM1, using solid-phase binding and thin-layer chromatography assays . Radioiodinated pili and flagella contaminated with lipopolysaccharide (LPS) bound to the glycolipid . When LPS was reduced to acceptable levels in pilus and flagellum samples, only pili specifically bound to the glycolipid . Commercial, radiolabeled LPS as well as whole bacteria of strain ATCC 19660 also bound to asialo GM1 . Binding was specific, competitive, and saturable . Organ cultures of whole mouse eyes and scanning electron microscopy techniques were used also, and strain ATCC 19660 was inhibited from corneal binding by exogenous pili or commercial LPS and inhibition was concentration dependent for both . Binding of radiolabeled strain ATCC 19660 bacteria to neutral lipids extracted from bovine corneal epithelial tissue showed that the bacteria bound to a glycolipid which migrated at a position similar to that of an asialo GM1 standard and that the glycolipid stained positively with an antibody specific for asialo GM1 . The data provide evidence that pili (reduced LPS) and LPS of P . aeruginosa bind to asialo GM1 glycolipid and that the glycolipid is not restricted to the mouse cornea. Infect Immun, 1994 Oct, 62(10), 4481 - 7 Cytotoxicity of Pseudomonas aeruginosa internal lectin PA-I to respiratory epithelial cells in primary culture; Bajolet-Laudinat O et al.; Pseudomonas aeruginosa is the most important bacterial pathogen associated with chronic airway infection, especially in cystic fibrosis . We addressed the question of whether the galactophilic internal lectin of P . aeruginosa (PA-I) could represent a virulence factor for the respiratory epithelium . PA-I lectin was localized in all the bacteria of P . aeruginosa ATCC 33347 as determined by immunofluorescence staining . We investigated the dose-dependent effect of P . aeruginosa PA-I lectin on the growth, the ciliary beating frequency, and the morphology of human respiratory cells in primary culture of nasal polyps collected from non-cystic fibrosis patients . PA-I lectin significantly (P < 0.01) inhibited the growth of respiratory cells at a concentration of > or = 10 micrograms/ml . The percentage of active ciliated cell surface of the cultures decreased significantly (P < 0.05) at a PA-I lectin concentration of 50 micrograms/ml . Exposed to a low concentration of PA-I lectin (10 micrograms/ml), respiratory epithelial cells showed intracytoplasmic vacuoles when examined by light and transmission electron microscopy . At a higher concentration of PA-I lectin (100 micrograms/ml), major cell damage and severe epithelial shedding occurred . These results demonstrate that the P . aeruginosa internal PA-I lectin has a dose-dependent cytotoxic effect on respiratory epithelial cells in vitro . The P . aeruginosa PA-I lectin may represent a virulence factor by contributing to the respiratory epithelial damage during P . aeruginosa respiratory infections. Infect Immun, 1994 Oct, 62(10), 4145 - 52 Role of interleukin-8 (IL-8) and an inhibitory effect of erythromycin on IL-8 release in the airways of patients with chronic airway diseases; Oishi K et al.; To evaluate of the role of interleukin-8 (IL-8), a chemotactic cytokine, in the continuous neutrophil accumulation in the airways of patients with chronic airway disease (CAD) and persistent Pseudomonas aeruginosa infection, we investigated the cell population, IL-8 levels, IL-1 beta levels, tumor necrosis factor (TNF) activities, and neutrophil elastase (NE) activities of bronchoalveolar lavage (BAL) fluids in 17 CAD patients (with P . aeruginosa infections {CAD+PA}, n = 9; without any bacterial infections {CAD-PA}, n = 8) and 8 normal volunteers . We found significant elevations of neutrophil numbers, IL-8/albumin ratios, and NE/albumin ratios in BAL fluids from CAD patients, in the following rank order: CAD+PA > CAD-PA > normal volunteers . IL-1 beta/albumin ratios were elevated only in CAD+PA, while no TNF bioactivity was detected in BAL fluids . The neutrophil numbers correlated significantly with the IL-8/albumin ratios and NE/albumin ratios in the BAL fluids of CAD patients . When anti-human IL-8 immunoglobulin G was used for neutralizing neutrophil chemotactic factor (NCF) activities in BAL fluids, the mean reduction rate of NCF activities in CAD+PA patients was significantly higher than that in CAD-PA patients . We also evaluated the effects of low-dose, long-term erythromycin therapy in BAL fluids from three CAD+PA and two CAD-PA patients . Treatment with erythromycin caused significant reductions of neutrophil numbers, IL-8/albumin ratios, and NE/albumin ratios in BAL fluids from these patients . To elucidate the mechanism of erythromycin therapy, we also examined whether erythromycin suppressed IL-8 production by human alveolar macrophages and neutrophils in vitro . We demonstrated a moderate inhibitory effect of erythromycin on IL-8 production in Pseudomonas-stimulated neutrophils but not in alveolar macrophages . Our data support the view that persistent P . aeruginosa infection enhances IL-8 production and IL-8-derived NCF activity, causing neutrophil accumulation in the airways and the progressive lung injuries observed in patients with CAD . The clinical efficacy of erythromycin therapy for CAD patients might be partly mediated through a reduced IL-8 production, diminishing neutrophil accumulation and NE release in the airways. Infect Immun, 1994 Oct, 62(10), 4118 - 23 Alteration of the pilin adhesin of Pseudomonas aeruginosa PAO results in normal pilus biogenesis but a loss of adherence to human pneumocyte cells and decreased virulence in mice; Farinha MA et al.; The disulfide loop domain of Pseudomonas aeruginosa PAO pilin was altered by insertion of a chloramphenicol acetyltransferase gene into the pilin gene so that the C-terminal nine amino acids were replaced with 11 new amino acids . The altered pilin gene was transferred into wild-type PAO by recombination, where it did not affect normal piliation as observed by transmission electron microscopy or change of sensitivity to f116, PO4, B9, and Pf1 pilus-specific bacteriophages . However, the binding to human pneumocyte A549 cells was markedly reduced when tested in an in vitro binding assay (2 to 6 bacteria bound per A549 cell for the mutant bacteria compared with 50 bacteria per A549 cell for the wild-type bacteria) . Additionally, when susceptible A.BY/SnJ mice were challenged with wild-type P . aeruginosa PAO and with P . aeruginosa PAO-MP (altered pilin gene), a 50% lethal dose of 3 x 10(6) bacteria per mouse was observed for PAO-MP compared with 7 x 10(4) bacteria per mouse for PAO . Approximately 90 of the adherence capability of P . aeruginosa PAO is seemingly attributable to the C-terminal disulfide loop adherence domain of pili . The pilus adherence function contributes significantly to the virulence of P . aeruginosa PAO in the A.BY/SnJ mouse infection model. Indian J Pathol Microbiol, 1994 Oct, 37(4), 409 - 14 Bacteriological study of bronchoalveolar lavage in immunocompromised hosts; Kumari GR et al.; Sixty bronchoalveolar lavage samples collected from lung cancer patients attending the Pulmonary Tuberculosis and Chest Diseases Unit of the Kasturba Medical College Hospital, Manipal; were cultured for both the aerobic and anaerobic organisms . Fifty nine samples yielded bacteria in pure culture . Pseudomonas aeruginosa (34.8%) was the commonest aerobe, Peptostreptococcus was the commonest anaerobe (45.2%) isolated . Bacteroides fragilis was isolated in (23.8%) of cases . Gentamycin was found to be effective against aerobes, Metronidazole and Rifampicin against anaerobes. New Microbiol, 1994 Oct, 17(4), 297 - 305 Adhesion of mucoid uropathogenic strains of Pseudomonas aeruginosa to HEp-2 cells; Zanetti S et al.; We studied a pili-independent adhesion mechanism to HEp-2 cells present in mucoid uropathogenic strains of Pseudomonas aeruginosa . Our data demonstrate that bacterial adhesion to HEp-2 cell surfaces is time dependent and that the phenotypes involved are influenced by bacterial growth conditions . Sonicated bacterial extracts competitively inhibit the adherence of homologous strains to HEp-2 cells . Adhesins that are heat and trypsin sensitive are located on the surface of the bacterial outer membrane . Immunogenic 55 kDa surface protein is required for the adherence to HEp-2 cell surfaces of non-piliated mucosal uropathogenic P . aeruginosa strains. J Ethnopharmacol, 1994 Oct, 44(2), 127 - 30 In vitro antibacterial screening of Cryptolepis sanguinolenta alkaloids; Paulo A et al.; The ethanol and aqueous crude extracts and five alkaloids isolated from the roots of Crytolepis sanguinolenta (Lindl.) Schlechter were screened for antibacterial activity against 7 reference strains by the twofold serial broth microdilution assay . The ethanol extract and the alkaloids cryptolepine and cryptoheptine inhibited the growth of all strains tested except that of Pseudomonas aeruginosa. Infect Control Hosp Epidemiol, 1994 Oct, 15(10), 652 - 7 Investigation of a pseudo-outbreak of orthopedic infections caused by Pseudomonas aeruginosa; Forman W et al.; OBJECTIVE: To report a pseudoepidemic of Pseudomonas aeruginosa infections discovered during an investigation of postoperative joint infections . DESIGN: A retrospective review of case patients' hospital charts, operative reports, and laboratory data, as well as environmental culturing, polymerase chain reaction (PCR) ribotyping of outbreak isolates, and in vitro analysis of P aeruginosa growth characteristics . SETTING: A 510-bed, university-affiliated adult tertiary care hospital . RESULTS: Between October 1 and December 1, 1992, seven postsurgical joint infections were diagnosed, including four caused by P aeruginosa . A bottle of "sterile" saline used to process tissue specimens was found to be contaminated with P aeruginosa . Further investigation revealed that P aeruginosa had grown from seven additional tissue cultures, all of which had been processed with the contaminated saline . PCR ribotypes of the contaminant matched those of the clinical isolates . In vitro, P aeruginosa strains were viable in commercial nonbacteriostatic saline, but never caused visible turbidity . Six patients received antibiotics for their presumed infections; four patients had peripherally inserted central catheters placed, and one experienced severe anaphylactic reactions to several antibiotics . CONCLUSIONS: Pseudoepidemics due to common organisms are often difficult to detect, and delayed recognition can result in substantial morbidity . This outbreak investigation illustrates the potential for contamination of diluents in the microbiology laboratory and emphasizes the need for meticulous quality control. Antimicrob Agents Chemother, 1994 Oct, 38(10), 2351 - 6 Pharmacodynamics of piperacillin alone and in combination with tazobactam against piperacillin-resistant and -susceptible organisms in an in vitro model of infection; Strayer AH et al.; The pharmacodynamics of dosage regimens of piperacillin alone or in combination with tazobactam against piperacillin-resistant or -susceptible bacteria were studied in an in vitro model of infection . Experiments were conducted by using a fixed daily exposure of 12 g of piperacillin, given as 3 g alone or in combination with tazobactam at 0.375 g every 6 h, or the same total dose of the combination given as 4 g of piperacillin plus 0.5 g of tazobactam every 8 h . The addition of tazobactam to piperacillin, irrespective of the dosing interval, did not alter the killing of piperacillin-susceptible organisms (Escherichia coli J53 and Pseudomonas aeruginosa ATCC 27853) . In contrast, experiments with an isogenic TEM-3-containing transconjugant of E . coli J53 (E . coli J53.2-TEM-3) that was resistant to piperacillin (MIC, 128 micrograms/ml) showed that the addition of tazobactam resulted in bacterial killing similar to that observed with the wild-type strain . Although tazobactam concentrations fell to less than 4 mg/liter (the concentration associated with a reduction in the piperacillin MIC from 128 to 2 mg/liter) 2 to 3 h after a dose, a similar degree of bacterial killing was observed when the same total 24-h dose of piperacillin-tazobactam was fractionated into dosing intervals of every 6 or 8 h . Investigations with Staphylococcus aureus 7176 (piperacillin MIC, 128 micrograms/ml) showed that the addition of tazobactam, again irrespective of dosing interval, also resulted in net bacterial killing which was not seen with piperacillin alone . These data support the use of extended dosing intervals (every 8 h) of piperacillin-tazobactam in the treatment of infections caused by piperacillin-resistant bacteria. Antimicrob Agents Chemother, 1994 Oct, 38(10), 2306 - 10 Antibodies against chromosomal beta-lactamase; Giwercman B et al.; A murine monoclonal anti-chromosomal beta-lactamase antibody was developed and an immunoblotting technique was used to study the presence of serum and sputum antibodies against Pseudomonas aeruginosa chromosomal group 1 beta-lactamase in patients with cystic fibrosis (CF) . The serum antibody response was studied with serum samples collected in 1992 from 56 CF patients in a cross-sectional study and with serum samples from 18 CF patients in a longitudinal study . Anti-beta-lactamase immunoglobulin G antibodies were present in all of the serum samples from the patients with chronic bronchopulmonary P . aeruginosa infection (CF + P) but in none of the CF patients with no or intermittent P . aeruginosa infection . Anti-beta-lactamase antibodies were present in serum from CF + P patients after six antipseudomonal courses (median) and correlated with infection with a beta-lactam-resistant strain of P . aeruginosa . The sputum antibody response and the beta-lactamase activity in sputum samples from 14 of the CF + P patients were also studied . beta-lactamase antibodies were present in 10 of these samples . P . aeruginosa strains isolated from these samples were partially derepressed, producing group 1 cephalosporinase . We found a wide range of chromosomal beta-lactamase activity in the sputum samples, with no correlation with basal or induced activity of beta-lactamase expression . The presence of anti-beta-lactamase antibodies in endobronchial sputum could be an important factor in the defense against the infection . On the other hand, immune complexes between the beta-lactamase and corresponding antibodies could play a role in the pathogenesis of bronchopulmonary injury in CF by mediating hyperimmune reactions. Eur Respir J, 1994 Oct, 7(10), 1754 - 8 Pseudomonas aeruginosa exotoxin A induces pulmonary endothelial cytotoxicity: protection by dibutyryl-cAMP; Bourke WJ et al.; In pseudomonal septicaemia, serum levels of antibody to exotoxin A have been demonstrated to be an important independent predictor of survival . Previously, we have demonstrated that exotoxin A directly injures pulmonary endothelial cells, and that dibutyryl-cyclic adenosine monophosphate (Db-cAMP) can attenuate this injury . The object of this study was to examine the mechanisms of this pulmonary endothelial cell injury and the mechanism of Db-cAMP protection . The effects of differing duration of ex