Clin Infect Dis, 2004 Aug 15, 39(4), 497 - 503 Epub 2004 Aug 02. Predicting hospital rates of fluoroquinolone-resistant Pseudomonas aeruginosa from fluoroquinolone use in US hospitals and their surrounding communities; Polk RE et al.; Rates of fluoroquinolone resistance among Pseudomonas aeruginosa in hospitals are increasing, but interhospital variability is great . We sought to determine whether this variability correlated to fluoroquinolone use in hospitals and in the surrounding community . Hospital quinolone use in 1999 (24 hospitals) through 2001 (35 hospitals) was determined from billing records . The number of fluoroquinolone prescriptions within a 10-mile (approximately 16-km) radius of each hospital was determined for 1999 and 2000 . Hospital fluoroquinolone use increased from 1999 through 2001, from 137 to 163 defined daily doses (DDD)/1000 patient-days (P=.01) . The rate of community fluoroquinolone use also increased, from 2.3 to 2.8 DDD/1000 inhabitant-days (P<.001) . Rates of fluoroquinolone-resistant P . aeruginosa increased from 29% in 1999 to 36% in 2001 (P=.003) . Both community and hospital fluoroquinolone use were predictive of rates of fluoroquinolone-resistant P . aeruginosa . Levofloxacin was associated with resistance, but ciprofloxacin was not . Most of the variability in resistance rates is explained by volume of fluoroquinolone use, both in the hospital and the surrounding community. J Antimicrob Chemother, 2004 Oct, 54(4), 772 - 9 Epub 2004 Sep 08. Pseudomonas aeruginosa-induced infection and degradation of human wound fluid and skin proteins ex vivo are eradicated by a synthetic cationic polymer; Werthen M et al.; OBJECTIVES: Antimicrobial peptides are important effectors of innate immunity . Bacteria display multiple defence mechanisms against these peptides . For example, Pseudomonas aeruginosa releases potent proteinases that inactivate the human cathelicidin LL-37 . Hence, in conditions characterized by persistent bacterial colonization, such as in P . aeruginosa-infected skin wounds, there is a need for efficient means of reducing bacterial load . Here, the effect of the cationic molecule polyhexamethylenebiguanide (PHMB) was evaluated . METHODS: Infection models in human wound fluid and human skin were established . Radial diffusion methods, bacterial growth and bactericidal assays were used for determination of effects of PHMB on bacteria in the presence of plasma, wound fluid or human skin . At the protein and tissue levels, SDS-PAGE, light microscopy and scanning electron microscopy were used to study the effects of P . aeruginosa infection before and after addition of PHMB . RESULTS: PHMB killed common ulcer-derived bacteria in the presence of human wound fluid . Furthermore, elastase-expressing P . aeruginosa completely degraded wound fluid proteins as well as human skin during infection ex vivo . The infection, and consequent protein degradation, was reversed by PHMB . CONCLUSIONS: The ex vivo infection models presented here should be helpful in the screening of novel antimicrobials and constitute a prerequisite for future clinical studies. Zhongguo Wei Zhong Bing Ji Jiu Yi Xue, 2004 Sep, 16(9), 540 - 3 {Characteristics and therapeutic strategies of nosocomial pneumonia in postoperative patients with acute hypertensive intracerebral hemorrhage}; Tong Y et al.; Objective: To investigate the characteristics of postoperative nosocomial pneumonia in patients with acute hypertensive intracerebral hemorrhage after evacuation of hematoma and the strategies of prevention and treatment . Methods: Retrospective analysis was performed on the clinical data of surgically treated patients with acute hypertensive intracerebral hemorrhage in the Department of Neurosurgery from 2000 to 2003 . The patients were divided into a group who developed nosocomial pneumonia to compare with a group who did not . Sputum was cultured B and drug sensitivity of isolated bacteria was performed . Appropriate treatment measures were given according to clinical manifestations . Results: In 112 post operative patients with acute hypertensive intracerebral hemorrhage, nosocomial pneumonia developed in 29 patients (25.89%), and 65.99% of microorganisms isolated from respiratory secretion was Gram-negative bacteria . The predominant bacteria were Pseudomonas aeruginosa and Staphylococcus aureus . The mortality was 34.48% (10/29 cases) . The prognosis of patients with nosocomial pneumonia was worse than the controls (7.23%, 6/83 cases, P<0.01) . Conclusion: The preventive and therapeutic strategies for nosocomial pneumonia in postoperative patients with acute hypertensive intracerebral hemorrhage include reduction of risk-factors, surveillance of pathogens, rational use of antibiotics, respiratory and nutrition support, and so on. Biomaterials, 2005 Mar, 26(8), 917 - 24 Nitric oxide-releasing sol-gels as antibacterial coatings for orthopedic implants; Nablo BJ et al.; To assess the benefits of nitric oxide (NO)-releasing sol-gels as potential antibacterial coatings for orthopedic devices, medical-grade stainless steel is coated with a sol-gel film of 40% N-aminohexyl-N-aminopropyltrimethoxysilane and 60% isobutyltrimethoxysilane . Upon converting the diamine groups in these films to diazeniumdiolate NO donors, the NO release from the sol-gel-coated stainless steel is evaluated at both ambient and physiological temperature . Sol-gel films incubated at 25 degrees C have a lower NO flux over the first 24 h compared to those at 37 degrees C, but release more than five times longer . The bacterial adhesion resistance of NO-releasing coatings is evaluated in vitro by exposing bare steel, sol-gel, and NO-releasing sol-gel-coated steel to cell suspensions of Pseudomonas aeruginosa, Staphylococcus aureus, and Staphylococcus epidermidis at 25 degrees C and 37 degrees C . Cell adhesion to bare and sol-gel-coated steel is similar, while NO-releasing surfaces have significantly less bacterial adhesion for all species and temperatures investigated. Arch Pediatr, 2004 Sep, 11(9), 1070 - 2 {Acute bartholinitis caused by Pseudomonas aeruginosa in an 18-month-old infant}; Touzot F et al.; Acute bartholinitis is a disease usually seen in women in the period of genital activity . Its occurence in a prepubertal child is an extremely rare event . CASE REPORT: We describe the case of a 18-month-old infant presenting a Bartholin's gland abces caused by Pseudomonas Aeruginosa with a resolutive evolution after antibiotherapy and surgical drainage . CONCLUSION: Diagnosis of Bartholinitis should be considered in any female infant with a labial enlargement. Biochemistry, 2004 Sep 14, 43(36), 11427 - 35 Crystallographic analysis of the Pseudomonas aeruginosa strain K122-4 monomeric pilin reveals a conserved receptor-binding architecture; Audette GF et al.; Adherence of pathogens to host cells is critical for the initiation of infection and is thus an attractive target for anti-infective therapeutics and vaccines . In the opportunistic human pathogen Pseudomonas aeruginosa, host-cell adherence is achieved predominantly by type IV pili . Analysis of several clinical strains of P . aeruginosa reveals poor sequence conservation between pilin genes, including the residues in the receptor-binding site . Interestingly, the receptor-binding sites appear to retain a conserved surface epitope because all Pseudomonas type IV pili recognize the same receptor on the host cell and cross-reactive antibodies specific for the receptor-binding site exist . Here, we present the crystallographic analysis of two crystal forms of truncated pilin from P . aeruginosa strain K122-4 (DeltaK122-4) at 1.54 and 1.8 A resolution, respectively . The DeltaK122-4 structure is compared to other crystallographically determined type IV pilin structures and an NMR structure of DeltaK122-4 pilin . A comparison with the structure of the highly divergent P . aeruginosa strain K (DeltaPAK) pilin indicates that the receptor-binding loop in both pilins forms a shallow depression with a surface that is formed by main-chain atoms . Conservation of this putative binding site is independent of the sequence as long as the main-chain conformation is conserved and could therefore explain the shared receptor specificity and antibody cross reactivity of highly divergent Pseudomonas type IV pilins. J Mater Sci Mater Med, 1998, 9(1), 17 - 22 The influence of substratum topography on bacterial adhesion to polymethyl methacrylate; Taylor RL et al.; The effect of substratum roughness on the adhesion of Pseudomonas aeruginosa and Staphylococcus epidermidis was investigated using PMMA . A small increase in Ra values (0.04-1.24 microm) resulted in a significant increase (P<0.05) in bacterial attachment . Subsequent increases in surface roughness (Ra=1.86-7.89 microm) resulted in a decrease in adhesion, although adhesion was still higher than to the smooth surface . When the PMMA surfaces were coated with protein (bovine serum albumin), no difference (P<0.05) could be determined in the amount of protein adsorbed, irrespective of surface topography . However, the influence of the underlying topography on adhesion was still evident . Substratum topography is an important parameter affecting bacterial adhesion to surfaces. J Mater Sci Mater Med, 2000 Dec, 11(12), 817 - 823 beta-Chitin-based wound dressing containing silver sulfurdiazine; Lee YM et al.; Physical and biological properties of some wound dressing materials based on beta-chitin were studied . Water vapor transmission rates (WVTR), oxygen permeabilities and biodegradation kinetics were examined for film-type samples . WVTR of samples was in the range 2400-2800 g/m^2/day . However, oxygen permeabilities of the samples were relatively low . To improve oxygen permeabilities, porous sponge-type wound dressing materials were prepared . In addition, these sponge-type samples contained antimicrobial agents, silver sulfadiazine (AgSD), in order to prevent bacteria infection on a wound surface . Anti-microbacterial tests on agar plate were carried out to confirm the bactericidal capacity of present materials . These materials impregnating AgSD had the complete bactericidal capacity against pseudomonas aeruginosa up to 7 days . Finally, a wound healing effect of beta-chitin-based semi-interpenetrating polymer networks was evaluated from the animal test using the wistar rat in vivo . Histological studies confirm the proliferation of fibroblasts in the wound bed and a distinct reduction in infectious cells . J Mater Sci Mater Med, 1999 Dec, 10(12), 853 - 5 Initial adhesion and surface growth of Pseudomonas aeruginosa on negatively and positively charged poly(methacrylates); Gottenbos B et al.; The infection risk of biomaterial implants is determined by an interplay of bacterial adhesion and surface growth of the adhering organisms . In this study, we compared initial adhesion and surface growth of Pseudomonas aeruginosa AK1 (zeta potential -7 mV) on negatively charged (PMMA/MAA, zeta potential -18 mV) and positively charged (PMMA/TMAEMA-Cl, zeta-potential +12 mV) methacrylate copolymers in situ in a parallel plate flow chamber . Initial adhesion was measured using phosphate-buffered saline and subsequent surface growth of the adhering bacteria using nutrient broth as growth medium . Initial adhesion was twice as fast on the positively charged methacrylate than on the negatively charged copolymer . Surface growth, however, was absent on the positively charged copolymer, while on the negatively charged methacrylate the number of bacteria increased exponentially during surface growth with a generation time of 32 min . From the results of this study it can be concluded that positively charged biomaterial surfaces might show reduced risks of biomaterials-centred infections, despite being more adhesive . Appl Environ Microbiol, 2004 Sep, 70(9), 5357 - 65 Localization and characterization of two novel genes encoding stereospecific dioxygenases catalyzing 2(2,4-dichlorophenoxy)propionate cleavage in Delftia acidovorans MC1; Schleinitz KM et al.; Two novel genes, rdpA and sdpA, encoding the enantiospecific alpha-ketoglutarate dependent dioxygenases catalyzing R,S-dichlorprop cleavage in Delftia acidovorans MC1 were identified . Significant similarities to other known genes were not detected, but their deduced amino acid sequences were similar to those of other alpha-ketoglutarate dioxygenases . RdpA showed 35% identity with TauD of Pseudomonas aeruginosa, and SdpA showed 37% identity with TfdA of Ralstonia eutropha JMP134 . The functionally important amino acid sequence motif HX(D/E)X(23-26)(T/S)X(114-183)HX(10-13)R/K, which is highly conserved in group II alpha-ketoglutarate-dependent dioxygenases, was present in both dichlorprop-cleaving enzymes . Transposon mutagenesis of rdpA inactivated R-dichlorprop cleavage, indicating that it was a single-copy gene . Both rdpA and sdpA were located on the plasmid pMC1 that also carries the lower pathway genes . Sequencing of a 25.8-kb fragment showed that the dioxygenase genes were separated by a 13.6-kb region mainly comprising a Tn501-like transposon . Furthermore, two copies of a sequence similar to IS91-like elements were identified . Hybridization studies comparing the wild-type plasmid and that of the mutant unable to cleave dichlorprop showed that rdpA and sdpA were deleted, whereas the lower pathway genes were unaffected, and that deletion may be caused by genetic rearrangements of the IS91-like elements . Two other dichlorprop-degrading bacterial strains, Rhodoferax sp . strain P230 and Sphingobium herbicidovorans MH, were shown to carry rdpA genes of high similarity to rdpA from strain MC1, but sdpA was not detected . This suggested that rdpA gene products are involved in the degradation of R-dichlorprop in these strains. Appl Environ Microbiol, 2004 Sep, 70(9), 5102 - 10 The gnyRDBHAL cluster is involved in acyclic isoprenoid degradation in Pseudomonas aeruginosa; Diaz-Perez AL et al.; Pseudomonas aeruginosa PAO1 mutants affected in the ability to degrade acyclic isoprenoids were isolated with transposon mutagenesis . The gny cluster (for geranoyl), which encodes the enzymes involved in the lower pathway of acyclic isoprenoid degradation, was identified . The gny cluster is constituted by five probable structural genes, gnyDBHAL, and a possible regulatory gene, gnyR . Mutations in the gnyD, gnyB, gnyA, or gnyL gene caused inability to assimilate acyclic isoprenoids of the citronellol family of compounds . Transcriptional analysis showed that expression of the gnyB gene was induced by citronellol and repressed by glucose, whereas expression of the gnyR gene had the opposite behavior . Western blot analysis of citronellol-grown cultures showed induction of biotinylated proteins of 70 and 73 kDa, which probably correspond to 3-methylcrotonoyl-coenzyme A (CoA) carboxylase and geranoyl-CoA carboxylase (GCCase) alpha subunits, respectively . The 73-kDa biotinylated protein, identified as the alpha-GCCase subunit, is encoded by gnyA . Intermediary metabolites of the isoprenoid pathway, citronellic and geranic acids, were shown to accumulate in gnyB and gnyA mutants . Our data suggest that the protein products encoded in the gny cluster are the beta and alpha subunits of geranoyl-CoA carboxylase (GnyB and GnyA), the citronelloyl-CoA dehydrogenase (GnyD), the gamma-carboxygeranoyl-CoA hydratase (GnyH), and the 3-hydroxy-gamma-carboxygeranoyl-CoA lyase (GnyL) . We conclude that the gnyRDBHAL cluster is involved in isoprenoid catabolism. Am J Chin Med, 2004, 32(3), 389 - 96 Shiunko promotes epithelization of wounded skin; Huang KF et al.; Shiunko is a traditional botanic formula (ointment) which is used clinically for the treatment of wounded skin caused by cuts, abrasions, frost or burn . The aim of this study was to evaluate the effect of Shiunko on epithelization of wounded skin . Experimental cutting wounds on the back skin of Sprague-Dawley rats were induced . Different bacterial inoculations (Pseudomonus aeruginosa and Staphylococcus aureus) and treatment (Shiunko, Povidone-iodine and saline) were arranged herein . The incidences of infection and the speed of epithelization were evaluated . We observed that the incidences of wound infection following Pseudomonas aeruginosa inoculation were lower on both the Shiunko-treated group (0%, p < 0.01) and Povidine-iodine-treated group (5%, p < 0.05), than the saline-treated group (40%) . The Shiunko-treated group reported higher percentages of complete epithelization not only on the sterilized wounds (100%) but also on the contaminated wounds (90%) when compared to the saline-treated group (60% sterilized wounds, 40% and 50% contaminated wounds) on day 7 (p < 0.01) . Povidone-iodine did not promote epithelization of wounded skin, whereas Shiunko did. J Am Vet Med Assoc, 2004 Aug 15, 225(4), 548 - 53 Evaluation of outcome of otitis media after lavage of the tympanic bulla and long-term antimicrobial drug treatment in dogs: 44 cases (1998-2002); Palmeiro BS et al.; OBJECTIVE: To evaluate the outcome of otitis media in dogs after video-otoscopic lavage of the tympanic bulla and long-term antimicrobial drug treatment . DESIGN: Retrospective study . ANIMALS: 44 dogs with otitis media treated in an academic referral practice . PROCEDURE: Medical records were reviewed for signalment, duration of ear canal disease, previous medical treatments, dermatologic diagnosis, results of cytologic examination and microbial culture of ear canal exudate, findings during video-otoscopy, medical treatment, days to resolution, and maintenance treatments prescribed . Four independent variables (age, duration of ear canal disease prior to referral, use of corticosteroids in treatment regimens, and infection with Pseudomonas aeruginosa) were evaluated statistically for potential influence on time to resolution . RESULTS: Mean +/- SD (range) duration of ear canal disease prior to referral was 24.9 +/- 21.6 (3 to 84) months . Otitis media in 36 dogs resolved after lavage of the tympanic bulla and medical management; mean +/- SD (range) time to resolution was 117 +/- 86.7 (30 to 360) days . Time to resolution was not significantly influenced by any variable evaluated . Three dogs were lost to follow-up, and 4 dogs eventually required surgical intervention . Seven of 36 dogs in which otitis had resolved relapsed; 4 required additional lavage procedures . CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that lavage of the tympanic bulla combined with medical management is an effective and viable option for treatment of otitis media in dogs. J Bacteriol, 2004 Sep, 186(18), 6142 - 9 Arginine biosynthesis in Thermotoga maritima: characterization of the arginine-sensitive N-acetyl-L-glutamate kinase; Fernandez-Murga ML et al.; To help clarify the control of arginine synthesis in Thermotoga maritima, the putative gene (argB) for N-acetyl-L-glutamate kinase (NAGK) from this microorganism was cloned and overexpressed, and the resulting protein was purified and shown to be a highly thermostable and specific NAGK that is potently and selectively inhibited by arginine . Therefore, NAGK is in T . maritima the feedback control point of arginine synthesis, a process that in this organism involves acetyl group recycling and appears not to involve classical acetylglutamate synthase . The inhibition of NAGK by arginine was found to be pH independent and to depend sigmoidally on the concentration of arginine, with a Hill coefficient (N) of approximately 4, and the 50% inhibitory arginine concentration (I0.5) was shown to increase with temperature, approaching above 65 degrees C the I0.50 observed at 37 degrees C with the mesophilic NAGK of Pseudomonas aeruginosa (the best-studied arginine-inhibitable NAGK) . At 75 degrees C, the inhibition by arginine of T . maritima NAGK was due to a large increase in the Km for acetylglutamate triggered by the inhibitor, but at 37 degrees C arginine also substantially decreased the Vmax of the enzyme . The NAGKs of T . maritima and P . aeruginosa behaved in gel filtration as hexamers, justifying the sigmoidicity and high Hill coefficient of arginine inhibition, and arginine or the substrates failed to disaggregate these enzymes . In contrast, Escherichia coli NAGK is not inhibited by arginine and is dimeric, and thus the hexameric architecture may be an important determinant of arginine sensitivity . Potential thermostability determinants of T . maritima NAGK are also discussed. Protein Sci, 2004 Oct, 13(10), 2706 - 15 Epub 2004 Aug 31. Probing the influence on folding behavior of structurally conserved core residues in P . aeruginosa apo-azurin; Engman KC et al.; The effects on folding kinetics and equilibrium stability of core mutations in the apo-mutant C112S of azurin from Pseudomonas aeruginosa were studied . A number of conserved residues within the cupredoxin family were recognized by sequential alignment as constituting a common hydrophobic core: I7, F15, L33, W48, F110, L50, V95, and V31 . Of these, I7, V31, L33, and L50 were mutated for the purpose of obtaining information on the transition state and a potential folding nucleus . In addition, residue V5 in the immediate vicinity of the common core, as well as T52, separate from the core, were mutated as controls . All mutants exhibited a nonlinear dependence of activation free energy of folding on denaturant concentration, although the refolding kinetics of the V31A/C112S mutant indicated that the V31A mutation destabilizes the transition state enough to allow folding via a parallel transition state ensemble . Phi-values could be calculated for three of the six mutants, V31A/C112S, L33A/C112S, and L50A/C112S, and the fractional values of 0.63, 0.33, and 0.50 (respectively) obtained at 0.5 M GdmCl suggest that these residues are important for stabilizing the transition state . Furthermore, a linear dependence of ln k(obs)(H2O) on DeltaG(U-N)(H2O) of the core mutations and the putative involvement of ground-state effects suggest the presence of native-like residual interactions in the denatured state that bias this ensemble toward a folding-competent state. Surg Today, 2004, 34(9), 725 - 31 Recovery of the susceptibility of isolated bacterium achieved by giving long-established antibiotics as prophylaxis against postoperative infections; Kusachi S et al.; PURPOSE: To evaluate the effectiveness of operative antibiotic therapy we changed the antibiotics and examined the susceptibility of the postoperative infection . METHODS: We studied 4 593 patients who underwent gastrointestinal surgery during the last 12.5 years, and examined the changes in intraoperative antibiotics, the return of bacteria isolated from infectious sites, the trends in frequency of Methicillin-resistant Staphylococcus aureus (MRSA) isolation, and changes in the antibiotic susceptibility of Pseudomonas aeruginosa and Bacterioides fragilis . We changed the antibiotics to Cefazolin (CEZ) for upper gastrointestinal tract surgery and cholecystectomy, and to Cefotiam (CTM) for colonic, liver, and pancreatic surgery . We also reduced the period of drug administration . RESULTS: The rate of MRSA infections decreased, the rate of P . aeruginosa infections was always under 20% and the rate of B . fragilis infections increased . After the guidelines of drug susceptibility were prepared, the minimum inhibitory concentrations (MICs) of Cefsulodin (CFS) and Piperacillin (PIPC) for P . aeruginosa and of Latamoxef (LMOX) and PIPC for B . fragilis, decreased . CONCLUSION: The use of antibiotics considered comparatively old for the prophylaxis of postoperative infection prevented the emergence of MRSA-like resistant strains, influenced changes in Gram-negative bacteria drug susceptibility, and led to an overall reduction in multiple drug resistance. Mol Ther, 2004 Sep, 10(3), 562 - 73 Effects of CFTR, interleukin-10, and Pseudomonas aeruginosa on gene expression profiles in a CF bronchial epithelial cell Line; Virella-Lowell I et al.; Mutations in CFTR lead to a complex phenotype that includes increased susceptibility to Pseudomonas infections, a functional deficiency of IL-10, and an exaggerated proinflammatory cytokine response . We examined the effects of CFTR gene correction on the gene expression profile of a CF bronchial epithelial cell line (IB3-1) and determined which CF-related gene expression changes could be reversed by IL-10 expression . We performed microarray experiments to monitor the gene expression profile of three cell lines over a time course of exposure to Pseudomonas . At baseline, we identified 843 genes with statistically different levels of expression in CFTR-corrected (S9) cells compared to the IB3-1 line or the IL-10-expressing line . K-means clustering and functional group analysis revealed a primary up-regulation of ubiquitination enzymes and TNF pathway components and a primary down-regulation of protease inhibitors and protein glycosylation enzymes in CF . Key gene expression changes were confirmed by real-time RT-PCR . Massive reprogramming of gene expression occurred 3 h after Pseudomonas exposure . Changes specific to CF included exaggerated activation of cytokines, blunted activation of anti-proteases, and repression of protein glycosylation enzymes . In conclusion, the CFTR genotype changes the expression of multiple genes at baseline and in response to bacterial challenge, and only a subset of these changes is secondary to IL-10 deficiency. Clin Ther, 2004 Jul, 26(7), 1046 - 54 Efficacy of ofloxacin otic solution once daily for 7 days in the treatment of otitis externa: a multicenter, open-label, phase III trial; Torum B et al.; BACKGROUND: Otitis externa (OE) is an infection of the external auditory canal that is typically treated with topically applied broad-spectrum antibiotics . Twice-daily topical treatment with ofloxacin otic 0.3% solution for 10 days has been reported to be as effective and well tolerated as the standard of care, neomycin sulfate/polymyxin B sulfate/hydrocortisone solution administered 4 times daily for 10 days . OBJECTIVE: This study evaluated the efficacy and safety profile of 7 days of a once-daily regimen of ofloxacin otic 0.3% solution in the treatment of OE . METHODS: This multicenter, open-label, Phase III study was conducted from June 12, 2002, to October 14, 2002 . Eligible patients were aged > or = 6 months and had OE of <2 weeks' duration with moderate to severe edema and tenderness involving 1 or both ears and sufficient exudate for microbiologic culture . Ofloxacin otic solution was instilled once daily for 7 days (5 drops for children aged 6 months to <13 years, 10 drops for adolescents/adults aged > or = 13 years) . Assessments were conducted at the end-of-treatment visit and 7 to 10 days later (the test-of-cure visit) . Medication was supplied free of charge to study participants who incurred no costs for physician visits . RESULTS: Of 489 patients enrolled at 58 sites in 3 countries, 439 were clinically evaluable (173 children, 266 adolescents/adults; 52 % males, 48% females; 47% Hispanic, 45% white; 5% black, and 3% other) . The cure rate among clinically evaluable patients was 91% (95% of children, 88% of adolescents/adults); 68% of patients were cured within 7 days . Forty-three potentially pathogenic strains were isolated from 253 microbiologically evaluable patients . Pseudomonas aeruginosa was isolated from 158 (62%) microbiologically evaluable patients and Staphylococcus aureus from 32 (13%) . Eradication rates were 96% overall . No serious adverse events were observed . Minor adverse events were experienced by 15 (3%) of 489 patients included in the safety population . The most common adverse events were pruritus (5 patients), increased earache (4 patients), and application-site reactions (3 patients) . Overall mean (SD) adherence to therapy was 98% (11.9) . CONCLUSIONS: Ofloxacin otic 0.3% solution administered once daily for 7 days was well tolerated and effective in achieving clinical and microbiologic cure of OE . The compliance rates in this study suggests that this regimen may be better accepted by patients than longer, more repetitive regimens. FEMS Microbiol Lett, 2004 Sep 1, 238(1), 49 - 55 The interaction of Pseudomonas aeruginosa PAK with human and animal respiratory tract cell lines; Hambrook J et al.; A major virulence factor of a common human pathogen Pseudomonas aeruginosa, was investigated to determine if it dominated attachment interactions in a variety of in vitro cell culture systems . It was found that Type-IV pilus-type mechanisms, which mediated the attachment of P . aeruginosa to three human respiratory tract cell lines (A549, BEAS-2B and RPMI 2650) also mediated attachment to two respiratory tract cell lines from mouse (C57) and rat (L-2) to a similar degree . Significant differences were found in the number of P . aeruginosa associated with the human, rat and mouse cell lines . Additionally, differences were also found between A547, C57 and L-2 cells with respect to the moieties that P . aeruginosa interacted with at the level of the cell surface, suggesting that asialo-GM1 ligands were not the only structure that this bacterium could interact with in order to associate with host cells. FEMS Microbiol Lett, 2004 Sep 1, 238(1), 23 - 8 MexZ-mediated regulation of mexXY multidrug efflux pump expression in Pseudomonas aeruginosa by binding on the mexZ-mexX intergenic DNA; Matsuo Y et al.; MexZ is a transcriptional regulator of the mexXY multidrug transporter operon, which confers aminoglycoside resistance on Pseudomonas aeruginosa . Highly purified MexZ showed direct binding with a specific site of the mexZ-mexX intergenic DNA when probed by a gel retardation assay . Both in vitro chemical cross-linking experiments and an in vivo two-hybrid expression system showed that the active form of MexZ, which is capable of binding the intergenic DNA, appeared to be a dimer . These results explain the mechanism by which MexZ represses transcription of the mexXY operon, but do not explain the substrate-induced hyperproduction of MexXY . The presence of inducer antibiotic in the gel-retardation assay mixture failed to detect altered MexZ-probe DNA interaction suggesting the possible involvement of an additional regulator. Pediatr Pulmonol, 2004 Oct, 38(4), 277 - 84 Longitudinal pulmonary status of cystic fibrosis children with meconium ileus; Li Z et al.; Although meconium ileus (MI) is the earliest manifestation of cystic fibrosis (CF), and is associated with poorer growth, the longitudinal pulmonary progression of CF children with MI is not clear . To test the hypothesis that MI is associated with worse pulmonary outcomes, we prospectively compared from diagnosis to 12 years of age 32 CF children with MI to 50 CF children without MI who were diagnosed during early infancy through neonatal screening . Pulmonary outcome measures included respiratory symptoms, respiratory infections, pathogens, antibiotic usage, hospitalizations, quantitative chest radiology, spirometry, and lung volume determinations . Obstructive lung disease was defined as percent predicted spirometry values below the lower limits of normal . Longitudinal analyses revealed no significant differences in cough, wheezing, respiratory infections, prevalence of and median times to acquisition of Pseudomonas aeruginosa or Staphylococcus aureus, antibiotic usage, and chest radiograph scores between the two groups . However, MI children showed significantly worse forced expiratory volume in 1 sec (FEV(1)), forced vital capacity (FVC), forced expiratory flow between 25-75% of FVC (FEF(25-75)), % predicted FEV(1), % predicted FEF(25-75), and total lung capacity (TLC) . These differences were particularly apparent beginning at age 8-10 years . MI children also had higher rates of and shorter median times to obstructive lung disease . Subgroup analyses showed MI children treated surgically and those treated medically had similar pulmonary outcomes . In conclusion, MI children have worse lung function and more obstructive lung disease than those without MI . Such abnormalities are accompanied by reduced lung volume . MI is a distinct CF phenotype with more severe pulmonary dysfunction. Am J Otolaryngol, 2004 Sep-Oct, 25(5), 323 - 8 The microbiology and antimicrobial resistance patterns in chronic rhinosinusitis; Kingdom TT et al.; OBJECTIVES: The purpose of this study was to review the microbiology of chronic rhinosinusitis (CRS) in patients undergoing endoscopic sinus surgery (ESS) and comment on antimicrobial resistance trends . METHODS: A retrospective review of 101 patients undergoing ESS during the period of 1997 to 2001 was performed . Patients were divided into groups based on their surgical history . Fifty-five patients without prior ESS history were placed in the primary group; 46 patients who had undergone prior ESS were placed in the revision group . Intraoperative microbiology culture data were reviewed and antimicrobial resistance data analyzed . RESULTS: Data on 101 patients were analyzed . There were 182 total cultures sent, yielding 257 isolates . The most common isolates were coagulase-negative Staphylococcus (SCN) (45% of cultures), gram-negative rods (25% of cultures), and Staphylococcus aureus (24% of cultures) . Pseudomonas aeruginosa was isolated in 9% of cultures . When comparing the 2 patient groups, we did not find consistent trends in the differences in the prevalence of these isolates . Antimicrobial resistance for SCN (P = .01) and S aureus (P < .001) was greater in the revision surgery . Overall, 62% of patients were found to have at least 1 isolate with decreased antibiotic sensitivity . CONCLUSION: The most prevalent microorganisms in patients with CRS are SCN, S aureus, and gram-negative rods . Perhaps more importantly, the antimicrobial sensitivities of these microorganisms appear to be a growing problem . These findings suggest increased antimicrobial resistance in patients undergoing revision ESS when compared with patients undergoing surgery for the first time. East Mediterr Health J, 2001 Jul-Sep, 7(4-5), 738 - 43 Infection following orthopaedic implants and bone surgery; Mousa HA; Forty-seven patients were investigated for early or late postoperative infections of orthopaedic implants and/or bone . A total of 88 isolates were recovered (64 aerobes and 24 anaerobes) . Pseudomonas aeruginosa and Staphylococcus epidermidis were the most common causative agents . Anaerobic bacteria were isolated from 16 (34%) patients; 50% of patients with late-onset infection and 10.5% with early-onset infection . In 6 (12.8%) patients, infection was with anaerobic organisms alone . All these patients had retained an extramedullary internal fixation device . Anaerobic microorganisms appear to play a significant role in the pathogenesis of late-onset postoperative infection, especially where there is an extramedullary internal fixation device. Eur Respir J, 2004 Aug, 24(2), 286 - 91 Airway iron and iron-regulatory cytokines in cystic fibrosis; Reid DW et al.; Iron availability is critical to Pseudomonas aeruginosa . The current authors determined sputum iron, ferritin, microalbumin levels and total cell counts (TCC) in 19 adult patients with cystic fibrosis (CF) during an acute exacerbation and repeated analyses following a median of 12 days antibiotic treatment . The current authors also determined sputum interleukin (IL)-1beta and tumour necrosis factor (TNF)-alpha levels because of their putative role in intracellular iron homeostasis . Additional data were obtained from 17 stable CF patients, eight patients with stable chronic obstructive pulmonary disease (COPD) and six normal subjects . Overall, sputum iron, ferritin, microalbumin, IL-1beta and TNF-alpha concentrations and TCCs were significantly elevated in the CF patients compared to those with COPD and normal controls . Sputum ferritin levels were significantly elevated in acute versus stable CF patients and there was a trend for sputum TCC to be higher, but all other inflammatory indices were similar . In the CF patients, sputum iron was positively and strongly related to IL-1beta, TNF-alpha, ferritin and microalbumin levels, but negatively related to forced expiratory volume in one second % predicted . In those acute patients who clinically improved with antibiotics (n=14), there were significant decreases in sputum TCC, iron, ferritin and IL-1beta content, but not TNF-alpha or albumin levels . However, changes in sputum TNF-alpha in acute patients were still closely related to changes in iron, ferritin and albumin content, and changes in IL-1beta were related to changes in sputum ferritin content . Iron and iron-regulatory cytokines may play a role in cystic fibrosis lung disease and the increased iron content may even facilitate Pseudomonas aeruginosa infection. J Neurooncol, 2004 Jul, 68(3), 245 - 8 Case report: Pseudomonas aeruginosa-related intervertebral discitis in a young boy with medulloblastoma; Mazza E et al.; We report a case of a 15-year-old boy with desmoplastic medulloblastoma of the posterior fossa (T3M3, according to Chang classification) incompletely resected, with leptomeningeal and nodular spread in the posterior fossa and in the cervical and thoracic tracts of the spine, treated with sequential high dose iv chemotherapy and with hyperfractionated cranio-spinal radiotherapy . While on maintenance chemotherapy, the boy developed fever and septic status caused by Pseudomonas aeruginosa, and 1 week later also low back pain . Magnetic resonance imaging (MRI) demonstrated abnormal signal in the fourth ventricle and in the dorso-lumbar tract suggesting medulloblastoma recurrence, so he started with a chemotherapy program . Due to a worsening of back pain, a second MRI of the spine was performed that showed a spondilodiscitis of T11-T12 and L1-L2 discs . The histological and cultural examination of a fine-needle biopsy of the L1-L2 disc revealed the presence of P . aeruginosa . So patient was treated with intensive antibiotic therapy with resolution of the infection . Spondilodiscitis is a rare complication in neoplastic patients, maybe due to either immunodeficient status or invasive procedures such as lumbar puncture . This case demonstrates that MRI is a useful method for differentiating between infection and malignancy in the spine, but sometimes it may be difficult to distinguish metastatic tumor from a lesion due to spondilodiscitis . In this case surgicopathological assessment is crucial and mandatory. Int J Hyg Environ Health, 2004 Jul, 207(3), 259 - 66 Surveillance of Pseudomonas aeruginosa-isolates in a neonatal intensive care unit over a one year-period; Zabel LT et al.; Outbreaks of gram-negative bacteria such as Pseudomonas aeruginosa in neonatal intensive care units (NICU) can be life-threatening to pre-term infants, which are highly susceptible to serious infections with bacteria . Forty-two ventilated neonates in the NICU of the University Children's Hospital of Tuebingen were found to be colonized (n = 40) or infected (n = 2) with P . aeruginosa within a sampling period of one year . To investigate the colonization patterns and identify potential outbreak sources, epidemiological investigations, environmental surveillance and typing by serotyping and pulsed-field gel electrophoresis of the recovered isolates were performed . The investigation demonstrated a genetically related cluster of P . aeruginosa isolates during the surveillance period in 39 neonates and a second cluster at the end of the period in two neonates . A third strain representing a genetically distinct group was found in only one patient . Environmental investigations demonstrated the presence of P . aeruginosa in the ventilation equipment of 22 patients: binasal prongs (n = 22), water reservoir (n = 9), and heater (n = 1) . In one case, P . aeruginosa was found in breast milk . Other environmental investigations revealed no P . aeruginosa . Although no evidence for a unique source was found, a series of intervention steps were initiated by the NICU personnel, medical microbiologists and infection control experts . The intervention steps included reinforced training of health care staff and a change from chemical to thermal disinfection of binasal prongs . Implementation of these measurements successfully stopped the recurrent occurrence of P . aeruginosa colonization. J Chemother, 2004 Jun, 16(3), 282 - 7 Microbiological activity and clinical efficacy of a colistin and rifampin combination in multidrug-resistant Pseudomonas aeruginosa infections; Tascini C et al.; The aim of the study was to assess the microbiological activity and clinical efficacy of colistin and rifampin combination against multidrug-resistant (MDR) Pseudomonas aeruginosa infections . The antimicrobial activity of the colistin/rifampin combination was evaluated using the checkerboard and time-kill curve methods against different MDR P . aeruginosa strains . The combination of rifampin and colistin resulted fully (1 strain) or partially (5 strains) synergistic for 6/7 strains and minimum inhibitory concentrations (MICs) in combination were reduced to easily obtainable therapeutic levels . The time-kill curves showed that the combination was bactericidal against the strains tested . The clinical efficacy of the combination was tested in four patients with difficult-to treat infections (sepsis or pneumonia) caused by MDR P . aeruginosa . All infections were successfully treated . Our microbiological and clinical observations suggest that the addition of rifampin to colistin may result in a synergistic bactericidal combination that may be useful in patients with infections caused by MDR P . aeruginosa which are difficult to cure. J Chemother, 2004 Jun, 16(3), 264 - 8 Susceptibility patterns in Pseudomonas aeruginosa causing nosocomial infections; Astal Z; Nosocomial infection caused by Pseudomonas aeruginosa has not been reported previously in the Gaza Strip . This study aims to determine the distribution of antimicrobial drug resistance in P . aeruginosa causing nosocomial infections . One hundred thirty-one P . aeruginosa isolates were collected from various nosocomial infection clinical samples . The study was conducted between April and October 2003 . The results of this study reveal that the most common resistance was to ampicillin, followed by cephalexin . The most effective antimicrobial agents were meropenem and amikacin, respectively . The highest resistance to ciprofloxacin was found among ICU and surgery sections . The data analysis shows that no remarkable difference was reported with respect to previous admission and prior antimicrobial treatment for most antibiotics . The results of this study emphasize the need for constant monitoring of antimicrobial effectiveness to correctly guide empiric therapy and local intervention programs in an attempt to reduce antimicrobial resistance. Acta Microbiol Pol, 2004, 53(1), 45 - 52 Purification and characterization of extracellular Pseudomonas aeruginosa urate oxidase enzyme; Saeed HM et al.; Urate oxidase (uricase) was isolated and purified from Pseudomonas aeruginosa to apparent homogeneity using ammonium sulphate precipitation followed by ion exchange and gel filtration chromatography . The specific activity of the purified uricase enzyme was found to be 636.36 with the use of uric acid as a substrate . The purified uricase enzyme is a monomeric protein with molecular weight of 64 kilodaltons . The optimal pH and temperature of the purified enzyme is 9.0 and 30 degrees C, respectively . The effect of some metal ions was studied . Sulphate forms of Fe+2, Zn+2 and Co+2 inhibit the uricolytic activity whereas; NaCl and CaCl2 enhance the enzyme activity . Moreover, the purified enzyme is inhibited by EDTA and KCN. Ann Acad Med Singapore, 2004 Jul, 33(4), 484 - 8 Contact lens microbial keratitis and prior topical steroid use: a disaster in the making? Wang JC, Su D, Lim L. INTRODUCTION: To review the best-corrected visual acuity, ulcer size, microbiological profile and morbidity of contact lens-related microbial keratitis with and without prior topical steroid use . MATERIALS AND METHODS: Retrospective case review of admitted cases of contact lens-related microbial keratitis in a tertiary hospital . Data pertaining to demographics, pre-admission treatment with or without topical steroids, ulcer size, duration of admission, Gram stain and culture results as well as the final best-corrected visual acuity were recorded . Patients are classified into 3 groups: Group 1 received no treatment prior to presentation, Group 2 received topical antibiotics only from their general practitioners and Group 3 prescribed both topical antibiotics and steroids . RESULTS: Forty-six cases were enrolled in the study, 41.3% had prior topical steroids (all dexamethasone) in combination with antibiotics . None of them had topical steroids alone . Large ulcers were associated with steroid use, odds ratio = 7.74 {95% confidence interval (CI), 1.18-50.56} and positivity of Gram stains odds ratio = 7.74 {95% CI, 1.18-50.56} whereas loss of more than 2 Snellen lines was associated with Pseudomonas aeruginosa infection, odds ratios of 21.70 {95% CI,2.09-225.03} and presence of central ulcer, 13.51 {95% CI, 2.33-78.3} . Prior topical steroid use was associated with longer duration of symptoms prior to admission but not duration of stay or surgical intervention . CONCLUSION: Patients with prior topical combined antibiotics-steroids present slightly later and with larger ulcers . However, the duration of stay, final visual acuity, treatment failure and complication rates were not statistically different from the non-treated group . This might be due to 1) early presentation and therefore early treatment of contact lens-related microbial keratitis and 2) the short duration of use of combined antibiotic-steroid eye drops. Nature, 2004 Aug 26, 430(7003), 1024 - 7 Cooperation and competition in pathogenic bacteria; Griffin AS et al.; Explaining altruistic cooperation is one of the greatest challenges for evolutionary biology . One solution to this problem is if costly cooperative behaviours are directed towards relatives . This idea of kin selection has been hugely influential and applied widely from microorganisms to vertebrates . However, a problem arises if there is local competition for resources, because this leads to competition between relatives, reducing selection for cooperation . Here we use an experimental evolution approach to test the effect of the scale of competition, and how it interacts with relatedness . The cooperative trait that we examine is the production of siderophores, iron-scavenging agents, in the pathogenic bacterium Pseudomonas aeruginosa . As expected, our results show that higher levels of cooperative siderophore production evolve in the higher relatedness treatments . However, our results also show that more local competition selects for lower levels of siderophore production and that there is a significant interaction between relatedness and the scale of competition, with relatedness having less effect when the scale of competition is more local . More generally, the scale of competition is likely to be of particular importance for the evolution of cooperation in microorganisms, and also the virulence of pathogenic microorganisms, because cooperative traits such as siderophore production have an important role in determining virulence. J Environ Sci Health A Tox Hazard Subst Environ Eng, 2002 Aug, 37(7), 1379 - 90 Antibacterial properties of chitosan in waterborne pathogen; Chen YM et al.; The antimicrobial properties of chitosan, a derivative of chitin, were investigated in the solid and liquid culture against bacteria associated with waterborne disease in order to assess the potential for using chitosan as a natural disinfectant . Six strains which included three gram-negative and three gram-positive bacteria were studied . The effects of the deacetylation degree, concentration, and molecular weight of chitosan on antibacterial activities were assessed . Chitosan exhibited the highest antibacterial activity against the Pseudomonas aeruginosa on the solid agar . Similar tendency was found when the bacteria were cultivated in liquid broth . The higher deacetylation degree and higher concentration of chitosan cause the higher antibacterial activities . The effect of molecular weight of chitosan on the inhibition efficacy of bacteria is dependent on the species of bacteria . Escherichia coli is sensitive to chitosan during its death phase and logarithmic phase . The antibacterial mechanism of chitosan was illustrated by the surface charge and persistence length . Results indicated that chitosan is potential as a natural disinfectant. Antimicrob Agents Chemother, 2004 Sep, 48(9), 3576 - 8 Isolation of an integron-borne blaVIM-4 type metallo-beta-lactamase gene from a carbapenem-resistant Pseudomonas aeruginosa clinical isolate in Hungary; Libisch B et al.; The first integron-borne metallo-beta-lactamase gene was isolated in Hungary . The bla(VIM-4) gene is located on a class 1 integron that also carries a novel bla(OXA)-like gene . The integron is harbored by a serotype O12 Pseudomonas aeruginosa strain and shows high structural similarity to integrons isolated in Greece and Poland. Antimicrob Agents Chemother, 2004 Sep, 48(9), 3573 - 5 Lack of association between hypermutation and antibiotic resistance development in Pseudomonas aeruginosa isolates from intensive care unit patients; Gutierrez O et al.; Hypermutation is a common feature of Pseudomonas aeruginosa isolates from chronically infected cystic fibrosis patients that is linked with antibiotic resistance development . In this work, using a large collection of sequential P . aeruginosa isolates from ICU patients, we found that despite the fact that mutational antibiotic resistance development is a frequent outcome, the prevalence of hypermutable strains is low (found in isolates from only 1 of 103 patients) and there is no evidence of coselection of the hypermutable and antibiotic resistance phenotypes. Antimicrob Agents Chemother, 2004 Sep, 48(9), 3457 - 61 Azithromycin inhibits MUC5AC production induced by the Pseudomonas aeruginosa autoinducer N-(3-Oxododecanoyl) homoserine lactone in NCI-H292 Cells; Imamura Y et al.; The features of chronic airway diseases, including chronic bronchitis, cystic fibrosis, bronchiectasis, and diffuse panbronchiolitis, include chronic bacterial infection and airway obstruction by mucus . Pseudomonas aeruginosa is one of the most common pathogens in chronic lung infection, and quorum-sensing systems contribute to the pathogenesis of this disease . The quorum-sensing signal molecule {N-(3-oxododecanoyl) homoserine lactone (3O-C(12)-HSL)} not only regulates bacterial virulence but also is associated with the immune response . In this study, we investigated whether 3O-C(12)-HSL could stimulate the production of a major mucin core protein, MUC5AC . The effect of a macrolide on MUC5AC production was also studied . 3O-C(12)-HSL induced NCI-H292 cells to express MUC5AC at both the mRNA and the protein levels in time- and dose-dependent manners . A 15-membered macrolide, azithromycin, inhibited MUC5AC production that was activated by 3O-C(12)-HSL . 3O-C(12)-HSL induced extracellular signal-regulated kinase (ERK) 1/2 and I-kappa B phosphorylation in cells, and this induction was suppressed by azithromycin . 3O-C(12)-HSL-induced MUC5AC production was blocked by the ERK pathway inhibitor PD98059 . Our findings suggest that the P . aeruginosa autoinducer 3O-C(12)-HSL contributes to excessive mucin production in chronic bacterial infection . Azithromycin seems to reduce this mucin production by interfering with intracellular signal transduction. Antimicrob Agents Chemother, 2004 Sep, 48(9), 3402 - 6 Sequence-selective recognition of extended-spectrum beta-lactamase GES-2 by a competitive, peptide nucleic acid-based multiplex PCR assay; Weldhagen GF; Extended-spectrum beta-lactamases (ESBLs) in Pseudomonas aeruginosa, such as GES-2, which compromises the efficacy of imipenem, tend to be geographically restricted . The CC-to-AA base pair substitution at positions 493 and 494 of the bla(GES-2)-coding region distinguishes this ESBL from bla(GES-1) and the bla(IBC)-type genes, making it an ideal target for the development of a novel sequence-specific, peptide nucleic acid (PNA)-based multiplex PCR detection method . By using two primer pairs in conjunction with a PNA probe, this method provided an accurate means of identification of bla(GES-2) compared to standard PCR and gene sequencing techniques when it was used to test 100 P . aeruginosa clinical isolates as well as previously published, well-described control strains encompassing all presently known genes in the bla(GES-IBC) ESBL family . This novel method has the potential to be used in large-scale, cost-effective screening programs for specific or geographically restricted ESBLs. Antimicrob Agents Chemother, 2004 Sep, 48(9), 3396 - 401 A mucoadhesive polymer extracted from tamarind seed improves the intraocular penetration and efficacy of rufloxacin in topical treatment of experimental bacterial keratitis; Ghelardi E et al.; Bacterial keratitis is a serious infectious ocular disease requiring prompt treatment to prevent frequent and severe visual disabilities . Standard treatment of bacterial keratitis includes topical administration of concentrated antibiotic solutions repeated at frequent intervals in order to reach sufficiently high drug levels in the corneal tissue to inhibit bacterial growth . However, this regimen has been associated with toxicity to the corneal epithelium and requires patient hospitalization . In the present study, a mucoadhesive polymer extracted from tamarind seeds was used for ocular delivery of 0.3% rufloxacin in the treatment of experimental Pseudomonas aeruginosa and Staphylococcus aureus keratitis in rabbits . The polysaccharide significantly increased the intra-aqueous penetration of rufloxacin in both infected and uninfected eyes . Rufloxacin delivered by the polysaccharide reduced P . aeruginosa and S . aureus in the cornea at a higher rate than that obtained by rufloxacin alone . In particular, use of the polysaccharide allowed a substantial reduction of S . aureus in the cornea to be achieved even when the time interval between drug administrations was extended . These results suggest that the tamarind seed polysaccharide prolongs the precorneal residence times of antibiotics and enhances drug accumulation in the cornea, probably by reducing the washout of topically administered drugs . The tamarind seed polysaccharide appears to be a promising candidate as a vehicle for the topical treatment of bacterial keratitis. Antimicrob Agents Chemother, 2004 Sep, 48(9), 3284 - 90 Functional and structural characterization of the genetic environment of an extended-spectrum beta-lactamase blaVEB gene from a Pseudomonas aeruginosa isolate obtained in India; Aubert D et al.; A Pseudomonas aeruginosa clinical strain isolated from a patient hospitalized in a New Delhi, India, hospital was resistant to expanded-spectrum cephalosporins, imipenem, and aztreonam . A bla(VEB-1)-like gene named bla(VEB-1a), which codes for the extended-spectrum beta-lactamase VEB-1a, was identified . The genetic environment of bla(VEB-1a) was peculiar: (i) no 5' conserved sequence (5'-CS) region was present upstream of the beta-lactamase gene, whereas bla(VEB-1)-like genes are usually associated with class 1 integrons; (ii) bla(VEB-1a) was inserted between two truncated 3'-CS regions in a direct repeat; and (iii) four 135-bp repeated DNA sequences (repeated elements) were located on each side of the bla(VEB-1a) gene . Expression of the bla(VEB-1a) gene was driven by a strong promoter located in one of these repeated sequences . In addition, cloning of the beta-lactamase content of this P . aeruginosa isolate followed by expression in Escherichia coli identified the naturally occurring AmpC beta-lactamase and a gene encoding an OXA-2-like beta-lactamase located in a class 1 integron, In78, in which an insertion sequence, ISpa7, was inserted within its 5'-CS region. Invest Ophthalmol Vis Sci, 2004 Sep, 45(9), 3177 - 84 Caspase-1 inhibitor reduces severity of pseudomonas aeruginosa keratitis in mice; Thakur A et al.; PURPOSE: To test an inhibitor of IL-1beta converting enzyme (ICE), with or without ciprofloxacin, in a C57BL/6 mouse model of keratitis induced by Pseudomonas aeruginosa in which corneal perforation is expected . METHODS: Clinical score, histopathology, myeloperoxidase (MPO) activity, bacterial counts, and ELISA analysis were used to assess the efficacy of treatment initiated at 18 hours postinfection (p.i.) with ICE inhibitor versus placebo; and with ICE inhibitor plus ciprofloxacin versus placebo plus ciprofloxacin . Efficacy of the ICE inhibitor was also tested and evaluated for clinical score in experimental corneal infection induced by a clinical isolate and a ciprofloxacin-resistant bacterial strain . RESULTS: Clinical scores were reduced at 3, 5, and 7 days p.i . in ICE inhibitor versus placebo-treated mice; reduced scores also were observed with a combined treatment (ICE inhibitor and ciprofloxacin) . Further testing (MPO assay) revealed reduced PMN number, particularly striking in ICE inhibitor and ciprofloxacin versus placebo and ciprofloxacin-treated mice . Corneal protein levels for IL-1beta and MIP-2 also were reduced in mice treated with the ICE inhibitor versus placebo and in ICE inhibitor and ciprofloxacin versus ciprofloxacin and placebo-treated mice . Treatment with ICE inhibitor also reduced clinical scores after corneal infection with a clinical isolate, KEI-1025, and with a ciprofloxacin-resistant P . aeruginosa strain . CONCLUSIONS: Downregulation of IL-1beta by ICE together with ciprofloxacin to kill bacteria may provide alternate therapy to current treatment . Copyright Association for Research in Vision and Ophthalmology Int J Antimicrob Agents, 2004 Sep, 24(3), 219 - 25 Effect of adjunctive treatment with gamma interferon against Pseudomonas aeruginosa pneumonia in neutropenic and non-neutropenic hosts; Babalola CP et al.; To evaluate the adjunctive effect of interferon-gamma (IFN-gamma) in treatment of Pseudomonas aeruginosa pneumonia, neutropenic and non-neutropenic mice received LD(100) of the organism intratracheally, followed by subcutaneous administration of IFN-gamma, ceftazidime (TAZ) or a combination of both agents 2 h post-inoculation . Treatment with IFN-gamma alone showed no significant increase in survival when compared with control . Addition of IFN-gamma to TAZ resulted in no significant change in survival compared with TAZ alone . Survival in TAZ and TAZ + IFN-gamma groups, was significantly higher than in control and IFN-gamma groups . This suggests that adjunctive treatment with IFN-gamma in combination with ceftazidime may not be more beneficial than the antibiotic alone when managing acute P . aeruginosa pneumonia in both immunocompromised and immunocompetent hosts. Biochem Biophys Res Commun, 2004 Sep 17, 322(2), 483 - 9 Role of the membrane fusion protein in the assembly of resistance-nodulation-cell division multidrug efflux pump in Pseudomonas aeruginosa; Mokhonov VV et al.; The tripartite xenobiotic-antibiotic transporter of Pseudomonas aeruginosa consists of the inner membrane transporter (e.g., MexB, MexY), the periplasmic membrane-fusion-protein (e.g., MexA, MexX), and the outer membrane channel protein (e.g., OprM) . These subunits were assumed to assemble into a transporter unit during export of the substrates . However, subunit interaction and their specificity in native form remained to be elucidated . To address these important questions, we analyzed the role of the individual subunits for the assembly of MexAB-OprM by pull-down assay tagging only one of the subunits . We found stable MexA-MexB-OprM complex without chemical cross-linking that withstand all purification procedures . Results of bi-partite interactions analysis showed tight association between MexA and OprM in the absence of MexB, whereas the expression systems lacking MexA failed to co-purify MexB or OprM . None of the heterologous subunit combinations such as MexA+MexY(his)+OprM and MexX+MexB(his)+OprM showed interaction . These results implied that the membrane fusion protein is central to the tripartite xenobiotic transporter assembly. J Immunol, 2004 Sep 1, 173(5), 2909 - 12 Cutting edge: 1,25-dihydroxyvitamin D3 is a direct inducer of antimicrobial peptide gene expression; Wang TT et al.; The hormonal form of vitamin D(3), 1,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)), is an immune system modulator and induces expression of the TLR coreceptor CD14 . 1,25(OH)(2)D(3) signals through the vitamin D receptor, a ligand-stimulated transcription factor that recognizes specific DNA sequences called vitamin D response elements . In this study, we show that 1,25(OH)(2)D(3) is a direct regulator of antimicrobial innate immune responses . The promoters of the human cathelicidin antimicrobial peptide (camp) and defensin beta2 (defB2) genes contain consensus vitamin D response elements that mediate 1,25(OH)(2)D(3)-dependent gene expression . 1,25(OH)(2)D(3) induces antimicrobial peptide gene expression in isolated human keratinocytes, monocytes and neutrophils, and human cell lines, and 1,25(OH)(2)D(3) along with LPS synergistically induce camp expression in neutrophils . Moreover, 1,25(OH)(2)D(3) induces corresponding increases in antimicrobial proteins and secretion of antimicrobial activity against pathogens including Pseudomonas aeruginosa . 1,25(OH)(2)D(3) thus directly regulates antimicrobial peptide gene expression, revealing the potential of its analogues in treatment of opportunistic infections. Infect Immun, 2004 Sep, 72(9), 5433 - 8 Transcriptome analysis of Pseudomonas aeruginosa after interaction with human airway epithelial cells; Frisk A et al.; The transcriptional profile of Pseudomonas aeruginosa after interactions with primary normal human airway epithelial cells was determined using Affymetrix GeneChip technology . Gene expression profiles indicated that various genes involved in phosphate acquisition and iron scavenging were differentially regulated. Infect Immun, 2004 Sep, 72(9), 5012 - 8 Microarray analysis reveals induction of lipoprotein genes in mucoid Pseudomonas aeruginosa: implications for inflammation in cystic fibrosis; Firoved AM et al.; The main cause of the high morbidity and mortality of cystic fibrosis (CF) is the progressive lung inflammation associated with Pseudomonas aeruginosa colonization . During the course of chronic CF infections, P . aeruginosa undergoes a conversion to a mucoid phenotype . The emergence of mucoid P . aeruginosa in CF is associated with increased inflammation, respiratory decline, and a poor prognosis . Here we show, by the use of microarray analysis, that upon P . aeruginosa conversion to mucoidy, there is a massive and preferential induction of genes encoding bacterial lipoproteins . Bacterial lipoproteins are potent agonists of Toll-like receptor 2 (TLR2) signaling . The expression of TLR2 in human respiratory epithelial cells was ascertained by Western blot analysis . Human respiratory epithelial cells responded in a TLR2-dependent manner to bacterial lipopeptides derived from Pseudomonas lipoproteins induced in mucoid strains . The TLR2 proinflammatory response was further augmented in CF cells . Thus, the excessive inflammation in CF is the result of a global induction in mucoid P . aeruginosa of lipoproteins that act as proinflammatory toxins (here termed lipotoxins) superimposed on the hyperexcitability of CF cells . Blocking the signaling cascade responding to bacterial lipotoxins may provide therapeutic benefits for CF patients. J Coll Physicians Surg Pak, 2004 Aug, 14(8), 459 - 61 Microbiology and drug sensitivity patterns of chronic suppurative otitis media; Aslam MA et al.; OBJECTIVE: To identify the commonest microorganisms associated with chronic discharging ears and their antimicrobial sensitivities . DESIGN: Descriptive study . PLACE AND DURATION OF STUDY: This study was carried out from August 2003 to February 2004 at the Department of Otorhinolaryngology and Head and Neck Surgery, Fauji Foundation Hospital, Rawalpindi . MATERIALS AND METHODS: A total of 124 patients with unilateral or bilateral active chronic suppurative otitis media attending the outpatient clinic were included in the study . All patients were evaluated through detailed history and clinical examination . Pus samples were collected from the discharging ear(s) and sent to the hospital laboratory where culture and sensitivity studies were done for aerobes, anaerobes and fungi and antibiotic sensitivity patterns . RESULTS: Overall microbiology of 142 samples was studied . Among them, 108 (76%) were pure cultures and 34 (23.9%) were mixed . There were 186 isolates including 182 (97.8%) aerobes, nil anaerobes and only 4 (2.1%) fungi . Pseudomonas aeruginosa 94(50.5%) was the most common isolate, followed by Staphylococcus aureus 44 (23.6%) . Drug sensitivities pattern of Pseudomonas aeruginosa showed that ciprofloxacin was active against majority 95.8% of isolates followed by amikacin 83.3%, gentamicin and tobarmycin 60% and cefotaxime 41.6% . Staphylococcus aureus isolates were resistant to penicillin, ampicillin and amoxicillin in 77.2% whereas majority was sensitive to coamoxiclav 81.8% and cephradine 86.3% . CONCLUSION: Commonest organisms isolated from chronic discharging ears were Pseudomonas aeruginosa and Staphylococcus aureus . Majority of isolates of Pseudomonas aeruginosa were sensitive to ciprofloxacin . Majority of strains of Staphylococcus aureus were resistant to penicillin . Cephradine and coamoxiclav were effective against most of the isolates of Staphylococcus aureus. Clin Exp Immunol, 2004 Sep, 137(3), 478 - 85 Faster activation of polymorphonuclear neutrophils in resistant mice during early innate response to Pseudomonas aeruginosa lung infection; Jensen PO et al.; Polymorphonuclear neutrophils (PMNs) are crucial for the outcome of Pseudomonas aeruginosa lung infection in patients with cystic fibrosis . We compared PMNs and inflammatory cytokines in the lungs and blood from susceptible BALB/c and resistant C3H/HeN mice 1 and 2 days after intratracheal challenge with alginate embedded P . aeruginosa . These parameters were correlated with the quantitative bacteriology and histopathology of the lungs . After challenge, the content of granulocyte colony-stimulating factor (G-CSF) and macrophage inflammatory protein-2 (MIP-2) was increased in the lungs and the sera and the percentage of PMNs was increased in the blood . However, 2 days after challenge the concentration of G-CSF and MIP-2 was higher in the lungs and sera of BALB/c mice . CD11b expression was higher on the PMNs of the C3H/HeN mice . The expression of CD62L on PMNs of both strains of mice was decreased 1 day after bacterial challenge, whereas the expression was increased after 2 days of challenge on PMNs of C3H/HeN mice only . These changes were accompanied by a more severe lung inflammation in BALB/c mice and faster clearance of the bacteria in C3H/HeN mice . In conclusion, the rapid early bacterial clearance in the lungs of C3H/HeN mice could be explained by faster activation of the PMNs, as indicated by the higher up-regulation of CD11b . The severe lung inflammation in BALB/c mice may be caused by the early higher content of G-CSF in the sera mobilizing PMNs from the bone marrow and the persistent chemotactic gradient provided by MIP-2 in the lungs. J Laryngol Otol, 2004 Jul, 118(7), 576 - 9 Use of magnetic resonance imaging as the primary imaging modality in the diagnosis and follow-up of malignant external otitis; Ismail H et al.; Malignant external otitis (MEO) is a severe infection of the external auditory meatus caused by Pseudomonas aeruginosa . Classical features include unrelenting deep otalgia, otorrhoea and granulations in the floor of the ear canal . Treatment is generally protracted antibiotic therapy and monitoring of inflammatory markers; the erythrocyte sedimentation rate (ESR) and C-reactive protein (CRP) . Traditionally computed tomography (CT) has been the imaging modality of choice . The authors present a case where magnetic resonance imaging (MRI) has been crucial in the diagnosis and follow up of a patient with MEO. J Biol Chem, 2004 Nov 19, 279(47), 48671 - 9 Epub 2004 Aug 17. Structure-activity relationships in defensin dimers: a novel link between beta-defensin tertiary structure and antimicrobial activity; Campopiano DJ et al.; Defensins are cationic antimicrobial peptides that have a characteristic six-cysteine motif and are important components of the innate immune system . We recently described a beta-defensin-related peptide (Defr1) that had potent antimicrobial activity despite having only five cysteines . Here we report a relationship between the structure and activity of Defr1 through a comparative study with its six cysteine-containing analogue (Defr1 Y5C) . Against a panel of pathogens, we found that oxidized Defr1 had significantly higher activity than its reduced form and the oxidized and reduced forms of Defr1 Y5C . Furthermore, Defr1 displayed activity against Pseudomonas aeruginosa in the presence of 150 mm NaCl, whereas Defr1 Y5C was inactive . By using nondenaturing gel electrophoresis and Fourier transform ion cyclotron resonance mass spectrometry, we observed Defr1 and Defr1 Y5C dimers . Two complementary fragmentation techniques (collision-induced dissociation and electron capture dissociation) revealed that Defr1 Y5C dimers form by noncovalent, weak association of monomers that contain three intramolecular disulfide bonds . In contrast, Defr1 dimers are resistant to collision-induced dissociation and are only dissociated into monomers by reduction using electron capture . This is indicative of Defr1 dimerization being mediated by an intermolecular disulfide bond . Proteolysis and peptide mass mapping revealed that Defr1 Y5C monomers have beta-defensin disulfide bond connectivity, whereas oxidized Defr1 is a complex mixture of dimeric isoforms with as yet unknown inter- and intramolecular connectivities . Each isoform contains one intermolecular and four intramolecular disulfide bonds, but because we were unable to resolve the isoforms by reverse phase chromatography, we could not assign each isoform with a specific antimicrobial activity . We conclude that the enhanced activity and stability of this mixture of Defr1 dimeric isoforms are due to the presence of an intermolecular disulfide bond . This first description of a covalently cross-linked member of the defensin family provides further evidence that the antimicrobial activity of a defensin is linked to its ability to form stable higher order structures. J Bacteriol, 2004 Sep, 186(17), 5672 - 84 Identification of AlgR-regulated genes in Pseudomonas aeruginosa by use of microarray analysis; Lizewski SE et al.; The Pseudomonas aeruginosa transcriptional regulator AlgR controls a variety of different processes, including alginate production, type IV pilus function, and virulence, indicating that AlgR plays a pivotal role in the regulation of gene expression . In order to characterize the AlgR regulon, Pseudomonas Affymetrix GeneChips were used to generate the transcriptional profiles of (i) P . aeruginosa PAO1 versus its algR mutant in mid-logarithmic phase, (ii) P . aeruginosa PAO1 versus its algR mutant in stationary growth phase, and (iii) PAO1 versus PAO1 harboring an algR overexpression plasmid . Expression analysis revealed that, during mid-logarithmic growth, AlgR activated the expression of 58 genes while it repressed the expression of 37 others, while during stationary phase, it activated expression of 45 genes and repression of 14 genes . Confirmatory experiments were performed on two genes found to be AlgR repressed (hcnA and PA1557) and one AlgR-activated operon (fimU-pilVWXY1Y2) . An S1 nuclease protection assay demonstrated that AlgR repressed both known hcnA promoters in PAO1 . Additionally, direct measurement of hydrogen cyanide (HCN) production showed that P . aeruginosa PAO1 produced threefold-less HCN than did its algR deletion strain . AlgR also repressed transcription of two promoters of the uncharacterized open reading frame PA1557 . Further, the twitching motility defect of an algR mutant was complemented by the fimTU-pilVWXY1Y2E operon, thus identifying the AlgR-controlled genes responsible for this defect in an algR mutant . This study identified four new roles for AlgR: (i) AlgR can repress gene transcription, (ii) AlgR activates the fimTU-pilVWXY1Y2E operon, (iii) AlgR regulates HCN production, and (iv) AlgR controls transcription of the putative cbb3-type cytochrome PA1557. J Bacteriol, 2004 Sep, 186(17), 5596 - 602 Functional characterization of an aminotransferase required for pyoverdine siderophore biosynthesis in Pseudomonas aeruginosa PAO1; Vandenende CS et al.; The fluorescent dihydroxyquinoline chromophore of the pyoverdine siderophore in Pseudomonas is a condensation product of D-tyrosine and l-2,4-diaminobutyrate . Both pvdH and asd (encoding aspartate beta-semialdehyde dehydrogenase) knockout mutants of Pseudomonas aeruginosa PAO1 were unable to synthesize pyoverdine under iron-limiting conditions in the absence of l-2,4-diaminobutyrate in the culture media . The pvdH gene was subcloned, and the gene product was hyperexpressed and purified from P . aeruginosa PAO1 . PvdH was found to catalyze an aminotransferase reaction, interconverting aspartate beta-semialdehyde and l-2,4-diaminobutyrate . Steady-state kinetic analysis with a novel coupled assay established that the enzyme adopts a ping-pong kinetic mechanism and has the highest specificity for alpha-ketoglutarate . The specificity of the enzyme toward the smaller keto acid pyruvate is 41-fold lower . The enzyme has negligible activity toward other keto acids tested . Homologues of PvdH were present in the genomes of other Pseudomonas spp . These homologues were found in the DNA loci of the corresponding genomes that contain other pyoverdine synthesis genes . This suggests that there is a general mechanism of l-2,4-diaminobutyrate synthesis in Pseudomonas strains that produce the pyoverdine siderophore. J Antimicrob Chemother, 2004 Oct, 54(4), 767 - 71 Epub 2004 Aug 18. Comparison of in vivo intrinsic activity of cefepime and imipenem in a Pseudomonas aeruginosa rabbit endocarditis model: effect of combination with tobramycin simulating human serum pharmacokinetics; Navas D et al.; OBJECTIVES: The purpose of this experimental study was first to compare the in vivo intrinsic activity of imipenem and cefepime administered as a continuous infusion and to determine their lowest effective serum steady-state concentration (LESSC) . Secondly, we studied the effect of combining therapy with tobramycin . METHODS: In a Pseudomonas aeruginosa (ATCC 27853) rabbit endocarditis model, beta-lactam antibiotics were administered by continuous infusion over a 24 h treatment period at different doses until the LESSC was reached, i.e . able to achieve a 2-log drop of cfu/g of vegetations versus untreated animals . The effect of adding tobramycin (3 mg/kg once daily) was then studied . RESULTS: The LESSC was between 3 x and 4 x MIC of cefepime for P . aeruginosa and about 0.2 5x MIC of imipenem . Combination of tobramycin with each of the two beta-lactams did not result in any further significant killing . CONCLUSION: The optimal Css/MIC ratio might differ from one molecule to another . The LESSC of imipenem is lower than that of cefepime, giving a better intrinsic activity in vivo, despite a higher MIC in vitro. Am J Respir Crit Care Med, 2004 Dec 1, 170(11), 1164 - 71 Epub 2004 Dec 1. Impact of cigarette smoke on clearance and inflammation after Pseudomonas aeruginosa infection; Drannik AG et al.; The object of this study was to investigate the impact of cigarette smoke on bacterial clearance and immune inflammatory parameters after infection with Pseudomonas aeruginosa in mice . We observed a delayed rate of bacterial clearance in smoke-exposed compared with sham-exposed mice . This was associated with increased inflammation characterized by greater numbers of neutrophils and mononuclear cells in the bronchoalveolar lavage . After infection, we observed increased levels of proinflammatory cytokines (tumor necrosis factor-alpha, interleukin-1beta, and interleukin-6) and chemokines (monocyte chemoattractant protein-1 {MCP-1} and macrophage inflammatory protein-2 {MIP-2}) as well as myeloperoxidase and proteolytic activity in the lungs of smoke-exposed compared with sham-exposed animals . Delayed clearance was associated with increased morbidity and greater weight loss of smoke-exposed mice . After delivery of inactivated bacteria, we observed a similar inflammatory response, clinical score, and tumor necrosis factor-alpha expression in smoke- and sham-exposed animals, suggesting that increased inflammation and altered clinical presentation are due to the delayed rate of bacterial clearance . Our findings suggest that cigarette smoke affects respiratory immune-inflammatory responses elicited by bacteria . We postulate that altered respiratory host defense may be implicated in smoking-related diseases such as chronic obstructive pulmonary disease. Hautarzt, 2004 Nov, 55(11), 1067 - 73 {Pseudomonas aeruginosa infection presenting as gonorrhea}; Schugt I et al.; A 68 year old man presented with urethritis and a purulent discharge, carrying the tentative diagnosis of gonorrhea . He had already been treated with multiple antibiotics . Microbiological investigation revealed Pseudomonas aeruginosa, a relatively frequent Gram-negative bacteria in hospitals, which can cause several nosocomial diseases such as pneumonia, wound infections and urogenital infections . Therapy can be difficult because of frequent antibiotic resistance . Guided by sensitivity studies, the patient was successfully treated with gyrase inhibitors . Pseudomonas aeruginosa-induced urogenital infections in ambulatory patients are extremely rare and usually not associated with a gonorrhea-like discharge. Dermatology, 2004, 209(2), 111 - 6 Early cutaneous alterations in experimental sepsis by Pseudomonas aeruginosa; Petropoulou H et al.; BACKGROUND: To evaluate whether histopathologic findings of skin in sepsis by Pseudomonas aeruginosa correlate with the clinical course . METHODS: Histological alterations after bacterial challenge by one susceptible (A) and two multidrug-resistant isolates (B and C) of P . aeruginosa were studied in 18 rabbits . Sepsis was induced by the intravenous infusion of 1 x 10(8) CFU by a catheter in the right jugular vein; blood was sampled for the estimation of tumor necrosis factor alpha (TNF-alpha) and malondialdehyde (MDA) . Skin biopsies were collected along with a subcutaneous fat specimen for culture . RESULTS: The mean survival was 0.85, 1.75 and 11.00 days after challenge by isolates A, B and C, respectively . The main histologic findings of skin were: inflammation and swelling of the dermis; thickening of the endothelium and infiltration of vessel wall and lumen by polymorphonuclear leukocytes; extravasation of red blood cells, and necrobiotic changes of the hair follicles . Serum TNF-alpha was elevated in animals challenged by isolate A compared to challenge by isolates B and C . Concentrations of MDA were similar for all isolates . Mean log(10) of viable cells isolated from subcutaneous fat were 5.74, 2.74 and 1.40 after challenge by isolates A, B and C, respectively . CONCLUSIONS: Prolongation of survival was accompanied by lower serum TNF-alpha, decreased viable cells from subcutaneous fat and intensified inflammatory response in the dermis and subcutaneous tissue . These findings might be of importance for immunomodulatory intervention. J Biol Chem, 2004 Oct 29, 279(44), 45919 - 25 Epub 2004 Aug 16. Crystal structure of ADP-ribosylated ribosomal translocase from Saccharomyces cerevisiae; Jorgensen R et al.; The crystal structure of ADP-ribosylated yeast elongation factor 2 in the presence of sordarin and GDP has been determined at 2.6 A resolution . The diphthamide at the tip of domain IV, which is the target for diphtheria toxin and Pseudomonas aeruginosa exotoxin A, contains a covalently attached ADP-ribose that functions as a very potent inhibitor of the factor . We have obtained an electron density map of ADP-ribosylated translation factor 2 revealing both the ADP-ribosylation and the diphthamide . This is the first structure showing the conformation of an ADP-ribosylated residue and confirms the inversion of configuration at the glycosidic linkage . Binding experiments show that the ADP-ribosylation has limited effect on nucleotide binding affinity, on ribosome binding, and on association with exotoxin A . These results provide insight to the inhibitory mechanism and suggest that inhibition may be caused by erroneous interaction of the translation factor with the codon-anticodon area in the P-site of the ribosome. J Med Microbiol, 2004 Sep, 53(Pt 9), 915 - 20 Effect of carbapenems on the transcriptional expression of the oprD, oprM and oprN genes in Pseudomonas aeruginosa; Kolayli F et al.; The effects of imipenem and meropenem on the transcriptional expression of resistance-related genes oprD, oprM and oprN in Pseudomonas aeruginosa were studied by quantitative real-time PCR . Four strains were examined: the type strain PT5 (PAO1), its derivatives M7 and PT149, and a clinical isolate, PaKT3 . The derivative M7 is a nalB mutant, overexpressing the MexAB-OprM pump, and the derivative PT149 is a nfxC-type mutant, overexpressing the MexEF-OprN pump while it is down-regulated for the OprD protein . After 18 h incubation in broth, the cultures were divided into three portions . Two were supplemented with antibiotics and the other was left antibiotic-free as the control . After a further 45 min incubation, total RNA was isolated from the strains by guanidine denaturation and acid-phenol/chloroform extraction . DNA-free total RNAs were immediately reverse-transcribed by MMuLV reverse transcriptase . Concentrations of mRNAs obtained by quantitative PCR were expressed relative to uninduced portions of the strains . The results showed that oprD was relatively stable against carbapenem antibiotics . oprM was induced significantly by imipenem in only one strain and oprN was induced by imipenem in most of the strains . The responses at the mRNA level found here were unexpected and suggested a chaotic, unpredictable regulatory mechanism. J Mol Biol, 2004 Jul 30, 341(1), 171 - 84 Characterization and manipulation of the Pseudomonas aeruginosa dimethylarginine dimethylaminohydrolase monomer--dimer equilibrium; Plevin MJ et al.; In mammals, the enzyme dimethylarginine dimethylaminohydrolase (DDAH) is implicated in the regulation of the cellular levels of asymmetric methylarginines, small molecule metabolites that themselves represent a family of endogenous inhibitors of nitric oxide synthase (NOS) . The involvement of DDAH function in the regulation of NOS makes this enzyme a potentially attractive therapeutic target . DDAH from the bacterium Pseudomonas aeruginosa (PaDDAH) is so far the only structurally tractable homologue of mammalian DDAH isoforms . To complement the recent crystal structure of this protein, we show by hydrodynamic measurements that PaDDAH exists in dynamic equilibrium between monomer (ca 29 kDa) and symmetric homodimer (ca 58 kDa) states with a dimer dissociation constant, K(d) approximately 500nM . For the purposes of NMR-based approaches to the study of this enzyme's interactions with substrate and inhibitor ligands, it would be useful to obtain the protein in monomeric form . Through detailed analysis of the homodimer PaDDAH crystal structure we identified key residues involved in the protomer-protomer interface and targeted these for mutation . The hydrodynamic and self-associative properties of a series of PaDDAH interface mutants were analyzed by concentration-dependent analytical size-exclusion chromatography and sedimentation equilibrium analytical ultracentrifugation . The individual substitution of several of the interface residues shifts the equilibrium position towards the monomer, which allowed the design of a double mutant variant (Arg40-->Glu, Arg98-->His) that behaves exclusively as a stable monomer, yet retains greater than 95% catalytic activity compared to wild-type . Comparative two-dimensional (1)H, (15)N heteronuclear NMR spectra indicate that the double mutant remains a monomer even at approximately 1 mM concentration . Accordingly, the double mutant PaDDAH is an attractive template for further NMR-based investigations of the enzyme mechanism and characterization of ligand-binding and inhibitor-binding profiles . These results indicate that dimerization of PaDDAH is not critical for the maintenance of the biological function of the protein . These results are discussed in the context of known modes of self-association between structurally related, but functionally distinct, members of the beta/alpha-propeller fold class. Chemosphere, 2004 Oct, 57(3), 165 - 9 Mechanism of Navitan Fast Blue S5R degradation by Pseudomonas aeruginosa; Valli Nachiyar C et al.; The mechanism by which Pseudomonas aeruginosa degraded Navitan Fast Blue S5R was studied using TLC, FTIR, HPLC and GC-MS analysis . Degradation started with the reduction of azo bonds producing metanilic acid and peri acid whose presence was confirmed by HPLC . Aniline, the desulfonated product from metanilic acid and salicylic acid could also be detected by HPLC . GC-MS analysis of the degradation products confirmed the presence of aniline and revealed the presence of beta-ketoadipic acid . Based on these products a probable pathway has been proposed for the degradation of Navitan Fast Blue S5R by Pseudomonas aeruginosa. Nat Struct Mol Biol, 2004 Sep, 11(9), 868 - 76 Epub 2004 Aug 15. How bacterial ADP-ribosylating toxins recognize substrates; Sun J et al.; ExoS and ExoT are bifunctional type III cytotoxins of Pseudomonas aeruginosa that contain an N-terminal RhoGAP domain and a C-terminal ADP-ribosylation domain . Although they share 76% amino acid identity, ExoS and ExoT ADP-ribosylate different substrates . Using protein modeling and site-directed mutagenesis, the regions of ExoS and ExoT that define substrate specificity were determined . Regions B (active site loop), C (ARTT motif) and E (PN loop) on ExoS are necessary and sufficient to recognize ExoS targets, whereas regions B, C and E on ExoT are necessary but not sufficient to recognize ExoT targets, such as the Crk proteins . A specific Crk recognition motif on ExoT was defined as region A (helix alpha1) . The electrostatic properties of regions A, B, C and E define the substrate specificity of ExoS and ExoT and these interactions can explain how other bacterial ADP-ribosylating toxins recognize their unique substrates. J Biol Chem, 2004 Oct 29, 279(44), 45791 - 802 Epub 2004 Aug 15. The heme oxygenase(s)-phytochrome system of Pseudomonas aeruginosa; Wegele R et al.; For many pathogenic bacteria like Pseudomonas aeruginosa heme is an essential source of iron . After uptake, the heme molecule is degraded by heme oxygenases to yield iron, carbon monoxide, and biliverdin . The heme oxygenase PigA is only induced under iron-limiting conditions and produces the unusual biliverdin isomers IXbeta and IXdelta . The gene for a second putative heme oxygenase in P . aeruginosa, bphO, occurs in an operon with the gene bphP encoding a bacterial phytochrome . Here we provide biochemical evidence that bphO encodes for a second heme oxygenase in P . aeruginosa . HPLC, (1)H, and (13)C NMR studies indicate that BphO is a "classic" heme oxygenase in that it produces biliverdin IXalpha . The data also suggest that the overall fold of BphO is likely to be the same as that reported for other alpha-hydroxylating heme oxygenases . Recombinant BphO was shown to prefer ferredoxins or ascorbate as a source of reducing equivalents in vitro and the rate-limiting step for the oxidation of heme to biliverdin is the release of product . In eukaryotes, the release of biliverdin is driven by biliverdin reductase, the subsequent enzyme in heme catabolism . Because P . aeruginosa lacks a biliverdin reductase homologue, data are presented indicating an involvement of the bacterial phytochrome BphP in biliverdin release from BphO and possibly from PigA. Microbes Infect, 2004 Aug, 6(10), 875 - 81 Role of alphavbeta5 integrins and vitronectin in Pseudomonas aeruginosa PAK interaction with A549 respiratory cells; Leroy-Dudal J et al.; Bacterial adherence to mammalian cells and their internalization are thought to participate in Pseudomonas aeruginosa pathogenicity . In this study, we explored the role of alpha5beta1 and alphavbeta5 integrins and their natural ligands, fibronectin (Fn) and vitronectin (Vn), in P . aeruginosa interaction with epithelial cells by using the PAK reference bacterial strain, A549 respiratory, and SKOV-3 human ovarian cell lines . The host cell cytoskeleton and cellular tyrosine kinases seem to be solicited during the PAK-respiratory cell interaction: cytochalasin D and genistein decreased the bacterial adherence and internalization . Blocking antibodies to alphavbeta5 integrins were the only antibodies tested to have inhibitory activity against PAK adherence to A549 cells . PAK internalization by A549 and SKOV-3 cells was markedly decreased in the presence of blocking antibodies to Vn and alphavbeta5 integrins . Addition of Vn in excess restored PAK invasion of both A549 and SKOV-3 cells in the presence of anti-Vn antibodies . Immunofluorescence experiments revealed that, in the presence of bacteria, the Vn fibrillar network disappeared, and alphavbeta5 staining was concentrated in sites where adherent bacteria were present . Taken together, these findings suggest that alphavbeta5 integrins, and their natural ligand Vn, are involved in PAK entry into human epithelial cells. Mol Microbiol, 2004 Aug, 53(4), 1135 - 46 Structure of the Pseudomonas aeruginosa acyl-homoserinelactone synthase LasI; Gould TA et al.; The LasI/LasR quorum-sensing system plays a pivotal role in virulence gene regulation of the opportunistic human pathogen, Pseudomonas aeruginosa . Here we report the crystal structure of the acyl-homoserine lactone (AHL) synthase LasI that produces 3-oxo-C12-AHL from the substrates 3-oxo-C12-acyl-carrier protein (acyl-ACP) and S-adenosyl-L-methionine . The LasI six-stranded beta sheet platform, buttressed by three alpha helices, forms a V-shaped substrate-binding cleft that leads to a tunnel passing through the enzyme that can accommodate the acyl-chain of acyl-ACP . This tunnel places no apparent restriction on acyl-chain length, in contrast to a restrictive hydrophobic pocket seen in the AHL-synthase EsaI . Interactions of essential conserved N-terminal residues, Arg23, Phe27 and Trp33, suggest that the N-terminus forms an enclosed substrate-binding pocket for S-adenosyl-L-methionine . Analysis of AHL-synthase surface residues identified a binding site for acyl-ACP, a role that was supported by in vivo reporter assay analysis of the mutated residues, including Arg154 and Lys150 . This structure and the novel explanation of AHL-synthase acyl-chain-length selectivity promise to guide the design of Pseudomonas aeruginosa-specific quorum-sensing inhibitors as antibacterial agents. Mol Microbiol, 2004 Aug, 53(4), 1089 - 98 A novel extracellular phospholipase C of Pseudomonas aeruginosa is required for phospholipid chemotaxis; Barker AP et al.; Pseudomonas aeruginosa and other bacterial pathogens express one or more homologous extracellular phospholipases C (PLC) that are secreted through the inner membrane via the twin arginine translocase (TAT) pathway . Analysis of TAT mutants of P . aeruginosa uncovered a previously unidentified extracellular PLC that is secreted via the Sec pathway (PlcB) . Whereas all presently known PLCs of P . aeruginosa (PlcH, PlcN and PlcB) hydrolyse phosphatidylcholine (PC), only PlcB is active on phosphatidylethanolamine (PE) . plcB candidates were identified based on deductions made from bioinformatics data and extant DNA microarray data . Among these candidates, a gene (PA0026) required for the expression of an extracellular PE-PLC was identified . The protein encoded by PA0026 has limited, but significant similarity, over a short region (approximately 60aa of 328), to a class of zinc-dependent prokaryotic PLCs . A conserved His residue of PlcB (His216) that is required for coordinate binding of zinc in this class of PLCs was mutated . Analysis of this mutant established that the protein encoded by PA0026 is PlcB . Three in-dependent recently published reports indicate that homoserine lactone-mediated quorum sensing regulates the expression of PA0026 (i.e . plcB) . PlcB, but not PlcH or PlcN, is required for directed twitching motility up a gradient of certain kinds of phospholipids . This response shows specificity for the fatty acid moiety of the phospholipid. Appl Biochem Biotechnol, 2004 Jul-Sep, 118(1-3), 243 - 51 Siderophore production by a marine Pseudomonas aeruginosa and its antagonistic action against phytopathogenic fungi; Manwar AV et al.; A marine isolate of fluorescent Pseudomonas sp . having the ability to produce the pyoverdine type of siderophores under low iron stress (up to 10 microM iron in the succinate medium) was identified as Pseudomonas aeruginosa by using BIOLOG Breathprint and siderotyping . Pyoverdine production was optimum at 0.2% (w/v) succinate, pH 6.0, in an iron-deficient medium . Studies carried out in vitro revealed that purified siderophores and Pseudomonas culture have good antifungal activity against the plant deleterious fungi, namely, Aspergillus niger, Aspergillus flavus, Aspergillus oryzae, Fusarium oxysporum, and Sclerotium rolfsii . Siderophore-based maximum inhibition was observed against A . niger . These in vitro antagonistic actions of marine Pseudomonas against phytopathogens suggest the potential of the organism to serve as a biocontrol agent. J Biol Chem, 2004 Oct 15, 279(42), 43595 - 603 Epub 2004 Aug 09. Amino acid residues involved in autophosphorylation and phosphotransfer activities are distinct in nucleoside diphosphate kinase from Mycobacterium tuberculosis; Tiwari S et al.; Nucleoside diphosphate kinase (NdK) is a ubiquitous enzyme in both prokaryotes and eukaryotes and is primarily involved in the maintenance of cellular nucleotide pools . We have cloned ndk from Mycobacterium tuberculosis strain H37Ra and expressed it in Escherichia coli as a fusion protein with glutathione S-transferase . The purified protein, following thrombin cleavage and gel permeation chromatography, was found to be hexameric with a monomeric unit molecular mass of approximately 16.5 kDa . The protein exhibited nucleotide binding, divalent cation-dependent autophosphorylation, and phosphate transfer ability from nucleoside triphosphate to nucleoside diphosphate . Although UDP inhibited the catalytic activity of the recombinant protein, the classic inhibitors, like cromoglycate, 5'-adenosine 3'-phosphate, and adenosine 3'-phosphate 5'-phosphosulfate, had no effect on the activity . Among three histidine residues in the protein, His-117 was found to be essential for autophosphorylation . However, in subsequent phosphate transfer, we observed that His-53 had a significant contribution . Consistent with this observation, substitution of His-53 with either Ala or Gln affected the ability of the recombinant protein to complement NdK function in Pseudomonas aeruginosa . Furthermore, mutational analysis established critical roles for Tyr-50 and Arg-86 of the M . tuberculosis protein in maintaining phosphotransfer ability. Chest, 2004 Aug, 126(2), 412 - 9 Lung function decline in cystic fibrosis patients and timing for lung transplantation referral; Rosenbluth DB et al.; STUDY OBJECTIVES: To determine risk factors associated with an accelerated decline in lung function in cystic fibrosis (CF), and whether longitudinal changes in FEV(1) would be a better predictor of the need for referral for lung transplantation than any single value for FEV(1.) DESIGN: The rate of decline in pulmonary function was determined by standard linear regression from each patient's calendar year's best percentage of predicted FEV(1) (%FEV(1)) over at least 4 years, and patients were classified into three cohorts based on their rate of decline . Differences between groups in age, weight-for-age z score, gender, genotype, pancreatic status, diabetes, and the presence of various lung microbial isolates were analyzed . A subset of 30 patients referred for lung transplantation were further analyzed, and a prediction model for lung transplantation referral was created using the patient's rate of decline in lung function, the mean waiting time for donor organs, and the average level of lung function of patients prior to lung transplantation . PATIENTS: One hundred fifty-three patients with CF followed up at the Washington University Adult Cystic Fibrosis Center . RESULTS: Younger age, malnutrition, and concurrent infection with both Pseudomonas aeruginosa and Staphylococcus aureus were significant (p < 0.05) risk factors for rapidly declining lung function . Among patients with rapidly declining lung function, referral for lung transplantation would have occurred 8.4 months earlier than actual referral age (p < 0.05) if the prediction model had been used, possibly resulting in additional patient salvage in several cases . CONCLUSIONS: Rate of decline in lung function should be routinely evaluated in patients with CF, and a prediction model utilizing the rate of decline in %FEV(1), and the median regional waiting period for donor lungs for patients with CF may assist in the timing of referral for lung transplantation and more rapidly declining lung function. Clin Microbiol Infect, 2004 Aug, 10(8), 705 - 8 Effect of chronic Pseudomonas aeruginosa infection on the development of atherosclerosis in a rat model; Turkay C et al.; In order to investigate the possible relationship between atherosclerosis and chronic Pseudomonas aeruginosa infection, 66 Wistar rats were given five separate intratracheal inoculations of either P . aeruginosa or sterile saline at 4-week intervals . The rats were divided into four groups: group 1 was infected with P . aeruginosa and fed a diet containing cholesterol 1% w/v; group 2 was infected with P . aeruginosa and fed a normal diet; group 3 was not infected and was fed a diet containing cholesterol 1% w/v; and group 4 (the control group) was not infected and was fed a normal diet . One month after the final inoculation, the rats were killed humanely; computerised image analysis was used to evaluate sections of the aorta and heart, and the maximal wall thickness of the aorta and coronary artery . The aortic wall thickness was significantly greater for group 1 (329.53 +/- 58.06 microm) compared to groups 2 (190.59 +/- 27.81 microm; p < 0.0001), 3 (262.90 +/- 61.12 microm; p < 0.0004) and 4 (158.00 +/- 30.30 microm; p < 0.0001) . Similarly, the coronary artery wall thickness was significantly greater for group 1 (72.96 +/- 10.67 microm) compared to groups 2 (35.07 +/- 8.53 microm; p < 0.0001), 3 (41.45 +/- 10.22 microm; p < 0.0001) and 4 (32.30 +/- 5.27 microm; p < 0.0001) . These findings strengthen the hypothesis that chronic infection plays a role in the pathogenesis of atherosclerosis. Biochemistry, 2004 Aug 17, 43(32), 10400 - 13 Real-time probing of membrane transport in living microbial cells using single nanoparticle optics and living cell imaging; Xu XH et al.; Membrane transport plays a leading role in a wide spectrum of cellular and subcellular pathways, including multidrug resistance (MDR), cellular signaling, and cell-cell communication . Pseudomonas aeruginosa is renowned for its intriguing membrane transport mechanisms, such as the interplay of membrane permeability and extrusion machinery, leading to selective accumulation of specific intracellular substances and MDR . Despite extensive studies, the mechanisms of membrane transport in living microbial cells remain incompletely understood . In this study, we directly measure real-time change of membrane permeability and pore sizes of P . aeruginosa at the nanometer scale using the intrinsic color index (surface plasmon resonance spectra) of silver (Ag) nanoparticles as the nanometer size index probes . The results show that Ag nanoparticles with sizes ranging up to 80 nm are accumulated in living microbial cells, demonstrating that these Ag nanoparticles transport through the inner and outer membrane of the cells . In addition, a greater number of larger intracellular Ag nanoparticles are observed in the cells as chloramphenicol concentration increases, suggesting that chloramphenicol increases membrane permeability and porosity . Furthermore, studies of mutants (nalB-1 and DeltaABM) show that the accumulation rate of intracellular Ag nanoparticles depends on the expression level of the extrusion pump (MexAB-OprM), suggesting that the extrusion pump plays an important role in controlling the accumulation of Ag nanoparticles in living cells . Moreover, the accumulation kinetics measured by Ag nanoparticles are similar to those measured using a small fluorescent molecule (EtBr), eliminating the possibility of steric and size effects of Ag nanoparticle probes . Susceptibility measurements also suggest that a low concentration of Ag nanoparticles (1.3 pM) does not create significant toxicity for the cells, further validating that single Ag nanoparticles (1.3 pM) can be used as biocompatible nanoprobes for the study of membrane transport kinetics in living microbial cells. South Med J, 2004 Jul, 97(7), 705 - 6 Pseudomonas sternoclavicular pyarthrosis; Kaw D et al.; Most cases of Pseudomonas pyarthrosis affecting the sternoclavicular joint have been reported in immunosuppressed intravenous drug users . We report a case of Pseudomonas pyarthrosis in a man who was otherwise immunocompetent, except for his age . A 66-year-old white man presented to the clinic with a 1-month history of right-sided shoulder and arm pain associated with swelling of the upper part of the chest in the region of the right sternoclavicular joint . The chest radiograph revealed opacity in the right superior mediastinum . Computed tomography scan of the chest confirmed a mass in the right sternoclavicular region with associated osteolysis of the clavicular head . A needle biopsy of the mass was negative for malignancy . An open biopsy specimen showed evidence of chronic inflammation without evidence of malignancy, and culture of the tissue grew Pseudomonas aeruginosa . The patient's symptoms improved after extensive incision and drainage of the affected area followed by treatment with antibiotics for 6 weeks. Niger Postgrad Med J, 2004 Jun, 11(2), 116 - 20 Profile of aerobic bacteria isolated in chronic maxillary sinusitis patients; Aneke EC et al.; OBJECTIVES: To determine the pattern of aerobic bacteria isolated in patients with chronic maxillary sinusitis at the Ear, Nose and Throat Clinic of the University of Nigeria Teaching Hospital, Enugu, and the antibiotics sensitivity pattern of these organisms . METHODS: A prospective hospital-based clinical study . RESULTS: Fifty - four patients with clinical diagnosis of chronic maxillary sinusitis were evaluated . Out of 54 maxillary sinus aspirate specimens studied, 31 yielded bacterial growths and 32 no bacterial growth . The common aerobic bacteria isolated were Staphylococcus aureus (32.3% ), Pseudomonas aeruginosa and Escherichia coli were 16.1% each . Staphylococcus aureus was sensitive to Erythromycin, and the gram-negative organisms to Gentamicin . CONCLUSION: Bacteria isolated in chronic maxillary sinusitis and their sensitivity patterns varied . Bacteriologic study of the antral washings / aspirates should be done in every patient with chronic maxillary sinusitis . Combination chemotherapy that included Erythromycin and Gentamicin was recommended. Acta Crystallogr D Biol Crystallogr, 1995 Sep, 51(Pt 5), 711 - 7 Structure of the azurin mutant nickel-Trp48Met from Pseudomonas aeruginosa at 2.2 A resolution; Tsai LC; The structure of the azurin mutant nickel-Trp48Met from Pseudomonas aeruginosa has been determined by difference Fourier synthesis using phases from the wild-type azurin model . The final crystallographic R value is 0.170 for 17 394 reflections to a resolution of 2.2 A . The mutant crystallized in the orthorhombic space group P2(1)2(1)2(1), a = 57.4, b = 80.4, c = 110.3 A . The four molecules in the asymmetric unit are packed as a dimer of dimers . The nickel metal site of this mutant structure is similar to the zinc metal site in the azurin Asp47 mutant . The site-specific mutation was performed at residue Trp48, which is located in the center of the protein in a highly hydrophobic environment, to investigate its suggested role in the long-range electron-transfer pathway between the disulfide bond on one side of the protein to the Cu centre . The structure around the mutation site Met48 showed no significant change compared with the wild-type structure. Acta Crystallogr D Biol Crystallogr, 1996 Sep, 52(Pt 5), 950 - 8 Mutant Met121Ala of Pseudomonas aeruginosa Azurin and Its Azide Derivative: Crystal Structures and Spectral Properties; Tsai LC; The crystal structures of the azurin mutant Met121Ala and its azide derivative Met121Ala-azide from Pseudomonas aeruginosa have been determined . The final crystallographic R values are 21.3 and 19.4% for the two structures, respectively . In the Met121Ala mutant, the distance between the copper ion and His117 increases by 0.34 A compared with the wild-type structure . The removal of the methionine in the apical position induces a shortening of the distance from the copper ion to the carbonyl O atom of Gly45 from 2.97 to 2.74 A . In the Met121Ala-azide structure, the azide anion occupies the cavity created by replacing the Met121 side chain with the smaller methyl group of Ala . The azide anion binds with a terminal N atom to the copper ion at a distance of about 2.04 A . In addition, the copper ion has moved out of the trigonal plane by about 0.26 A towards the azide anion . Thus, the copper site in this structure has a distorted tetrahedral arrangement . The spectroscopic characteristics show, in addition, that the copper sites in the two structures are distinctively different . The Met121Ala mutant still maintains the properties of an ordinary type 1 copper site while the Met121Ala-azide derivative has an absorption maximum at about 409 nm and the copper hyperfine coupling has increased to a value intermediate between those of type 2 copper and the wild-type azurin. Acta Crystallogr D Biol Crystallogr, 1993 Sep, 49(Pt 5), 449 - 57 Structure of Pseudomonas aeruginosai zinc azurin mutant Asn47Asp at 2.4 A resolution; Sjolin L; The Pseudomonas aeruginosa azurin mutant Asn47Asp has been isolated, its spectroscopic and kinetic properties characterized, and the X-ray crystal structure of its zinc derivative determined . While the optical and electron paramagnetic resonance spectra as well as the electron-transfer activity of the mutant are very similar to the wild-type values, the Asn47Asp reduction potential is slightly increased by 20 mV . The mutant crystallized in the orthorhombic space group P2(1)2(1)2(1) with cell dimensions a = 57.8, b = 81.5 and c = 112.6 A . There are four molecules in the asymmetric unit, packed as a tetramer which consists of two independent dimers . The zinc site of this mutant structure is similar to the wild-type zinc azurin and, in particular, the metal-binding site is almost identical to the site found in the wild-type zinc-azurin structure {Nar, Huber, Messerschmidt, Filippou, Barth, Jaquinod, Kamp & Canters (1992) . Eur . J . Biochem . 205, 1123-1129} . The Asp47 side chain at that mutation site takes on a very similar orientation to Asn47 in the wild-type structure preserving the two hydrogen bonds with the neighbouring Thr113 NH and O(gamma)H . Therefore, the increased reduction potential of the mutant is probably a result of an altered charge distribution close to the metal site. Acta Crystallogr D Biol Crystallogr, 1994 Jan, 50(Pt 1), 37 - 9 Crystallization and preliminary crystallographic data for the azurin mutant End-121 from Pseudomonas aeruginosa; Strange RW; Pseudomonas aeruginosa azurin has been crystallized from a mutant where residues from Met 121 to Lys128 have been deleted from the protein . The crystals form pale-blue well formed prisms in the orthorhombic space group P2(1)2(1)2(1), with cell dimensions a = 60.79 (5), b = 123.47 (5), c = 187.77 (5) A . The crystals diffract to 3.0 A and there are eight molecules in the asymmetric unit. Curr Microbiol, 2004 Aug, 49(2), 108 - 14 Isolation, identification, and characterization of a novel, oil-degrading bacterium, Pseudomonas aeruginosa T1; Hasanuzzaman M et al.; A novel, oil-degrading bacterium (strain T1) was isolated from a hot spring in Hokkaido, Japan . It efficiently degrades different types of fats and oils, including edible oil waste . When grown in a mineral salt medium containing 1% triacylglycerol (as salad oil), hydrolysis products were 1,3- and 1,2-diacylglycerols, monoacylglycerol, and free fatty acid . However, these products were almost completely consumed during cultivation at 30 degrees C for 5 days, indicating that extracellular lipase acts randomly at different sn-positions of acylglycerols and that strain T1 has a high capacity to utilize free fatty acids . Secreted lipase activity was induced by salad oil and oleic acid . This strain was a Gram-negative straight rod shaped, aerobic, with a polar flagellum, capable of growing in temperature ranges between 15 degrees C and 55 degrees C . The 16S rRNA gene sequence analysis and DNA-DNA hybridization revealed it as a new strain of Pseudomonas aeruginosa . The type strain was T1. J Clin Microbiol, 2004 Aug, 42(8), 3857 - 60 V-antigen genotype and phenotype analyses of clinical isolates of Pseudomonas aeruginosa; Allmond LR et al.; The pcrV genotype was analyzed in clinical isolates of Pseudomonas aeruginosa which showed a negative phenotype for secretion of V-antigen PcrV . The suppression of PcrV secretion in these isolates was due not to a lack of the pcrV gene but rather to suppression of PcrV expression. Biotechnol Prog, 2004 Jul-Aug, 20(4), 1233 - 6 Study of the attachment of Pseudomonas aeruginosa on gold and modified gold surfaces using surface plasmon resonance; Jenkins AT et al.; This paper describes how the technique of surface plasmon resonance (SPR) can be utilized to follow (in real time) the attachment of Pseudomonas aeruginosa bacteria on bare gold and gold modified with a self-assembled monolayer (SAM) of mercaptounadecanoic acid . We show that SPR is able to discriminate between the adsorption of live versus dead (thermally shocked) bacteria . Moreover, the SPR distinguishes between the adsorption of wild-type versus mutant bacteria (single gene knockouts), the concentration of the bacterial suspension, and between bacteria adsorbing on SAM-modified and bare gold . SPR is able to measure bacterial adsorption within seconds of the bacterial suspension being introduced . Finally, a qualitative correlation between results from SPR with a crystal violet staining assay for different mutant bacteria was observed. Commun Agric Appl Biol Sci, 2003, 68(2 Pt A), 207 - 10 Comparative biodegradation examination of Pseudomonas aeruginosa (ATCC 27853) and other oil degraders on hydrocarbon contaminated soil; Szoboszlay S et al.; The soil that we investigated in our experiment was extremly high contaminated with