Nutrition, 1999 Jan, 15(1), 11 - 7 Phenotypic changes in the lipopolysaccharide of Pseudomonas aeruginosa and Escherichia coli grown in milk-based enteral nutrition solutions; Hodgson I et al.; Previous studies have shown enteral nutritional solutions (ENS) contaminated with large numbers of microorganisms from the environment or gastrointestinal (GI) tract of patients have caused respiratory infections, acute and chronic enteritis, and septicemia . The introduction of "closed" enteral feeding systems has been used to prevent contaminating organisms from entering enteral feeding systems in large numbers . However, there is some discussion as to whether this has been an effective measure in reducing ENS-related infections because there is anecdotal evidence to suggest that disease processes resulting from enteral feeding are still commonplace in the hospital and home . This is because there is very little information about the growth of microorganisms in ENS and whether growth in ENS may affect the virulence and pathogenicity of microorganisms . This study shows that Escherichia coli and Pseudomonas aeruginosa may grow at 25 degrees C from either high or low initial numbers to up to 9.2 log colony-forming units per mL in a range of milk-based ENS . However, these organisms did not grow in the fruit-based ENS . The effect on the lipopolysaccharide (LPS) of culturing E . coli and P . aeruginosa in milk-based ENS as opposed to standard laboratory media was examined using polyacrylamide gel electrophoresis . We found that there were significant qualitative changes in the phenotype of O-polysaccharide side chains of the LPS from these organisms . O-polysaccharide is known to mediate in the complement, antibiotic and bile resistance, and affect adherence . Therefore, changes in the virulence and pathogenicity of these microorganisms when cultured in ENS may be indicated . Thus, the study provides further evidence for reevaluating the microbiologic and immunologic effects of enteral feeding, especially on the microbial flora of the GI tract. Zentralbl Hyg Umweltmed, 1998 Dec, 201(4-5), 349 - 55 Detection of Pseudomonas aeruginosa in bronchial and tracheal aspirates by PCR by amplification of the exotoxin A gene}; Hummel A et al.; For the first time, a PCR test based on the amplification of the Exotoxin A was evaluated for its ability to rapidly detect Pseudomonas aeruginosa in tracheal and bronchial aspirates from mechanically ventilated patients . The reaction is based on the amplification of a 396 bp region within the Exotoxin A gene . The results show that this PCR-method is suitable for the detection of Pseudomonas aeruginosa even in clinical samples such as aspirates . Among the 380 clinical samples tested in this way, 57 were found to be positive while only 36 were positive using routine culture . In conclusion, these results suggest that the PCR-method mentioned above can be used to provide a specific, rapid, simple, and highly sensitive detection of Pseudomonas aeruginosa in clinical samples. Infect Immun, 1999 Feb, 67(2), 972 - 5 Pseudomonas aeruginosa keratitis in knockout mice deficient in intercellular adhesion molecule 1; Hobden JA et al.; In this study, the role of intercellular adhesion molecule 1 (ICAM-1) in the pathogenesis of Pseudomonas aeruginosa keratitis was examined by using inbred ICAM-1-deficient knockout mice . These mice had significantly less (P </= 0.02) ocular disease than wild-type mice, suggesting that ICAM-1 contributes to a more severe disease response following P . aeruginosa infection. Infect Immun, 1999 Feb, 67(2), 914 - 20 Regulation of ExoS production and secretion by Pseudomonas aeruginosa in response to tissue culture conditions; Vallis AJ et al.; This study was initiated to characterize the regulation and secretion of ExoS by Pseudomonas aeruginosa during contact with eukaryotic cells . The production of ExoS was monitored by a sensitive ADP-ribosyltransferase activity assay, and specific activities were calculated for supernatant and cell-associated fractions . Time course analysis indicated that ExoS was produced after a lag period, suggesting that induction of the regulon is necessary for the expression of detectable amounts of enzyme activity . Under tissue culture growth conditions, ExoS was induced when P . aeruginosa was in contact with Chinese hamster ovary (CHO) cells or after growth in tissue culture medium with serum . The serum induction of ExoS appeared to result in generalized type III secretion, while induction by contact with CHO cells appeared to result in polarized type III secretion . Mutants in the type III secretory system that express a null phenotype for ExoS production in bacteriological medium produced but did not secrete the enzyme when P . aeruginosa was grown under inducing conditions in tissue culture medium . These results suggest that both induction and secretion of ExoS may differ when the bacteria are exposed to different growth environments . The putative type III translocation proteins and secretion apparatus of P . aeruginosa were required for translocation of bacterial factors that mediate changes in CHO cell morphology during infection. Infect Immun, 1999 Feb, 67(2), 717 - 25 The R-type pyocin of Pseudomonas aeruginosa C is a bacteriophage tail-like particle that contains single-stranded DNA; Lee FK et al.; Pseudomonas aeruginosa R-type pyocin particles have been described as bacteriocins that resemble bacteriophage tail-like structures . Because of their unusual structure, we reexamined whether they contained nucleic acids . Our data indicated that pyocin particles isolated from P . aeruginosa C (pyocin C) contain DNA . Probes generated from this DNA by the random-primer extension method hybridized to distinct bands in restriction endonuclease-digested P . aeruginosa C genomic DNA . These probes also hybridized to genomic DNA from 6 of 18 P . aeruginosa strains that produced R-type pyocins . Asymmetric PCR, complementary oligonucleotide hybridization, and electron microscopy indicated that pyocin C particles contained closed circular single-stranded DNA, approximately 4.0 kb in length . Examination of total intracellular DNA from mitomycin C-induced cultures revealed the presence of two extrachromosomal DNA molecules, a double-stranded molecule and a single-stranded molecule, which hybridized to pyocin DNA . Sequence analysis of 7,480 nucleotides of P . aeruginosa C chromosomal DNA containing the pyocin DNA indicated the presence of pyocin open reading frames with similarities to open reading frames from filamentous phages and cryptic phage elements . We did not observe any similarities to known phage structural proteins or previously characterized pseudomonal prt genes expressing R-type pyocin structural proteins . These studies demonstrate that pyocin particles from P . aeruginosa C are defective phages that contain a novel closed circular single-stranded DNA and that this DNA was derived from the chromosome of P . aeruginosa C. Biol Chem, 1998 Dec, 379(12), 1427 - 32 Pterin-4a-carbinolamine dehydratase from Pseudomonas aeruginosa: characterization, catalytic mechanism and comparison to the human enzyme; Koster S et al.; The three-dimensional structure of pterin-4a-carbinolamine dehydratase (PCD) from Pseudomonas aeruginosa has been solved . Based on this we have investigated the roles of putative active center residues through functional replacement by site-directed mutagenesis . Three histidines, His73, His74 and His91, appear to be involved in dehydration catalysis . The three-dimensional positions of these residues match those of corresponding histidines at the active center of human PCD . Based on the coincidence of catalytic parameters, and on the similar effects induced by the mutations, it is concluded that the substrate binding mode and the reaction mechanisms of bacterial and human PCD are basically identical. Proc Natl Acad Sci U S A, 1999 Jan 19, 96(2), 715 - 20 Killing of Caenorhabditis elegans by Pseudomonas aeruginosa used to model mammalian bacterial pathogenesis; Tan MW et al.; We show that a single clinical isolate of the human opportunistic pathogen Pseudomonas aeruginosa (strain PA14), which previously was shown to be pathogenic in mice and plants, also kills Caenorhabditis elegans . The rate of PA14-mediated killing of C . elegans depends on the composition of the agar medium on which PA14 is grown . When PA14 is grown on minimal medium, killing occurs over the course of several days and is referred to as "slow" killing . When PA14 is grown on high-osmolarity medium, killing occurs over the course of several hours and is referred to as "fast" killing . Several lines of evidence, including the fact that heat-killed bacteria are still capable of fast but not slow killing of C . elegans, indicate that fast and slow killing occur by distinct mechanisms . Slow killing involves an infection-like process and correlates with the accumulation of PA14 within worm intestines . Among 10 PA14 virulence-related mutants that had been shown previously to affect pathogenicity in plants and mice, 6 were less effective in killing C . elegans under both fast- and slow-killing conditions, indicating a high degree of commonalty among the P . aeruginosa factors required for pathogenicity in disparate eukaryotic hosts . Thus, we show that a C . elegans pathogenicity model that is genetically tractable from the perspectives of both host and pathogen can be used to model mammalian bacterial pathogenesis. J Exp Med, 1999 Jan 18, 189(2), 341 - 6 Targeted disruption of migration inhibitory factor gene reveals its critical role in sepsis; Bozza M et al.; To study the biologic role of migration inhibitory factor (MIF), a pleiotropic cytokine, we generated a mouse strain lacking MIF by gene targeting in embryonic stem cells . Analysis of the role of MIF during sepsis showed that MIF-/- mice were resistant to the lethal effects of high dose bacterial lipopolysaccharide (LPS), or Staphylococcus aureus enterotoxin B (SEB) with D-galactosamine and had lower plasma levels of tumor necrosis factor alpha (TNF-alpha) than did wild-type mice, but normal levels of interleukin (IL)-6 and IL-10 . When stimulated with LPS and interferon gamma, macrophages from MIF-/- mice showed diminished production of TNF-alpha, normal IL-6 and IL-12, and increased production of nitric oxide . MIF-/- animals cleared gram-negative bacteria Pseudomonas aeruginosa instilled into the trachea better than did wild-type mice and had diminished neutrophil accumulation in their bronchoalveolar fluid compared to the wild-type mice . Thioglycollate elicited peritoneal exudates in uninfected MIF-/- mice, but showed normal neutrophil accumulation . Finally, the findings of enhanced resistance to P . aeruginosa and resistance to endotoxin-induced lethal shock suggest that the counteraction or neutralization of MIF may serve as an adjunct therapy in sepsis. Eur J Med Res, 1999 Jan 26, 4(1), 27 - 30 Pseudomonas aeruginosa orbital phlegmon in a patient treated for myelodysplastic syndrome with concomitant Sjögren's syndrome; Giagounidis AA et al.; Pseudomonas aeruginosa orbital infections have been described very rarely in patients with neutropenia after chemotherapy . We report the case of a woman with the unusual association of Sjogren's disease and myelodysplasia, who suffered from a Pseudomonas aeruginosa orbital phlegmon after chemotherapy for her myelodysplastic syndrome . Partial intestinal antibiotic decontamination with ciprofloxacine did not prevent the infection . She was treated successfully with intravenous ceftazidime, netilmicin and granulocyte-colony stimulating factor (G-CSF) . The normalization of the granulocyte count seems to play a crucial role for recovery . We present the clinical and radiological findings, discuss the therapy and review the literature concerning ocular infections due to Pseudomonas . Other infections due to this germ in immunocompromised hosts are briefly reviewed. J Infect, 1998 Nov, 37(3), 217 - 23 The effect of subinhibitory concentrations of antibiotics on adherence of Pseudomonas aeruginosa to cystic fibrosis (CF) and non-CF-affected tracheal epithelial cells; Wolter JM et al.; OBJECTIVES: this project investigated the proposition that Pseudomonas aeruginosa binds preferentially to cystic fibrosis (CF) respiratory cells . In addition, disadherence of P . aeruginosa by subinhibitory concentrations of antibiotics was examined as a possible explanation for clinical improvement seen in chronically colonized patients when treated with anti-pseudomonal agents . METHODS: we used a distinctive HPV-transformed respiratory cell line to compare adherence of two strains of P . aeruginosa to CF and non-CF respiratory cells . The effect of subinhibitory concentrations of antipseudomonal antibiotics on adherence of P . aeruginosa to cell monolayers was measured . RESULTS: piliated P . aeruginosa bound significantly better to CF than non-CF-affected cells (P=0.003) . Adherence was significantly reduced when organisms were preincubated in subinhibitory concentrations of antibiotics overnight (P<0.001) . CONCLUSIONS: these results support the presence of an altered receptor present on CF-affected cells that binds piliated strains of P . aeruginosa . Bacterial disadherence due to subinhibitory concentrations of antibiotics may partially explain why clinical improvement is observed in CF patients with acute respiratory exacerbations. Lett Appl Microbiol, 1998 Dec, 27(6), 372 - 4 Effect of divalent cations on permeabilizer-induced lysozyme lysis of Pseudomonas aeruginosa; Ayres HM et al.; The presence of divalent (Mg2+) ions greatly reduced the lysis of Pseudomonas aeruginosa strain G48 in a system at pH 7.8 or 9.0 consisting of ethylenediamine tetraacetic acid (EDTA), lysozyme and tris . Similar reductions in lysis occurred when EDTA was replaced by nitrilotriacetic acid, sodium citrate or sodium polyphosphate . The effect depended on the cation concentration . Mg2+ may replace cations removed from the outer membrane, or may effectively remove the permeabilizer from the system . The results suggest that the permeabilizing activity associated with these agents against this organism has a common basis in affecting the outer membrane. Tidsskr Nor Laegeforen, 1998 Nov 20, 118(28), 4370 - 5 {Bacteremia with granulocytopenia--microbiology and empiric antibiotic treatment}; Hammerstrom J et al.; This article sums up a retrospective analysis of 84 episodes of bacteraemia in acute leukaemia patients with severe neutropenia in a Norwegian teaching hospital during the period 1990-95 . Gram negative bacteria represented 54% of the blood culture isolates, all of which were susceptible to aminoglycosides, and nearly all to ceftazidime and imipenem . Penicillin/aminoglycoside was used as initial therapy in 43% of the episodes . Initial empiric therapy was modified in 52% of the events . Only 15% of patients receiving the penicillin/aminoglycoside combination actually had infections with organisms susceptible to penicillin . Only 2% of patients with gram negative infections received initial synergistic treatment with two effective drugs . Mortality from infections was 8% in acute leukaemia patients with documented bacteraemia . Deaths mainly occurred in patients with terminal leukaemia disease . Late breakthrough bacteraemias with Stenotrophomonas maltophilia and Pseudomonas aeruginosa caused 50% of all fatal infections . The analysis suggests that no patients died during initial bacteraemia with penicillin-resistant organisms treated with penicillin/aminoglycoside . The antibiotic susceptibility of the isolated bacteria was favourable compared to what has been found in other countries . For the time being, we believe that the ecological advantages of using penicillin/aminoglycoside as initial empiric treatment of febrile neutropenia are greater than the disadvantages. Pediatr Pulmonol, 1998 Dec, 26(6), 371 - 9 Severe viral respiratory infections in infants with cystic fibrosis; Armstrong D et al.; Limited data in children with cystic fibrosis (CF) suggest that respiratory viral infections during infancy result in substantial morbidity . Eighty of 101 (79%) infants with CF diagnosed by neonatal screening during 1991-1996 were recruited into a prospective, multiple-birth cohort study . We aimed to perform an initial, then annual bronchoalveolar lavage (BAL) for bacterial and viral culture, cytology, IL-8, and elastolytic activity over the following 2 years . When possible, BAL was also performed during any hospitalization for a pulmonary exacerbation, and additional specimens for viral culture were collected by nasopharyngeal aspiration . Thirteen infants undergoing bronchoscopy for congenital stridor served as disease controls . During infancy, 31 children (39%) were hospitalized for respiratory disease and 20 (65%) cases had an etiologic agent identified . Respiratory viruses were detected in 16/31 (52%) cases, including four with simultaneous bacterial infection . Another four were infected with Staphylococcus aureus . Respiratory syncytial virus predominated and was found in seven infants . In the absence of bacteria, those with viral infections had acute onset of respiratory distress, were not treated with antibiotics, and had an uncomplicated hospital course . Compared to noninfected CF subjects and controls, infected infants had elevated BAL inflammatory indices (P < 0.01) . Eleven of 31 (35%) hospitalized infants followed for 12-60 months acquired Pseudomonas aeruginosa, compared with only three of 49 (6%) subjects not hospitalized for respiratory symptoms during infancy (risk ratio 5.8, CI 1.9, 24) . We conclude that respiratory viruses are important causes of hospitalization in CF infants . While viral infections were self-limited, they were accompanied by airway inflammatory changes, and admission to hospital was associated with early acquisition of Pseudomonas aeruginosa and persistent respiratory symptoms. J Immunol, 1999 Jan 1, 162(1), 392 - 9 IL-10 is a major mediator of sepsis-induced impairment in lung antibacterial host defense; Steinhauser ML et al.; To explore the mechanism of immunosuppression associated with sepsis, we developed a murine model of sepsis-induced Pseudomonas aeruginosa pneumonia . CD-1 mice underwent either cecal ligation and 26-gauge needle puncture (CLP) or sham surgery, followed by the intratracheal (i.t.) administration of P . aeruginosa or saline . Survival in mice undergoing CLP followed 24 h later by the i.t . administration of saline or P . aeruginosa was 58% and 10%, respectively, whereas 95% of animals undergoing sham surgery followed by P . aeruginosa administration survived . Increased mortality in the CLP/P . aeruginosa group was attributable to markedly impaired lung bacterial clearance and the early development of P . aeruginosa bacteremia . The i.t . administration of bacteria to CLP-, but not sham-, operated mice resulted in an impressive intrapulmonary accumulation of neutrophils . Furthermore, P . aeruginosa challenge in septic mice resulted in a relative shift toward enhanced lung IL-10 production concomitant with a trend toward decreased IL-12 . The i.p., but not i.t., administration of IL-10 Abs given just before P . aeruginosa challenge in septic mice significantly improved both survival and clearance of bacteria from the lungs of septic animals administered P . aeruginosa . Finally, alveolar macrophages isolated from animals undergoing CLP displayed a marked impairment in the ability to ingest and kill P . aeruginosa ex vivo, and this defect was partially reversed by the in vivo neutralization of IL-10 . Collectively, these observations indicate that the septic response substantially impairs lung innate immunity to P . aeruginosa, and this effect is mediated primarily by endogenously produced IL-10. Microbiology, 1998 Dec, 144 ( Pt 12), 3379 - 86 Mutual stabilization of the XcpZ and XcpY components of the secretory apparatus in Pseudomonas aeruginosa; Michel G et al.; Protein secretion in gram-negative bacteria is often dependent on the general secretory pathway (GSP) . In Pseudomonas aeruginosa, this system requires at least 12 Xcp (Gsp) proteins, which are proposed to constitute a multiprotein complex localized in the bacterial envelope . Hitherto, little was known about the mutual interactions between Xcp proteins . In this study, mutants affected in the xcpZ gene encoding a bitopic inner-membrane protein were analysed to investigate the role of this protein in the architecture of the secretory machinery . The absence of XcpZ resulted in a decreased amount of XcpY . Reciprocally, XcpZ was not detectable in a xcpY mutant, demonstrating a mutual stabilization of these two proteins . These results strongly suggest that XcpZ and XcpY interact within the functional secretory apparatus. Public Health, 1998 Nov, 112(6), 415 - 7 Methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa isolation from pharyngeal swab cultures among the Japanese elderly at admission to a geriatric hospital; Washio M et al.; The isolation rate of Methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa from pharyngeal swab cultures in the Japanese elderly was studied at admission to a geriatric hospital which had long-term care units . The subjects were 233 consecutive patients who were admitted to K Hospital in the time period April 1994 to March 1996 . The isolation rate of MRSA and of Pseudomonas aeruginosa was 10.3% and 8.2% respectively . The proportions of the patients with severely to moderately limited Activities of Daily Living (ADL) (ADL score = 0-1) (P < 0.01), those with fever (P < 0.01), those with CRP positive (P = 0.04) and those with hypoalbuminemia (serum albumin < 3.5 g/dl) (P < 0.01) were higher in the MRSA positive patients than in the negative patients while the proportion of the patients with fever was higher in the Pseudomonas aeruginosa positive patients than in the negative patients (P < 0.02) . In the multiple logistic regression analysis, the limitation of ADL (ADL score 0-1 vs 2-3, OR = 1.54, 95% CI = 1.02-2.33) and fever (with vs without, OR = 1.77, 95% CI = 1.18-2.66) remained as risk factors for the isolation of MRSA while only fever (with vs without, OR = 1.67, 95% CI = 1.11-2.53) remained as a risk factor for the isolation of Pseudomonas aeruginosa. J Bacteriol, 1999 Jan, 181(2), 627 - 31 Dissecting the VanRS signal transduction pathway with specific inhibitors; Ulijasz AT et al.; The VanRS two-component signal transduction pathway from Enterococcus faecium was reconstituted in vitro from partially purified components and shown to be inhibited by the halophenyl isothiazolone LY-266,400, inhibitor A, a compound shown previously to reduce expression of the AlgR1-AlgR2 two-component signal transduction pathway in Pseudomonas aeruginosa (S . Roychoudhury, N . A . Zielinski, A . J . Ninfa, N . E . Allen, L . N . Jungheim, T . I . Nicas, and A . M . Chakrabarty, Proc . Natl . Acad . Sci . USA 90:965-969, 1993) . Inhibitor A attenuates phosphoryl transfer from VanS approximately P to VanR by its action on the ability of VanR to accept . We observed an apparent stimulatory effect of inhibitor A on VanS autophosphorylation which is attributable to the accumulation of VanS approximately P as an intermediate unable to transfer Pi to the inhibited VanR . Thus, inhibitor A acts on the second of two sequential steps which lead to transcriptional activation of the VanHAXYZ gene cluster and the resultant expression of vancomycin resistance. J Bacteriol, 1999 Jan, 181(2), 382 - 8 Assembly of XcpR in the cytoplasmic membrane is required for extracellular protein secretion in Pseudomonas aeruginosa; Ball G et al.; A broad range of extracellular proteins secreted by Pseudomonas aeruginosa use the type II or general secretory pathway (GSP) to reach the medium . This pathway requires the expression of at least 12 xcp gene products . XcpR, a putative nucleotide-binding protein, is essential for the secretion process across the outer membrane even though the protein contains no hydrophobic sequence that could target or anchor it to the bacterial envelope . For a better understanding of the relationship between XcpR and the other Xcp proteins which are located in the envelope, we have studied its subcellular localization . In a wild-type P . aeruginosa strain, XcpR was found associated with the cytoplasmic membrane . This association depends on the presence of the XcpY protein, which also appears to be necessary for XcpR stability . Functional complementation of an xcpY mutant required the XcpY protein to be expressed at a low level . Higher expression precluded the complementing activity of XcpY, although membrane association of XcpR was restored . This behavior suggested that an excess of free XcpY might interfere with the secretion by formation of inactive XcpR-XcpY complexes which cannot properly interact with their natural partners in the secretion machinery . These data show that a precise stoichiometric ratio between several components may be crucial for the functioning of the GSP. Burns, 1998 Nov, 24(7), 637 - 41 Pseudomonas infections in Tohid Burn Center, Iran; Rastegar Lari A et al.; Burn injury is a major public health problem in many areas of the world . Pseudomonas aeruginosa is one of the most common causes of burn wound infection in burn patients . Septicemia due to this organism is a major cause of mortality among burn patients . This study analyzed P . aeruginosa infections in the Tohid Burn Center in Tehran during 1995-1997 in order to estimate their frequency, antibiotic susceptibility and their role in burn morbidity . Among 2122 patients who were admitted during this study period, 3365 bacterial strains were isolated and the frequency of P . aeruginosa was 73.9% . This was followed by Staphylococcus aureus (9.1%) and other organisms (17%) in frequency . The frequency of P . aeruginosa resistant to gentamicin, carbenicillin, co-trimoxazole, ceftizoxime and tetracycline was over 95% and resistance to amikacin which was 49% in 1995, increased to 90% in 1997 . With the introduction of ciprofloxacin at our burn center, the frequency of P . aeruginosa resistance increased from 45% in 1995, to 82% in 1997 . P . aeruginosa was found more frequently in the ICU than in the wards . These findings show that P . aeruginosa remains the leading cause of nosocomial infections in our burn center . It is necessary to introduce urgent measures for restriction of the spread of P . aeruginosa infections in our burn center. Ann Fr Anesth Reanim, 1998, 17(10), 1239 - 42 {Intrathoracic drainage of a compressive pulmonary bulla in a patient receiving mechanical ventilation}; Sleth JC et al.; A lung suppuration may result in a lung bulla with its own course . We report such a case following a Pseudomonas aeruginosa pneumonia of the upper right lobe, after aspiration of gastric contents, in a 21-year-old tracheotomized patient in chronic post-traumatic coma . Mechanical ventilation (IPPV) was indicated because of respiratory insufficiency . The pneumonia was followed by an abscess and later a lung bulla, increasing in size under the effect of mechanical ventilation with progressive mediastinal compression . Surgery was contraindicated because of poor physical status . An acute episode of cardiac tamponade was controlled with an emergency transthoracic drain insertion into the bulla . The course was favourable after a drainage for 23 days and a persisting small cavity in the lung apex . All weaning attempts being unsuccessful, the patient was discharged under home mechanical ventilation . A CT-scan control 6 months later showed a normal lung parenchyma . The various alternative techniques to surgery for treatment of a lung bulla are discussed. J Med Microbiol, 1998 Feb, 47(2), 129 - 34 Effects of characterised Pseudomonas aeruginosa exopolysaccharides on adherence to human tracheal cells; Marty N et al.; This study evaluated, in vitro, the role of different Pseudomonas aeruginosa exopolysaccharides (EPS) in mediating adherence to human respiratory epithelial cells . Two mucoid and non-mucoid isogenic pairs of P aeruginosa strains isolated from patients with cystic fibrosis (CF) and bronchiectasis were used . Adherence was tested with human tracheal epithelial cell lines from CF and normal fetuses . The CF cells bound significantly more bacteria than the normal cells . The strain from the bronchiectasis patient was significantly more adherent than that from the CF patient and this difference was consistently most marked with the non-mucoid variant and with normal epithelial cells . The differing behaviour of mucoid CF and non-mucoid bronchiectasis strains reflected the chemical composition of their EPS: mainly alginate in the former and neutral polysaccharides in the latter . Additive inhibition experiments with chemically characterised EPS indicated that neutral polysaccharides associated with alginate may act as ligands for the adherence of P . aeruginosa to CF epithelial cells. N Engl J Med, 1999 Jan 7, 340(1), 23 - 30 Intermittent administration of inhaled tobramycin in patients with cystic fibrosis . Cystic Fibrosis Inhaled Tobramycin Study Group; Ramsey BW et al.; BACKGROUND AND METHODS: We conducted two multicenter, double-blind, placebo-controlled trials of intermittent administration of inhaled tobramycin in patients with cystic fibrosis and Pseudomonas aeruginosa infection . A total of 520 patients (mean age, 21 years) were randomly assigned to receive either 300 mg of inhaled tobramycin or placebo twice daily for four weeks, followed by four weeks with no study drug . Patients received treatment or placebo in three on-off cycles for a total of 24 weeks . The end points included pulmonary function, the density of P . aeruginosa in sputum, and hospitalization . RESULTS: The patients treated with inhaled tobramycin had an average increase in forced expiratory volume in one second (FEV1) of 10 percent at week 20 as compared with week 0, whereas the patients receiving placebo had a 2 percent decline in FEV1 (P<0.001) . In the tobramycin group, the density of P . aeruginosa decreased by an average of 0.8 log10 colony-forming units (CFU) per gram of expectorated sputum from week 0 to week 20, as compared with an increase of 0.3 log10 CFU per gram in the placebo group (P<0.001) . The patients in the tobramycin group were 26 percent (95 percent confidence interval, 2 to 43 percent) less likely to be hospitalized than those in the placebo group . Inhaled tobramycin was not associated with detectable ototoxic or nephrotoxic effects or with accumulation of the drug in serum . The proportion of patients with P . aeruginosa isolates for which the minimal inhibitory concentration of tobramycin was 8 microg per milliliter or higher increased from 25 percent at week 0 to 32 percent at week 24 in the tobramycin group, as compared with a decrease from 20 percent at week 0 to 17 percent at week 24 in the placebo group . CONCLUSIONS: In a 24-week study of patients with cystic fibrosis, intermittent administration of inhaled tobramycin was well tolerated and improved pulmonary function, decreased the density of P . aeruginosa in sputum, and decreased the risk of hospitalization. Int Arch Allergy Immunol, 1998 Dec, 117(4), 270 - 5 Flow-cytometric evaluation of oxidative burst in phagocytic cells of children with cystic fibrosis; Fruhwirth M et al.; OBJECTIVES: The objective of this study was to assess the dye 2', 7'-dichlorofluorescein (DCF) assay in screening for alterations in polymorphonuclear cell (PMN) and monocyte (MC) oxidative burst of cystic fibrosis (CF) patients . STUDY DESIGN: 56 CF patients aged between 2 and 20 years were investigated . Purified cells were stimulated with phorbolmyristate acetate (PMA) and zymosan (ZX) . A range for DCF fluorescence for PMA- and ZX-stimulated and non-stimulated cells was established based on data from 60 healthy controls . RESULTS: PMNs showed both enhancement and impairment . A deficient oxidative burst was detected in a total of 14 CF patients caused by abnormally high mean fluorescence intensity (MFI) of resting cells . Enhanced oxidative burst was seen in 6 CF patients . CF patients responded differently to PMA or ZX stimulation . Pseudomonas aeruginosa colonization significantly enhanced (p<0.005) the MFI of resting PMNs . MCs of CF patients showed a significantly (p<0.05) enhanced oxidative burst after stimulation with PMA compared to healthy controls, but no differences could be observed after stimulation with ZX . Serum concentrations of interleukin-6 were elevated in all CF patients, in particular in those with activation of both PMNs and MCs . CONCLUSION: The DCF assay shows for the first time the heterogeneity of the oxidative burst reaction in CF patients . In our opinion, the DCF assay is a reliable method for detecting pathological oxidative burst in CF patients. Eur Respir J, 1998 Dec, 12(6), 1388 - 96 Characterizing alterations in the pulmonary surfactant system in a rat model of Pseudomonas aeruginosa pneumonia; Vanderzwan J et al.; Bacterial pneumonia remains a significant cause of patient morbidity and mortality worldwide . Pulmonary surfactant serves to maintain homeostasis in the lung through the maintenance of alveolar stability and the regulation of the alveolar immune response . The purpose of this study was to characterize the lung injury and associated surfactant alterations in a rat model of acute Pseudomonas aeruginosa pneumonia . Pneumonia was induced in male Sprague-Dawley rats via intratracheal injection of 0.2 ml, phosphate-buffered saline (PBS) containing P . aeruginosa (6x10(8) colony-forming units x mL(-1)) . Control animals received 0.2 mL sterile PBS . Twenty-four hours after inoculation, the pneumonia group (PN) exhibited clinical signs of pneumonia including deficits in gas exchange, leukopenia and elevated arterial lactate levels . Morphological assessment confirmed the presence of pneumonia with airspaces filled with polymorphonuclear cells . Lung homogenate analysis demonstrated evidence of bacterial colonization of pneumonic lung tissue . Lung compliance was also significantly lower in the PN group . Lung lavage analysis of PN rats revealed the pooled surfactant levels to be lower and the surfactant function reduced compared to control rats . Surfactant composition was also found to be altered in PN rats . These results demonstrate that in Pseudomonas aeruginosa pneumonia, the pulmonary surfactant system is both poorly functioning and reduced in quantity . These alterations may contribute to the lung dysfunction characteristic of this disorder. Braz J Med Biol Res, 1998 Oct, 31(10), 1329 - 34 Endothelium-dependent vasodilation in response to Pseudomonas aeruginosa lipopolysaccharide: an in vitro study on canine arteries; Evora PR et al.; Early systemic arterial hypotension is a common clinical feature of Pseudomonas septicemia . To determine if Pseudomonas aeruginosa endotoxin induces the release of endothelium-derived nitric oxide (EDNO), an endogenous nitrovasodilator, segments of canine femoral, renal, hepatic, superior mesenteric, and left circumflex coronary arteries were suspended in organ chambers (physiological salt solution, 95% O2/5% CO2, pH 7.4, 37 degrees C) to measure isometric force . In arterial segments contracted with 2 microM prostaglandin F2 alpha, Pseudomonas endotoxin (lipopolysaccharide (LPS) serotype 10(Habs) from Pseudomonas aeruginosa (0.05 to 0.50 mg/ml) induced concentration-dependent relaxation of segments with endothelium (P < 0.05) but no significant change in tension of arteries without endothelium . Endothelium-dependent relaxation in response to Pseudomonas LPS occurred in the presence of 1 microM indomethacin, but could be blocked in the coronary artery with 10 microM NG-monomethyl-L-arginine (L-NMMA), a competitive inhibitor of nitric oxide synthesis from L-arginine . The inhibitory effect of L-NMMA on LPS-mediated vasorelaxation of the coronary artery could be reversed by exogenous 100 microM L-arginine but not by 100 microM D-arginine . These experiments indicate that Pseudomonas endotoxin induces synthesis of nitric oxide from L-arginine by the vascular endothelium . LPS-mediated production of EDNO by the endothelium, possibly through the action of constitutive nitric oxide synthase (NOSc), may decrease systemic vascular resistance and may be the mechanism of early hypotension characteristic of Pseudomonas septicemia. Arch Pharm Res, 1998 Oct, 21(5), 570 - 5 Formation, properties and antimicrobial activities of cotton xanthate-Cu(II)-homosulfamine complex; Ha NJ et al.; To develop a cotton derivatives with prolonged antimicrobial activities, homosulfamine (Hs) was coupled to cotton xanthate (CX) via chelate bond in the presence of Cu(II) ion by one- and two-bath processes . In one-bath process, CX was treated with Cu(II)-Hs solution . In two-bath process, CX was treated with Cu(II) ion solution to produce CX-Cu(II) complex, which was isolated and treated in turn with Hs solution . Effects of concentration, Cu(II)/Hs ratio, and pH on the binding of Hs were investigated at 10 degrees C . In one-bath process, binding of Hs took place readily with optimum pH around 5-6 . The amount of binding increased to give a maximum within 5 min and decreased slowly to establish an equilibrium within an hour . In two-bath process, binding of Hs was much lower than that of one-bath process . Release of Hs from CX-Cu(II)-Hs was investigated by batch and flow method . Antimicrobial activities of CX-Cu(II)-Hs were tested against Staphylococcus aureus, Bacillus subtilis, Pseudomonas aeruginosa and Escherichia coli and it showed prolonged activity compared to that of free Hs. Scand J Immunol, 1998 Dec, 48(6), 672 - 8 Small cytoplasmic antigens from Pseudomonas aeruginosa stimulate gammadelta T lymphocytes; Julia MR et al.; Gammadelta T lymphocytes respond to different bacterial antigens and transformed cells . The antigenic molecules responsible for this activity have been studied extensively in antigenic preparations from Mycobacterium . We describe here the in vitro effect of Pseudomonas aeruginosa on gammadelta T lymphocytes and the properties of the implicated compounds . We found a preferential gammadelta T-cell expansion when we used heat-treated P . aeruginosa preparations and a dose-dependent inhibition of proliferation of peripheral blood mononuclear cells (PBMC) when non-heat-treated antigens were studied . This expansion corresponded to a Vgamma9-positive subpopulation . In contrast to alphabeta T lymphocytes, the highest stimulatory activity was restricted to very small cytosolic compounds . This activity was protease resistant and phosphatase sensitive and always dependent on interleukin (IL)-2 or alphabeta T-cell activation . We concluded that the antigenic molecules from P . aeruginosa that activated gammadelta T lymphocytes were small, non-peptidic, phosphorylated compounds, similar to those previously described from Mycobacterium. Am J Respir Crit Care Med, 1999 Jan, 159(1), 258 - 61 Unopposed neutrophil elastase in bronchoalveolar lavage from transplant recipients with cystic fibrosis; Nunley D et al.; Large numbers of neutrophils with unopposed neutrophil elastase (NE) proteolytic activity are found in lower respiratory tract secretions from most patients with advanced cystic fibrosis (CF) . To determine whether antielastase defenses may be overwhelmed in epithelial lining fluid after lung transplantation, we measured NE activity (cleavage of the specific substrate, MeO-Suc-Ala-Ala-Pro-Val-pNA) in bronchoalveolar lavage fluids (BALF) obtained for surveillance or diagnostic purposes at various intervals (1 mo to 7 yr after transplantation) from 52 recipients who had undergone double or bilateral lung transplantation for end-stage CF . Unopposed NE activity was found in BALF from 14 recipients, most of whom also had >= 10(5) colony forming units (cfu) of Pseudomonas aeruginosa in BALF . Ten of the 14 recipients with unopposed NE in bronchoalveolar lavage (BAL) had developed obliterative bronchiolitis (OB), but only 8 of the 38 subjects without unopposed NE activity had OB (p = 0 . 002; Fisher exact test) . We conclude that antiprotease defenses in lower respiratory tract secretions of CF patients receiving lung allografts are sufficient in the majority of patients to prevent unopposed NE activity . However, the presence of unopposed NE activity in BAL from lung allografts of patients with CF is associated with progressive, irreversible OB and graft failure. J Appl Microbiol, 1998 Dec, 85(6), 925 - 32 Efficacy and mechanisms of action of sodium hypochlorite on Pseudomonas aeruginosa PAO1 phage F116; Maillard JY et al.; The Pseudomonas aeruginosa PAO1 phage F116 was used to investigate the viricidal activity and the mechanism of action of sodium hypochlorite . The bacteriophage was inactivated with a low concentration (0.0005% available chlorine) of the biocide prepared in tap water but it was less sensitive to a sodium hypochlorite solution prepared in ultra-pure water (0.0075% available chlorine) . For all the effective concentrations of sodium hypochlorite (i.e . producing at least 4 log reduction in phage titre), F116 was readily inactivated within 30 s . Electron microscopical investigations of the phage particles challenged with sodium hypochlorite showed a wide variety of deleterious effects, some of which have not been previously observed with other biocides . The wide range of structural alterations observed suggested that sodium hypochlorite has multiple target sites against F116 bacteriophage . A 30 s exposure to sodium hypochlorite (0.001% available chlorine) produced severe damage, the number and severity of which increased with a higher concentration (0.0075% available chlorine) and with a longer contact time . These observations suggested that sodium hypochlorite inactivated F116 bacteriophage by causing structural alterations to the phage head, tail and overall structure, hence possibly releasing the viral genome from damaged capsids in the surrounding media. Antimicrob Agents Chemother, 1999 Jan, 43(1), 129 - 33 Evaluation of several dosing regimens of cefepime, with various simulations of renal function, against clinical isolates of Pseudomonas aeruginosa in a pharmacodynamic infection model; Cappelletty DM; The objectives of this study were as follows: (i) to examine the killing activity of 2-g doses of cefepime against two clinical isolates (mucoid and nonmucoid) of Pseudomonas aeruginosa in a pharmacodynamic in vitro infection model, (ii) to compare the percentage of time above the MIC (T > MIC) for each of the regimens against P . aeruginosa, and (iii) to evaluate the area under the bactericidal curve for each regimen . Cefepime was administered at intervals of 8, 12, and 24 h with and without tobramycin, and two different levels of renal function were simulated: normal (creatinine clearance {CLCR} = 90 ml/min) and decreased (CRCL = 60 ml/min) . Also, the killing activity of cefepime with and without tobramycin was compared to the killing activity of ceftazidime (2 g every 8 h) with and without tobramycin . The T > MIC was 100% in the central chamber except for the regimen in which cefepime was administered every 12 h and the CLCR was 90 ml/min, which provided concentrations above the MIC for 92% of the dosing interval against the C31 (mucoid; MIC of cefepime, 4 microg/ml) isolate and for 75% of the interval against the C34 (nonmucoid; MIC of cefepime, 8 microg/ml) isolate . All cefepime and ceftazidime monotherapy simulations resulted in 99.9% killing of the nonmucoid isolate within 4 to 8 h and within 4 to 6 h, respectively . Against the mucoid isolate, 99.9% killing was achieved only with combination therapy . The results of this study indicate that cefepime dosed at 2 g every 12 h under conditions of normal renal function and every 24 h with decreased creatinine clearance (60 ml/min) is effective both as monotherapy and in combination therapy against a nonmucoid strain of P . aeruginosa . With cefepime MICs of 4 and 8 microg/ml, the single-agent regimens provided T > MIC values in the central chamber for 92 and >/=75% of the dosing interval against the mucoid and nonmucoid isolates, respectively . Cefepime dosed at 2 g every 12 h, with a creatinine clearance of 90 ml/min, and every 24 h, with a creatinine clearance of 60 ml/min, resulted in killing activity equivalent to that of ceftazidime dosed at 2 g every 8 h . None of the monotherapies provided adequate killing of the mucoid strain of P . aeruginosa despite drug concentrations being above the MIC for >/=92% of all dosing intervals . Finally, combination therapy with tobramycin and either cefepime or ceftazidime enhanced the killing of both the mucoid and nonmucoid P . aeruginosa isolates. Antimicrob Agents Chemother, 1999 Jan, 43(1), 62 - 6 Type II topoisomerase mutations in ciprofloxacin-resistant strains of Pseudomonas aeruginosa; Mouneimne H et al.; We determined the sequences of the quinolone resistance-determining regions of gyrA, gyrB, and parC genes for 30 clinical strains of Pseudomonas aeruginosa resistant to ciprofloxacin that were previously complemented by wild-type gyrA and gyrB plasmid-borne alleles and studied for their coresistance to imipenem (E . Cambau, E . Perani, C . Dib, C . Petinon, J . Trias, and V . Jarlier, Antimicrob . Agents Chemother . 39:2248-2252, 1995) . In the present study, we found mutations in type II topoisomerase genes for all strains . Twenty-eight strains had a missense mutation in gyrA (codon 83 or 87) . Ten of them had an additional mutation in parC (codon 80 or 84), including a novel mutation of Ser-80 to Trp, but all were fully complemented by a plasmid-borne wild-type gyrA allele . The remaining two strains harbored the first gyrB mutation described in P . aeruginosa, leading to the substitution of phenylalanine for serine 464 . The strains which had two mutations in type II topoisomerase genes (i.e., gyrA and parC) were significantly more resistant to fluoroquinolones than those with a single mutation in gyrA or gyrB (geometric mean MICs of ciprofloxacin, 39.4 versus 10.9 microg/ml, P < 0.01; geometric mean MICs of sparfloxacin, 64.0 versus 22.6, P < 0 . 01) . No mutant with a parC mutation alone was observed, which favors DNA gyrase being the primary target for fluoroquinolones . These results demonstrate that gyrA mutations are the major mechanism of resistance to fluoroquinolones for clinical strains of P . aeruginosa and that additional mutations in parC lead to a higher level of quinolone resistance. J Hosp Infect, 1998 Dec, 40(4), 313 - 23 Antimicrobial efficacy of biocides tested on skin using an ex-vivo test; Maillard JY et al.; An ex-vivo test was used to evaluate the activity of antimicrobials against three microorganisms, Escherichia coli, Pseudomonas aeruginosa and Staphylococcus aureus . The ex-vivo test is a carrier test using freshly excised animal skin samples maintained in viable conditions for a short period of time . Skin samples came from a veterinary practice and were excised from either dogs or cats . The antimicrobial activity of povidone iodine, chlorhexidine diacetate, cetrimide and benzalkonium chloride was also evaluated with suspension and glass-carrier tests . Generally, the activity of the antimicrobials tested was reduced when applied to the skin surface . Apart from povidone iodine (2%) against S . aureus, the biocides investigated failed to achieve a 5 log10 reduction in bacterial titre when tested with the ex-vivo method . There was no significant difference in reduction of bacterial titres after treatment with antimicrobials between the glass-carrier and the suspension tests . Furthermore, the drying process of bacterial inoculum was less detrimental on skin than on glass surfaces . This study confirmed that the activity of a biocide tested in suspension or on an inanimate surface did not reflect its activity when tested on skin . Further development of the ex-vivo test may be useful, especially for testing the antimicrobial activity of formulations with antiseptic properties. Arch Pharm Res, 1998 Dec, 21(6), 671 - 6 Ofloxacin resistance mechanism in PA150 and PA300-clinical isolates of Pseudomonas aeruginosa in Korea; Lee S et al.; Five hundred and seventy clinical strains of Pseudomonas aeruginosa were isolated from August 1993 to August 1994 in Korea and screened for their resistance to ciprofloxacin, norfloxacin, and ofloxacin . Among these, two P . aeruginosa strains (PA150 and PA300) were selected based on their strong resistance (MICs > 50 micrograms/ml) to all three quinolones . The susceptible strain as well as two resistant strains had proton gradient-dependent efflux system . Efflux system in PA300 showed different specificities to ofloxacin and ciprofloxacin while PA 150 had less permeability for ofloxacin . Ofloxacin had a less inhibitory action on DNA synthesis in permeabilized cells of PA150 and PA300 than 1771M . When quinolone resistance determining region (QRDR) in gyrA was sequenced, PA300 had one missense mutation, Asn 116Tyr, which was newly reported in this work . The results showed that PA150 became ofloxacin resistant by reduced ofloxacin accumulation due to the existence of efflux system and low permeability, while resistance of PA300 was due to the efflux system and a mutation in QRDR of gyrA-the target site of quinolone. Am J Epidemiol, 1998 Dec 15, 148(12), 1175 - 83 Neutropenia is a risk factor for gram-negative bacillus bacteremia in human immunodeficiency virus-infected patients: results of a nested case-control study; Mathews WC et al.; A previous cohort study demonstrated a relation between neutropenia and bacteremia due to gram-negative bacilli among patients infected with human immunodeficiency virus (HIV) . To explore further the relation between neutropenia and bacteremia due to Escherichia coli, Klebsiella pneumoniae, or Pseudomonas aeruginosa among HIV-infected patients, controlling for confounding factors, the authors conducted a nested case-control study with matching and risk-set sampling of controls . The cohort included 1,645 HIV-infected patients followed at the University of California, San Diego, Medical Center in San Diego, California, between 1991 and 1995 . Absolute neutrophil count (ANC) was summarized as mean ANC during the 7-day interval preceding the index date of bacteremia . Covariates were ascertained by medical record review . The matching ratio was 6:1 (controls:cases) . Odds ratios were estimated using conditional logistic regression . Forty-four incident cases of bacteremia were identified . After adjustment for covariates, the estimated odds ratio for the effect of neutropenia (ANC=500 vs . >500/microl) during the 7 days preceding the index date was 8.1 (95% confidence interval confidence interval 1.5-43.1) . The rate of bacteremia due to E . coli, K . pneumoniae, or P . aeruginosa is increased eightfold if the average current-week ANC is less than or equal to 500/microl compared with more than 500/microl. Emerg Infect Dis, 1998 Oct-Dec, 4(4), 551 - 60 Cell-to-cell signaling and Pseudomonas aeruginosa infections; Van Delden C et al.; Pseudomonas aeruginosa is a bacterium responsible for severe nosocomial infections, life-threatening infections in immunocompromised persons, and chronic infections in cystic fibrosis patients . The bacterium's virulence depends on a large number of cell-associated and extracellular factors . Cell-to-cell signaling systems control the expression and allow a coordinated, cell-density-dependent production of many extracellular virulence factors . We discuss the possible role of cell-to-cell signaling in the pathogenesis of P . aeruginosa infections and present a rationale for targeting cell-to-cell signaling systems in the development of new therapeutic approaches. J Bacteriol, 1999 Jan, 181(1), 141 - 8 The Azotobacter vinelandii response regulator AlgR is essential for cyst formation; Nunez C et al.; Azotobacter vinelandii produces the exopolysaccharide alginate, which is essential for the encystment process . In Pseudomonas aeruginosa, as well as in A . vinelandii, the sigmaE factor encoded by algU is required for transcription of algD, which encodes a key enzyme of the alginate biosynthetic pathway . The P . aeruginosa response regulator AlgR activates transcription of algD . fimS, located upstream algR, is proposed to encode the AlgR cognate sensor kinase . We have cloned and characterized the A . vinelandii algR gene; the deduced amino acid sequence of the protein encoded by this gene shows 79% identity with its P . aeruginosa homolog . Sequence analysis around the algR gene revealed the absence of a fimS homolog . Inactivation of A . vinelandii algR diminished alginate production by 50%, but did not affect algD transcription, and completely impaired the capacity to form mature cysts . Electron microscopy of the cyst structures formed by the algR mutant revealed that the encystment process is blocked at the step of exine formation . The transcriptional regulation of the A . vinelandii algR gene and the role of AlgR in alginate production differ significantly from those of its P . aeruginosa counterparts . These differences could be due to the fact that in A . vinelandii, alginate plays a role in encystment, a function not found in P . aeruginosa. J Bacteriol, 1999 Jan, 181(1), 107 - 16 Identification of an Escherichia coli pepA homolog and its involvement in suppression of the algB phenotype in mucoid Pseudomonas aeruginosa; Woolwine SC et al.; Strains of Pseudomonas aeruginosa isolated from the respiratory tracts of patients with cystic fibrosis often display a mucoid morphology due to high levels of expression of the exopolysaccharide alginate . The response regulator AlgB is required for full transcription of the alginate biosynthetic operon . Repeated attempts to demonstrate a direct interaction between AlgB and the promoter region of algD, the first gene in the alginate operon, have thus far been unsuccessful . The possibility that AlgB exerts its effect on algD indirectly exists . To identify putative genes under the control of AlgB which affect algD transcription, transposon mutagenesis of nonmucoid algB derivatives of the mucoid strain FRD1 was employed . Of approximately 3,000 transposon mutants screened, 6 were found to display phenotypes which were mucoid relative to the phenotype of the parental algB strain . The phenotypes of these mutants ranged from being only slightly mucoid to being indistinguishable from that of the original FRD1 strain . One of the particularly mucoid transposon mutants was chosen for further study . This strain was found to be disrupted in a previously uncharacterized open reading frame with 56% amino acid identity to PepA of Escherichia coli . PepA is classified as a leucine aminopeptidase, and homologs have been detected in a number of bacterial, plant, and animal species . This novel gene has been designated phpA (P . aeruginosa homolog of pepA) . The insertional inactivation of phpA was found to correlate with the mucoid phenotype and an increase in algD transcription in the algB strain . Expression of phpA from an ectopic chromosomal locus compensated for the transposon insertion in the native phpA gene, restoring algD transcription to levels similar to those observed in the parental algB strain . While phpA expression did not appear to be under the control of AlgB at the transcriptional level, this study demonstrates that loss of phpA in an algB genetic background had a positive effect on alginate expression and, more specifically, on transcription of the alginate biosynthetic operon. Infect Immun, 1999 Jan, 67(1), 16 - 21 Glucose stimulates phagocytosis of unopsonized Pseudomonas aeruginosa by cultivated human alveolar macrophages; Wong SY et al.; Glucose has previously been shown to increase the in vitro phagocytosis of unopsonized Pseudomonas aeruginosa by freshly explanted murine peritoneal macrophages (PM) and cultivated alveolar macrophages (AM) . This study examined the effect of glucose on the same phagocytosis process in human AM in order to determine whether this phenomenon is conserved among species . Freshly explanted human AM phagocytosed unopsonized P . aeruginosa at a low level (2 bacteria/macrophage/30 min), whereas mouse AM ingested a negligible number of P . aeruginosa (0.01 bacterium/macrophage/30 min) . Glucose had no effect on this or other phagocytic processes in freshly explanted mouse or human AM . However, following in vitro cultivation for 72 h, human AM phagocytosed three to four times more unopsonized P . aeruginosa than did freshly explanted cells, but only in the presence of glucose . This glucose-inducible phagocytic response had also been observed in cultivated murine AM . Although similar increases were also detected for the phagocytosis of latex particles and complement-coated sheep erythrocytes by cultivated human AM, these processes were not glucose dependent . The lack of response to glucose in freshly explanted mouse AM was attributed to insufficient glucose transport; however, freshly explanted human AM exhibited significant facilitative glucose transport activity that was inhibitable by cytochalasin B and phloretin . Taken together, these results suggest that the process of glucose-inducible phagocytosis of unopsonized P . aeruginosa is conserved among macrophages from different species, including humans, and that AM, but not PM, required cultivation for this glucose effect to occur . Glucose transport by AM appears to be necessary but not sufficient for phagocytosis of unopsonized P . aeruginosa. FEBS Lett, 1998 Nov 27, 440(1-2), 93 - 8 Metal coordination of azurin in the unfolded state; Romero C et al.; 1H NMR data applied to the paramagnetic cobalt(II) derivative of azurin from Pseudomonas aeruginosa have made it possible to show that the metal ion is bound to the protein in the unfolded state . The relaxation data as well as the low magnetic anisotropy of the metal ion indicate that the cobalt ion is tetrahedral in the unfolded form . The cobalt ligands have been identified as the residues Gly45, His46, Cys112 and His117 . Met121 is not coordinated in the unfolded state . In this state, the metal ion is not constrained to adopt a bipyramidal geometry, as imposed by the protein when it is folded . This is clear confirmation of the rack-induced bonding mechanism previously proposed for the metal ion in azurin. Gene, 1998 Nov 26, 223(1-2), 247 - 55 Recognition of binding sites I and II by the TrpI activator protein of pseudomonas aeruginosa: efficient binding to both sites requires InGP even when site II is replaced by site I; Olekhnovich I et al.; TrpI protein, the activator of transcription of the trpBA operon of three species of fluorescent Pseudomonads, bends the DNA when it forms either of two well-characterized complexes with the trpBA regulatory region . In complex 1, TrpI is bound only to its strong binding site (site I), whereas in complex 2, which is required for activation of the trpBA promoter, TrpI is bound both to site I and to the weaker site II . Indoleglycerol phosphate (InGP) strongly stimulates formation of complex 2 and is required for activation . The present study focuses on the binding of TrpI to DNA containing a duplication of site I and the effect of the duplication on TrpI-induced DNA bending . We find that even on DNA containing a tandem (direct or inverted) duplication of site I, the formation of DNA-TrpI complexes with both sites occupied is strongly stimulated by InGP . Thus, even when TrpI binding to two adjacent sites needs not be cooperative, InGP significantly promotes the formation of complex 2 . Gel binding data indicate that InGP can have several effects: (1) TrpI molecules bound to either of two adjacent strong binding sites appear to interfere with binding to the other site; InGP relieves this apparent interference . (2) InGP increases the intrinsic affinity of TrpI for sites I and II and/or enhances cooperative TrpI binding to adjacent DNA sites . Furthermore, a third molecule of TrpI can form a footprint adjacent to the duplication on DNA containing a direct (but not inverted) repeat of site I, indicating that TrpI bound to site I is oriented asymmetrically in spite of the quasi-symmetry of the binding site . The calculated bending angle for DNA in complex 2 is increased by approximately 20 degrees when site I is substituted in either orientation for site II; thus, on DNA containing a site I duplication, the bending angle of complex 2 is nearly twice that of complex 1. Wien Klin Wochenschr, 1998 Oct 30, 110(20), 715 - 20 {Imipenem resistance in Pseudomonas aeruginosa}; Fille M et al.; In 1997 in western Austria, 9.9% of Pseudomonas aeruginosa strains from patients of general practitioners were resistant to imipenem as well as 18.2% of the isolates from hospitals and 20.2% of the strains at a university teaching hospital . Within the hospital the imipenem resistance varied from 9.9% among out-patients to 28.7% in isolates from intensive care units . In medical/surgical words, up to 15.1% of P . aeruginosa strains were resistant to imipenem . The incidence of imipenem-resistant P . aeruginosa strains correlates to the use of carbapenems . In June 1997, 10 consecutive isolates from 8 patients were obtained and typed using restriction fragment length polymorphism analysis (RFLP) and Pyocin typing . All 10 isolates were resistant to meropenem as well as to imipenem . The finding (by RFLP and Pyocin typing) of individual bacterial types in each isolate strongly contradicts the spread of infection by cross infection . However, all patients were proven to have been treated with imipenem during the 3 months prior to testing . In 1997, 13,880 g of imipenem were used at the university hospital in Innsbruck . The use of carbapenems appears to be the main cause for the increased incidence of imipenem-resistant P . aeruginosa strains. Int J Mol Med, 1998 Jan, 1(1), 249 - 56 Construction and in vitro evaluation of RFT5(scFv)-ETA', a new recombinant single-chain immunotoxin with specific cytotoxicity toward CD25+ Hodgkin-derived cell lines; Barth S et al.; The data of a closed phase I/II trial in patients with resistant Hodgkin's lymphoma indicate promising results using a chemically linked anti-CD25 ricin-A immunotoxin (IT) (RFT5-SMPT-dgA) . This IT is based on the high-affinity moab RFT5 . Since recombinant DNA technology permits the readier production of large amounts of ITs, we constructed a new RFT5-based fusion toxin {RFT5(scFv)-ETA'} . We isolated mRNA from the hybridoma cell line RFT5, synthesized first strand cDNA and performed RT-PCR . Amplified coding regions of the light and heavy chain variable domains were joined together with a synthetic (Gly4-Ser)3 linker . The resulting single chain variable fragment (scFv) was fused to a modified Pseudomonas aeruginosa exotoxin A (ETA') lacking its cell-binding domain I . After IPTG-induced expression in Escherichia coli, the 70 kDa His-tagged fusion protein {RFT5(scFv)-ETA'} was isolated by osmotic shock and sonication under denaturing conditions . The recombinant toxin was purified on a Ni2+-NTA chelating sepharose and eluted with 250 mM imidazole . Pooled protein was renatured, dialyzed and concentrated by precipitation . Binding properties of RFT5(scFv)-ETA' were assessed on the CD25-expressing cell line L540cy by ELISA, immunohistochemistry and FACS analysis . CD25-specific binding was confirmed by immunoprecipitation experiments with recombinant human IL-2 receptor alpha . The in vitro toxicity of the chimeric protein was tested on the Hodgkin-derived cell lines L540cy, L428, L1236, a monocyte cell line U937 and a Burkitt lymphoma cell line BL38 . RFT5(scFv)-ETA' inhibited protein biosynthesis of L540cy and L428 cells by 50% at concentrations (IC50) of 18 and 12 ng/ml, respectively . CD25-specific toxicity was confirmed by competitive toxicity assays . These data confirm for the first time binding specificity and toxicity of a recombinant anti-CD25 immunotoxin, against Hodgkin-derived cell lines; its applicability on Hodgkin's lymphoma needs yet to be evaluated in vivo. J Bacteriol, 1998 Dec, 180(24), 6784 - 8 The fur-regulated gene encoding the alternative sigma factor PvdS is required for iron-dependent expression of the LysR-type regulator ptxR in Pseudomonas aeruginosa; Vasil ML et al.; We previously identified a novel regulator of the exotoxin A gene (toxA) in Pseudomonas aeruginosa, PtxR, that belongs to the LysR family of prokaryotic regulatory proteins . Preliminary data also suggest that PtxR affects the expression of siderophores in P . aeruginosa . Because toxA expression and siderophore production in this organism are coordinately regulated by the ferric uptake regulator (Fur) and the Fur-regulated alternative sigma factor PvdS, regulation of ptxR itself in the context of these regulators was examined . RNase protection analyses of ptxR transcription revealed that there are two independent transcription initiation sites (T1 and T2) . While transcription from the promoter of T1 is constitutive throughout the growth cycle of PAO1, transcription from the second promoter (P2) is negatively affected by iron . Transcription from the P2 promoter is constitutive in a fur mutant under microaerobic conditions but still iron regulated during aerobic growth . High concentrations (>100 nM) of the ferric uptake regulatory protein (Fur) failed to bind to either of the promoter regions of ptxR in either gel mobility shift assays or DNase I footprint experiments . These results indicate that Fur indirectly regulates the iron-dependent expression of ptxR . Iron-regulated transcription of ptxR from the P2 promoter, but not constitutive expression from the P1 promoter, was dependent on the Fur-regulated alternative sigma factor gene pvdS, even under aerobic conditions . Consequently, there are two levels of iron-regulated expression of ptxR . The iron-regulated expression of ptxR under microaerobic conditions from the P2 promoter of ptxR is mediated indirectly by Fur through the iron-regulated expression of pvdS . In contrast, pvdS-mediated iron regulation of ptxR under aerobic conditions is Fur independent. J Bacteriol, 1998 Dec, 180(24), 6764 - 8 A novel serine/threonine protein kinase homologue of Pseudomonas aeruginosa is specifically inducible within the host infection site and is required for full virulence in neutropenic mice; Wang J et al.; A genetic locus of Pseudomonas aeruginosa was identified that is highly and specifically inducible during infection of neutropenic mice . This locus, ppkA, encodes a protein that is highly homologous to eukaryote-type serine/threonine protein kinases . A ppkA null mutant strain shows reduced virulence in neutropenic mice compared to the wild type . Overexpression of the PpkA protein greatly inhibited the growth of Escherichia coli or P . aeruginosa . However, a single amino acid change at the catalytic site of the kinase domain eliminated the toxic effect of PpkA on bacterial cells, suggesting that the kinase domain of PpkA is functional within bacterial cells. J Bacteriol, 1998 Dec, 180(24), 6753 - 6 Insertion mutagenesis of the ferric pyoverdine receptor FpvA of Pseudomonas aeruginosa: identification of permissive sites and a region important for ligand binding; Kilburn L et al.; Insertion of an 18-amino-acid-encoding sequence within the fpvA gene identified permissive sites at residues Y350, A402, R451, R521, and R558, consistent with these residues occurring in extramembranous loop regions of the protein . Insertions at R451, R521, and R558 did not adversely affect receptor function, although insertions at Y350 and A402 compromised ferric pyoverdine binding and uptake . The latter region likely contributes to or interacts with the ligand-binding site. Mol Genet Metab, 1998 Nov, 65(3), 203 - 12 Modification of development by the CFTR gene in utero; Morrow SL et al.; The in utero infection of rats at 16-17 days gestation with a recombinant adenovirus carrying the human cystic fibrosis transmembrane conductance regulator (cftr) gene resulted in altered lung development and morphology . These structural alterations prompted an evaluation of concurrent functional changes in the cftr-treated lung . CFTR protein could be detected in treated lungs for up to 30 days postinfection, although it was not detected in the intestines at this time . Increased levels of secreted glycoconjugates and lipids were found in lungs treated in utero with human cftr and large vacuoles containing glycoconjugates were detected within cells of the intestines . The scope and durability of these changes suggested that in utero cftr treatment influenced the activity of secretory cells in the developing lung . Altered secretory products in the lungs of cystic fibrosis patients are thought to be associated with increased susceptibility to Pseudomonas aeruginosa infection . We challenged 3-month-old rats (treated in utero with the human cftr gene) with a lethal, intratrachial dose of this bacteria . Rats treated with cftr exhibited enhanced resistance to Pseudomonas infection when compared to controls . These animals displayed little or no associated inflammatory response . No evidence of the adenovirus transgene was detectable at the time of P . aeruginosa inoculation, indicating that continuous ectopic expression of hcftr was not required for enhanced protection . These data demonstrate that in utero, cftr expression influenced the development and function of cells involved in the primary host defense against bacterial infection in the lung . FEMS Microbiol Lett, 1998 Dec 1, 169(1), 179 - 83 Construction of improved plasmid vectors for promoter characterization in Pseudomonas aeruginosa and other gram-negative bacteria; Rist M et al.; We report the construction of two broad host range promoter-probe plasmid vectors for rapid analysis of promoters in Gram-negative bacteria . The new vectors, pME4507 and pME4510, carry carbenicillin and gentamycin resistance genes, respectively, and are small sized (4 kb) with a flexible multiple cloning site to facilitate directional cloning of putative promoter elements . The vectors allow rapid plate-based screening for promoter activities, using beta-galactosidase as the reporter enzyme . In the absence of an inserted promoter fragment, they display very low background activity, making them a useful tool for analysis of low expression level promoters. Bone Marrow Transplant, 1998 Nov, 22(10), 1023 - 6 Fournier's gangrene: a clinical presentation of necrotizing fasciitis after bone marrow transplantation; Martinelli G et al.; Three patients with ANLL developed Fournier's gangrene as an early complication after allo-BMT (two cases) and auto-BMT (one case); two patients were in first CR, the third had resistant disease . Patients developed fever, perineal pain, swelling and blistering of the genital area . Pseudomonas aeruginosa was isolated from the lesions and patients received systemic antibiotic therapy, surgical debridement and medication with potassium permanganate solution . Two patients made a complete recovery although one died of sepsis . The third had progressive involvement of the abdominal wall and later died of leukemia . Early diagnosis of this disorder and prompt initiation of appropriate therapy can prevent progression of this acute necrotizing infection. Electrophoresis, 1998 Nov, 19(15), 2665 - 76 Enhancement of sample loadings for the analysis of oligosaccharides isolated from Pseudomonas aeruginosa using transient isotachophoresis and capillary zone electrophoresis - electrospray - mass spectrometry; Auriola S et al.; The analysis of underivatized core oligosaccharides arising from mild acid hydrolysis of lipopolysaccharides from Pseudomonas aeruginosa serotype 05 was achieved using a transient isotachophoretic preconcentration method coupled to capillary zone electrophoresis-electrospray-mass spectrometry (tCITP-CZE-ES-MS) . The combination of a tCITP preconcentration step provided a 10- to 50-fold enhancement of sample loading and a corresponding improvement in sensitivity compared to the conventional zone electrophoresis format . Electrophoretic conditions, enabling the separation of these anionic analytes, were developed to determine possible sites of heterogeneity on either the core or the O-chain structures . The tCITP-CZE-ES-MS technique provided unparalleled resolution of the different core glycoforms and oligosaccharides obtained from the acid cleavage of the native endotoxins whether isolated following conventional gel permeation chromatography or obtained from direct hydrolysis of the bacterial isolates . These investigations also highlighted the highly phosphorylated nature of these complex cell membrane components, where the heptose residues of the core oligosaccharide can bear up to six phosphate groups. Am J Respir Crit Care Med, 1998 Dec, 158(6), 1702 - 8 Protective effect of endotoxin instillation on subsequent bacteria-induced acute lung injury in rats; Jean D et al.; The phagocytic capability afforded by neutrophil influx into the lungs is essential to ward off invading bacteria . The objective of this study was to evaluate the effect of prior neutrophil recruitment induced by alveolar instillation of endotoxin (LPS, 200 micrograms/kg) 16 h before a pulmonary infection caused by instillation of live Pseudomonas aeruginosa ({PYO}: 1.5 x 10(8) colony-forming units {cfu}/kg) in rats . A first series of experiments showed that lipopolysaccharide (LPS) instillation induced recruitment of alveolar neutrophils that were capable, ex vivo, of elastase exocytosis, reactive oxygen species secretion, and PYO killing . In a second set of experiments, LPS followed by PYO was compared with PYO alone (n = 11 surviving rats in each group) . Parameters were studied 24 h after the bacterial challenge . As compared with PYO alone, pretreatment with LPS followed by PYO was associated with decreased mortality (0% versus 54%, p < 0.05), decreased protein leakage into bronchoalveolar lavage (BAL) fluid (1.8 +/- 0.4 versus 13.5 +/- 2.2 mg/ml, p < 0.001), and improved bacterial clearance from BAL (4.0 +/- 1.4 x 10(2) versus 1.2 +/- 0.5 x 10(4) cfu/ml, p < 0.05) and from pulmonary parenchyma (8.5 +/- 6.4 x 10(5) versus 1.9 +/- 0.8 x 10(7) cfu/ml, p < 0.05) . We conclude that prior alveolar endotoxin instillation induces local recruitment of functionally active neutrophils, and that this is associated with resistance to subsequent experimental pneumonia. Microbiology, 1998 Nov, 144 ( Pt 11), 3135 - 48 Dihydroaeruginoic acid synthetase and pyochelin synthetase, products of the pchEF genes, are induced by extracellular pyochelin in Pseudomonas aeruginosa; Reimmann C et al.; The siderophore pyochelin of Pseudomonas aeruginosa is derived from one molecule of salicylate and two molecules of cysteine . Two cotranscribed genes, pchEF, encoding peptide synthetases have been identified and characterized . pchE was required for the conversion of salicylate to dihydroaeruginoate (Dha), the condensation product of salicylate and one cysteine residue and pchF was essential for the synthesis of pyochelin from Dha . The deduced PchE (156 kDa) and PchF (197 kDa) proteins had adenylation, thiolation and condensation/cyclization motifs arranged as modules which are typical of those peptide synthetases forming thiazoline rings . The pchEF genes were coregulated with the pchDCBA operon, which provides enzymes for the synthesis (PchBA) and activation (PchD) of salicylate as well as a putative thioesterase (PchC) . Expression of a translational pchE'-'lacZ fusion was strictly dependent on the PchR regulator and was induced by extracellular pyochelin, the end product of the pathway . Iron replete conditions led to Fur (ferric uptake regulator)-dependent repression of the pchE'-'lacZ fusion . A translational pchD'-'lacZ fusion was also positively regulated by PchR and pyochelin and repressed by Fur and iron . Thus, autoinduction by pyochelin (or ferric pyochelin) and repression by iron ensure a sensitive control of the pyochelin pathway in P . aeruginosa. Microbiology, 1998 Nov, 144 ( Pt 11), 3127 - 34 PhhC is an essential aminotransferase for aromatic amino acid catabolism in Pseudomonas aeruginosa; Gu W et al.; The phhC gene of Pseudomonas aeruginosa encodes a protein which is a member of the Family I aminotransferases . At high expression levels in the heterologous Escherichia coli system, PhhC can compensate for the absence of AspC (which functions in L-aspartate biosynthesis) and TyrB (which functions in aromatic biosynthesis) . In the native organism, PhhC is essential for catabolism of either L-tyrosine or L-phenylalanine, as demonstrated by gene inactivation . This catabolic function of PhhC is consistent with its inclusion as the distal gene in the inducible phenylalanine hydroxylase operon . The presence of PhhC for catabolism of aromatic amino acids is required in spite of an existing multiplicity of other P . aeruginosa aminotransferases having a similar pattern of broad substrate specificity in vitro . This implies a spatial orientation of PhhC that effectively specializes it for aromatic amino acid catabolism. Chem Pharm Bull (Tokyo), 1998 Nov, 46(11), 1710 - 5 Synthesis and antibacterial activity of novel pyridobenzoxazine analogues; Kawakami K et al.; A series of novel LVFX (7) analogues bearing 4,4-dialkyl-3-aminopyrrolidines at the C-10 position of pyridobenzoxazine was synthesized and their antibacterial activities, pharmacokinetics and acute toxicities in animals were evaluated . Non-alkylated pyrrolidine derivative 26a showed greater activity than LVFX (7) against gram-positive and gram-negative bacteria including Pseudomonas aeruginosa, but 26a possessed high acute toxicity in mice and unfavorable pharmacokinetics in rats . When compared with 26a, 4,4-dialkylated derivatives 26c, e.g . showed more potent activity against gram-positive bacteria along with an improvement of pharmacokinetics and reduction of acute toxicity . Increases in lipophilicity by alkylation on the pyrrolidine ring resulted in a good influence on the above profiles. Am J Respir Cell Mol Biol, 1998 Dec, 19(6), 950 - 8 Effect of inhibition of nitric oxide synthase on Pseudomonas aeruginosa infection of respiratory mucosa in vitro; Dowling RB et al.; We studied the effect of the nitric oxide synthase (NOS) inhibitor asymmetric dimethyl arginine (ADMA) and the inactive enantiomer N G-methyl-D-arginine (D-NMMA) on Pseudomonas aeruginosa infection of the respiratory mucosa in nasal turbinate organ cultures . We also investigated the effect of P . aeruginosa culture filtrate on the expression of inducible NOS (iNOS) messenger RNA (mRNA) by an epithelial cell line (A549) . Organ cultures were preincubated with ADMA (0.1 to 4 x 10(-4) M) or D-NMMA (2 x 10(-4) M) for 30 min prior to bacterial infection . Infected organ cultures (8 h) had significantly (P <= 0.05) greater epithelial damage and fewer ciliated and unciliated cells than did control cultures . There was an increased level of nitrite in the medium feeding infected organ cultures as compared with control cultures . ADMA significantly (P <= 0.05) reduced both bacterially induced epithelial damage and loss of ciliated cells in a concentration-dependent manner . D-NMMA did not influence the effect of P . aeruginosa infection of the mucosa . ADMA, but not D-NMMA, significantly (P <= 0.04) reduced total bacterial numbers adherent to the respiratory mucosa . P . aeruginosa culture filtrates (24 h and 36 h) significantly (P = 0.02) increased iNOS with respect to glyceraldehyde-3-phosphate dehydrogenase mRNA expression . These results show that P . aeruginosa stimulates iNOS expression by a cell line and NO production by an organ culture . ADMA reduces mucosal damage and loss of ciliated cells, which suggests that NO may be a mediator of epithelial damage caused by P . aeruginosa. Am J Respir Cell Mol Biol, 1998 Dec, 19(6), 920 - 8 Effects of ion composition and tonicity on human neutrophil antibacterial activity; Verghese MW et al.; Infants with cystic fibrosis (CF) often are infected with Staphylococcus aureus (S . aur.), which is followed by colonization with Pseudomonas aeruginosa (P . aerug.) . In spite of an excessive, neutrophil-dominated inflammatory response in the respiratory tract, patients with CF often succumb to pulmonary infections with P . aerug . Because peripheral blood neutrophils of these patients have normal functions, we examined whether hypothesized alterations of the airway surface liquids (ASL) in these patients significantly impair neutrophil bactericidal activity in the microenvironment of the CF lung . The ionic composition of CF ASL is still not entirely defined and has been speculated to be abnormally high or abnormally low in Na+ and Cl- concentrations; estimates of osmolarities have ranged from 200 (hypo-osmolar) to 285 (iso-osmolar) to > 300 meq/L (hyper-osmolar) . Our data indicate that bacterial killing activity of human peripheral blood neutrophils against P . aerug . or S . aur . is not decreased in buffers in which NaCl was replaced with equimolar concentrations of choline Cl, KCl, or N-methyl-D-glucamine chloride to maintain isotonicity . Amiloride or benzamil, known modulators of Na+ transport in neutrophils, did not interfere with this neutrophil function . Deviations from isotonicity of +/- 50% also failed to diminish bactericidal activity of neutrophils significantly . In contrast, superoxide production and enzyme secretion in response to the chemotactic peptide N-formylmethionylleucylphenylalanine appeared to be sensitive to the ionic milieu of the assay buffers . Our results suggest that the postulated alterations in the ionic composition of ASL in CF lungs are insufficient to explain why neutrophils fail to clear infections with P . aerug . in these patients. Am J Respir Cell Mol Biol, 1998 Dec, 19(6), 853 - 66 The relationship of chronic mucin secretion to airway disease in normal and CFTR-deficient mice; Cressman VL et al.; In the cystic fibrosis (CF) patient, lung function decreases throughout life as a result of continuous cycles of infection, particularly with Pseudomonas aeruginosa and Staphylococcus aureus . The mechanism underlying the pathophysiology of the disease in humans has not been established . However, it has been suggested that abnormal, tenacious mucus, resulting perhaps from improper hydration from loss of Cl- secretion via the cystic fibrosis transmembrane conductance regulator (CFTR) protein, impairs clearance of bacteria from the CF airway and provides an environment favorable to bacterial growth . If this hypothesis is correct, it could explain the absence of respiratory disease in CFTR-deficient mice, since mice have only a single submucosal gland and display few goblet cells in their lower airways, even when exposed to bacteria . To test this hypothesis further, we induced allergic airway disease in CFTR-deficient mice . We found that induction of allergic airway disease in mice, unlike bacterial infection, results in an inflammatory response characterized by goblet cell hyperplasia, increased mucin gene expression, and increased production of mucus . However, we also found that disease progression and resolution is identical in Cftr-/- mice and control animals . Furthermore, we show that the presence of mucus in the Cftr-/- airway does not lead to chronic airway disease, even upon direct inoculation with S . aureus and P . aeruginosa . Therefore, factors in addition to the absence of high levels of mucus secretion protect the mouse from the airway disease seen in human CF patients. Acta Virol, 1998 Jun, 42(3), 175 - 9 Two high-frequency-transduction phage isolates from lysogenic strains of Pseudomonas aeruginosa transducing antibiotic resistance; Blahova J et al.; Two high frequency transduction (HFT) phage isolates, obtained from seriously ill patients, transducing individual determinants of antibiotic resistance with a frequency of 10(-5) (phage isolate AP-103) and 10(-6) (phage isolate AP-343), are described . The frequency of transduction depended on the transduced determinant(s) of resistance used for the detection of transductants and on the individual recipient antibiotic-susceptible strain of Pseudomonas aeruginosa (PAO and/or ML series) . A multiple-antibiotic resistance was transduced by the phage isolate AP-343 to all tested recipient strains . The appearance of such phages in clinical conditions with an unusually high frequency of transduction might contribute to the dissemination of antibiotic resistance genes among nosocomial strains of P . aeruginosa . The existence of HFT phages might reflect an increased efficiency of transduction of antibiotic resistance among P . aeruginosa strains, and thus an increased risk of spread of antibiotic resistance even to recently introduced anti-pseudomonadal antibiotics among pseudomonads with unfavourable and unwanted epidemiological consequences in hospital conditions. J Infect Dis, 1999 Jan, 179(1), 151 - 62 Priming of blood neutrophils in children with cystic fibrosis: correlation between functional and phenotypic expression of opsonin receptors before and after platelet-activating factor priming; Witko-Sarsat V et al.; Blood phagocyte opsonin receptor CR1 (CD35) and CR3 (CD11b) functions were examined in cystic fibrosis (CF) patients with endobronchial Staphylococcus aureus or Pseudomonas aeruginosa chronic infection, CF patients without infection, heterozygous, non-CF patients with chronic pulmonary infection, and healthy controls . Circulating and platelet-activating factor (PAF)-primed phagocyte luminol luminescence responses to complement-opsonized zymosan were increased in both groups of infected CF and non-CF children relative to uninfected CF children and healthy control children and adults . The ratio between circulating and PAF-primed phagocyte responses was significantly elevated in all children with CF, and in these, the ratio could serve as an indicator of response to antibiotic treatment . The ratios of circulating and PAF-primed phenotypic expression for CR1, CR3, and FcgammaRIII (CD16), but not FcgammaRII (CD32), correlated with the functional ratios . Phagocyte opsonin receptor response capacity might be used for evaluation of inflammation and infection in CF patients. J Appl Physiol, 1998 Dec, 85(6), 2298 - 304 Mechanism of pyocyanin- and 1-hydroxyphenazine-induced lung neutrophilia in sheep airways; Lauredo IT et al.; Pyocyanin (Pyo) and 1-hydroxyphenazine (1-HP) are extracellular products of Pseudomonas aeruginosa . To test whether these products were capable of producing an inflammatory response in the airways, combinations of Pyo and 1-HP at concentrations of 10(-4) and 10(-5) M were instilled into sheep airways, and indexes of inflammation were assessed by bronchoalveolar lavage (BAL) 24 h later . Challenge with the phenazines caused a significant dose-dependent increase in the number of cells and neutrophils recovered by BAL . Control challenges produced no such changes . The lung neutrophilia was accompanied by an increased concentration of albumin in BAL . The increases in BAL neutrophils and albumin could be blocked by treating the sheep with the 5-lipoxygenase inhibitor zileuton . Neither 1-HP nor Pyo was chemotactic to neutrophils when tested in vitro, but when alveolar macrophages (AM) were cultured in vitro in the presence of both Pyo and 1-HP (1 microM), the supernatants caused neutrophil chemotaxis . Analysis of AM culture supernatants incubated with the combination of pigments showed significant increases in leukotriene B4 and interleukin-8, and blocking these mediators separately or together reduced AM supernatant-induced neutrophil chemotaxis . We conclude that local instillation of Pyo and 1-HP can initiate an inflammatory response in the airways of sheep in vivo . This effect can be explained, in part, by the release of chemotactic factors produced by AM. Antimicrob Agents Chemother, 1998 Dec, 42(12), 3269 - 75 Evaluation of antimicrobial and lipopolysaccharide-neutralizing effects of a synthetic CAP18 fragment against Pseudomonas aeruginosa in a mouse model; Sawa T et al.; CAP18 (cationic antimicrobial protein; 18 kDa) is a neutrophil-derived protein that can bind to and inhibit various activities of lipopolysaccharide (LPS) . The 37 C-terminal amino acids of CAP18 make up the LPS-binding domain . A truncated 32-amino-acid C-terminal fragment of CAP18 had potent activity against Pseudomonas aeruginosa in vitro . We studied the antimicrobial and LPS-neutralizing effects of this synthetic truncated CAP18 peptide (CAP18106-137) on lung injury in mice infected with cytotoxic P . aeruginosa . To determine its maximal effect, the CAP18106-137 peptide was mixed with bacteria just prior to tracheal instillation, and lung injury was evaluated by determining the amount of leakage of an alveolar protein tracer (125I-albumin) into the circulation and by the quantification of lung edema . The lung injury caused by the instillation of 5 x 10(5) CFU of P . aeruginosa was significantly reduced by the concomitant instillation of CAP18106-137 . However, the administration of CAP18106-137 alone, without bacteria, induced lung edema, suggesting that it has some toxicity . Also, the peptide did not significantly reduce the number of bacteria that had been simultaneously instilled, nor did it significantly improve the survival of the infected mice . The addition of CAP18106-137 to aztreonam along with the bacteria did decrease the level of antibiotic-induced release of inflammatory mediators including tumor necrosis factor alpha, interleukin-6, and nitric oxide and also improved the survival of the mice . Therefore, more investigations are needed to confirm the toxicities and the therapeutic benefits of CAP18106-137 as an adjunctive therapy to antibiotics in the treatment of infections caused by gram-negative bacteria. Antimicrob Agents Chemother, 1998 Dec, 42(12), 3173 - 8 Expression of the Pseudomonas aeruginosa gentamicin resistance gene aacC3 in Escherichia coli; van Boxtel RA et al.; The Pseudomonas aeruginosa aacC3 gene was expressed in Escherichia coli after cloning of the single gene behind the strong tac promoter . In the original Pseudomonas strain, aacC3 is preceded by cysC; together they form a single transcription unit . The ribosome-binding site and start codon of aacC3 are involved in a putative intercistronic hairpin, the stability of which interfered with the aminoglycoside resistance level . In Northern blots, full-length transcripts comprising both cysC and aacC3 could not be detected either in the original Pseudomonas strain or in E . coli harboring a plasmid with the cloned operon . In contrast, cysC transcripts were abundant . Cloning of the operon between the tac promoter and a transcription termination signal resulted in higher mRNA levels and phenotypic expression in E . coli . The absence of a transcription termination signal in the wild-type cysC-aacC3 sequence is associated with transcripts of heterogeneous size that were undetected in Northern blots . Our results shed more light on the expression of this gentamicin resistance determinant, although the discrepancies between its expression in E . coli and Pseudomonas are not fully solved. Antimicrob Agents Chemother, 1998 Dec, 42(12), 3117 - 22 OXA-16, a further extended-spectrum variant of OXA-10 beta-lactamase, from two Pseudomonas aeruginosa isolates; Danel F et al.; Two extended-spectrum mutants of the class D beta-lactamase OXA-10 (PSE-2) from Pseudomonas aeruginosa isolates obtained in Ankara, Turkey, were described previously and were designated OXA-11 and -14 . P . aeruginosa 906 and 961, isolated at the same hospital, were highly resistant to ceftazidime (MIC >/= 128 microgram/ml) and produced a beta-lactamase with a pI of 6.2 . The MICs of ceftriaxone, cefoperazone, cefsulodin, and cefepime were 4- to 16-fold above the typical values for P . aeruginosa, whereas the MICs of penicillins and cefotaxime were raised only marginally . Ceftazidime MICs were not significantly reduced by clavulanate or tazobactam at 4 microgram/ml . Ceftazidime resistance did not transfer conjugatively but was mobilized to P . aeruginosa PU21 by plasmid pUZ8 . Both isolates gave similar DNA restriction patterns, suggesting that they were replicates; moreover, they yielded identically sized BamHI fragments that hybridized with a blaOXA-10 probe . DNA sequencing revealed that both isolates had the same new beta-lactamase, designated OXA-16, which differed from OXA-10 in having threonine instead of alanine at position 124 and aspartate instead of glycine at position 157 . The latter change is also present in OXA-11 and -14 and seems critical to ceftazidime resistance . Kinetic parameters showed that OXA-16 enzyme was very active against penicillins, cephaloridine, cefotaxime, and ceftriaxone, but hydrolysis of ceftazidime was not detected despite the ability of the enzyme to confer resistance. Antimicrob Agents Chemother, 1998 Dec, 42(12), 3113 - 6 Novel OXA-10-derived extended-spectrum beta-lactamases selected in vivo or in vitro; Mugnier P et al.; A clinical isolate of Pseudomonas aeruginosa, PAe191, was found to be highly resistant to all anti-Pseudomonas beta-lactam antibiotics (except imipenem) and resistant also to aminoglycosides . It produced a beta-lactamase (with an apparent pI of 7.6) which was not inhibited by clavulanic acid . Cloning and characterization of the beta-lactamase gene showed that it coded for a novel extended-spectrum OXA-10 variant, called OXA-19, which differed from OXA-10 by nine amino acids and from OXA-13 by two, i.e., Asn in position 73 (Asn73) instead of Ser and Asp157 instead of Gly . Asparagine in position 157 is implicated in resistance to ceftazidime, while the amino acid in position 73, in this variant, seems to condition the level of resistance to penicillins . The oxa19 gene was found to be inserted, in a typical integron structure, immediately downstream from an aac(6')-Ib gene coding for an aminoglycoside acetyltransferase variant, which was called AAC(6')-Ib9. Alcohol Clin Exp Res, 1998 Nov, 22(8), 1640 - 5 Effect of alcohol on bacterial translocation in rats; Mason CM et al.; Bacterial translocation has been proposed to be important in the pathophysiology of sepsis, as well as to be a consequence of sepsis . To study the effect of alcohol on bacterial translocation from the gut, normal Sprague-Dawley rats were administered alcohol by gavage by two regimens: Acute (3.7 g/kg, one dose) or Subacute (1 of 2 doses, 2.4 or 3.7 g/kg/day once daily for 14 days) . Mesenteric lymph node cultures were performed, and portal venous blood was assayed for endotoxin . Ileal and cecal permeability studies were performed in the Acute and Subacute groups using fluorescein isothiocyanate-labeled dextrans of either 4,000 or 70,000 kDa size . As an index of the effect of systemic endotoxin, tissues from mesenteric lymph nodes, liver, and intestinal Peyer's patches were assayed for the presence of mRNA for tumor necrosis factor . Additionally, because extrapulmonary sepsis has been shown to suppress pulmonary antibacterial defenses, animals in the Subacute group were challenged by aerosol inoculation with Pseudomonas aeruginosa to determine bacterial clearance and alveolar cellular responses . The results show that neither of the alcohol regimens resulted in bacterial growth from mesenteric lymph nodes or portal blood . Animals in the Subacute group had more endotoxin present in portal blood than did the Control group (92.9 pg/ml vs . 40.2 pg/ml; p < 0.02) . None of the animals had demonstrable mRNA for tumor necrosis factor in any of the tissues assayed . There were no demonstrable increases in ileal or cecal permeability for either the small or large molecular weight dextran in either alcohol group . Furthermore, there was no delay in the clearance of P . aeruginosa from the lung in the Subacute group, but these animals recruited fewer neutrophils into the airspaces in response to this challenge than did the Control animals . Thus, alcohol intoxication does not result in bacterial translocation from the gut in this model . Despite higher levels of portal venous endotoxin in the animals in the Subacute alcohol group, no adverse systemic consequences of this phenomenon could be demonstrated. J Pept Res, 1998 Oct, 52(4), 289 - 99 The use of synthetic peptides in the design of a consensus sequence vaccine for Pseudomonas aeruginosa; Cachia PJ et al.; Pseudomonas aeruginosa employs pili to mediate adherence to epithelial cell surfaces . Research has shown that the C-terminal region of the pilin monomer contains the epithelial cell binding domain, which is semiconserved in seven different strains of this bacterium . Antibodies to this region of the pilin molecule are also able to block and prevent the infection process . As there is a degree of sequence and structural homology in the C-terminal region and all strains examined have been shown to bind to the same cell surface receptor, we reasoned that it should be possible to produce a synthetic peptide consensus sequence which would provide cross-reactive antiserum from a single peptide immunogen inhibiting the adherence of the known strains of P . aeruginosa . In this article we examine the cross-reactivity of five rabbit polyclonal antisera . One has been raised against the cell-surface receptor binding domain of native PAK strain pilin (residues 128-144) while the others have been raised to analogues of this region . Analysis of the cross-reactivity of these antisera, using competitive ELISA assay, has shown that it is possible to manipulate the amino acid sequence of a peptide immunogen to generate antiserum, which exhibits enhanced cross-reactivity to various strains of P . aeruginosa . Furthermore, when this peptide is conjugated to tetanus toxoid and used to vaccinate mice it provided cross-reactive protection against heterologous challenge with PAO strain bacteria . The results of these experiments are analyzed, and the applicability of our hypothesis and the implications of this approach to the design of a strain-independent consensus vaccine for immunization against Pseudomonas aeruginosa are discussed. Int J Antimicrob Agents, 1998 Aug, 10(3), 223 - 8 Susceptibility of adherent organisms from Pseudomonas aeruginosa and Staphylococcus aureus strains isolated from burn wounds to antimicrobial agents; Trafny EA; To assess the bactericidal effects of ciprofloxacin, netilymicin, and polymyxin B on adherent Pseudomonas aeruginosa organisms and also the bactericidal effects of ciprofloxacin, vancomycin and teicoplanin on adherent Staphylococcus aureus cells, a simple end-point microplate assay, based on the method described by Miyake et al . was used in the present study . As results of the assay, the minimal inhibitory concentration (MICADH) values are taken, which express the susceptibility of the bacterial cells spontaneously released from the surface of adherent microcolonies to antimicrobial agents . Also, a minimal bactericidal concentration (MBCADH) value was read, which is defined as the lowest antibiotic concentration required to kill the sessile bacterial cells . For twenty P . aeruginosa strains and nineteen S . aureus strains isolated from burn wounds, an enhanced resistance against bactericidal action of the applied antibiotics was observed when bacterial cells were attached to polystyrene surface . The MICADH values were comparable with the conventional MIC values only for ciprofloxacin and netilmicin for P . aeruginosa strains . The MBCADH values exceeded many times the conventional MBC values for the majority of strains . The validity of the assay was estimated in the experiment designed to determine the concentration of ciprofloxacin that should be released topically from the collagen dressing to prevent the biomaterial from microbial colonization and allow the decontamination of the wound. Int J Antimicrob Agents, 1998 Aug, 10(3), 207 - 14 Management of fluoroquinolone resistance in Pseudomonas aeruginosa--outcome of monitored use in a referral hospital; Peterson LR et al.; We evaluated a strategy designed to improve useful activity of ciprofloxacin against Pseudomonas aeruginosa . Following changes in antimicrobial agent use made by the institutional Pharmacy and Therapeutic Drug Committee, monthly drug usage and microbial susceptibility records from June 1992 through October 1995 were reviewed . From July 1992 through October 1992 (Period 1), ciprofloxacin and ofloxacin usage represented 95 and 5% of total quinolone doses; from December 1992 to March 1993 (Period 2), ciprofloxacin represented 19%; from July 1993 to October 1993 (Period 3), ciprofloxacin usage represented 85%; from July 1994 to October 1994 (Period 4), ciprofloxacin represented 95%; and from July 1995 to October 1995 (Period 5), ciprofloxacin and ofloxacin respectively represented 98 and 2% of total quinolone doses . Comparison of the anti-pseudomonal activity of the two fluoroquinolones to ofloxacin use, ciprofloxacin use and total quinolone use during the entire observational period showed the highest (negative) correlation with ofloxacin use versus ofloxacin activity (y = -15.04x + 1367.99, r2 = 0.06, w = 0.126) . Increased use of quinolones plus a change to primarily ofloxacin usage appeared to adversely affect the activity of both ofloxacin and ciprofloxacin against P . aeruginosa. Infect Control Hosp Epidemiol, 1998 Nov, 19(11), 853 - 5 Colonization with Pseudomonas aeruginosa in patients developing ventilator-associated pneumonia; Bergmans DC et al.; To determine routes of colonization and genotypic variation of Pseudomonas aeruginosa leading to ventilator-associated pneumonia, colonization of the rectum, stomach, oropharynx, and trachea was studied chronologically in 10 patients . Ninety-one isolates of P aeruginosa were genotyped; seven different genotypes were identified . Patients developing ventilator-associated pneumonia caused by P aeruginosa were colonized at multiple body sites and may be colonized with multiple genotypes . The upper respiratory tract is the predominant initial site of colonization with P aeruginosa. Zhonghua Yi Xue Za Zhi (Taipei), 1998 Oct, 61(10), 589 - 95 Prevalence of nosocomial respiratory tract infections in the surgical intensive care units of a medical center; Wang FD et al.; BACKGROUND: The intensive care unit (ICU) is one of the most common locations in the hospital for the development of nosocomial respiratory tract infections (RTIs) . The purpose of this study was to better understand and compare the rate of nosocomial RTIs, their distribution and the frequency of infective organisms in three different adult surgical ICUs at a medical center in Taiwan . METHODS: This retrospective study covered the period from 1991 to 1996, and targeted patients who had acquired nosocomial RTIs more than 72 hours after admission to the surgical, cardiovascular surgical, or neurosurgical ICU . Infection control nurses made routine weekly rounds to the units and conducted comprehensive patient-based surveys . Data collected were subjected to descriptive and deductive statistical analyses . RESULTS: A total of 277 episodes of nosocomial RTIs were encountered (3.8 episodes per 1,000 patient-days of hospitalization) . Neurosurgical ICU patients had the highest infection rate (4.8%) (p < 0.01) . The median length of hospital stay of patients acquiring nosocomial RTIs was 61 days; the infection occurred 19 days (median) after admission to the ICU and the length of postinfection stay was 38 days (median) . Pneumonia accounted for 91.3% of these infections . The overall mortality rate was 42.6%, with the surgical ICU having the highest mortality rate (61.5%), which was significantly higher compared with the other two ICUs studied . The majority of patients with nosocomial RTIs had medical catheters or devices in place . A total of 297 isolates were cultured from 277 infected patients . Pseudomonas aeruginosa was the predominant organism isolated (29.6%), followed by Staphylococcus aureus (26.6%), of which 88.2% were methicillin-resistant S aureus (MRSA) . CONCLUSIONS: The rate of nosocomial RTIs was significantly higher in neurosurgical ICU patients; the majority of infected patients had medical catheters or devices in place . S aureus played an increasingly important role as a nosocomial pathogen; hence, control of MRSA should be the focus of infection control policies. J Appl Microbiol, 1998 Nov, 85(5), 799 - 806 Resistance of Pseudomonas aeruginosa PAO1 phage F116 to sodium hypochlorite; Maillard JY et al.; The development of viral resistance to sodium hypochlorite was investigated using the Pseudomonas aeruginosa bacteriophage F116 as a model system . This phage was chosen because of its structural characteristics and former investigations conducted in this laboratory . F116 was shown to be sensitive to a sodium hypochlorite concentration of 0.0075 gl-1 (available chlorine) which produced a 5 log10 reduction in titre in a suspension test . Survival bacteriophages challenged with this sodium hypochlorite concentration were isolated, propagated and challenged again with the same and higher concentrations of the biocide . It was observed that progeny virions were becoming increasingly resistant to sodium hypochlorite challenges up to a concentration of 0.0175 gl-1 of available chlorine . It was also noticed that 1-2 log10 of F116 virions from resistant phage lysates remained sensitive to the biocide . An electron microscopical investigation of F116 resistant lysates showed that the phage resistance to sodium hypochlorite was not caused by F116 particles aggregation . Furthermore, no morphological difference between the sensitive and resistant F116 particles to sodium hypochlorite was identified. J Biol Chem, 1998 Dec 4, 273(49), 32369 - 72 Hydrocarbon rulers in UDP-N-acetylglucosamine acyltransferases; Wyckoff TJ et al.; UDP-GlcNAc acyltransferase (LpxA), the first enzyme of lipid A biosynthesis, catalyzes the transfer of an acyl chain activated on acyl carrier protein (ACP) to UDP-GlcNAc . LpxAs are very selective for the lengths of their acyl donor substrates . Escherichia coli LpxA prefers R-3-hydroxymyristoyl-ACP to R-3-hydroxydecanoyl-ACP by a factor of approximately 1000, whereas Pseudomonas aeruginosa LpxA prefers the opposite . E . coli G173M LpxA and the reciprocal P . aeruginosa M169G LpxA show reversed substrate selectivity in vitro and in vivo, demonstrating the existence of precise hydrocarbon rulers in LpxAs. Infect Immun, 1998 Dec, 66(12), 5777 - 84 Pseudomonas pyocyanin increases interleukin-8 expression by human airway epithelial cells; Denning GM et al.; Pseudomonas aeruginosa, an opportunistic human pathogen, causes acute pneumonia in patients with hospital-acquired infections and is commonly associated with chronic lung disease in individuals with cystic fibrosis (CF) . Evidence suggests that the pathophysiological effects of P . aeruginosa are mediated in part by virulence factors secreted by the bacterium . Among these factors is pyocyanin, a redox active compound that increases intracellular oxidant stress . We find that pyocyanin increases release of interleukin-8 (IL-8) by both normal and CF airway epithelial cell lines and by primary airway epithelial cells . Moreover, pyocyanin synergizes with the inflammatory cytokines tumor necrosis factor alpha and IL-1alpha . RNase protection assays indicate that increased IL-8 release is accompanied by increased levels of IL-8 mRNA . The antioxidant n-acetyl cysteine, general inhibitors of protein tyrosine kinases, and specific inhibitors of mitogen-activated protein kinases diminish pyocyanin-dependent increases in IL-8 release . Conversely, inhibitors of protein kinases C (PKC) and PKA have no effect . In contrast to its effects on IL-8 expression, pyocyanin inhibits cytokine-dependent expression of the monocyte/macrophage/T-cell chemokine RANTES . Increased release of IL-8, a potent neutrophil chemoattractant, in response to pyocyanin could contribute to the marked infiltration of neutrophils and subsequent neutrophil-mediated tissue damage that are observed in Pseudomonas-associated lung disease. Eur J Biochem, 1998 Oct 15, 257(2), 409 - 19 Quinoprotein ethanol dehydrogenase of Pseudomonas aeruginosa is a homodimer--sequence of the gene and deduced structural properties of the enzyme; Diehl A et al.; The gene coding for the periplasmic quinoprotein ethanol dehydrogenase of Pseudomonas aeruginosa ATCC 17933 was cloned and sequenced . The deduced amino acid sequence contained a signal peptide of 34 residues and the major protein of 589 amino acids showed high similarities to pyrroloquinoline-quinone-dependent periplasmic and membrane-bound dehydrogenases acting on alcohols, glucose and quinate or shikimate . It was demonstrated by alignment with the amino acid sequence of the large subunit of the quinoprotein methanol dehydrogenase from Methylobacterium extorquens, whose X-ray structure is known, that the amino acid residues involved in the binding of pyrroloquinoline quinone and Ca2+ at the active site are conserved in the quinoprotein ethanol dehydrogenase of P . aeruginosa . Also, the glycine/tryptophan docking motifs involved in stabilizing the superbarrel structure of the quinoprotein methanol dehydrogenase of M . extorquens were conserved . The known sequences of pyrroloquinoline-quinone-dependent dehydrogenases were used to derive new, more specific sequence motifs for detecting members of this family of enzymes . Despite the sequence similarity between the large a subunit of quinoprotein methanol dehydrogenase from M . extorquens and the quinoprotein ethanol dehydrogenase from P . aeruginosa, the two enzyme systems were quite different . In the presence of the prosthetic group, pyrroloquinoline quinone expression of the Pseudomonas gene encoding the 60-kDa subunit of quinoprotein ethanol dehydrogenase in Escherichia coli resulted in formation of active enzyme . The formation of active quinoprotein methanol dehydrogenase, however, is known to require, in addition to the large alpha subunit, the expression of a small beta subunit, and helper proteins {Lidstrom, M . E . (1995) Genetics of bacterial quinoproteins, Methods Enzymol . 258, 217-227}. Epidemiol Infect, 1998 Oct, 121(2), 349 - 56 Removal of airborne bacteria by filtration using a composite microporous membrane made of a pyridinium-type polymer showing strong affinity with microbial cells; Kawabata N et al.; A composite microporous membrane made of poly(N-benzyl-4-vinylpyridinium chloride) that showed strong affinity with bacterial cells was prepared as a filter material for removing airborne bacteria . Thickness, pore diameter and porosity of the membrane were 0.72 mm, 14.5 microm and 63%, respectively . Electron micrographic analysis revealed that the membrane consisted of a very large number of connected beads of 1.4 microm in diameter made of the pyridinium-type polymer . Filtration using the membrane was performed easily at low flow rates with insignificant pressure drop across the membrane . Filtration at 63.7 cm/sec gave 99.98% and 99.996% removal (3.7 and 4.4 log10-unit reduction in concentration) of Escherichia coli and Pseudomonas aeruginosa, respectively . Staphylococcus aureus was not detected in filtrates . Since pores of the membrane were much larger than these bacteria, the efficient removal was best explained in terms of the affinity of the polymer with bacterial cells. Zh Mikrobiol Epidemiol Immunobiol, 1998 Sep-Oct, (5), 43 - 7 {Epidemiologic features of hospital pyo-septic infection in a cardiosurgical hospital}; Shkarin VV et al.; Th