J Pediatr Surg, 2001 Jun, 36(6), 948 - 50 Surgical implications of pseudomonas aeruginosa necrotizing fasciitis in a child with acute lymphoblastic leukemia; Jaing TH et al.; Necrotizing fasciitis caused by Pseudomonas aeruginosa is extremely rare . Only 4 cases were reported in the literature . The authors report the occurrence of P aeruginosa necrotizing fasciitis starting out as a vulval abscess in a girl before induction chemotherapy for acute lymphoblastic leukemia . To our knowledge, this is the second case described in association with leukemia . In this case, the outcome was favorable because of early surgical intervention, confirming the diagnosis . J Pediatr Surg 36:948-950 . Invest Ophthalmol Vis Sci, 2001 Jun, 42(7), 1561 - 7 Plasminogen Activators and Inhibitors in the Corneas of Mice Infected with Pseudomonas aeruginosa; Berk RS et al.; PURPOSE: To characterize the presence of plasminogen activators and their inhibitors in the corneas during the inflammatory response in naive and immunized mice intracorneally infected with Pseudomonas aeruginosa . METHODS: RT-PCR was used to detect gene expression for plasminogen activators and their inhibitors in naive and immunized mice . Immunoblot analysis, zymography, and ELISA were used to demonstrate the syntheses of these proteins . RESULTS: Naive mice intracorneally infected with P . aeruginosa showed a temporally enhanced expression of tissue-type plasminogen activator (tPA), urokinase-type plasminogen activator (uPA), its receptor (uPAR), and plasminogen activator inhibitors 1 and 2 (PAI-1 and PAI-2), over a several-day holding period . Immunized mice demonstrated a lower and shorter expression of these factors over the same period . Expression of these factors at the mRNA and protein levels may have been due to enzymes and inhibitors present in inflammatory cells and in resident corneal cells . CONCLUSIONS: These results show a correlation between the overexpression of the PA system in infected naive mice as part of the inflammatory response, with eventual ocular destruction . Immunized mice exhibit a more balanced and shorter expression of the PA system, which may contribute to the restoration of corneal clarity. J Ethnopharmacol, 2001 Jun, 76(1), 39 - 44 Screening of antibiotic resistant inhibitors from local plant materials against two different strains of Pseudomonas aeruginosa; Aburjai T et al.; The methanolic extracts of 19 Jordanian plants were combined with seven different antibiotics and applied to check the inhibitory effects of the combination on the resistance of Pseudomonas aeruginosa . A resistant strain of Ps . aeruginosa, which was isolated from a patient and a standard strain of the same microorganism were used in this study . Our results showed that there are significant variations in the effects of some combinations used on the resistant and the standard strains probably due to structural changes . Almost all the plant materials used in combination with penicillin G and erythromycin allowed full growth of the standard strain, while the combination with some plant materials like Gundelia tournefortii L . and Lepidium sativum L . inhibited the growth of the resistant strain . Chloramphenicol, gentamicin and cephalosporin can be given advantageously with almost all the plant materials used with few exceptions on the resistant strain . Nalidixic acid activity was improved significantly when combined with all plant materials and tested on standard strain . On the other hand, its activity on the resistant strain was slightly improved using the same combinations. J Clin Microbiol, 2001 Jun, 39(6), 2072 - 8 Nosocomial outbreak due to a multiresistant strain of Pseudomonas aeruginosa P12: efficacy of cefepime-amikacin therapy and analysis of beta-lactam resistance; Dubois V et al.; Over a 3-year period, 67 patients of the Hospital of Pau (Pau, France), including 64 patients hospitalized in the adult intensive care unit (ICU), were colonized and/or infected by strains of Pseudomonas aeruginosa P12, resistant to all potentially active antibiotics except colistin . Most patients were mechanically ventilated and presented respiratory tract infections . Since cefepime and amikacin were the least inactive antibiotics by MIC determination, all ICU patients were treated with this combination, and most of them benefited . Cefepime-amikacin was found highly synergistic in vitro . Ribotyping and arbitrary primer-PCR analysis confirmed the presence of a single clonal isolate . Isoelectrofocusing revealed that the epidemic strain produced large amounts of the chromosomal cephalosporinase and an additional enzyme with a pI of 5.7, corresponding to PSE-1, as demonstrated by PCR and sequencing . Outer membrane protein profiles on sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed the absence of a ca . 46-kDa protein, likely to be OprD, and increased production of two ca . 49- and 50-kDa proteins, consistent with the outer membrane components of the efflux systems, MexAB-OprM and MexEF-OprN . Thus, we report here a nosocomial outbreak due to multiresistant P . aeruginosa P12 exhibiting at least four mechanisms of beta-lactam resistance, i.e., production of the penicillinase PSE-1, overproduction of the chromosomal cephalosporinase, loss of OprD, and overexpression of efflux systems, associated with a better activity of cefepime than ceftazidime. Crit Care Med, 2001 Mar, 29(3), 548 - 56 The importance of bacterial sepsis in intensive care unit patients with acquired immunodeficiency syndrome: implications for future care in the age of increasing antiretroviral resistance; Rosenberg AL et al.; OBJECTIVE: To describe the clinical characteristics and outcomes of patients with acquired immunodeficiency syndrome (AIDS) admitted to the intensive care unit (ICU) . DESIGN: An observational cohort study with retrospective chart review . SETTING: ICU of an urban university medical center . PATIENTS: Consecutive ICU admissions of patients with AIDS at an urban university medical center between December 1993 and June 1996 . INTERVENTIONS: None . MEASUREMENTS AND MAIN RESULTS: For each patient, we recorded ICU admission diagnosis, clinical characteristics, and outcome . Among 129 ICU admissions of patients with AIDS, 102 (79%) were admitted for infections, of which (45%) had infections caused by bacteria . Pseudomonas aeruginosa, Staphylococcus aureus, and other enteric pathogens were the most frequent isolates . Pneumonia accounted for 65% of 102 admissions for infections . Overall hospital mortality was 54%, but mortality was higher (68%) for patients with bacterial sepsis . Neutropenia was associated with differences in unadjusted survival rates, whereas CD4 counts were not . Independent predictors of hospital mortality included increasing acute physiology scores and severity of sepsis . CONCLUSIONS: In our ICU, among patients with AIDS, sepsis resulting from bacterial infection is now a more frequent cause of admission than Pneumocystis carinii pneumonia . Severity of illness and the presence of severe sepsis were the clinical predictors most associated with increased mortality . Patients who are not receiving or responding to highly active antiretroviral therapy may become as likely to be admitted to an ICU with a treatable bacterial infection as with classic opportunistic infections . Therefore, broad-spectrum empirical antibacterial therapy is particularly important when the etiology of infection is uncertain. J Infect Dis, 2001 Jun 15, 183(12), 1767 - 74 Epub 2001 May 17. Type III protein secretion is associated with death in lower respiratory and systemic Pseudomonas aeruginosa infections; Roy-Burman A et al.; The ability of Pseudomonas aeruginosa to secrete specific toxins using the type III-mediated pathway has been reported . To determine the association of this phenotype with human illness, immunoblot analysis was used to detect expression of type III secretory proteins in P . aeruginosa isolates from respiratory tract or blood cultures of 108 consecutive patients . Relative risk of mortality was 6-fold greater with expression of the type III secretory proteins ExoS, ExoT, ExoU, or PcrV . Phenotype was independently correlated with toxicity in cellular and murine models . Prevalence of this phenotype was significantly higher in acutely infected patients than in chronically infected patients with cystic fibrosis . These results suggest that the type III protein secretion system is integral to increased P . aeruginosa virulence . A positive phenotype is a predictor of poor clinical outcome . In the future, such analyses may help distinguish potentially lethal infection from colonization and help determine appropriate therapy for critically ill patients. J Bacteriol, 2001 Jun, 183(12), 3712 - 20 Regulation of rpoS gene expression in Pseudomonas: involvement of a TetR family regulator; Kojic M et al.; The rpoS gene encodes the sigma factor which was identified in several gram-negative bacteria as a central regulator during stationary phase . rpoS gene regulation is known to respond to cell density, showing higher expression in stationary phase . For Pseudomonas aeruginosa, it has been demonstrated that the cell-density-dependent regulation response known as quorum sensing interacts with this regulatory response . Using the rpoS promoter of P . putida, we identified a genomic Tn5 insertion mutant of P . putida which showed a 90% decrease in rpoS promoter activity, resulting in less RpoS being present in a cell at stationary phase . Molecular analysis revealed that this mutant carried a Tn5 insertion in a gene, designated psrA (Pseudomonas sigma regulator), which codes for a protein (PsrA) of 26.3 kDa . PsrA contains a helix-turn-helix motif typical of DNA binding proteins and belongs to the TetR family of bacterial regulators . The homolog of the psrA gene was identified in P . aeruginosa; the protein showed 90% identity to PsrA of P . putida . A psrA::Tn5 insertion mutant of P . aeruginosa was constructed . In both Pseudomonas species, psrA was genetically linked to the SOS lexA repressor gene . Similar to what was observed for P . putida, a psrA null mutant of P . aeruginosa also showed a 90% reduction in rpoS promoter activity; both mutants could be complemented for rpoS promoter activity when the psrA gene was provided in trans . psrA mutants of both Pseudomonas species lost the ability to induce rpoS expression at stationary phase, but they retained the ability to produce quorum-sensing autoinducer molecules . PsrA was demonstrated to negatively regulate psrA gene expression in Pseudomonas and in Escherichia coli as well as to be capable of activating the rpoS promoter in E . coli . Our data suggest that PsrA is an important regulatory protein of Pseudomonas spp . involved in the regulatory cascade controlling rpoS gene regulation in response to cell density. Acta Pol Pharm, 2001 Jan-Feb, 58(1), 17 - 20 Changes in the disintegration properties of some brands of paracetamol tablets inoculated with four bacterial species; Obuekwe CO et al.; Four most common brands of paracetamol (4-aceta-midophenol) tablets were examined for the changes in their disintegration properties after inoculation with Staphylococcus aureus, Bacillus cereus, Klebsiella aerogenes and Pseudomonas aeruginosa and incubating for 5 weeks . The disintegration times varied from one brand to the other, reaching maximum values of 72 min., 82 min., 110 min . and 120 min . for S . aureus, B . cereus, P . aeruginosa and Klebsiella aerogenes, respectively . All brands of paracetamol tablets revealed the presence of cotton wool-like fibrils which were seen to be interwoven within the tablets' matrices and these were believed to have caused the higher disintegration times. Drugs, 2001, 61(5), 553 - 64 Emerging strategies in infectious diseases: new carbapenem and trinem antibacterial agents; Sader HS et al.; Beta-lactam antibiotics represent the most commonly prescribed antibacterial agents . New beta-lactams have been introduced continuously as many bacteria have developed resistance to older agents . In the late 1970s, a new class of exceptionally broad spectrum beta-lactams, the carbapenems, was identified . Despite being a very potent compound, the antibacterial activity of the first carbapenem, imipenem, was compromised because of hydrolysis by the renal dehydropeptidase enzyme (DHP-1), and it is now coadministered with a potent competitive inhibitor of the DHP-1 enzyme, cilastin . Molecular modifications in the carbapenem nucleus were able to increase stability to DHP-1 and retain the antibacterial activity . However, some important pathogenic bacteria were found to be resistant to this new class of agents . In addition, other clinically important gram-negative species, such as Pseudomonas aeruginosa, developed resistance mainly by the production of potent beta-lactamases and reduced permeability of the outer membrane . Since the discovery of imipenem/cilastatin, a great number of carbapenems have been developed, and a few of them have been marketed . Stability to hydrolysis by DHP-1 and decrease in toxicity were achieved by meropenem and biapenem . However, only a slight increase in the antibacterial potency and spectrum has been accomplished with either the new marketed or experimental parenteral compounds . In addition, compounds that can be administered orally, such as the carbapenens faropenem, CS-834 and MK-826, and the trinem sanfetrinem, have been developed . However, when compared with the parenterally administered compounds, the oral agents seem to lose some in vitro antibacterial activity, especially against P . aeruginosa. Saudi Med J, 2000 Nov, 21(11), 1081 - 4 Bronchiolitis obliterans organizing pneumonia associated with pseudomonas aeruginosa infection; Akbar DH et al.; Bronchiolitis obliterans organizing pneumonia is a rare disease characterized by the presence of granulation tissue within alveolar ducts and alveoli . Most cases are idiopathic, but it may also be seen during resolution of viral or bacterial pneumonia (mycoplasma, legionella and chlamydia) . It may present as a community acquired pneumonia which does not respond to antibiotics, which make the diagnosis very difficult . We described a 53-year old patient who presented with Bronchiolitis obliterans organizing pneumonia during the resolution of pseudomonas aeruginosa pneumonia . Initially there was slight improvement on antibiotics but later he became severely hypoxic and placed on mechanical ventilator . Diagnosis of Bronchiolitis obliterans organizing pneumonia was obtained with bronchoscopic lung biopsy . He showed an excellent response to steroid treatment . To our knowledge this is the first case of Bronchiolitis obliterans organizing pneumonia secondary to Pseudomonas aeruginosa pneumonia. Mol Microbiol, 2001 May, 40(3), 708 - 18 Cloning and functional characterization of the Pseudomonas aeruginosa rhlC gene that encodes rhamnosyltransferase 2, an enzyme responsible for di-rhamnolipid biosynthesis; Rahim R et al.; Pseudomonas aeruginosa is an opportunistic pathogen capable of producing a wide variety of virulence factors, including extracellular rhamnolipids and lipopolysaccharide . Rhamnolipids are tenso-active glycolipids containing one (mono-rhamnolipid) or two (di-rhamnolipid) L-rhamnose molecules . Rhamnosyltransferase 1 (RhlAB) catalyses the synthesis of mono-rhamnolipid from dTDP-L-rhamnose and beta-hydroxydecanoyl-beta-hydroxydecanoate, whereas di-rhamnolipid is produced from mono-rhamnolipid and dTDP-L-rhamnose . We report here the molecular characterization of rhlC, a gene encoding the rhamnosyltransferase involved in di-rhamnolipid (L-rhamnose-L-rhamnose-beta-hydroxydecanoyl-beta-hydroxydecanoate) production in P . aeruginosa . RhlC is a protein consisting of 325 amino acids with a molecular mass of 35.9 kDa . It contains consensus motifs that are found in other glycosyltransferases involved in the transfer of L-rhamnose to nascent polymer chains . To verify the biological function of RhlC, a chromosomal mutant, RTII-2, was generated by insertional mutagenesis and allelic replacement . This mutant was unable to produce di-rhamnolipid, whereas mono-rhamnolipid was unaffected . In contrast, a null rhlA mutant (PAO1-rhlA) was incapable of producing both mono- and di-rhamnolipid . Complementation of mutant RTII-2 with plasmid pRTII-26 containing rhlC restored the level of di-rhamnolipid production in the recombinant to a level similar to that of the wild-type strain PAO1 . The rhlC gene was located in an operon with an upstream gene (PA1131) of unknown function . A sigma54-type promoter for the PA1131-rhlC operon was identified, and a single transcriptional start site was mapped . Expression of the PA1131-rhlC operon was dependent on the P . aeruginosa rhl quorum-sensing system, and a well-conserved lux box was identified in the promoter region . The genetic regulation of rhlC by RpoN and RhlR was in agreement with the observed increasing RhlC rhamnosyltransferase activity during the stationary phase of growth . This is the first report of a rhamnosyltransferase gene responsible for the biosynthesis of di-rhamnolipid. J Hosp Infect, 2001 May, 48(1), 55 - 65 An evaluation of the use of chlorine dioxide (Tristel One-Shot) in an automated washer/disinfector (Medivator) fitted with a chlorine dioxide generator for decontamination of flexible endoscopes; Coates D; Microbiological tests were carried out to evaluate a new chlorine dioxide sterilant: Tristel OneShot . Preliminary in vitro suspension tests showed that solutions containing around 140 ppm chlorine dioxide achieved a reduction factor exceeding 10(6) of Staphylococcus aureus in 1min and of Bacillus subtilis spores in 2.5 min in the presence of 3g/L bovine albumin . Subsequent tests evaluated the effectiveness of Tristel One-Shot in a Medivator washer/disinfector fitted with a Tristel Generator for processing flexible endoscopes . Each test run involved three stages . In the first, the instrument and air-water channels of a gastroscope were inoculated with a suspension of Pseudomonas aeruginosa (10(8)cfu/ml) in 10% sodium glutamate and serum (0, 5 or 10%) and then drained, partially dried, and saline flushed through for total viable counts (TVCs) . In the second stage, the channels were re-inoculated with test organisms; detergent was flushed through the channels which were then brushed; and saline was flushed through for TVCs . In the third stage, the channels were re-inoculated; detergent was flushed through the channels which were then brushed; the endoscope was processed in the Medivator; and saline was flushed through for TVCs . Carrying out all three stages enabled determination of (1) the contribution played by manual cleaning of channels prior to processing in the Medivator, and (2) the combined effect of manual cleaning followed by processing . Two series of test runs were done . In the first, the Tristel Generator was set to generate 230ppm chlorine dioxide, and in the second 150ppm . In the first, cleaning followed by processing in the Medivator consistently achieved a >/= 10(6)-fold reduction of test organisms, and in the second a >/= 10(5)-fold reduction . Pre-cleaning of channels was very important-when done the initial concentration of serum in the inoculum (0-10%) had no affect on the results obtained after processing . Intensive Care Med, 2001 Mar, 27(3), 503 - 12 Prospective study of nosocomial colonization and infection due to Pseudomonas aeruginosa in mechanically ventilated patients; Berthelot P et al.; OBJECTIVE: To investigate the respective contribution of endogenous and exogenous transmission of Pseudomonas aeruginosa in the colonization of lungs in the mechanically ventilated patient, to estimate the role of P . aeruginosa colonization in the occurrence of severe infections, and to extrapolate appropriate control measures for the prevention of P . aeruginosa ventilator-associated pneumonia . DESIGN: Prospective study of the presence of P . aeruginosa (in stomach fluid, throat specimens, stool, and sputum) on admission, twice a week throughout the patient's stay, and in their environment . O-serotyping, pulsed-field gel electrophoresis, and arbitrarily-primed polymerase chain reaction were used to characterize the strains . SETTING: The two intensive care units (ICUs 1 and 2) of a university hospital . PATIENTS: During a 6-month period, 59 patients were included (21 in ICU 1 and 38 in ICU 2) . RESULTS: P . aeruginosa was isolated in 26 patients, including ten pneumonia cases and seven colonizations on admission . The incidence of acquired colonization was statistically different between the two ICUs: 5.5 and 20.5 per 1000 days of mechanical ventilation, in ICUs 1 and 2, respectively . Endogenous acquisition was the main origin of P . aeruginosa colonization (21 of 26 patients) and the upper respiratory tract was the main bacterial reservoir in broncho-pulmonary colonization and infection . However, during the 6-month period of the study, a multidrug-resistant strain of P . aeruginosa O:11, isolated in the sink of the room of 12 patients, was found responsible for two colonizations (1 digestive, 1 throat/lungs) and one pneumonia . As a whole, from 26 cases of colonization/infection with P . aeruginosa, 5 were related to an exogenous contamination (environmental reservoir in 4 patients and cross-contamination in one patient) . CONCLUSIONS: These results emphasize the need for applying various infection control measures to prevent colonization of patients with P . aeruginosa, including strategies to limit the potential of sinks from acting as a source or reservoir for this bacterium. Intensive Care Med, 2001 Mar, 27(3), 493 - 502 Efficacy and tolerability of piperacillin/tazobactam versus ceftazidime in association with amikacin for treating nosocomial pneumonia in intensive care patients: a prospective randomized multicenter trial; Alvarez-Lerma F et al.; OBJECTIVE: To compare clinical and bacteriological efficacy as well as tolerability of two regimens of broad-spectrum antibiotics (ceftazidime versus piperacillin/tazobactam) combined with amikacin in the treatment of nosocomial pneumonia in intensive care patients . DESIGN: Open label, prospective, multicenter, and randomized phase III clinical trial . SETTING: Medical or surgical intensive care units (ICUs) of nine acute-care teaching hospitals in Spain . PATIENTS AND PARTICIPANTS: One hundred and twenty-four ICU patients with nosocomial pneumonia and requiring mechanical ventilation were included . They were randomized to receive amikacin (15 mg/day divided into two doses) combined with either piperacillin (4 g every 6 h) and tazobactam (0.5 g every 6 h) (n = 88) or ceftazidime (2 g every 8 h) (n = 36) . MEASUREMENTS AND RESULTS: The causative pathogen was determined in 60.2% of patients in the group of amikacin plus piperacillin/tazobactam and in 76.9% in the group of amikacin plus ceftazidime . A total of 94 bacterial organisms were isolated among which gram-negative bacilli predominated, Pseudomonas aeruginosa being the most frequent . Clinical response at the end of antibiotic therapy was considered satisfactory (cure and/or improvement) in 63.9% of patients in the amikacin plus piperacillin/tazobactam group and in 61.5% in the amikacin plus ceftazidime (odds ratio 1.1; 95% confidence interval 0.44-2.75) . Eradication or presumptive eradication rates for each pathogen and for either gram-negative or gram-positive bacteria were similar in both antibiotic combinations (odds ratio 1.2; 95% confidence interval 0.39-3.66) . A total of 21 adverse effects (23.9%) occurred in the amikacin plus piperacillin and tazobactam group and six (16.7%) in the amikacin plus ceftazidime group, thrombocytosis, renal dysfunction, and hepatic cytolysis being the most common . The efficacy and tolerability of the two therapeutic regimens were similar not only in the whole study population, but also in the subset of P . aeruginosa-related pneumonia (odds ratio 1; 95% confidence interval 0.08-13.37) . CONCLUSIONS: Amikacin associated with either ceftazidime or piperacillin and tazobactam has shown comparable efficacy and tolerability in the treatment of ICU patients with nosocomial pneumonia. Rev Pneumol Clin, 2001 Apr, 57(2), 67 - 72 {Prevention of infection transmitted by bronchial fibroscopes}; Belleguic C et al.; A growing number of infectious complications are reported after bronchial fibroscopy procedures . The risk of nosocomial patient-to-patient or environment-to-patient infection is real via contaminated fibroscopes . Cross transmission can be caused by several microorganisms, the most frequently identified being Pseudomonas aeruginosa, Mycobacterium tuberculosis and other atypical mycobacteria . Fibroscopes can be contaminated via different mechanisms, generally related to poorly adapted cleaning and decontamination protocols . Identified errors include an insufficient cleaning phase, an inappropriate disinfecting agent (iodine derivatives, chlorhexidine), defective cleaning or disinfection of accessory equipment, or use of tap water for rinsing . Finally several episodes of Pseudomonas and atypical mycobacteria infections have been found to result from the use of automatic cleaning machines . Particular attention must be paid to the use of these devices . Official guidelines for the disinfection of endoscopic equipment must be rigorously applied in all centers . The personnel should have adequate training . It is also important to take regular samples and make regular bacteriology controls of the water and fibroscopic equipment. Proc Natl Acad Sci U S A, 2001 May 22, 98(11), 5981 - 5 Epub 2001 May 15. Designing surfaces that kill bacteria on contact; Tiller JC et al.; Poly(4-vinyl-N-alkylpyridinium bromide) was covalently attached to glass slides to create a surface that kills airborne bacteria on contact . The antibacterial properties were assessed by spraying aqueous suspensions of bacterial cells on the surface, followed by air drying and counting the number of cells remaining viable (i.e., capable of growing colonies) . Amino glass slides were acylated with acryloyl chloride, copolymerized with 4-vinylpyridine, and N-alkylated with different alkyl bromides (from propyl to hexadecyl) . The resultant surfaces, depending on the alkyl group, were able to kill up to 94 +/- 4% of Staphylococcus aureus cells sprayed on them . A surface alternatively created by attaching poly(4-vinylpyridine) to a glass slide and alkylating it with hexyl bromide killed 94 +/- 3% of the deposited S . aureus cells . On surfaces modified with N-hexylated poly(4-vinylpyridine), the numbers of viable cells of another Gram-positive bacterium, Staphylococcus epidermidis, as well as of the Gram-negative bacteria Pseudomonas aeruginosa and Escherichia coli, dropped more than 100-fold compared with the original amino glass . In contrast, the number of viable bacterial cells did not decline significantly after spraying on such common materials as ceramics, plastics, metals, and wood. Antimicrob Agents Chemother, 2001 Jun, 45(6), 1930 - 3 Azithromycin inhibits quorum sensing in Pseudomonas aeruginosa; Tateda K et al.; We report that 2 microg of azithromycin/ml inhibits the quorum-sensing circuitry of Pseudomonas aeruginosa strain PAO1 . Addition of synthetic autoinducers partially restored the expression of the trancriptional activator-encoding genes lasR and rhlR but not that of the autoinducer synthase-encoding gene lasI . We propose that azithromycin interferes with the synthesis of autoinducers, by an unknown mechanism, leading to a reduction of virulence factor production. Antimicrob Agents Chemother, 2001 Jun, 45(6), 1780 - 7 C-terminal region of Pseudomonas aeruginosa outer membrane porin OprD modulates susceptibility to meropenem; Epp SF et al.; We investigated the unusual susceptibility to meropenem observed for seven imipenem-resistant clinical isolates of Pseudomonas aeruginosa . These strains were genetically closely related, expressed OprD, as determined by Western blot analyses, and were resistant to imipenem (>5 microg/ml) but susceptible to meropenem (<1 microg/ml) . The oprD genes from two isolates were entirely sequenced, and their deduced protein sequences showed 93% identity with that of OprD of strain PAO1 . The major alteration consisted of the replacement of a stretch of 12 amino acids, located in putative external loop L7 of OprD, by a divergent sequence of 10 amino acid residues . The oprD gene variants and the wild-type oprD gene were cloned and expressed in a defined oprD mutant . The meropenem MICs for strains carrying the oprD genes from clinical isolates were four times lower than that for the strain carrying the wild-type oprD gene . Imipenem activities, however, were comparable for all strains . Furthermore, meropenem hypersusceptibility was obtained with a hybrid OprD porin that consisted of the PAO1 oprD gene containing loop L7 from a clinical isolate . These results show that the C-terminal portion of OprD, in particular, loop L7, was responsible for the unusual meropenem hypersusceptibility . Competition experiments suggested that the observed OprD modifications in the clinical isolates did not affect antagonism between imipenem and the basic amino acid L-lysine . We further propose that shortening of putative loop L7 of the OprD porin by 2 amino acid residues sufficiently opens the porin channel to allow optimal penetration of meropenem and increase its activity . In contrast, this alteration would not affect susceptibility to a smaller carbapenem molecule, such as imipenem. Antimicrob Agents Chemother, 2001 Jun, 45(6), 1615 - 20 Oxacillinase-mediated resistance to cefepime and susceptibility to ceftazidime in Pseudomonas aeruginosa; Aubert D et al.; Pseudomonas aeruginosa clinical isolate SOF-1 was resistant to cefepime and susceptible to ceftazidime . This resistance phenotype was explained by the expression of OXA-31, which shared 98% amino acid identity with a class D beta-lactamase, OXA-1 . The oxa-31 gene was located on a ca . 300-kb nonconjugative plasmid and on a class 1 integron . No additional efflux mechanism for cefepime was detected in P . aeruginosa SOF-1 . Resistance to cefepime and susceptibility to ceftazidime in P . aeruginosa were conferred by OXA-1 as well. CLAO J, 2001 Apr, 27(2), 89 - 93 Comparative radiolabel and ATP analyses of adhesion of Pseudomonas aeruginosa and Staphylococcus epidermidis to hydrogel lenses; Ahanotu EN et al.; PURPOSE: A comparative assessment of the relative primary adhesion of cells of Pseudomonas aeruginosa, its lux transformant, and of slime and non-slime producing strains of Staphylococcus epidermidis to various hydrogel lenses was conducted . METHODS: Hydrogel lenses were placed in cell suspensions with bacteria with or without a tritiated leucine label . After 2 hours exposure, the lenses were rinsed vigorously and densities of cells on the lenses were determined via scintillation counting or ATP analyses . RESULTS: The radiolabel procedure indicated greater numbers than the ATP analyses of adhered cells per lens per common inoculum of all strains . All strains exhibited greater primary adhesion to the 38% water content contact lens, with the lux transformant of P . aeruginosa showing the greatest degree of adhesion . Primary adhesion by P . aeruginosa was typically at least ten-fold greater per lens than that observed with S . epidermidis . CONCLUSIONS: Both a radiolabel-cell procedure and bioluminescent ATP analyses demonstrated similar patterns of primary adhesion of bacteria to hydrogel lenses . Generally the adhesion increased inversely to the water content of the lenses but the chemical composition of the lenses, particularly surface properties, altered this pattern for lenses of similar water content . The magnitude of primary adhesion varied with the species and strain of bacterium. CLAO J, 2001 Apr, 27(2), 81 - 3 Low-power microwave disinfection of soft contact lenses; Kastl PR et al.; PURPOSE: To evaluate the effectiveness of low-power microwave irradiation on the disinfection of contaminated contact lenses . METHODS: We infected 24 soft contact lenses with Staphylococcus epidermidis, Staphylococcus aureus, and Pseudomonas aeruginosa . Each contact lens was placed into a vial containing 10 mL of contaminated, non-preserved saline . Each vial was then exposed to 2, 4, 6, 9, 12, 15, or 18 minutes of microwave irradiation in an oven (2,450 MHz 600W) set at 10% power . RESULTS: We found all contact lenses to be disinfected after 15 minutes of exposure . All lenses remained hydrated and revealed no apparent damage when examined under the slit-lamp biomicroscope . CONCLUSIONS: Low level microwaving in a home microwave oven can result in effective and non-damaging disinfection of soft contact lenses. J Biol Chem, 2001 Jul 13, 276(28), 26030 - 5 Epub 2001 May 11. Maturation of Pseudomonas aeruginosa elastase . Formation of the disulfide bonds; Braun P et al.; Elastase of Pseudomonas aeruginosa is synthesized as a preproenzyme . After propeptide-mediated folding in the periplasm, the proenzyme is autoproteolytically processed, prior to translocation of both the mature enzyme and the propeptide across the outer membrane . The formation of the two disulfide bonds present in the mature enzyme was examined by studying the expression of the wild-type enzyme and of alanine for cysteine mutant derivatives in the authentic host and in dsb mutants of Escherichia coli . It appeared that the two disulfide bonds are formed successively . First, DsbA catalyzes the formation of the disulfide bond between Cys-270 and Cys-297 within the proenzyme . This step is essential for the subsequent autoproteolytic processing to occur . The second disulfide bond between Cys-30 and Cys-57 is formed more slowly and appears to be formed after processing of the proenzyme, and its formation is catalyzed by DsbA as well . This second disulfide bond appeared to be required for the full proteolytic activity of the enzyme and contributes to its stability. Am J Respir Cell Mol Biol, 2001 May, 24(5), 608 - 15 Alveolar macrophages that phagocytose apoptotic neutrophils produce hepatocyte growth factor during bacterial pneumonia in mice; Morimoto K et al.; Hepatocyte growth factor (HGF) is postulated to play an important role in the repair of pulmonary epithelium in acute lung injury . To evaluate the role of HGF in bacterial pneumonia, the kinetics of HGF production and the cellular sources of HGF have been examined in the lungs of mice that had been intratracheally challenged with Pseudomonas aeruginosa . Neutrophil accumulation in the airway occurred immediately, reached a peak at 36 h, and then progressively declined by 14 d after infection . We found a biphasic pattern of HGF messenger RNA expression and protein synthesis in the lung after bacterial infection . The first peak for HGF production was found at 6 h after infection, and the primary source of HGF was shown to be bronchial epithelial cells . Interestingly, the second peak for HGF production, which was found around 48 to 72 h after infection, was closely associated with the increase in the percentage of alveolar macrophages (AMs) that became positive for myeloperoxidase, indicating phagocytosis of apoptotic neutrophils . The cellular source of the second peak was found to be AMs . Further, murine AMs which phagocytosed apoptotic neutrophils induced higher levels of HGF production in vitro . These results strongly indicate a novel mechanism of HGF production by AMs, which are phagocytosing apoptotic neutrophils, and the pivotal role of AMs in the healing and repair of damaged pulmonary epithelium through the production of HGF. Infect Immun, 2001 Jun, 69(6), 4116 - 9 Effects of exogenous interleukin-6 during Pseudomonas aeruginosa corneal infection; Cole N et al.; Lack of interleukin-6 (IL-6) during Pseudomonas aeruginosa corneal infection leads to more severe disease with changes in neutrophil recruitment . Exogenous IL-6 leads to increased efficiency of neutrophil recruitment and reduced bacterial loads in corneal infection in both IL-6 gene knockout and wild-type mice . This may be mediated by IL-6 increasing the production of corneal macrophage inflammatory protein 2 and intercellular cell adhesion molecule 1 . We conclude that effective recruitment of neutrophils into the cornea is dependent on the production of IL-6 and that early augmentation of IL-6 may be protective in corneal infection. New Microbiol, 2001 Apr, 24(2), 157 - 63 AP-PCR typing of Pseudomonas aeruginosa isolated from patients with cystic fibrosis; Giordano A et al.; Arbitrarily Primed Polymerase Chain Reaction has been used for an epidemiological evaluation of 42 strains of P . aeruginosa isolated from nine cystic fibrosis patients during a three-year investigation period . The resistance patterns of the same strains have also been evaluated . The AP-PCR type fingerprinting was perfomed with primers 10514 and 208 . Resistance was evaluated by the Minimal Inhibitory Concentration method . With 10514 eleven different genotypes could be evidenced, while with 208 only five of them could be detected . During the investigation period patients were always colonised by the same genotype . A possible correlation between resistance pattern and genotype with both primers has shown, within the same patient, a correspondence of about 20% for 10514 and a correspondence of only 10% for 208 . Patients are colonised by one or two strains of P . aeruginosa and there is no relation between genotype and resistance pattern. Wiad Lek, 2001, 54(1-2), 105 - 9 {Deafness, as a complication of Pseudomonas aeruginosa septicemia in a 9-year old boy with acute lymphoblastic leukemia}; Bubala H et al.; We described natural history of Pseudomonas aeruginosa septicemia in 9 year old boy, who was treated for acute lymphoblastic leukemia (ALL) . After 14 day treatment of ALL the following signs and symptoms occurred: fever, earache with otorhoea, deafness, bilateral peripheral paralysis of n . VII, erythema, pneumonia, paralytic ileus . After 4 weeks of antimicrobial and supportive therapy, in the 10th week of chemotherapy, he achieved haematological remission . During continuation therapy, two-stage bilateral myringoplasty was performed . At present the maintenance therapy is continued, and in the future hearing aid and cochlear implant, will be applied. J Clin Lab Anal, 2001, 15(3), 131 - 7 Real-time identification of Pseudomonas aeruginosa direct from clinical samples using a rapid extraction method and polymerase chain reaction (PCR); Jaffe RI et al.; Pseudomonas aeruginosa has emerged as one of the most problematic Gram-negative nosocomial pathogens . Bacteremia caused by P . aeruginosa is clinically indistinguishable from other Gram-negative infections although the mortality rate is higher . This microorganism is also inherently resistant to common antibiotics . Standard bacterial identification and susceptibility testing is normally a 48-hour process and difficulty sometimes exists in rapidly and accurately identifying antimicrobial resistance . The Polymerase Chain Reaction (PCR) is a rapid and simple process for the amplification of target DNA sequences . However, many sample preparation methods are unsuitable for the clinical laboratory because they are not cost effective, take too long to perform, or do not provide a good template for PCR . Our goal was to provide same-day results to facilitate rapid diagnosis . In this report, we have utilized our rapid DNA extraction method to generate bacterial DNA direct from clinical samples for PCR . The lower detection level for P . aeruginosa was estimated to be 10 CFU/ml . In addition, we wanted to compare the results of a new rapid-cycle DNA thermocycler that uses continuous fluorescence monitoring with the results of standard thermocycling . We tested 40 clinical isolates of P . aeruginosa and 18 non-P . aeruginosa isolates received in a blinded fashion . Coded data revealed that there was 100% correlation in both the rapid-cycle DNA thermocycling and standard thermocycling when compared to standard clinical laboratory results . In addition, total results turn-around time was less than 1 hour . Specific identification of P . aeruginosa was determined using intragenic primer sets for bacterial 16S rRNA and Pseudomonas outer-membrane lipoprotein gene sequences . The total cost of our extraction method and PCR was $2.22 per sample . The accuracy and rapidness of this DNA-extraction method, with its PCR-based identification system, make it an ideal candidate for use in the clinical laboratory. J Bacteriol, 2001 Jun, 183(11), 3345 - 52 Adenylate kinase as a virulence factor of Pseudomonas aeruginosa; Markaryan A et al.; Adenylate kinase (AK; ATP:AMP phosphotransferase, EC 2.7.4.3) is a ubiquitous enzyme that contributes to the homeostasis of adenine nucleotides in eukaryotic and prokaryotic cells . AK catalyzes the reversible reaction Mg . ATP + AMP <--> Mg . ADP + ADP . In this study we show that AK secreted by the pathogenic strains of Pseudomonas aeruginosa appears to play an important role in macrophage cell death . We purified and characterized AK from the growth medium of a cystic fibrosis isolate strain of P . aeruginosa 8821 and hyperproduced it as a fusion protein with glutathione S-transferase . We demonstrated enhanced macrophage cell death in the presence of both the secreted and recombinant purified AK and its substrates AMP plus ATP or ADP . These data suggested that AK converts its substrates to a mixture of AMP, ADP, and ATP, which are potentially more cytotoxic than ATP alone . In addition, we observed increased macrophage killing in the presence of AK and ATP alone . Since the presence of ATPase activity on the macrophages was confirmed in the present work, external macrophage-effluxed ATP is converted to ADP, which in turn can be transformed by AK into a cytotoxic mixture of three adenine nucleotides . Evidence is presented in this study that secreted AK was detected in macrophages during infection with P . aeruginosa . Thus, the possible role of secreted AK as a virulence factor is in producing and keeping an intact pool of toxic mixtures of AMP, ADP, and ATP, which allows P . aeruginosa to exert its full virulence. J Pediatr, 2001 May, 138(5), 699 - 704 Clinical outcome after early Pseudomonas aeruginosa infection in cystic fibrosis; Nixon GM et al.; OBJECTIVE: To determine the clinical consequences of acquiring Pseudomonas aeruginosa infection during early childhood in children with cystic fibrosis (CF) . DESIGN: Prospective, observational cohort study of 56 children with CF identified by newborn screening during 1990-92 . Each child underwent an annual bronchial lavage during the first 2 to 3 years of life . Clinical outcome was determined at 7 years of age . RESULTS: P aeruginosa infection was diagnosed in 24 (43%) cohort subjects . Four children died before 7 years of age, all of whom had been infected with a multi-resistant, mucoid strain of P aeruginosa (P =.04) . In survivors, P aeruginosa infection was associated with significantly increased morbidity as measured by lower National Institutes of Health scores, greater variability in lung function, increased time in the hospital, and higher rates of recombinant human deoxyribonuclease therapy (P <.01) . In this young CF cohort, best forced expiratory volume in 1 second was an insensitive measure of increased morbidity . CONCLUSIONS: Acquisition of P aeruginosa was common by 7 years of age in this CF birth cohort and was associated with increased morbidity and mortality . An improved disease severity score would improve the evaluation and study of early CF lung disease. J Biol Chem, 2001 Jul 13, 276(28), 26479 - 85 Epub 2001 May 07. Structural characterization of the Pseudomonas aeruginosa 1244 pilin glycan; Castric P et al.; An antigenic similarity between lipopolysaccharide (LPS) and glycosylated pilin of Pseudomonas aeruginosa 1244 was noted . We purified a glycan-containing molecule from proteolytically digested pili and showed it to be composed of three sugars and serine . This glycan competed with pure pili and LPS for reaction with an LPS-specific monoclonal antibody, which also inhibited twitching motility by P . aeruginosa bearing glycosylated pili . One-dimensional NMR analysis of the glycan indicated the sugars to be 5N beta OHC(4)7NfmPse, Xyl, and FucNAc . The complete proton assignments of these sugars as well as the serine residue were determined by COSY and TOCSY . Electrospray ionization mass spectrometry (MS) determined the mass of this molecule to be 771.5 . The ROESY NMR spectrum, tandem MS/MS analysis, and methylation analysis provided information on linkage and the sequence of oligosaccharide components . These data indicated that the molecule had the following structure: alpha-5N beta OHC(4)7NFmPse-(2-->4)beta-Xyl-(1-->3)-beta-FucNAc-(1-->3)-beta-Ser. J Pharm Pharmacol, 2001 Apr, 53(4), 579 - 82 Buchu (Agathosma betulina and A . crenulata, Rutaceae) essential oils: their pharmacological action on guinea-pig ileum and antimicrobial activity on microorganisms; Lis-Balchin M et al.; The mode of action of essential oils from two buchu species (Agathosma betulina and A . crenulata, Rutaceae) from the Cape region of South Africa has been studied on smooth muscle in-vitro using guinea-pig ileum . At high concentration, the oils had an initial spasmogenic activity followed by spasmolysis . The spasmolytic action was post-synaptic, not atropine-like and did not involve adrenoceptor or guanylyl cyclase activation . In the presence of the phosphodiesterase inhibitor rolipram, the spasmolytic action of A . betulina was significantly increased whilst that due to A . crenulata was also increased but not to a significant level . These results suggested a mode of action for the oils involving cyclic adenosine monophosphate . In addition, A . betulina appeared to block calcium channels but this was not seen with A . crenulata, possibly because the initial spasmogenic activity complicated the study of its spasmolytic action . Neither essential oil (10 microL, undiluted) demonstrated antimicrobial action against Enterococcus hirae and Pseudomonas aeruginosa but very low activity was observed against Escherichia coli, Saccharomyces cerevisiae and Staphylococcus aureus, suggesting little potential for these oils as antimicrobial agents/preservatives. Jpn J Antibiot, 2001 Feb, 54(2), 79 - 87 {Antimicrobial susceptibility of Pseudomonas aeruginosa isolated in Fukushima Prefecture}; Niitsuma K et al.; We investigated the susceptibility of Pseudomonas aeruginosa (isolated from the sputum of patients with respiratory infection in 4 medical institutions in Fukushima Prefecture) to 8 beta-lactam antibiotics including three carbapenems and relationships among MICs of antibiotics tested . The MIC90 values for a total of 216 strains were 6.25 micrograms/ml for meropenem, 12.5 micrograms/ml for imipenem and ceftazidime, 25 micrograms/ml for panipenem and cefsulodin, 50 micrograms/ml for cefpirome and over than 200 micrograms/ml for cefoperazone and piperacillin . The frequency of resistance of these strains to each antibiotic was as follows: The resistant strains were 19 (8.8%) for meropenem, 34 (15.7%) for imipenem and ceftazidime, 50 (23.1%) for cefsulodin, 72 (33.3%) for panipenem, 76 (35.2%) for piperacillin and 90 (41.7%) for cefpirome . Eighteen strains (18.3%) of 19 meropenem resitant straisn were resistant to imipenem and panipenem, but 16 strains of the 34 imipenem-resistant strains and 54 strains of the 72 panipenem-resistant strains were susceptible to meropenem . In investigation of isolation of multi-resistant Pseudomonas aeruginosa, the susceptibility of strains tested to 7 antibiotics except cefoperazone was as follows: The strains susceptible to all the 7 antibiotics were 92 strains (42.6%), and 33 strains (15.2%) were resistant to 2 antibiotics, 31 strains (14.4%) were resistant to 1 antibiotic, 21 strains (9.7%) were resistant to 3 antibiotics, 13 strains (6.0%) were resistant to 5 antibiotics, 9 (4.2%) were resistant to 4 and 7 antibiotics, and 8 strains (3.7%) were reistant to 6 antibiotics . Since the emergence of these multi-resistant strains is closely related to frequent use of antibiotics for nosocomial infections, special attention should be paid to the antimicrobial susceptibility of Pseudomonas aeruginosa and the situation of antibiotic resistant strains. Jpn J Antibiot, 2001 Feb, 54(2), 69 - 78 {Combination effect of arbekacin and cefepime on mixed culture of MRSA and P . aeruginosa}; Araake M et al.; We investigated the in vitro combination effects of arbekacin (ABK), vancomycin (VCM) or teicoplanin (TEIC) and cefepime (CFPM) on methicillin-resistant Staphylococcus aureus (MRSA) and Pseudomonas aeruginosa isolated from various clinical specimens . Using checkerboard titration technique by agar dilution, combinations of ABK, VCM or TEIC and CFPM exhibited synergistic effects on 25 MRSA strains . The similar effects were also observed on ABK-resistant MRSA . Combination of ABK and CFPM exhibited a good effect on P . aeruginosa, but combinations of VCM or TEIC and CFPM exhibited no synergistic effect on P . aeruginosa . In vitro bactericidal activities of ABK, VCM or TEIC and CFPM against mixed cultures of MRSA with P . aeruginosa were examined at concentration of each drug that corresponds to the serum concentration at 3 hours after the usual therapeutic dosage . VCM or TEIC alone showed bacteriostatic effects against MRSA, and no enhancements were observed when combined with CFPM . ABK alone showed good bactericidal activity against MRSA and combination with CFPM enhanced the bactericidal activity . Against P . aeruginosa, ABK or CFPM alone showed the bactericidal activity, and strong bactericidal activity was induced by the combination of ABK and CFPM . VCM and TEIC showed no bactericidal activities against P . aeruginosa . When CFPM combined with VCM or TEIC, the bactericidal activity of CFPM was not enhanced against P . aeruginosa. Nature, 2001 May 3, 411(6833), 98 - 102 Exploitation of syndecan-1 shedding by Pseudomonas aeruginosa enhances virulence; Park PW et al.; Cell-surface heparan sulphate proteoglycans (HSPGs) are ubiquitous and abundant receptors/co-receptors of extracellular ligands, including many microbes . Their role in microbial infections is poorly defined, however, because no cell-surface HSPG has been clearly connected to the pathogenesis of a particular microbe . We have previously shown that Pseudomonas aeruginosa, through its virulence factor LasA, enhances the in vitro shedding of syndecan-1-the predominant cell-surface HSPG of epithelia . Here we show that shedding of syndecan-1 is also activated by P . aeruginosa in vivo, and that the resulting syndecan-1 ectodomains enhance bacterial virulence in newborn mice . Newborn mice deficient in syndecan-1 resist P . aeruginosa lung infection but become susceptible when given purified syndecan-1 ectodomains or heparin, but not when given ectodomain core protein, indicating that the ectodomain's heparan sulphate chains are the effectors . In wild-type newborn mice, inhibition of syndecan-1 shedding or inactivation of the shed ectodomain's heparan sulphate chains prevents lung infection . Our findings uncover a pathogenetic mechanism in which a host response to tissue injury-syndecan-1 shedding-is exploited to enhance microbial virulence apparently by modulating host defences. Pediatr Infect Dis J, 2001 Apr, 20(4), 452 - 4 Nonopsonic phagocytosis of Pseudomonas aeruginoas: insights from an infant with leukocyte adhesion deficiency; Pollard AJ et al.; Children with leukocyte adhesion deficiency type I are at risk for overwhelming infection because their neutrophils lack surface beta 2 integrins (CD18/CD11) that normally interact with endothelial cell adhesion molecules and mediate migration to sites of bacterial invasion . In vitro studies of phagocytic cells from an infant with leukocyte adhesion deficiency type I demonstrated that complement receptor 3 (CD18/CD11b) mediates nonopsonic phagocytosis of some Pseudomonas aeruginosa strains and might play a control role in the control of Pseudomonas infections at sites where there are low levels of opsonins. Mol Cell Biochem, 2001 Feb, 218(1-2), 105 - 11 All-D-cecropin B: synthesis, conformation, lipopolysaccharide binding, and antibacterial activity; Bland JM et al.; Cecropin B (LCB) is a natural peptide with antibacterial and antifungal properties . The enantiomer of LCB, containing all-D amino acids (DCB), was synthesized to examine its antibacterial and binding properties . The conformation of DCB was compared to its enantiomer by circular dichroism . Both the L- and D-peptides showed an identical induction of alpha-helical secondary structure . However, binding studies between Lipopolysaccharide (LPS) and DCB or LCB were studied with a dimethylmethylene blue spectrophotometric assay, showing the two enantiomeric peptides differed in their interaction with LPS . Antibacterial activity of DCB was determined against three Gram-negative bacteria, Pantoea agglomerans (ATCC 27996), Escherichia coli (ATCC 8739), and Pseudomonas aeruginosa (ATCC 17648), giving comparable results to LCB. Appl Microbiol Biotechnol, 2001 Mar, 55(2), 205 - 9 Heterologous expression of the acyl-acyl carrier protein thioesterase gene from the plant Umbellularia californica mediates polyhydroxyalkanoate biosynthesis in recombinant Escherichia coli; Rehm BH et al.; The acyl-acyl carrier protein (ACP) thioesterase cDNA from the plant Umbellularia californica was functionally expressed in various recombinant Escherichia coli strains in order to establish a new metabolic route toward medium-chain-length polyhydroxyalkanoate (PHA(MCL)) biosynthesis from non-related carbon sources . Coexpression of the PHA synthase genes from Ralstonia eutropha and Pseudomonas aeruginosa, or only the PHA synthase gene from P . aeruginosa, respectively, showed PHA(MCL) accumulation when the type II PHA synthase from P . aeruginosa was produced . Both wild-type E . coli and various fad mutants were investigated; and only when the beta-oxidation pathway was impaired PHA(MCL) accumulation from gluconate was observed, contributing to about 6% of cellular dry weight . Thus coexpression of type II PHA synthase gene with cDNA encoding the medium-chain acyl-ACP thioesterase from U . californica established a new PHA(MCL) biosynthesis pathway, connecting fatty acid de novo biosynthesis with fatty acid beta-oxidation, using a non-related carbon source. Transplantation, 2001 Mar 27, 71(6), 744 - 5 Continuous beta-lactam antibiotic therapy in a double-lung transplanted patient with a multidrug-resistant Pseudomonas aeruginosa infection; Domenig C et al.; BACKGROUND: It is well known that the bactericidal effect of beta-lactam antibiotics is closely related to the time which the serum concentration of the antibiotic remains above the minimal inhibitory concentration of the target pathogen . Thus, the optimal administration of beta-lactam antibiotics would be the continuous infusion of the drug . METHODS: We present a case report with a critically ill double-lung transplanted patient with pneumonia due to a multidrug-resistant Pseudomonas aeruginosa who received continuously 8 g meropenem/24 hr . Based on a previous pharmacokinetic study showing that continuous infusion of meropenem is at least equivalent to intermittent administration this case report is reported to demonstrate the clinical efficacy of continuous infusion . RESULTS: C-reactive protein and pneumonia decreased rapidly when clinical conditions were improved significantly . Continuous administration of meropenem did not interfere with cyclosporine, no side effects were seen, and the patient's renal function was not impaired during the whole period of treatment . CONCLUSION: The continuous administration of beta-lactam antibiotics is a powerful application in critically ill patients to intensify antimicrobial therapy. Clin Diagn Lab Immunol, 2001 May, 8(3), 632 - 6 Elastase deficiency phenotype of Pseudomonas aeruginosa canine otitis externa isolates; Petermann SR et al.; Pseudomonas aeruginosa veterinary isolates were assayed for elastase and total matrix protease activity . The elastase activity of canine ear isolates was much less than that of strain PAO1 and that of all other veterinary isolates (P < 0.0001) . The results indicate that canine ear isolates have a distinct elastase phenotype. J Antimicrob Chemother, 2001 May, 47(5), 665 - 70 In vitro activity of garlic oil and four diallyl sulphides against antibiotic-resistant Pseudomonas aeruginosa and Klebsiella pneumoniae; Tsao S et al.; The in vitro antibacterial activities of garlic oil and four diallyl sulphides naturally occurring in this oil were studied against Pseudomonas aeruginosa and Klebsiella pneumoniae (total 237 clinical isolates) . Garlic oil at 4 x MIC could reduce original inoculum to <or=2 log(10) in both P . aeruginosa and K . pneumoniae within 8 h . The MIC values of four diallyl sulphides against these two pathogens followed the order diallyl monosulphide > diallyl disulphide > diallyl trisulphide (DAT) > diallyl tetrasulphide (DATS) (P < 0.05) . Most interactions of ceftazidime, gentamicin, imipenem and meropenem with DAT or DATS, determined according to the fractional inhibitory concentration index, showed synergic or additive effects . These results suggest that garlic oil, DAT and DATS may have potential for the prevention or treatment of nosocomial, antibiotic-resistant bacterial infections. J Antimicrob Chemother, 2001 May, 47(5), 617 - 22 In vivo efficacy of continuous infusion versus intermittent dosing of ceftazidime alone or in combination with amikacin relative to human kinetic profiles in a Pseudomonas aeruginosa rabbit endocarditis model; Robaux MA et al.; Ceftazidime and amikacin were administered in a Pseudomonas aeruginosa rabbit endocarditis model using computer-controlled intravenous (iv) infusion pumps to simulate human serum concentrations for the following regimens: continuous (constant rate) infusion of 4, 6 or 8 g of ceftazidime over 24 h or intermittent dosing of 2 g every 8 h either alone or in combination with amikacin (15 mg/kg once daily) . The in vivo activities of these regimens were tested on four Pseudomonas aeruginosa strains . Animals were killed 24 h after the beginning of treatment . Efficacy was assessed by comparing the effects of the different groups on bacterial counts in vegetations for each strain tested . For a susceptible reference strain (ATCC 27853; MICs of ceftazidime and amikacin 1 and 2 mg/L, respectively), continuous infusion of 4 g alone or with amikacin was as effective as intermittent dosing with amikacin . For a clinical isolate producing an oxacillinase (MICs of ceftazidime and amikacin 8 and 32 mg/L, respectively), continuous infusion of 6 g was equivalent to intermittent dosing . For a clinical isolate producing a TEM-2 penicillinase (MIC of ceftazidime and amikacin 4 mg/L), continuous infusion of 6 g, but not intermittent dosing, had a significant in vivo effect . For a clinical isolate producing an inducible, chromosomally encoded cephalosporinase (MIC of ceftazidime and amikacin 8 and 4 mg/L, respectively), neither continuous infusion nor intermittent dosing proved effective . Determination of ceftazidime concentrations in vegetations showed that continuous infusion produced tissue concentrations at the infection site far greater than the MIC throughout the treatment . It is concluded that continuous infusion of the same total daily dose provides significant activity as compared with fractionated infusion . This study confirms that a concentration of 4-5 x MIC is a reasonable therapeutic target in most clinical settings of severe P . aeruginosa infection. Invest Ophthalmol Vis Sci, 2001 May, 42(6), 1247 - 53 Stimulatory effect of pseudomonal elastase on collagen degradation by cultured keratocytes; Nagano T et al.; PURPOSE . The pathobiology of corneal ulceration induced by Pseudomonas aeruginosa was investigated by characterization of the pseudomonal pathogenic factors responsible for degradation of the collagen matrix . METHODS . Three-dimensional gels of type I collagen containing (or not) rabbit keratocytes were incubated in the presence of either culture supernatant of P . aeruginosa strain PAO1 or pseudomonal pathogenic factors (elastase, lipopolysaccharide, or exotoxin A), and the extent of collagen degradation was assessed after 24 hours by measurement of released hydroxyproline . Activation of matrix metalloproteinases (MMPs) produced by keratocytes was also examined by gelatin zymography and immunoblot analysis . RESULTS . In the absence of keratocytes, the PAO1-conditioned medium increased the extent of collagen degradation . The conditioned medium also promoted keratocyte-mediated collagen degradation . Of the pseudomonal pathogenic factors examined, only elastase degraded collagen directly as well as stimulated keratocyte-mediated collagen degradation . Culture supernatant of elastase-deficient P . aeruginosa (lasR or lasB) mutants had no effect on collagen degradation in the absence or presence of keratocytes . Elastase also induced the conversion of the inactive precursors of MMP-1, -2, -3, and -9 produced by keratocytes to the active forms of the enzymes . CONCLUSIONS . These results suggest that pseudomonal elastase both degrades type I collagen directly and promotes collagen degradation mediated by keratocytes, the latter effect being likely attributable, at least in part, to the activation of proMMPS: Cancer Res, 2001 May 1, 61(9), 3698 - 703 Cancer immunotherapy using a DNA vaccine encoding the translocation domain of a bacterial toxin linked to a tumor antigen; Hung CF et al.; Certain domains of bacterial toxins have been shown to facilitate translocation from extracellular and vesicular compartments into the cytoplasm . This feature represents an opportunity to enhance class I presentation of exogenous antigen to CD8(+) T cells . We investigated this notion by creating a novel fusion of the translocation domain (domain II) of Pseudomonas aeruginosa exotoxin A (ETA(dII)) with a model tumor antigen, human papillomavirus type 16 E7, in the context of a DNA vaccine . Our in vitro studies indicated that cells transfected with ETA(dII)/E7 DNA or dendritic cells pulsed with lysates containing ETA(dII)/E7 protein exhibited enhanced MHC class I presentation of E7 antigen . Vaccination of mice with ETA(dII)/E7 DNA generated a dramatic increase in the number of E7-specific CD8(+) T cell precursors ( approximately 30-fold compared with wild-type E7 DNA) and converted a less effective DNA vaccine into one with significant potency against human papillomavirus type 16 E7-expressing murine tumors via a CD8-dependent pathway . These results indicate that fusion of the translocation domain of a bacterial toxin to an antigen may greatly enhance vaccine potency. Biophys J, 2001 May, 80(5), 2431 - 8 Structural perturbations of azurin deposited on solid matrices as revealed by trp phosphorescence; Gabellieri E et al.; The phosphorescence emission of Cd-azurin from Pseudomonas aeruginosa was used as a probe of possible perturbations in the dynamical structure of the protein core that may be induced by protein-sorbent and protein-protein interactions occurring when the macromolecule is deposited into amorphous, thin solid films . Relative to the protein in aqueous solution, the spectrum is unrelaxed and the phosphorescence decay becomes highly heterogeneous, the average lifetime increasing sharply with film thickness and upon its dehydration . According to the lifetime parameter, adsorption of the protein to the substrate is found to produce a multiplicity of partially unfolded structures, an influence that propagates for several protein layers from the surface . Among the substrates used for film deposition, hydrophilic silica, dextran, DEAE-dextran, dextran sulfate, and hydrophobic octodecylamine, the perturbation is smallest with dextran sulfate and largest with octodecylamine . The destabilizing effect of protein-protein interactions, as monitored on 50-layer-thick films, is most evident at a relative humidity of 75% . Stabilizing agents were incorporated to attenuate the deleterious effects of protein aggregation . Among them, the most effective in preserving a more native-like structure are the disaccharides sucrose and trehalose in dry films and the polymer dextran in wet films . Interestingly, the polymer was found to achieve maximum efficacy at sensibly lower additive/protein ratios than the sugars. Clin Infect Dis, 2001 May 15, 32 Suppl 2, S146 - 55 Characterization of Pseudomonas aeruginosa isolates: occurrence rates, antimicrobial susceptibility patterns, and molecular typing in the global SENTRY Antimicrobial Surveillance Program, 1997-1999; Gales AC et al.; During 1997-1999, a total of 70,067 isolates (6631 Pseudomonas aeruginosa isolates) were analyzed in the SENTRY program by geographic region and body site of infection . The respiratory tract was the most common source of P . aeruginosa . P . aeruginosa isolation rates increased during the study interval . Europe was the only region to show a significant decline in beta-lactam and aminoglycoside susceptibility rates . There was a reduction in the rates of susceptibility of Canadian isolates to imipenem and of Latin American isolates to meropenem . A total of 218 multidrug-resistant P . aeruginosa isolates (MDR-PSA; resistant to piperacillin, ceftazidime, imipenem, and gentamicin) were observed; MDR-PSA occurrence rates (percentages of all isolates) ranged from 8.2% (Latin America) to 0.9% (Canada) . No antimicrobial inhibited >50% of MDR-PSA strains . Molecular characterization of selected, generally resistant strains was performed . Isolates showing unique ribogroups were found in Europe, Latin America, and the United States, but clonal spread was documented in several medical centers. Microbiology, 2001 May, 147(Pt 5), 1105 - 13 Pseudomonas aeruginosa mutations in lasI and rhlI quorum sensing systems result in milder chronic lung infection; Wu H et al.; To understand the importance of quorum sensing in chronic Pseudomonas aeruginosa lung infection, the in vivo pathogenic effects of the wild-type P . aeruginosa PAO1 and its double mutant, PAO1 lasI rhlI, in which the signal-generating parts of the quorum sensing systems are defective were compared . The rat model of P . aeruginosa lung infection was used in the present study . The rats were killed on days 3, 7, 14 and 28 after infection with the P . aeruginosa strains . The results showed that during the early stages of infection, the PAO1 double mutant induced a stronger serum antibody response, higher production of pulmonary interferon gamma, and more powerful blood polymorphonuclear leukocyte (PMN) chemiluminescence compared to its wild-type counterpart . On days 14 and 28 post-infection, significantly milder lung pathology, a reduction in the number of mast cells present in the lung foci, a reduced number of lung bacteria, and minor serum IgG and IgG1 responses but increased lung interferon gamma production were detected in the group infected with the PAO1 double mutant when compared with the PAO1-infected group . Delayed immune responses were observed in the PAO1-infected group and they might be associated with the production of virulence factors that are controlled by the quorum sensing systems . The conclusion of this study is that functional lasI and rhlI genes of P . aeruginosa PAO1 play a significant role during lung infection. Anal Biochem, 2001 May 1, 292(1), 26 - 33 Application of a fluorometric assay for characterization of the catalytic competency of a domain III fragment of Pseudomonas aeruginosa exotoxin A; Armstrong S et al.; Pseudomonas aeruginosa exotoxin A (ETA) is a member of the family of bacterial ADP-ribosylating toxins that use NAD(+) as the ADP-ribose donor . The reaction catalyzed by ETA involves the nucleophilic attack of the diphthamide residue on the anomeric carbon of the nicotinamide ribose forming a new glycosidic bond . A fluorometric assay involving the use of etheno-beta-nicotinamide adenine dinucleotide (epsilon-NAD(+)), an analog of NAD(+), has been found to provide a rapid, reliable, and sensitive procedure for assessing the kinetic parameters of this class of enzymes including ETA and its C-terminal fragment, PE24 . Furthermore, application of this new assay facilitated the determination of the kinetic parameters for the protein substrate of ETA, elongation factor, which has previously been difficult to characterize . These findings provide new insights into catalytic mechanism of dipthamide-specific ribosyltransferases . In addition, this assay should also prove valuable for the study of NADases or NAD(+)-glycohydrolase enzymes (B . Weng, W . C . Thompson, H . J . Kim, R . L . Levine, and J . Moss, 1999, J . Biol . Chem . 274, 31797-31803; Y . S . Cho, M . K . Han, O . S . Kwark, M . S . Phoe, Y . S . Cha, N . H . An, and U . H . Kim, 1998, Comp . Physiol . B: Biochem . Mol . Biol . 120, 175-181) and the poly-ADP-ribosyltransferases (A . A . Pieper, A . Verma, J . Zhang, S . H . Snyder, 1999, Trends Pharmacol . Sci . 20, 171-181; M . K . Jacobson and E . L . Jacobson, 1999, Trends Biochem . Sci . 24, 415-417). Clin Microbiol Infect, 2001 Mar, 7(3), 144 - 51 Effect of beta-lactam antibiotics on the in vitro development of resistance in Pseudomonas aeruginosa; Carsenti-Etesse H et al.; OBJECTIVE: To investigate whether stepwise selection of resistance mutations may mirror the continued bacterial exposure to antibiotics that occurs in the clinical setting . METHODS: We examined the in vitro development of resistance to a number of commonly used antibiotics (cefepime, cefpirome, ceftazidime, cefotaxime, piperacillin and imipenem) in Pseudomonas aeruginosa, a significant nosocomial pathogen . Stepwise resistance was assessed by serial passage of colonies located nearest to the inhibition zone on antibiotic-containing gradient plates . RESULTS: The lowest frequencies of spontaneous resistance mutations were found with cefepime and imipenem; these drugs also resulted in the slowest appearance of resistance of spontaneous resistance mutations . In five wild-type P . aeruginosa strains, cefepime-selected isolates required a mean of 30 passages to reach resistance; resistance occurred more rapidly in strains selected with other cephalosporins . P . aeruginosa strains that produced beta-lactamase or non-enzymatic resistance generally developed resistance more rapidly than wild-type strains . For most strains, resistance to all antibiotics except imipenem correlated with increased levels of beta-lactamase activity . Cross-resistance of cephalosporin-selected resistant mutants to other cephalosporins was common . Cephalosporin-resistant strains retained susceptibility to imipenem and ciprofloxacin . CONCLUSIONS: From our in vitro study, we can conclude that the rate of development of resistance of P . aeruginosa is lower with cefepime compared with other cephalosporines. Am J Ther, 2000 Sep, 7(5), 309 - 12 Epidemiology of ciprofloxacin-resistant Pseudomonas aeruginosa in a veterans affairs hospital; Khayr W et al.; This study was performed to identify risk factors for the nosocomial acquisition of ciprofloxacin-resistant Pseudomonas aeruginosa (CRPA) in a Veterans Administration hospital between January 1994, and March 1995 . The study was a retrospective comparison of host factors and in-hospital exposures of patients who acquired nosocomially CRPA and ciprofloxacin-sensitive P . aeruginosa (CSPA) . Participants included 42 adult patients with nosocomial CRPA acquisition and 52 adult patients with nosocomial CSPA acquisition . Before pseudomonal acquisition, antecedent ciprofloxacin receipt (50% compared with 8%; odds ratio {OR}, 12; p = 0.001), the presence of an indwelling airway (36% compared to 17%; OR, 2.6; p = 0.04), and documented antecedent infection (74% compared to 52%; OR, 2.6; p = 0.03) were significantly associated with acquisition of CRPA . On multivariate analysis, antecedent ciprofloxacin receipt (OR, 16.8; p = 0.0001) and presence of an indwelling airway (OR, 10.5; p = 0.009) remain as significant associations . Furthermore, the test of significance confirmed synergy between these two factors . Antecedent ciprofloxacin therapy and indwelling airway act independently and synergistically to promote CRPA acquisition. Eur J Hum Genet, 2001 Apr, 9(4), 273 - 8 An alpha1-antitrypsin enhancer polymorphism is a genetic modifier of pulmonary outcome in cystic fibrosis; Henry MT et al.; Lung disease is the direct cause of death in over 90% of cystic fibrosis (CF) patients . Excess neutrophil elastase is an important determinant of pulmonary disease in CF . alpha1-antitrypsin (AAT), also known as alpha1-proteinase inhibitor (alpha1PI) is a major modulator of elastase activity . We investigated the hypothesis that an enhancer polymorphism in the AAT gene would contribute to pulmonary prognosis in CF . Respiratory function, chest X-ray scores, bacterial colonisation and infective exacerbation were assessed to evaluate pulmonary disease severity in the CF group . Sixteen patients were found to have the 1237A allele, and 108 the more frequent G allele . Contrary to expectation, the patients with the 1237A allele were found to have better indices of pulmonary disease progression than those without, as indicated by less change in X-ray score (1237A: 0.2+/-0.1; 1237G: 1.2+/-0.1; P = 0.002) and fewer infective exacerbations (1237A: 2.8+/-0.6; 1237G: 4.6+/-0.3; P = 0.03) over the preceding 2 years . Also, a higher proportion of the 1237A (25%) than the 1237G (6.5%) were not colonised by Pseudomonas Aeruginosa (P = 0.04) . Prospective monitoring of infections for a further 2 years confirmed a lesser propensity to infection in patients with the 1237A allele . These trends were also observed in a tightly matched sub-set of CF genotypes of similar age and sex, thus confirming that these effects were independent of the CF genotype . These results indicate that this AAT enhancer polymorphism is associated with better pulmonary prognosis in CF . Though the number of CF patients with the polymorphism is small, and these data need to be confirmed in larger studies, they suggest that a cautious approach should perhaps be taken to treatment of CF patients with supplemental AAT. Thorax, 2001 May, 56(5), 379 - 87 Pulmonary infiltrates in non-HIV immunocompromised patients: a diagnostic approach using non-invasive and bronchoscopic procedures; Rano A et al.; BACKGROUND: The development of pulmonary infiltrates is a frequent life threatening complication in immunocompromised patients, requiring early diagnosis and specific treatment . In the present study non-invasive and bronchoscopic diagnostic techniques were applied in patients with different non-HIV immunocompromised conditions to determine the aetiology of the pulmonary infiltrates and to evaluate the impact of these methods on therapeutic decisions and outcome in this population . METHODS: The non-invasive diagnostic methods included serological tests, blood antigen detection, and blood, nasopharyngeal wash (NPW), sputum and tracheobronchial aspirate (TBAS) cultures . Bronchoscopic techniques included fibrobronchial aspirate (FBAS), protected specimen brush (PSB), and bronchoalveolar lavage (BAL) . Two hundred consecutive episodes of pulmonary infiltrates were prospectively evaluated during a 30 month period in 52 solid organ transplant recipients, 53 haematopoietic stem cell transplant (HSCT) recipients, 68 patients with haematological malignancies, and 27 patients requiring chronic treatment with corticosteroids and/or immunosuppressive drugs . RESULTS: An aetiological diagnosis was obtained in 162 (81%) of the 200 patients . The aetiology of the pulmonary infiltrates was infectious in 125 (77%) and non-infectious in 37 (23%); 38 (19%) remained undiagnosed . The main infectious aetiologies were bacterial (48/125, 24%), fungal (33/125, 17%), and viral (20/125, 10%), and the most frequent pathogens were Aspergillus fumigatus (n=29), Staphylococcus aureus (n=17), and Pseudomonas aeruginosa (n=12) . Among the non-infectious aetiologies, pulmonary oedema (16/37, 43%) and diffuse alveolar haemorrhage (10/37, 27%) were the most common causes . Non-invasive techniques led to the diagnosis of pulmonary infiltrates in 41% of the cases in which they were used; specifically, the diagnostic yield of blood cultures was 30/191 (16%); sputum cultures 27/88 (31%); NPW 9/50 (18%); and TBAS 35/55 (65%) . Bronchoscopic techniques led to the diagnosis of pulmonary infiltrates in 59% of the cases in which they were used: FBAS 16/28 (57%), BAL 68/135 (51%), and PSB 30/125 (24%) . The results obtained with the different techniques led to a change in antibiotic treatment in 93 cases (46%) . Although changes in treatment did not have an impact on the overall mortality, patients with pulmonary infiltrates of an infectious aetiology in whom the change was made during the first 7 days had a better outcome (29% mortality) than those in whom treatment was changed later (71% mortality; p=0.001) . CONCLUSIONS: Non-invasive and bronchoscopic procedures are useful techniques for the diagnosis of pulmonary infiltrates in immunocompromised patients . Bronchial aspirates (FBAS and TBAS) and BAL have the highest diagnostic yield and impact on therapeutic decisions. J Microbiol Methods, 2001 Jun, 45(2), 95 - 101 Effects of cell surface damage on surface properties and adhesion of Pseudomonas aeruginosa; Bruinsma GM et al.; Bacterial cell surfaces play a crucial role in their adhesion to surfaces . In the present study, physico-chemical cell surface properties of Pseudomonas aeruginosa, isolated from a case of contact lens associated keratitis, are determined for mid-exponential and early stationary phase cells and for cells after exposure to a lens care solution or after mechanical damage by sonication . Exposure to a lens care solution and mechanical cell surface damage reduced the cell surface hydrophobicity and water contact angles decreased from 129 degrees to 96 degrees and 83 degrees, respectively . Zeta potentials in saline (-9 mV) were hardly affected after mechanical damage, but tri-modal zeta potential distributions, with subpopulation zeta potentials at -11, -28 and -41 mV, were observed after exposure of bacteria to a lens care solution . X-ray photoelectron spectroscopy indicated changes in the amounts of oxygen-, nitrogen- and phosphorus-rich cell surface components . Mid-exponential phase cells had more nitrogen-rich cell surface components than early stationary phase cells, but water contact angles and zeta potentials were not very different . In addition, mid-exponential phase cells adhered better than early stationary phase cells to hydrophobic and hydrophilic substrata in a parallel plate flow chamber . The capacity of P . aeruginosa to adhere was decreased after inflicting cell surface damage . Exposure to a lens care solution yielded a larger reduction in adhesion capacity than sonication, likely because sonication left most of the cells in a viable state, in contrast to exposure to a lens care solution . It is argued that for clinically relevant experiments, it may be preferable to work with surface damaged cells rather than with gently harvested organisms. ILAR J, 1999 Mar, 40(2), 43 - 50 Use of an Animal Model in Studies of Bacterial Corneal Infection; Cowell BA et al.; Despite medical advancements in available therapies, bacterial corneal infection frequently results in vision loss . Contact lens wear is a common predisposing factor for corneal infection; other reported risk factors are dry eye syndrome, blepharitis, trauma, and surgery . Both the immune defense against infection and the pathogenic mechanisms bacteria employ have been studied in vitro . However, there are complex interactions between the pathogen, the immune system, and the corneal tissue in vivo . Animal models allow the researcher to take the results of in vitro assays and validate their role in corneal infection in a living organism . A murine model is frequently used for studies of the pathogenesis of corneal infection caused by Pseudomonas aeruginosa . In this study, a modified scoring system is introduced that was designed to increase the information derived from this infection model . The new system includes evaluation of area, density, and surface characteristics of the ulceration . Results of in vitro experiments had previously indicated that ExsA, a transcriptional regulator of virulence-associated proteins, was important in pathogenesis of corneal infection caused by P . aeruginosa . Here we use the new scoring system to demonstrate in vivo that ExsA is involved. Eur Respir J, 2001 Jan, 17(1), 27 - 35 Inflammation in cystic fibrosis airways: relationship to increased bacterial adherence; Scheid P et al.; It is unclear whether inflammation in the cystic fibrosis (CF) lung relates predominantly to bacterial infection, or occurs as a direct consequence of mutant cystic fibrosis transmembrane conductance regulator (CFTR) protein . Interleukin (IL)-8 secretion from CF and non-CF cell lines, and from CF and non-CF human primary nasal epithelial cells incubated with or without Pseudomonas aeruginosa, was measured . Activation of nuclear factor-kappaB (NF-kappaB) in unstimulated CF and non-CF nasal epithelial cells, cell lines and murine tissues was measured by gel-shift assays . No significant difference in basal IL-8 production or NF-kappaB activation was observed between CF and non-CF primary nasal cells . However, CF cells exhibited a significantly (p<0.01) increased IL-8 secretion following P . aeruginosa stimulation . Equalization of the increased P . aeruginosa adherence observed in CF cells, to non-CF levels, resulted in comparable IL-8 secretion . Further, IL-8 production did not differ with mutations which result in either correctly localized CFTR, or in partial/total mislocalization of this protein . Similar levels of NF-kappaB activation were observed in a number of organs of wildtype and CF mice . Finally, IL-8 secretion and NF-kappaB activity were not consistently increased in CF cell lines . Cos-7 cell transfection with plasmids expressing deltaF508 or G551D mutant CFTR protein resulted in increased activation of a p50-containing NF-kappaB complex, but IL-8 secretion was similar to wild-type cells . The authors conclude that the stimulus produced by Pseudomonas aeruginosa is the predominant inflammatory trigger in their models. Nippon Rinsho, 2001 Apr, 59(4), 701 - 6 {Metallo-beta-lactamase producing bacteria}; Iyobe S; Metallo-beta-lactamases are the carbapenemases belonging to the class B beta-lactamases on a molecular level . Two types of the metallo-beta-lactamase, IMP-1 and VIM-1, which have been detected mainly from Pseudomonas aeruginosa, respectively, in Japan and in Europe are especially important, because of the broad substrate specificity . These enzymes confer resistance to not only the carbapenems but also almost all beta-lactam antibiotics expect for monobactams . Clinical strains producing IMP-1 type enzymes are easily detectable by the PCR method or by the double-disk method using the enzyme inhibitor of mercapto-compound and the substrate antibiotic such as imipenem or ceftazidime . The gene blaIMP is frequently identified as a cassette inserted in the integron structure on the transmissible plasmids, disseminating among various gram-negative bacteria. Shock, 2001 Apr, 15(4), 285 - 90 Evaluation of the safety of recombinant P-selectin glycoprotein ligand-immunoglobulin G fusion protein in experimental models of localized and systemic infection; Opal SM et al.; P-selectin is a major component in the early interaction between platelets, endothelial cells, and inflammatory cells in the initial phases of the innate immune response . The major ligand for P-selectin is P-selectin glycoprotein ligand-1 (PSGL-1) and this ligand is expressed on the surface of monocyte, lymphocyte, and neutrophil membranes . A truncated form of recombinant human P-selectin glycoprotein ligand-1 has been covalently linked to immunoglobulin G (rPSGL-Ig) and this fusion peptide functions as a competitive inhibitor of PSGL-1 . As an inhibitor of neutrophil-endothelial cell adherence, rPSGL-Ig is in early clinical development for the treatment of ischemia reperfusion injury . To determine the potential for deleterious effects from inhibition in P-selectin-mediated neutrophil attachment in the presence of bacterial infection, the effects of therapeutic doses of rPSGL-Ig were tested in three standard laboratory sepsis models . The experimental models included: the murine systemic Listeria monocytogenes infection model, the Pseudomonas aeruginosa bacteremia model in neutropenic rats, and the cecal ligation and puncture (CLP)-induced peritonitis model in rats . Recombinant human PSGL-Ig had no adverse effects on mortality or immune clearance in systemic bacterial infection in any of the three infection models . The PSGL-1 inhibitor did significantly decrease local neutrophil infiltration and bacterial clearance in the peritoneum following CLP, but this did not increase the systemic levels of proinflammatory cytokines, the quantitative levels of bacteremia, or the overall mortality rate following CLP . The results indicate that rPSGL-Ig did not exacerbate infection in these experimental sepsis models. J Chromatogr B Biomed Sci Appl, 2001 Mar 25, 753(1), 151 - 6 Quantification of gentamicin in Mueller-Hinton agar by high-performance liquid chromatography; Arcelloni C et al.; The aim of this study was to optimise a method for gentamicin determination in an agar matrix and to investigate if and how agar composition can affect the gentamicin diffusion kinetics during the agar diffusion tests for antibiotics sensitivity . Gentamicin was separated by RP-HPLC and detected at 365 nm after pre-column derivatization with 1-fluoro-2,4-dinitrobenzene . Recovery (> or = 79%), linearity (r2 > or = 0.997) and sensitivity (1 microg/ml) were assessed using four different agar matrices . The kinetics of gentamicin diffusion tested on BioMerieux and DID manufacturers' products showed in uninoculated agar plates significant differences that were even more pronounced in the presence of Pseudomonas aeruginosa metabolism. Drugs Aging, 2001, 18(3), 189 - 200 Antibacterial treatment of invasive mechanical ventilation-associated pneumonia; Barcenilla F et al.; Patients admitted to intensive care units (ICU) are at higher risk of acquiring nosocomial infections than patients in other hospital areas . This is the consequence of both a greater severity of illness with its implications (manipulation, invasiveness) and crossed infection from reservoirs inside the ICU . The most frequent nosocomial infection is invasive ventilation-associated pneumonia (VAP) which leads to an important increase in morbidity and mortality . The most important aetiological agents in VAP are bacteria, with a marked predominance of Staphylococcus aureus and Pseudomonas aeruginosa . These aetiologies may be different depending upon the type of ICU (medical, surgical, coronary) or the presence of certain risk factors (duration of mechanical ventilation before onset of pneumonia, previous exposure to antibacterials) . Susceptibilities of the aetiological agents to antibacterials may also vary according to the type of ICU and over time . Data from global studies show an increase in multiresistant bacteria but these data may not be applied to a local ICU . The availability of accurate and updated information on the most frequently encountered organisms in each ICU and their susceptibilities is very important in order to provide the most adequate treatment . A controversial issue is the selection of antibacterials . According to the latest evidence the most adequate approach is a prompt administration of empirical treatment . Based on knowledge of bacterial flora in our own ICU, the choice of an adequate therapeutic regimen will decrease both morbidity and mortality . A second issue is monotherapy versus combined therapy . The most common recommendation, with a few exceptions, is to use combined therapy until microbiological results are received . Another controversy is the choice of antibacterials in the combined regimen . The most commonly recommended combination is that of a beta-lactam with an aminoglycoside, except in early-onset pneumonia without risk factors . The use of monotherapy with a cefalosporin without antipseudomonal activity or amoxicillin-clavulanic acid is the recommended regimen . Treatment should be modified based on microbiological results . There are no well documented recommendations on the prophylactic duration of treatment and it must be based on the aetiological agent and the clinical course . In summary treatment of VAP must be prompt, empirical and combined (beta-lactam plus aminoglycoside ) . However, the choice of the antibacterial regimen should follow local guidelines of treatment based upon the knowledge of the most frequently isolated bacterial flora and their susceptibilities in different clinical settings. Biochemistry, 2001 Apr 3, 40(13), 3985 - 95 Backbone dynamics of receptor binding and antigenic regions of a Pseudomonas aeruginosa pilin monomer; Suh JY et al.; Pilin is the major structural protein that forms type IV pili of various pathogenic bacteria, including Pseudomonas aeruginosa . Pilin is involved in attachment of the bacterium to host cells during infection, in the initiation of immune response, and serves as a receptor for a variety of bacteriophage . We have used (15)N nuclear magnetic resonance relaxation measurements to probe the backbone dynamics of an N-terminally truncated monomeric pilin from P . aeruginosa strain K122-4 . (15)N-T(1), -T(2), and {(1)H}-(15)N nuclear Overhauser enhancement measurements were carried out at three magnetic field strengths . The measurements were interpreted using the Lipari-Szabo model-free analysis, which reveals the amplitude of spatial restriction for backbone N-NH bond vectors with respect to nano- to picosecond time-scale motions . Regions of well-defined secondary structure exhibited consistently low-amplitude spatial fluctuations, while the terminal and loop regions showed larger amplitude motions in the subnano- to picosecond time-scale . Interestingly, the C-terminal disulfide loop region that contains the receptor binding domain was found to be relatively rigid on the pico- to nanosecond time-scale but exhibited motion in the micro- to millisecond time-scale . It is notable that this disulfide loop displays a conserved antigenic epitope and mediates binding to the asialo-GM(1) cell surface receptor . The present study suggests that a rigid backbone scaffold mediates attachment to the host cell receptor, and also maintains the conformation of the conserved antigenic epitope for antibody recognition . In addition, slower millisecond time-scale motions are likely to be crucial for conferring a range of specificity for these interactions . Characterization of pilin dynamics will aid in developing a detailed understanding of infection, and will facilitate the design of more efficient anti-adhesin synthetic vaccines and therapeutics against pathogenic bacteria containing type IV pili. Plant Physiol, 2001 Apr, 125(4), 2059 - 67 Expression of a Pseudomonas aeruginosa citrate synthase gene in tobacco is not associated with either enhanced citrate accumulation or efflux; Delhaize E et al.; Aluminum (Al) toxicity and poor phosphorus (P) availability are factors that limit plant growth on many agricultural soils . Previous work reported that expression of a Pseudomonas aeruginosa citrate synthase gene in tobacco (Nicotiana tabacum; CSb lines) resulted in improved Al tolerance (J.M . de la Fuente, V . Ramirez-Rodriguez, J.L . Cabrera-Ponce, L . Herrera-Estrella {1997} Science 276: 1566-1568) and an enhanced ability to acquire P from alkaline soils (J . Lopez-Bucio, O . Martinez de la Vega, A . Guevara-Garcia, L . Herrera-Estrella {2000} Nat Biotechnol 18: 450-453) . These effects were attributed to the P . aeruginosa citrate synthase increasing the biosynthesis and efflux of citrate from roots . To verify these findings we: (a) characterized citrate efflux from roots of wild-type tobacco; (b) generated tobacco lines expressing the citrate synthase gene from P . aeruginosa; and (c) analyzed selected CSb lines described above . Al stimulated citrate efflux from intact roots of wild-type tobacco and root apices were found to be responsible for most of the efflux . Despite generating transgenic tobacco lines that expressed the citrate synthase protein at up to a 100-fold greater level than the previously described CSb lines, these lines did not show increased accumulation of citrate in roots or increased Al-activated efflux of citrate from roots . Selected CSb lines, similarly, failed to show differences compared with controls in either citrate accumulation or efflux . We conclude that expression of the P . aeruginosa citrate synthase gene in plants is unlikely to be a robust and easily reproducible strategy for enhancing the Al tolerance and P-nutrition of crop and pasture species. Lett Appl Microbiol, 2001 Apr, 32(4), 257 - 61 Intracellular nickel accumulation by Pseudomonas aeruginosa and its chemical nature; Sar P et al.; AIMS: To investigate intracellular localization of nickel and its chemical nature in Pseudomonas aeruginosa . METHODS AND RESULTS: Transmission electron micrographs of Ni-loaded bacteria exhibited a darkened electron opaque zone throughout the cell periphery . Energy dispersive X-ray analysis confirmed the deposition of metallic nickel . Cell fractionation revealed that 88% of the accumulated nickel was restricted to the periplasm and membrane . X-ray diffraction patterns ascertained the chemical nature of cellular Ni as phosphide (Ni5P4, NiP2 and Ni12P5) and carbide (Ni3C) crystals . CONCLUSION: Pseudomonas aeruginosa accumulated nickel as its phosphide and carbide crystal mostly in the cell envelope region, indicating the predominant role of phosphoryl and carboxyl/carbonyl groups of cell wall/membrane components in cation sequestration . SIGNIFICANCE AND IMPACT OF THE STUDY: The data contribute significantly to a better understanding of bacteria-metal interaction and will be useful in developing biotechnological strategies for toxic metal bioremediation. Cell Microbiol, 2001 Apr, 3(4), 223 - 36 Pseudomonas aeruginosa ExoT inhibits in vitro lung epithelial wound repair; Geiser TK et al.; The nosocomial pathogen Pseudomonas aeruginosa causes clinical infection in the setting of pre-existing epithelial tissue damage, an association that is mirrored by the increased ability of P . aeruginosa to bind, invade and damage injured epithelial cells in vitro . In this study, we report that P . aeruginosa inhibits the process of epithelial wound repair in vitro through the type III-secreted bacterial protein ExoT, a GTPase-activating protein (GAP) for Rho family GTPases . This inhibition primarily targets cells at the edge of the wound, and causes actin cytoskeleton collapse, cell rounding and cell detachment . ExoT-dependent inhibition of wound repair is mediated through the GAP activity of this bacterial protein, as mutations in ExoT that alter the conserved arginine (R149) within the GAP domain abolish the ability of P . aeruginosa to inhibit wound closure . Because ExoT can also inhibit P . aeruginosa internalization by phagocytes and epithelial cells, this protein may contribute to the in vivo virulence of P . aeruginosa by allowing organisms both to overcome local host defences, such as an intact epithelial barrier, and to evade phagocytosis by immune effector cells. Mol Microbiol, 2001 Apr, 40(1), 76 - 85 Pore-forming activity of type III system-secreted proteins leads to oncosis of Pseudomonas aeruginosa-infected macrophages; Dacheux D et al.; The Pseudomonas aeruginosa cystic fibrosis isolate CHA induces type III secretion system-dependent but ExoU-independent oncosis of neutrophils and macrophages . Time-lapse microscopy of the infection process revealed the rapid accumulation of motile bacteria around infected cells undergoing the process of oncosis, a phenomenon we termed pack swarming . Characterization of the non-chemotactic CHAcheZ mutant showed that pack swarming is a bacterial chemotactic response to infected macrophages . A non-cytotoxic mutant, lacking the type III-secreted proteins PcrV, PopB and PopD, was able to pack swarm only in the presence of the parental strain CHA or when macrophages were pretreated with the pore-forming toxin streptolysin O . Interaction of P . aeruginosa with red blood cells (RBCs) showed that the contact-dependent haemolysis provoked by CHA requires secretion via the type III system and the PcrV, PopB/PopD proteins . The pore inserted into RBC membrane was estimated from osmoprotection experiments to be between 2.8 and 3.5 nm . CHA-infected macrophages could be protected from cell lysis with PEG3350, indicating that the pore introduced into RBC and macrophage membranes is of similar size . The time course uptake of the vital fluorescent dye, Yo-Pro-1, into infected macrophages confirmed that the formation of transmembrane pores by CHA precedes cellular oncosis . Therefore, CHA-induced macrophage death results from a pore-forming activity that is dependent on the intact pcrGVHpopBD operon. J Biol Chem, 2001 Jun 29, 276(26), 24186 - 93 Epub 2001 Apr 09. Structure of a pilin monomer from Pseudomonas aeruginosa: implications for the assembly of pili; Keizer DW et al.; Type IV pilin monomers assemble to form fibers called pili that are required for a variety of bacterial functions . Pilin monomers oligomerize due to the interaction of part of their hydrophobic N-terminal alpha-helix . Engineering of a truncated pilin from Pseudomonas aeruginosa strain K122-4, where the first 28 residues are removed from the N terminus, yields a soluble, monomeric protein . This truncated pilin is shown to bind to its receptor and to decrease morbidity and mortality in mice upon administration 15 min before challenge with a heterologous strain of Pseudomonas . The structure of this truncated pilin reveals an alpha-helix at the N terminus that lies across a 4-stranded antiparallel beta-sheet . A model for a pilus is proposed that takes into account both electrostatic and hydrophobic interactions of pilin subunits as well as previously published x-ray fiber diffraction data . Our model indicates that DNA or RNA cannot pass through the center of the pilus, however, the possibility exists for small organic molecules to pass through indicating a potential mechanism for signal transduction. J Biol Inorg Chem, 2001 Feb, 6(2), 182 - 8 Probing copper ligands in denatured Pseudomonas aeruginosa azurin: unfolding His117Gly and His46Gly mutants; Pozdnyakova I et al.; Azurin is a single-domain beta-barrel protein with a redox-active copper cofactor . Upon Pseudomonas aeruginosa azurin unfolding, the cofactor remains bound to the polypeptide, coordinating three ligands: cysteine-112, one histidine imidazole, and a third, unknown ligand . In order to identify which histidine (histidine-117 and histidine-46 both coordinate copper in native azurin) is involved in copper coordination in denatured azurin, two single-site (histidine to glycine) mutants, His117Gly and His46Gly azurin, are investigated here . Equilibrium denaturation experiments of His46Gly azurin loaded with copper demonstrate that copper remains bound to this mutant in high urea concentrations where the protein's secondary structure is lost . In contrast, for copper-loaded His117Gly azurin, copper does not stay coordinated upon polypeptide unfolding . The copper absorption at 370 nm in denatured His46Gly azurin agrees with that for copper in complex with a peptide corresponding to residues 111-123 in azurin, suggesting similar metal coordination . We conclude that histidine-117 (and not histidine-46) is the histidine copper ligand in denatured azurin . This is also in accord with the proximity of histidine-117 to cysteine-112 in the primary sequence. Infect Immun, 2001 May, 69(5), 3510 - 5 Protection against Pseudomonas aeruginosa chronic lung infection in mice by genetic immunization against outer membrane protein F (OprF) of P . aeruginosa; Price BM et al.; The Pseudomonas aeruginosa major constitutive outer membrane porin protein OprF, which has previously been shown to be a protective antigen, was targeted as a DNA vaccine candidate . The oprF gene was cloned into plasmid vector pVR1020, and the plasmid vaccines were delivered to mice by biolistic (gene gun) intradermal inoculation . Antibody titers in antisera from immunized mice were determined by enzyme-linked immunosorbent assay, and the elicited antibodies were shown to be specifically reactive to OprF by immunoblotting . The immunoglobulin G (IgG) immune response was predominantly of the IgG1 isotype . Sera from DNA vaccine-immunized mice had significantly greater opsonic activity in opsonophagocytic assays than did sera from control mice . Following the initial immunization and two consecutive boosts, each at 2-week intervals, protection was demonstrated in a mouse model of chronic pulmonary infection by P . aeruginosa . Eight days postchallenge, both lungs were removed and examined . A significant reduction in the presence of severe macroscopic lesions, as well as in the number of bacteria present in the lungs, was seen . Based on these findings, genetic immunization with oprF has potential for development as a vaccine to protect humans against infection by P . aeruginosa. Infect Immun, 2001 May, 69(5), 3295 - 304 Effector mechanisms of protection against Pseudomonas aeruginosa keratitis in immunized rats; Thakur A et al.; Pseudomonas aeruginosa is an opportunistic pathogen which causes sight-threatening corneal infections in humans . The purpose of this study was to evaluate various immunization routes that may provide protection against Pseudomonas keratitis and to define the molecular mechanisms involved in the protection . Sprague-Dawley rats (10 to 12 weeks old) were immunized using paraformaldehyde-killed P . aeruginosa (strain 6206) via oral, nasal, and intra-Peyer's patch (IPP) routes followed by an ocular topical booster dose . Scratched corneas were challenged with an infective dose of P . aeruginosa . Following clinical examination, eyes were enucleated for histology, polymorphonuclear leukocyte (PMN) quantitation, bacterial count, enzyme-linked immunosorbent assay, and RNase protection assay . PMN infiltration was higher early (4 h) during the infection in immunized rats than in nonimmunized rats . Later during the infection, the number of PMNs diminished in immunized rats while in nonimmunized animals the number of PMNs continued to increase . Bacteria were cleared much faster from immunized groups than from the nonimmunized group, and the nasally immunized group had the most efficacious response among the immunized groups . Nasal and IPP immunization groups had increased cytokine expression of interleukin-2 (IL-2) and IL-5 and differed from each other for IL-6 . All three immunized groups had significantly reduced IL-1 beta levels when compared with the nonimmunized rats and a significantly altered profile for CINC-1 expression . This study has shown that the route of immunization modulates the inflammatory response to ocular P . aeruginosa infection, thus affecting the severity of keratitis and adverse pathology, with nasal immunization being the most effective. J Hosp Infect, 2001 Apr, 47(4), 325 - 7 Heterogeneity among infecting strains of Pseudomonas aeruginosa in diverse departments of a large Tunisian hospital; Ben Slama K et al.; Clinical isolates of Pseudomonas aeruginosa were obtained during a half-year screening period of five different wards of the La Rabta Hospital (Tunis) . Distinct clinical isolates (N= 82) were obtained from patients, 40 (48%) of which originated from the Department of Otolaryngology . In order to define the local epidemiology of this opportunistic organism, all strains were serotyped, analysed for pyocin production and genetically characterized with the help of pulsed-field gel electrophoresis (PFGE) . The data show that, despite the frequent occurrence of identical serotypes, most of the isolates represent unique pyocin types (N= 53) and genotypes (N= 64) . A combination of the pyocin and PFGE data showed that nearly all strains were of unique types, except for two pairs of strains . A limited number of strain clusters was observed on the basis of DNA typing data alone . This involved eight genotypes, some of which were clustered with respect to clinical environment or time . Genotype 22 occurred most frequently (6/83, 7%) and independently of time and locale, indicating that it may represent either a clonal type constituting a major fraction of all P . aeruginosa isolates in the region or a more prevalent organism . Despite a relatively high incidence of P . aeruginosa infections, the polyclonality of these strains shows that, in La Rabta Hospital, pseudomonal infections are not primarily due to excessive spread of a single bacterial genotype . J Hosp Infect, 2001 Apr, 47(4), 321 - 4 Ventilator-associated pneumonias in a cardiothoracic surgery centre postoperative intensive care unit; Simsek S et al.; Cases of ventilator-associated pneumonia (VAP) were investigated in a cardiothoracic surgery postoperative intensive care unit between 1 January 1999 and 31 December 1999 . A total of 1716 patients who had undergone cardiothoracic operations and admitted to the intensive care unit (ICU) were included in the study . Patient- and laboratory-based prospective surveillance of VAP was done along with other hospital-acquired infections . During the study period a total of 26 585 patient-days with 2708 ventilator-days were recorded . Forty-six cases of VAP occurred in 36 of 1716 patients who had undergone cardiothoracic operations (2.09%, 1.3 episodes of pneumonia per patient) . The ventilator utilization rate at our institution was 0.10 . There were 16.4 VAPs per 1000 ventilation days . Thirty-eight percent of VAP were caused by Gram-negative enteric rods, 34% by Pseudomonas aeruginosa, and 17% by Staphylococcus aureus . VAP was polymicrobial in 9% of cases . No causative micro-organism was identified in 2% of cases . The same bacteria were isolated in both blood and endotracheal aspirate cultures in 10 of 46 pneumonia cases (22%) . The crude mortality rate of VAP was calculated as 30% Ceska Slov Farm, 2001 Mar, 50(2), 83 - 5 {Bergenia crassifolia (L.) Fritsch in vitro}; Duskova J et al.; Sterile germinating plants were used to derive a tissue culture . The greatest stimulating effect on the growth of the culture was exerted by NAA cultures in all concentrations tested, by IAA in concentrations of 1.0 and 10.0 mg.l-1, by IBA in a concentration of 0.1 mg.l-1, and by the combination of IBA + K . The difference between the values of growth on these media was statistically insignificant . TLC analysis of callus extracts demonstrated the presence of bergenin, arbutin, hydroquinone, and methylarbutine . HPLC analysis confirmed the findings (arbutin 0.25%, hydroquinone 0.05%, methylarbutin 0.28%) . The results of biotransformation tests show that the highest increase in arbutin took place after addition of 4-hydroxybenzoic acid and withdrawal of the sample after 24 hours (3.82%) . No transformation of arbutin or 4-methoxyphenol to produce methylarbutin took place in the culture under the given conditions . The highest increase in the summary content of phenolic substances occurred with the use of the elicitor Pseudomonas aeruginosa--88% (conc . 0.0001 g/100 ml, 12-hour action). Am J Infect Control, 2001 Apr, 29(2), 79 - 84 Incidence and determinants of Pseudomonas aeruginosa infection among persons with HIV: association with hospital exposure; Sorvillo F et al.; BACKGROUND: Little information exists on risk factors for Pseudomonas aeruginosa infection in persons with HIV . We assessed the incidence and factors associated with P aeruginosa among persons with HIV enrolled in a large observational cohort study in Los Angeles . METHODS: Data were analyzed from 4825 persons aged > or =13 years with HIV infection enrolled from 4 outpatient facilities from 1990 to 1998 . The association between P aeruginosa infection and demographic, risk behavior, and clinical factors was assessed . RESULTS: P aeruginosa was diagnosed in 72 (1.5%) patients representing a crude incidence rate of 0.74 per 100 person-years . The most frequent site of infection was pulmonary (47%) . In multivariate analysis, prior hospitalization (adjusted rate ratio = 7.9, 95% CI, 3.8-16.2), and both dapsone (adjusted rate ratio = 4.0, 95% CI, 2.2-7.4) and trimethoprim-sulfamethoxazole (adjusted rate ratio = 2.5, 95% CI, 1.2-5.3) use were independently associated with higher rates of infection . Increasing days of inpatient stay (P <.01) and decreasing CD4(+) counts (P <.01) were strongly associated with P aeruginosa . Azithromycin use decreased the risk of infection by nearly 70% . CONCLUSION: Although the overall observed incidence of P aeruginosa was low, hospital exposure, declining CD4(+) levels, and the use of dapsone or trimethoprim-sulfamethoxazole increased the risk of P aeruginosa disease, and azithromycin use was protective in this population . These findings may assist in the early recognition and diagnosis of persons likely to be at increased risk of P aeruginosa infection. Proc Natl Acad Sci U S A, 2001 Apr 10, 98(8), 4426 - 30 Epub 2001 Apr 03. Deuterium isotope effect on the intramolecular electron transfer in Pseudomonas aeruginosa azurin; Farver O et al.; Intramolecular electron transfer in azurin in water and deuterium oxide has been studied over a broad temperature range . The kinetic deuterium isotope effect, k(H)/k(D), is smaller than unity (0.7 at 298 K), primarily caused by the different activation entropies in water (-56.5 J K(-1) mol(-1)) and in deuterium oxide (-35.7 J K(-1) mol(-1)) . This difference suggests a role for distinct protein solvation in the two media, which is supported by the results of voltammetric measurements: the reduction potential (E(0')) of Cu(2+/+) at 298 K is 10 mV more positive in D(2)O than in H(2)O . The temperature dependence of E(0') is also different, yielding entropy changes of -57 J K(-1) mol(-1) in water and -84 J K(-1) mol(-1) in deuterium oxide . The driving force difference of 10 mV is in keeping with the kinetic isotope effect, but the contribution to DeltaS from the temperature dependence of E(0') is positive rather than negative . Isotope effects are, however, also inherent in the nuclear reorganization Gibbs free energy and in the tunneling factor for the electron transfer process . A slightly larger thermal protein expansion in H(2)O than in D(2)O (0.001 nm K(-1)) is sufficient both to account for the activation entropy difference and to compensate for the different temperature dependencies of E(0') . Thus, differences in driving force and thermal expansion appear as the most straightforward rationale for the observed isotope effect. Ann Trop Paediatr, 2001 Mar, 21(1), 20 - 5 Neonatal sepsis and mortality in a regional hospital in Trinidad: aetiology and risk factors; Orrett FA et al.; A total of 132 neonatal deaths among 627 infants admitted to the neonatal ward of the San Fernando General Hospital, Trinidad over a 2-year period were reviewed . The most common cause of death was prematurity (43.9%) . Infection was the second most common cause (21.2%) . Pseudomonas aeruginosa and Staphylococcus aureus were the most frequently isolated organisms (43%) . The major drugs used empirically in suspected cases of sepsis were ampicillin or ceftazidime plus gentamicin . About 85% of S . aureus were resistant to ampicillin, and P . aeruginosa resistance to ceftazidime and gentamicin was 76.7% and 72.1%