Klebsiella

Infect Immun, 2003 Jul, 71(7), 4182 - 5
Leptin modulates neutrophil phagocytosis of Klebsiella pneumoniae; Moore SI et al.; Leptin is a pleiotropic hormone-cytokine known to regulate energy homeostasis and immune function . Neutrophils from leptin-deficient mice exhibited impaired phagocytosis of Klebsiella pneumoniae opsonized with serum containing complement and reduced CD11b expression that could be restored with exogenous leptin . These results suggest that leptin is required for normal neutrophil complement-mediated phagocytosis of bacteria.

J Am Med Dir Assoc, 2002 Mar-Apr, 3(2), 71 - 2
Rhinoscleroma in an octogenarian woman; Leibovitz A et al.; Rhinoscleroma, a granulomatous infection caused by Klebsiella rhinoscleromatis and usually afflicting young adults, was diagnosed in an 81-year-old woman . Treatment was started with orally administered antibiotics but was soon halted because of side effects . She was treated instead by local spread of a 3% rifampin ointment . After 6 weeks, the lesion cleared up and she improved remarkably . There was no recurrence during 18 months of follow-up . The current report is the second published case of rhinoscleroma in an elderly person and the first report on its cure by local treatment with a 3% rifampin ointment . This favorable experience contributes to the limited knowledge on rhinoscleroma in the elderly.

West Indian Med J, 2003 Mar, 52(1), 37 - 40
Hepatic abscesses at the University Hospital of the West Indies . A 24-year autopsy review; Gaskin DA et al.; A retrospective review of all hepatic abscesses identified at autopsy over the 24-year period 1977-2000 at the University Hospital of the West Indies (UHWI), Kingston, Jamaica, was conducted . Post mortem and microbiology records were reviewed . Data collected included age, gender, predisposing factors, organisms isolated, number of abscesses and associated conditions, such as diabetes mellitus . Data for the adults was analyzed separately . Thirty-nine cases of pyogenic abscesses were identified from 7480 post-mortems . Thirty-three occurred in adults of mean age 59.5 years . Sixty per cent of the abscesses were solitary . Biliary tract disease was the predisposing factor in 33% of cases . Six per cent were cryptogenic: 11/33 patients were diabetic and these were significantly older than non-diabetics (p < 0.014) Klebsiella pneumoniae was the most common organism isolated from diabetics (6.9) . Only 2/33 abscesses were diagnosed ante-mortem . The abscesses in children were more frequently multiple and associated with extra-abdominal infection . Gram positive cocci were the commonly isolated organisms in children . There were no cases of amoebic abscess . The prevalence of hepatic abscess was low . Diabetes mellitus was a significant contributing factor . A high index of clinical suspicion is therefore warranted particularly in elderly diabetics.

FEBS Lett, 2003 Jun 12, 545(1), 9 - 17
Proton pumping by NADH:ubiquinone oxidoreductase . A redox driven conformational change mechanism?
Brandt U, Kerscher S, Drose S, Zwicker K, Zickermann V.
The modular evolutionary origin of NADH:ubiquinone oxidoreductase (complex I) provides useful insights into its functional organization . Iron-sulfur cluster N2 and the PSST and 49 kDa subunits were identified as key players in ubiquinone reduction and proton pumping . Structural studies indicate that this 'catalytic core' region of complex I is clearly separated from the membrane . Complex I from Escherichia coli and Klebsiella pneumoniae was shown to pump sodium ions rather than protons . These new insights into structure and function of complex I strongly suggest that proton or sodium pumping in complex I is achieved by conformational energy transfer rather than by a directly linked redox pump.

Biosci Biotechnol Biochem, 2003 Apr, 67(4), 933 - 6
Microbial production of 2-deoxyribose 5-phosphate from acetaldehyde and triosephosphate for the synthesis of 2'-deoxyribonucleosides; Ogawa J et al.; 2-Deoxyribose 5-phosphate was produced from acetaldehyde and dihydroxyacetone phosphate via D-glyceraldehyde 3-phosphate by Klebsiella pneumoniae B-4-4 through deoxyriboaldolase- and triosephosphate isomerase-catalyzing reactions . Under the optimum conditions, 98.7 mM 2-deoxyribose 5-phosphate was produced from 200 mM acetaldehyde and 117 mM dihydroxyacetone phosphate in 2 h with a molar yield of 84% . The 2-deoxyriobse 5-phosphate produced was directly transformed to 2'-deoxyribonucleoside by phosphopentomutase- and nucleoside phosphorylase-catalyzing reactions.

Int J Immunopathol Pharmacol, 1999 May, 12(2), 103 - 111
Effect of bacterial extracts on the immunologic profile in chronic relapsing brucellosis patients; Boura P et al.; Brucellosis is an intracellular bacterial disease of common incidence in Greece . Existing therapy is inadequate and a considerable proportion of patients become chronically ill and are immunocompromised . Defects of the monocyte-macrophage system and T-lymphocytes have been described in chronic brucellosis and can be restored after immunopotentiation therapy . Bacterial (Klebsiella pneumoniae) extracts exert immunostimulating effects on the monocyte-macrophage system and have already been used successfully in the prevention of common infections of the respiratory track . So we decided to investigate: 1) Leukocyte Migration Index (LMI), 2) Monocyte-macrophage random and directed migration against both nonspecific leukoattractant (casein) and disease specific antigens (Brucella melitensis, Brucella abortus), 3) Monocyte-macrophage phagocytosis index, 4) Delayed-type hypersensitivity (skin tests) against seven antigens, before (TO), during (T2), and after (T3) oral administration of bacterial (Klebsiella pneumoniae) extracts at conventional doses plus antibiotics or not . Our results show that: 1) Concerning the LMI, 4 out of 19 remained anergic at time T3 of the study, 2) Random migration was not affected during treatment, 3) Directed migration increased significantly without reaching control group values, 4) Phagocytosis index increased significantly and reached normal values at T3, 5) Delayed type hypersensitivity reactions (skin tests) increased significantly at the end of the study period . Reaction against Tuberculin and Candida antigens showed the most pronounced increase in skin reactivity . In conclusion, bacterial (Klebsiella pneumoniae) extracts improve peripheral monocyte locomotion and restore phagocytosis index, thus enhancing cellular immunity parameters in immunocompromised chronic brucellosis patients.

J Formos Med Assoc, 2003 Mar, 102(3), 189 - 92
Therapeutic effect of interferon-gamma for prevention of severe infection in X-linked chronic granulomatous disease; Ma HR et al.; Chronic granulomatous disease (CGD) is an inherited phagocytic disorder caused by defective oxidative burst activity of neutrophils and monocytes . Patients with CGD may present with recurrent, life-threatening bacterial or fungal infections and often need repeated hospitalization as early as infancy . We report a case of a boy aged 3 years and 1 month with a history of oral thrush, chronic diarrhea, skin abscesses, multiple small joints osteomyelitis, and multiple liver abscesses since 2 months of age . X-linked chronic granulomatous disease was diagnosed by nitroblue tetrazolium test and further confirmed by genetic study using single strand conformation polymorphism (SSCP) analysis and genomic DNA sequencing . Two episodes of aspergillosis with severe aspergillus lung empyema and life-threatening Klebsiella pneumoniae infections were noted in the first 2 years of life despite long-term prophylaxis with trimethoprim-sulfamethoxazole and itraconazole . Recombinant human interferon-gamma (rINF-gamma) 50 microg/m(2) subcutaneous injection 3 times per week was added after his last episode of severe infection . Thereafter, the boy had normal growth and development with no evidence of severe infection during 18 months of follow-up at our outpatient clinic.

Klin Monatsbl Augenheilkd, 2003 May, 220(5), 357 - 61
{Long-term follow-up of bilateral endogenous Klebsiella endophthalmitis}; Holak H et al.; BACKGROUND: Endophthalmitis subsequent to Klebsiella sepsis leads to functional blindness in most cases and is very difficult to treat . Every successful therapeutic modality can therefore help in creating an optimal therapeutic plan . CASE REPORT: A 69-year old diabetic patient exhibited bilateral Klebsiella endophthalmitis with sepsis after a pneumonia . Two intravenous antibiotics were used: aminoglycosides (Gentamycin) and cephalosporins (Cefotaxim or Cefuroxim) with local parabulbar injections of Prednisolon . The long-term follow-up of four years provided some overview of morphological aspects of the development of endophthalmitis . Characteristic greyish hypopyon was seen in both eyes, which was more pronounced in the left eye than in the right . The left eye became phthisic . After resorption of the hypopyon in the right eye and prolonged resorption of the subretinal abscess for 9 months a useful visual acuity at 0.2 was achieved . Two years after the endophthalmitis a cataract surgery with implantation of a posterior chamber silicon lens was performed and good visual acuity (0.6) was achieved . After four years, the subretinal abscess left an extremely large, sharp bordered, unpigmented scar up to the sclera . CONCLUSION: An early diagnosis and adequate long-time antibiotic therapy under the co-operative supervision of an ophthalmologist with internist appears to be most important for the therapeutic success in Klebsiella endophthalmitis.

Zhonghua Er Bi Yan Hou Ke Za Zhi, 2001 Feb, 36(1), 42 - 3
{Clinicopathologic analysis of rhinoscleroma}; Zhang S et al.; OBJECTIVE: To investigate the clinicopathologic characteristics and reason of tissue injury caused by klebsiella rhinoscleromatis(KR) infection . METHODS: Twenty-four cases of rhinoscleromas in the nasal and pharyngeal region in this hospital from March 1983 to March 1998 were studied retrospectively . Warthin-Starry (W-S) stain was used in all cases and CD43, CD20, CD68, Lysozyme mark were observed in eleven specimens by ABC immunochemical method . Two cases were studied further with transmission electron microscopy . RESULTS: A great number of KR were found in Mikulicz cells and were confirmed further in the phagosomes in cytoplasm of Mikulicz cells, where a small number of endoplasmic reticulums and lysozymes was squeezed to the side of cells . A few of T-lymphocytes was found within the granuloma . CONCLUSION: Cellular immunity function of the patient with rhinoscleroma was depressed and the response to immunization of host which regards to KR was the major cause of tissue injury.

FEMS Microbiol Lett, 2003 May 16, 222(1), 93 - 8
Sequence analysis and biochemical characterisation of chromosomal CAV-1 (Aeromonas caviae), the parental cephalosporinase of plasmid-mediated AmpC 'FOX' cluster; Fosse T et al.; Aeromonas caviae CIP 74.32 was resistant to amoxicillin, ticarcillin and cephalothin, and susceptible to cefoxitin, cefotaxime, ceftazidime, aztreonam and imipenem . This strain produced a cephalosporinase (pI 7.2) and an oxacillinase (pI 8.5) . The cephalosporinase gene cav-1 was cloned and sequenced . Unlike A . caviae donor, Escherichia coli pNCE50 transformant producing CAV-1 beta-lactamase was resistant to cefoxitin . The deduced protein sequence CAV-1 contained 382 amino acids, and shared >96% homology with FOX-1 to FOX-5 cephalosporinase . CAV-1 presented only two amino acid substitutions (Thr270Ser and Arg271Ala) with FOX-1 . CAV-1 is the chromosomal putative ancestor of the FOX family, a cluster of class C/group 1 plasmidic cephalosporinases spreading in Klebsiella and E . coli clinical isolates via conjugative plasmids.

J Bacteriol, 2003 Jun, 185(11), 3416 - 28
Type IV-like pili formed by the type II secreton: specificity, composition, bundling, polar localization, and surface presentation of peptides; Vignon G et al.; The secreton or type II secretion machinery of gram-negative bacteria includes several type IV pilin-like proteins (the pseudopilins) that are absolutely required for secretion . We previously reported the presence of a bundled pilus composed of the pseudopilin PulG on the surface of agar-grown Escherichia coli K-12 cells expressing the Klebsiella oxytoca pullulanase (Pul) secreton genes at high levels (N . Sauvonnet, G . Vignon, A . P . Pugsley, and P . Gounon, EMBO J . 19:2221-2228, 2000) . We show here that PulG is the only pseudopilin in purified pili and that the phenomenon is not restricted to the Pul secreton reconstituted in E . coli or to PulG . For example, high-level expression of the endogenous E . coli gsp secreton genes caused production of bundled pili composed of the pseudopilin GspG, and the Pul secreton was able to form pili composed of PulG-like proteins from secreton systems of other bacteria . PulG derivatives in which the C terminus was extended by the addition of eight different peptides were also assembled into pili and functioned in secretion . Three of the C-terminal peptides were shown to be exposed along the entire length of the assembled pili . Hence, the C terminus of PulG may represent a permissive site for the insertion of immunogenic epitopes or other peptide sequences . One of these PulG variants, with a six-histidine tag at its C terminus, formed nonpolar, nonbundled pili, suggesting that bundle formation and polar localization are not correlated with the ability of PulG to function in secretion . We propose that the PulG pilus is an artifactual manifestation of a periplasmic "pseudopilus" and that cycles of pseudopilus extension and retraction within the periplasm propel pullulanase through secretin channels in the outer membrane . Abnormally long pili that extend beyond the outer membrane are produced only when pilus length control and retraction are deregulated by overproduction of the major pseudopilus subunit (PulG).

Natl Toxicol Program Tech Rep Ser, 1986 Oct, 257, 1 - 222
NTP Toxicology and Carcinogenesis Studies of Diglycidyl Resorcinol Ether (Technical Grade) (CAS No . 101-90-6) In F344/N Rats and B6C3F1 Mice (Gavage Studies); National Toxicology Program ; Diglycidyl resorcinol ether (DGRE), a pale, yellow, translucent, amorphous solid at room temperature, is used as a liquid spray epoxy resin, as a diluent in the production of other epoxy resins used in electrical, tooling, adhesive, and laminating applications, and as a curing agent for polysulfide rubber . Approximately 3,000 workers are exposed to DGRE . The quantity of DGRE produced in the United States is not known . Toxicology and carcinogenesis studies of technical grade diglycidyl resorcinol ether (81% pure) were conducted by administering the chemical in corn oil by gavage to groups of 50 male and 50 female F344/N rats at doses of 25 or 50 mg/kg and to groups of 50 male and female B6C3F1 mice at doses of 50 or 100 mg/kg . A supplemental study of similar design in male and female rats (0 or 12 mg/kg) was started approximately 12 months later because of high mortality in the 50 mg/kg dose groups . Doses were administered five times per week for 103 weeks . Groups of 50 rats and 50 mice of each sex received corn oil by gavage on the same dosing schedule and served as vehicle controls . Throughout most of the primary study, mean body weights of high dose male and female rats and female mice were lower than those of the corresponding vehicle controls . In the supplemental study, body weights of both sexes of the dosed rats were unaffected by administration of DGRE . Survival of dosed rats of each sex in the primary study was dose related and was shorter (P<0.001) than that of the vehicle controls . No high dose male rats and only 1/50 high dose female rats lived to the end of the study . Bronchopneumonia was the most frequent cause of early death among the rats and may have resulted from the animals' aspiration of corn oil containing diglycidyl resorcinol ether . Survival of the dosed male rats in the supplemental study was reduced (P<0.005) when compared to controls . There was no significant difference in survival between dosed and control female rats in the supplemental study . Survival of dosed and control mice was comparable but poorer in females, with 20/50 (40%) of the controls, 13/50 (26%) of the low dose, and 10/50 (20%) of the high dose groups alive at the end of 2 years . These early deaths were due to suppurative and necrotizing inflammation of the reproductive tract, possibly caused by a Klebsiella sp . infection . The incidences of rats and mice with hyperkeratosis and hyperplasia of the forestomach were compound related . For rats and mice of each sex, incidences of animals with squamous cell papillomas, squamous cell carcinomas, or both occurred with statistically significant positive trends and the incidences observed in other organs in dosed groups relative to the controls . An audit of the experimental data was conducted for the 2-year studies of diglycidyl resorcinol ether . No data discrepancies were found that influenced the final interpretations . Under the conditions of these 2-year gavage studies, technical grade diglycidyl resorcinol ether caused hyperkeratosis and hyperplasia of the forestomach in rats and mice . DGRE was carcinogenic for male and female F344/N rats and for male and female B6C3F1 mice, causing both benign and malignant neoplasms of the forestomach . Levels of Evidence of Carcinogenicity: Male Rats: Positive Female Rats: Positive Male Mice: Positive Female Mice: Positive Synonym: DGRE

Int J Food Microbiol, 2003 Jun 25, 83(3), 295 - 305
The effect of abrupt shifts in temperature on the lag phase duration of Escherichia coli and Klebsiella oxytoca; Mellefont LA et al.; The effect of temperature of incubation on lag times of two gram-negative foodborne bacteria was investigated . Bacteria were instantaneously transferred between temperatures within and beyond the normal physiological temperature range (NPTR) . Abrupt temperature shifts induced lag phases, but the degree of the response was dependent on the direction and magnitude of the shift . Temperature downshifts induced larger relative lag times (RLT; the ratio of lag time to generation time), than equivalent upshifts . The hypothesis of Robinson et al . {Int . J . Food Microbiol . 44 (1998) 83} that lag time can be understood in terms of the amount of work to be done to adjust to new environmental conditions and the rate at which that work is done was supported . Deviation of the reported proportionality between lag time and generation time was observed when late-exponential phase cells were subjected to abrupt temperature shifts from beyond the normal physiological range.

West Afr J Med, 2002 Jul-Sep, 21(3), 252 - 5
Klebsiella-induced purpura fulminans in a Nigerian child: case report and a review of literature; Olowu WA; Purpura fulminans (PPF) is a very severe but rare acute thrombohaemorrhagic illness of infants and young children . It occurs mainly, in patients with either congenital or acquired deficiencies of proteins C and S and antithrombin III . Features of PPF include disseminated intravascular coagulopathy, symmetrical necrotic purpura and/or ecchymoses and symmetrical peripheral gangrene; digital and/ or limb(s) amputations and end-organ failure(s) may also occur . The case of a 3.5 year-old Nigerian girl, who developed PPF following Klebsiella-rhinoscleromatis septicaemia is reported to illustrate the seriousness of the disease and the need for early diagnosis and management.

Biochemistry, 2003 May 20, 42(19), 5917 - 24
Identification of basic amino acid residues important for citrate binding by the periplasmic receptor domain of the sensor kinase CitA; Gerharz T et al.; The sensor kinase CitA and the response regulator CitB of Klebsiella pneumoniae form the paradigm of a subfamily of bacterial two-component regulatory systems that are capable of sensing tri- or dicarboxylates in the environment and then induce transporters for the uptake of these compounds . We recently showed that the separated periplasmic domain of CitA, termed CitAP (encompasses residues 45-176 supplemented with an N-terminal methionine residue and a C-terminal hexahistidine tag), is a highly specific citrate receptor with a K(d) of 5.5 microM at pH 7 . To identify positively charged residues involved in binding the citrate anion, each of the arginine, lysine, and histidine residues in CitAP was exchanged for alanine, and the resulting 17 muteins were analyzed by isothermal titration calorimetry (ITC) . In 12 cases, the K(d) for citrate was identical to that of wild-type CitAP or slightly changed (3.9-17.2 microM) . In one case (R98A), the K(d) was 6-fold decreased (0.8 microM), whereas in four cases (R66A, H69A, R107A, and K109A) the K(d) was 38- to >300-fold increased (0.2 to >1 mM) . The secondary structure of the latter five proteins in their apo-form as deduced from far-UV circular dichroism (CD) spectra did not differ from the apo-form of wild-type CitAP; however, all of them showed an increased thermostability . Citrate increased the melting point (T(m)) of wild-type CitAP and mutein R98A by 6.2 and 9.5 degrees C, respectively, but had no effect on the T(m) of the four proteins with disturbed binding . Three of the residues important for citrate binding (R66, H69, and R107) are highly conserved in the CitA subfamily of sensor kinases, indicating that they might be involved in ligand binding by many of these sensor kinases.

Salud Publica Mex, 2003 Mar-Apr, 45(2), 90 - 5
{Neonatal sepsis morbidity and mortality in a tertiary care hospital.}; Rodriguez-Weber MA et al.; OBJECTIVE: To compare the epidemiological, clinical and microbiological profiles between patients with neonatal sepsis who lived or died . MATERIAL AND METHODS: The medical records of patients with neonatal sepsis were retrospectively reviewed at Instituto Nacional de Pediatria (National Pediatric Institute) of Secretaria de Salud (Ministry of Health) in Mexico City, between 1992 and 2000 . Neonatal sepsis cases were classified as surviving or not after 90 days of postnatal follow-up . The survivor and decreased groups were compared using Mann-Whitney's U test for continuous variables, and the chi-squared test or the Fisher's exact test for categorical variables . Significantly associated variables were included in a Cox proportional hazards model . A p-value < 0.05 was considered statistically significant for all analyses . RESULTS: A total of 116 patients with neonatal sepsis were included (65 live and 51 dead) . Multivariate analysis showed that fetal distress, respiratory distress, a delayed capillary fill up, a low platelet count, and a positive hemoculture for Klebsiella pneumoniae were significant risk factors for death . CONCLUSIONS: Epidemiological, clinical, laboratory, and microbiological variables are significant predictors of death in newborns with neonatal sepsis . The English version of this paper is available at: http://www.insp.mx/salud/index.html.

Curr Microbiol, 2003 Jun, 46(6), 423 - 31
Altered kinetic properties of tyrosine-183 to cysteine mutation in glutamine synthetase of anabaena variabilis strain SA1 is responsible for excretion of ammonium ion produced by nitrogenase; Healy FG et al.; A L-methionine- D, L-sulfoximine-resistant mutant of the cyanobacterium Anabaena variabilis, strain SA1, excreted the ammonium ion generated from N(2) reduction . In order to determine the biochemical basis for the NH(4)(+)-excretion phenotype, glutamine synthetase (GS) was purified from both the parent strain SA0 and from the mutant . GS from strain SA0 (SA0-GS) had a pH optimum of 7.5, while the pH optimum for GS from strain SA1 (SA1-GS) was 6.8 . SA1-GS required Mn(+2) for optimum activity, while SA0-GS was Mg(+2) dependent . SA0-GS had the following apparent K(m) values at pH 7.5: glutamate, 1.7 m M; NH(4)(+), 0.015 m M; ATP, 0.13 m M . The apparent K(m) for substrates was significantly higher for SA1-GS at its optimum pH (glutamate, 9.2 m M; NH(4)(+), 12.4 m M; ATP, 0.17 m M) . The amino acids alanine, aspartate, cystine, glycine, and serine inhibited SA1-GS less severely than the SA0-GS . The nucleotide sequences of glnA (encoding glutamine synthetase) from strains SA0 and SA1 were identical except for a single nucleotide substitution that resulted in a Y183C mutation in SA1-GS . The kinetic properties of SA1-GS isolated from E . coli or Klebsiella oxytoca glnA mutants carrying the A . variabilis SA1 glnA gene were also similar to SA1-GS isolated from A . variabilis strain SA1 . These results show that the NH(4)(+)-excretion phenotype of A . variabilis strain SA1 is a direct consequence of structural changes in SA1-GS induced by the Y183C mutation, which elevated the K(m) values for NH(4)(+) and glutamate, and thus limited the assimilation of NH(4)(+) generated by N(2) reduction . These properties and the altered divalent cation-mediated stability of A . variabilis SA1-GS demonstrate the importance of Y183 for NH(4)(+) binding and metal ion coordination.

Fukuoka Igaku Zasshi, 2003 Feb, 94(2), 31 - 6
Endophthalmitis with Klebsiella pneumoniae liver abscess; Dohmen K et al.; Endogenous endophthalmitis is a rare, but devastating complication of septicemia . The prognosis of maintaining visual acuity in patients with septic endophthalmitis is poor in spite of an early diagnosis and the timely start of conventional therapeutic procedures because the intravitreous drug concentration remains low after the systemic administration of antibiotics due to the blood-ocular barrier . We treated an elderly female patient with endogenous endophthalmitis complicated with disseminated intravascular coagulation associated with a Klebsiella pneumoniae liver abscess . Endophthalmitis developed rapidly and we thus had to perform an enucleation of both eyeballs even though we made an early diagnosis and performed liver abscess drainage as well as the prompt systemic and subconjunctival administration of antibiotics . Our experience in treating this case emphasizes the need to perform the timely intravitreous infusion of antibiotics with a support therapy consisting of the systemic and subconjunctival administration of antibiotics for endogenous endophthalmitis associated with a Klebsiella pneumoniae liver abscess.

J Immunol, 2003 May 1, 170(9), 4432 - 6
Cutting edge: roles of Toll-like receptor 4 and IL-23 in IL-17 expression in response to Klebsiella pneumoniae infection; Happel KI et al.; Local production of IL-17 is a significant factor in effective host defense against Gram-negative bacteria . However, the proximal events mediating IL-17 elaboration by T cells remain unclear . In this study, we show in vivo that intact Toll-like receptor 4 signaling in the lung is required for induction of both the p19 transcript of IL-23 and IL-17 protein elaboration in response to Klebsiella pneumoniae . Although IL-17 is widely considered a CD4(+) T cell product, we also demonstrate significant in vitro IL-17 production by CD8(+) T cells after culture in medium from dendritic cells exposed to these bacteria . The dominant portion of this IL-17-inducing activity for both CD4(+) and CD8(+) T cells is IL-23 . These data demonstrate the critical signaling pathway for IL-17 induction in the host response to Gram-negative pulmonary infection and suggest a direct role for IL-23 in CD8(+) T cell IL-17 production.

Carbohydr Res, 2003 May 1, 338(10), 1033 - 7
First synthesis of beta-D-Galf-(1-->3)-D-Galp--the repeating unit of the backbone structure of the O-antigenic polysaccharide present in the lipopolysaccharide (LPS) of the genus Klebsiella; Wang H et al.; beta-D-Galactofuranosyl-(1-->3)-D-galactopyranose (1), the repeating unit of the backbone structure of the O-antigenic polysaccharide present in the lipopolysaccharide (LPS) of the genus Klebsiella, has been efficiently synthesized using 1,2:5,6-di-O-isopropylidine-alpha-D-galactofuranose (3) as the glycosyl acceptor and 2,3,5,6-tetra-O-benzoyl-beta-D-galactofuranosyl trichloroacetimidate (6) as the glycosyl donor with TMSOTf as catalyst by the well-known Schmidt glycosylation method . The preparation of 3 was improved by increasing the ratio of DMF to acetone and employing a solid-supported catalyst.

J Bacteriol, 2003 May, 185(9), 2920 - 6
Nitrogen regulation of the codBA (cytosine deaminase) operon from Escherichia coli by the nitrogen assimilation control protein, NAC; Muse WB et al.; Transcription of the cytosine deaminase (codBA) operon of Escherichia coli is regulated by nitrogen, with about three times more codBA expression in cells grown in nitrogen-limiting medium than in nitrogen-excess medium . Beta-galactosidase expression from codBp-lacZ operon fusions showed that the nitrogen assimilation control protein NAC was necessary for this regulation . In vitro transcription from the codBA promoter with purified RNA polymerase was stimulated by the addition of purified NAC, confirming that no other factors are required . Gel mobility shifts and DNase I footprints showed that NAC binds to a site centered at position -59 relative to the start site of transcription and that mutants that cannot bind NAC there cannot activate transcription . When a longer promoter region (positions -120 to +67) was used, a double footprint was seen with a second 26-bp footprint separated from the first by a hypersensitive site . When a shorter fragment was used (positions -83 to +67), only the primary footprint was seen . Nevertheless, both the shorter and longer fragments showed NAC-mediated regulation in vivo . Cytosine deaminase expression in Klebsiella pneumoniae was also regulated by nitrogen in a NAC-dependent manner . K . pneumoniae differs from E . coli in having two cytosine deaminase genes, an intervening open reading frame between the codB and codA orthologs, and a different response to hypoxanthine which increased cod expression in K . pneumoniae but decreased it in E . coli.

J Appl Physiol, 2003 Aug, 95(2), 491 - 6 Epub 2003 Apr 11.
Active hexose correlated compound enhances resistance to Klebsiella pneumoniae infection in mice in the hindlimb-unloading model of spaceflight conditions; Aviles H et al.; Previous studies have demonstrated that resistance to infection is decreased in Swiss Webster female mice maintained in the hindlimb-unloading model (Aviles H, Belay T, Fountain K, Vance M, and Sonnenfeld G . J Appl Physiol 95: 73-80, 2003; Belay T, Aviles H, Vance M, Fountain K, and Sonnenfeld G . J Allergy Clin Immunol 110: 262-268, 2002) . This is a model of some of the aspects of spaceflight conditions, including lack of load bearing on hindlimbs and a fluid shift to the head . Active hexose correlated compound (AHCC), extracted from Basidiomycete mushrooms, has been shown to induce enhancement of immune responses, including enhanced natural killer activity . In the present study, AHCC was orally administered to mice to determine whether the treatment could decrease immunosuppression and mortality of mice maintained in the hindlimb-unloaded model and infected with Klebsiella pneumoniae . The results of the present study showed that administration of AHCC by gavage for 1 wk (1 g/kg body wt) before suspension and throughout the 10-day suspension period yielded significant beneficial effects for the hindlimb-unloaded group, including 1) . decreased mortality, 2) . increased time to death, and 3) . increased ability to clear bacteria . The results suggest that AHCC can decrease the deleterious effects of the hindlimb-unloading model on immunity and resistance to infection.

Conn Med, 2003 Mar, 67(3), 149 - 52
Utilization of extended-spectrum beta-lactamase (ESBL) detection systems in microbiology laboratories: survey of Connecticut hospitals from 1998-2002; Dandekar PK et al.; There has been recent concern raised over the detection, prevalence, and clinical implications of infection with Escherichia coli and Klebsiella spp . which produce extended-spectrum beta-lactamases (ESBLs) . These enzymes hydrolyze beta-lactams including the cephalosporins (i.e., ceftazidime, ceftriaxone, cefotaxime), thus frequently leading to treatment failures . Standard in vitro testing may report these isolates as susceptible when in fact they are resistant in vivo . As a result of this phenomenon, additional testing for suspected isolates is recommended nationally . We surveyed 28 Connecticut hospitals from 1998-2002 to determine if these institutions utilized screening and confirmation methods for suspected isolates . The number of hospitals which have implemented ESBL detection systems doubled from 11 to 22 over the study period . Currently, 15 of the 22 laboratories conduct both screening and confirmatory testing . This expanded testing will be of great assistance to clinicians in optimizing the clinical care of patients with Gramnegative infections.

J Biol Chem, 2003 Jun 20, 278(25), 22717 - 25 Epub 2003 Apr 08.
Structural rationalization for the lack of stereospecificity in coenzyme B12-dependent diol dehydratase; Shibata N et al.; Adenosylcobalamin-dependent diol dehydratase of Klebsiella oxytoca is apparently not stereospecific and catalyzes the conversion of both (R)- and (S)-1,2-propanediol to propionaldehyde . To explain this unusual property of the enzyme, we analyzed the crystal structures of diol dehydratase in complexes with cyanocobalamin and (R)- or (S)-1,2-propanediol . (R)- and (S)-isomers are bound in a symmetrical manner, although the hydrogen-bonding interactions between the substrate and the active-site residues are the same . From the position of the adenosyl radical in the modeled "distal" conformation, it is reasonable for the radical to abstract the pro-R and pro-S hydrogens from (R)- and (S)-isomers, respectively . The hydroxyl groups in the substrate radicals would migrates from C(2) to C(1) by a suprafacial shift, resulting in the stereochemical inversion at C(1) . This causes 60 degrees clockwise and 70 degrees counterclockwise rotations of the C(1)-C(2) bond of the (R)- and (S)-isomers, respectively, if viewed from K+ . A modeling study of 1,1-gem-diol intermediates indicated that new radical center C(2) becomes close to the methyl group of 5'-deoxyadenosine . Thus, the hydrogen back-abstraction (recombination) from 5'-deoxyadenosine by the product radical is structurally feasible . It was also predictable that the substitution of the migrating hydroxyl group by a hydrogen atom from 5'-deoxyadenosine takes place with the inversion of the configuration at C(2) of the substrate . Stereospecific dehydration of the 1,1-gem-diol intermediates can also be rationalized by assuming that Asp-alpha335 and Glu-alpha170 function as base catalysts in the dehydration of the (R)- and (S)-isomers, respectively . The structure-based mechanism and stereochemical courses of the reaction are proposed.

Infect Control Hosp Epidemiol, 2003 Mar, 24(3), 172 - 9
Use of an alcohol-based hand rub and quality improvement interventions to improve hand hygiene in a Russian neonatal intensive care unit; Brown SM et al.; BACKGROUND: Hand hygiene (HH) is critical to infection control, but compliance is low . Alcohol-based antiseptics may improve HH . HH practices in Russia are not well described, and facilities are often inadequate . SETTING: Four 6-bed units in a neonatal intensive care unit in St . Petersburg, Russia . METHODS: Prospective surveillance of HH compliance, nosocomial colonization, and antibiotic administration was performed from January until June 2000 . In February 2000, alcohol-based hand rub was provided for routine HH use . Eight weeks later, a quality improvement intervention was implemented, consisting of review of interim data, identification of opinion leaders, posting of colonization incidence rates, and regular feedback . Means of compliance, colonization, and antibiotic use were compared for periods before and after each intervention . RESULTS: A total of 1,027 events requiring HH were observed . Compliance was 44.2% before the first intervention, 42.3% between interventions, and 48% after the second intervention . Use of alcohol rose from 15.2% of HH indications to 25.2% between interventions and 41.5% after the second intervention . The incidence of nosocomial colonization (per 1,000 patient-days) with Klebsiella pneumoniae was initially 21.5, decreased to 4.7, and then was 3.2 in the final period . Rates of antibiotic and device use also decreased . CONCLUSIONS: HH may have increased slightly, but the largest effect was a switch from soap and water to alcohol which may have been associated with decreased cross-transmission of Klebsiella, although this may have been confounded by lower device use . Alcohol-based antiseptic may be an improvement over current practices, but further research is required.

J Antimicrob Chemother, 2003 Apr, 51(4), 957 - 62 Epub 2003 Feb 25.
Clinical features and in vitro antimicrobial susceptibilities of community-acquired Klebsiella pneumoniae meningitis in Taiwan; Lee PY et al.; Twenty-seven adult patients were identified as having community-acquired Klebsiella pneumoniae meningitis . The K . pneumoniae isolates, collected from cerebrospinal fluid samples, were tested for in vitro antimicrobial susceptibilities . The prognostic factors of these 27 patients were also analysed . All of the third- and fourth-generation cephalosporins tested, as well as monobactam, carbapenem and ciprofloxacin, had good activities against the isolated K . pneumoniae strains . None of the clinical isolates was detected as being an extended-spectrum beta-lactamase-producing pathogen . Among the third- and fourth-generation cephalosporins, ceftizoxime, cefepime, ceftriaxone and cefotaxime had superior activities, with MIC90s about four- to eight-fold lower than those of ceftazidime and moxalactam . Mortality rates of patients classified by different antimicrobial agents were as follows: ceftazidime 38% (8/21) and cefepime 16.7% (1/6) . The presence of septic shock and the initial level of consciousness at the start of appropriate antimicrobial therapy were the major determinants of survival and neurological outcomes in these 27 patients . Early diagnosis and choice of appropriate antibiotics according to antimicrobial susceptibilities may improve therapeutic outcomes.

Am J Respir Cell Mol Biol, 2003 Apr, 28(4), 443 - 50
Hyperoxia impairs antibacterial function of macrophages through effects on actin; O'Reilly PJ et al.; Oxidative stress may impair alveolar macrophage function in patients with inflammatory lung diseases or those exposed to high concentrations of oxygen . We investigated putative mechanisms of injury to macrophages by oxidative stress, using RAW 264.7 cells exposed to 95% oxygen for 48 h . Hyperoxia-exposed macrophages were less able to phagocytose and kill Klebsiella pneumoniae than normoxic controls, despite increased production of nitric oxide, a free radical important in pathogen killing . Exposure of macrophages to hyperoxia had marked effects on the actin cytoskeleton, including increased actin polymerization, loss of cortical actin, formation of stress fibers, de novo synthesis of actin, and actin oxidation . Hyperoxia induced changes in cell morphology, with increased cell size and pseudopod formation . Exposure of macrophages to jasplakinolide, an agent that increases actin polymerization, also impaired their ability to phagocytose Klebsiella . Alveolar macrophages isolated from mice exposed to 100% oxygen for 84 h also demonstrated impaired phagocytic function, as well as similar effects on the actin cytoskeleton and cell morphology to macrophages exposed to hyperoxia in vitro . We conclude that oxidative stress in vitro and in vivo impairs macrophage antibacterial function through effects on actin.

Eur J Biochem, 2003 Apr, 270(7), 1555 - 66
Oxygen control of nif gene expression in Klebsiella pneumoniae depends on NifL reduction at the cytoplasmic membrane by electrons derived from the reduced quinone pool; Grabbe R et al.; In Klebsiella pneumoniae, the flavoprotein, NifL regulates NifA mediated transcriptional activation of the N2-fixation (nif) genes in response to molecular O2 and ammonium . We investigated the influence of membrane-bound oxidoreductases on nif-regulation by biochemical analysis of purified NifL and by monitoring NifA-mediated expression of nifH'-'lacZ reporter fusions in different mutant backgrounds . NifL-bound FAD-cofactor was reduced by NADH only in the presence of a redox-mediator or inside-out vesicles derived from anaerobically grown K . pneumoniae cells, indicating that in vivo NifL is reduced by electrons derived from membrane-bound oxidoreductases of the anaerobic respiratory chain . This mechanism is further supported by three lines of evidence: First, K . pneumoniae strains carrying null mutations of fdnG or nuoCD showed significantly reduced nif-induction under derepressing conditions, indicating that NifL inhibition of NifA was not relieved in the absence of formate dehydrogenase-N or NADH:ubiquinone oxidoreductase . The same effect was observed in a heterologous Escherichia coli system carrying a ndh null allele (coding for NADH dehydrogenaseII) . Second, studying nif-induction in K . pneumoniae revealed that during anaerobic growth in glycerol, under nitrogen-limitation, the presence of the terminal electron acceptor nitrate resulted in a significant decrease of nif-induction . The final line of evidence is that reduced quinone derivatives, dimethylnaphthoquinol and menadiol, are able to transfer electrons to the FAD-moiety of purified NifL . On the basis of these data, we postulate that under anaerobic and nitrogen-limited conditions, NifL inhibition of NifA activity is relieved by reduction of the FAD-cofactor by electrons derived from the reduced quinone pool, generated by anaerobic respiration, that favours membrane association of NifL . We further hypothesize that the quinol/quinone ratio is important for providing the signal to NifL.

J Bacteriol, 2003 Apr, 185(7), 2267 - 76
Mutations which uncouple transport and phosphorylation in the D-mannitol phosphotransferase system of Escherichia coli K-12 and Klebsiella pneumoniae 1033-5P14; Otte S et al.; Mutants of Escherichia coli K-12 were isolated which lack the normal phosphotransferase system-dependent catabolic pathway for D-mannitol (Mtl) . In some mutants the pts genes for the general proteins enzyme I and histidine protein of the phosphoenolpyruvate-dependent carbohydrate phosphotransferase systems were deleted . Other mutants expressed truncated mannitol-specific enzymes II (II(Mtl)) which lacked the IIA(Mtl) or IIBA(Mtl) domain(s), and the mtlA genes originated either from E . coli K-12 or from Klebsiella pneumoniae 1033-5P14 . The dalD gene from Klebsiella oxytoca M5a1 was cloned on single-copy plasmids and transformed into the strains described above . This gene encodes an NAD-dependent D-arabinitol dehydrogenase (DalD) which converts D-arabinitol into D-xylulose and also converts D-mannitol into D-fructose . The different strains were used to isolate mutations which allow efficient transport of mannitol through the nonphosphorylated II(Mtl) complexes by selecting for growth on this polyhydric alcohol . More than 40 different mutants were analyzed to determine their ability to grow on mannitol, as well as their ability to bind and transport free mannitol and, after restoration of the missing domain(s), their ability to phosphorylate mannitol . Four mutations were identified (E218A, E218V, H256P, and H256Y); all of these mutations are located in the highly conserved loop 5 of the IIC membrane-bound transporter, and two are located in its GIHE motif . These mutations were found to affect the various functions in different ways . Interestingly, in the presence of all II(Mtl) variants, whether they were in the truncated form or in the complete form, in the phosphorylated form or in the nonphosphorylated form, and in the wild-type form or in the mutated form, growth occurred on the low-affinity analogue D-arabinitol with good efficiency, while only the uncoupled mutated forms transported mannitol at a high rate.

Clin Imaging, 2003 Mar-Apr, 27(2), 129 - 31
Clinically occult isolated right iliac mycotic aneurysm with duodenal involvement in a diabetic elderly man: multislice CT diagnosis; Ko SF et al.; A diabetic old man presented with vague abdominal discomfort and intermittent tarry stools for 2 days and gastric ulcers with bleeding was diagnosed after endoscopy . Multislice computed tomography (MSCT) clearly depicted an isolated right iliac mycotic aneurysm with retroperitoneal extension and duodenal involvement . Timely operation and effective antibiotic treatment resulted in complete recovery . To our knowledge, this is the first report of an isolated mycotic iliac artery aneurysm (IAA) complicated with an aneurysmo-duodenal fistula induced by Klebsiella pneumoniae .

Int J Antimicrob Agents, 2003 Mar, 21(3), 285 - 8
Extended-spectrum beta-lactamase types in Klebsiella pneumoniae and Escherichia coli in two Greek hospitals; Tzelepi E et al.; Seventy-nine Klebsiella pneumoniae and 124 Escherichia coli clinical strains, isolated consecutively during August-October 2001 in two Greek hospitals, were examined for production of extended-spectrum beta-lactamases (ESBLs) . Seventy-one (35%) isolates (46 K . pneumoniae and 25 E . coli) were ESBL-positive by phenotypic methods . Isoelectric focusing of beta-lactamases and PCR assays for bla genes showed that SHV-5-type ESBLs were the most frequent (45 isolates, 22%) followed by CTX-M (24 isolates, 12%) and IBC (three isolates, 1.5%) . The latter two ESBL types may have been established recently in this setting.

FEMS Immunol Med Microbiol, 2003 Mar 20, 35(2), 93 - 8
K-antigen-specific, but not O-antigen-specific natural human serum antibodies promote phagocytosis of Klebsiella pneumoniae; Lepper PM et al.; Infections due to Klebsiella pneumoniae and other Klebsiella spp . are a leading cause of hospital-associated morbidity, especially in the intensive care setting . In this study, the hypothesis that normal human sera contain sufficient concentrations of K-antigen-specific antibodies to promote phagocytic killing of encapsulated, highly virulent Klebsiella organisms was tested . K2-antigen-specific IgG and IgM antibodies were detected in each of 10 normal sera, and such antibodies were functionally active in a phagocytic killing assay . Phagocytosis depended critically on sufficient numbers of neutrophils and was impaired by the presence of soluble Klebsiella capsular polysaccharide (CPS) . Thus, insufficient numbers of neutrophils and circulation of soluble CPS but not lack of K-specific antibodies may be detrimental in Klebsiella sepsis . The efficacy of hyperimmune sera might be based not on enhancement of phagocytosis but on the neutralization of these detrimental effects of circulating CPS and LPS.

Microbiology, 2003 Feb, 149(Pt 2), 451 - 7
Identification of the essential histidine residue for high-affinity binding of AlbA protein to albicidin antibiotics; Weng LX et al.; The albA gene from Klebsiella oxytoca encodes a protein that binds albicidin phytotoxins and antibiotics with high affinity . Previously, it has been shown that shifting pH from 6 to 4 reduces binding activity of AlbA by about 30%, indicating that histidine residues might be involved in substrate binding . In this study, molecular analysis of the albA coding region revealed sequence discrepancies with the albA sequence reported previously, which were probably due to sequencing errors . The albA gene was subsequently cloned from K . oxytoca ATCC 13182(T) to establish the revised sequence . Biochemical and molecular approaches were used to determine the functional role of four histidine residues (His(78), His(125), His(141) and His(189)) in the corrected sequence for AlbA . Treatment of AlbA with diethyl pyrocarbonate (DEPC), a histidine-specific alkylating reagent, reduced binding activity by about 95 % . DEPC treatment increased absorbance at 240-244 nm by an amount indicating conversion to N-carbethoxyhistidine of a single histidine residue per AlbA molecule . Pretreatment with albicidin protected AlbA against modification by DEPC, with a 1 : 1 molar ratio of albicidin to the protected histidine residues . Based on protein secondary structure and amino acid surface probability indices, it is predicted that His(125) might be the residue required for albicidin binding . Mutation of His(125) to either alanine or leucine resulted in about 32 % loss of binding activity, and deletion of His(125) totally abolished binding activity . Mutation of His(125) to arginine and tyrosine had no effect . These results indicate that His(125) plays a key role either in an electrostatic interaction between AlbA and albicidin or in the conformational dynamics of the albicidin-binding site.

J Clin Microbiol, 2003 Mar, 41(3), 1161 - 6
Patterns of resistance associated with integrons, the extended-spectrum beta-lactamase SHV-5 gene, and a multidrug efflux pump of Klebsiella pneumoniae causing a nosocomial outbreak; Gruteke P et al.; Multiresistant Klebsiella pneumoniae caused a nosocomial outbreak . Resistance patterns of the presumed outbreak isolates varied among and within patients . In order to control the outbreak, screening for extended-spectrum beta-lactamase (ESBL)-producing K . pneumoniae was commenced . A number of susceptible K . pneumoniae strains were stored to serve as controls in genetic strain typing . Typing by pulsed-field gel electrophoresis proved the clonality of the strains in the recognized outbreak patients . Typing of the control strains by pulsed-field gel electrophoresis showed that at least one patient had been missed by the ESBL screening procedure . Further genetic typing confirmed the presence of the SHV-5 ESBL gene in all but one of the outbreak strains . Variable presence of integrons that carried the aminoglycoside resistance genes aadB and aadA2 was found . A gyrA mutation in codon 83 was present in all outbreak strains tested, despite considerable differences in ciprofloxacin MICs . The MICs of ciprofloxacin and the chemically unrelated drug cefoxitin were correlated (r = 0.86, P < 0.01) and were compatible with the overexpression of an efflux pump in a subset of the outbreak strains . We conclude that outbreak strains that express an ESBL gene only at a low level may pass unnoticed in a screening procedure, when the laboratory is unaware of variable ESBL expression . In this particular outbreak, screening for strains for which ciprofloxacin MICs were > or =0.25 micro g/ml would in retrospect have been the most sensitive method for detection of the K . pneumoniae outbreak strain.

J Antimicrob Chemother, 2003 Mar, 51(3), 711 - 4
Plasmid-mediated, carbapenem-hydrolysing beta-lactamase, KPC-2, in Klebsiella pneumoniae isolates; Smith Moland E et al.; Four isolates of Klebsiella pneumoniae obtained from patients at a Maryland medical centre exhibited reduced susceptibility to carbapenems and were found to produce the novel, class A, plasmid-mediated, carbapenem-hydrolysing enzyme, KPC-2 . This enzyme has 99% identity with the plasmid-mediated, carbapenem-hydrolysing enzyme KPC-1, reported previously in a North Carolina K . pneumoniae isolate . The KPC-2-producing isolates were either susceptible or intermediate to imipenem and meropenem, unlike the KPC-1-producing isolate, which was resistant to these agents . Detection of KPC-2 may be a problem for clinical laboratories because in this study it was associated with positive extended-spectrum beta-lactamase (ESBL) confirmation tests (clavulanate-potentiated activities of ceftriaxone, ceftazidime, cefepime and aztreonam) . Therefore, a failure to recognize the significance of reduced carbapenem susceptibility in the isolates that remained susceptible to imipenem or meropenem could have resulted in the isolates being incorrectly identified as ESBL producers.

J Antimicrob Chemother, 2003 Mar, 51(3), 605 - 12
Variable susceptibility to piperacillin/tazobactam amongst Klebsiella spp . with extended-spectrum beta-lactamases; Babini GS et al.; MICs of piperacillin/tazobactam are conventionally determined by varying the concentration of piperacillin in the presence of a fixed 4 mg/L tazobactam . When tested in this way, the MIC distribution for Klebsiella isolates with extended-spectrum beta-lactamases (ESBLs) is strongly bimodal, such that many producers are inhibited at 16 + 4 mg/L whilst others require MICs of > or =512 + 4 mg/L . When, however, piperacillin/tazobactam was tested as a fixed 8:1 ratio, the MIC distribution became unimodal . If clavulanate 4 mg/L was combined with piperacillin, a unimodal MIC distribution was seen for ESBL-producing Klebsiella spp . but a bimodal distribution arose if the clavulanate concentration was reduced to 0.25 mg/L . These data for alternative combinations suggested that the bimodal MIC distribution seen for piperacillin + tazobactam 4 mg/L was a titration effect, not a reflection of some ESBLs being resistant to tazobactam . Even within single strains, as defined by serotype and DNA fingerprints, there was considerable variation in susceptibility to piperacillin + tazobactam 4 mg/L, with some representatives highly susceptible and others highly resistant . Some of the more resistant representatives produced more of their ESBL, or had a greater number of beta-lactamase types, but these associations were not universal . Elevated resistance to piperacillin + tazobactam was not associated with porin change in any ESBL producer examined, but has been found by others.

Biophys J, 2003 Mar, 84(3), 1651 - 9
The citrate carrier CitS probed by single-molecule fluorescence spectroscopy; Kastner CN et al.; A prominent region of the Na(+)-dependent citrate carrier (CitS) from Klebsiella pneumoniae is the highly conserved loop X-XI, which contains a putative citrate binding site . To monitor potential conformational changes within this region by single-molecule fluorescence spectroscopy, the target cysteines C398 and C414 of the single-Cys mutants (CitS-sC398, CitS-sC414) were selectively labeled with the thiol-reactive fluorophores AlexaFluor 546/568 C(5) maleimide (AF(546), AF(568)) . While both single-cysteine mutants were catalytically active citrate carriers, labeling with the fluorophore was only tolerated at C398 . Upon citrate addition to the functional protein fluorophore conjugate CitS-sC398-AF(546), complete fluorescence quenching of the majority of molecules was observed, indicating a citrate-induced conformational change of the fluorophore-containing domain of CitS . This quenching was specific for the physiological substrate citrate and therefore most likely reflecting a conformational change in the citrate transport mechanism . Single-molecule studies with dual-labeled CitS-sC398-AF(546/568) and dual-color detection provided strong evidence for a homodimeric association of CitS.

J Liposome Res, 2002 Feb-May, 12(1-2), 121 - 7
Liposome-enabled synergistic interaction of antimicrobial agents; Schiffelers RM et al.; Antimicrobial agents may interact synergistically when both drugs are present at the infected site for an adequate period of time at sufficient concentrations . Generally speaking, the agents in the combination show different tissue distributions and pharmacokinetics . By co-encapsulation of the drugs in a drug carrier, like liposomes, parallel tissue distributions of both drugs may be ensured and drug concentrations at the site of infection may be increased . In this presentation therapeutic efficacy of liposome-co-encapsulated gentamicin (GN) and ceftazidime (CZ) will be shown in a GN-CZ-susceptible and GN-CZ-resistant Klebsiella pneumoniae-pneumonia in rats.

Zhejiang Da Xue Xue Bao Yi Xue Ban, 2002 Aug, 31(6), 457 - 460
{Encoding genes and genotypes of gamma-lactamases produced by a multiple resistant Klebsiella pneumoniae}; Yu YS et al.; OBJECTIVE: To determine the sequence of gene for encoding beta-lactamase produced by Klebsiella pneumoniae E3 isolated from Jiaxing Area in Zhejiang Province . METHODS The Klebsiella pneumoniae strain E3 was identified as an ESBLs-producing bacterium by inhibitor-potentiated broth dilution test . The gene encoding gamma-lactamase of the strain was amplified by PCR . The purified PCR product was cloned and sequenced by Sanger's dideoxy chain termination composition method . RESULTS The Klebsiella pneumoniae strain E3 produced both TEM and SHV gamma lactamases . The SHV encoding gene had 812 nucleotide residues responsible for encoding SHV-11 gamma-lactamase and the TEM encoding gene had 973 nucleotide residues responsible for encoding TEM-1 gamma-lactamase . CONCLUSION The Klebsiella pneumoniae strain E3 isolated from a patient in Jiaxing Area in Zhejiang Province is able to produce both TEM-1 and SHV-11 gamma-lactamases.

Hua Xi Yi Ke Da Xue Xue Bao, 2001 Jun, 32(2), 157 - 62
{K . pneumoniea endotoxin induced mice beta-defensin-4 mRNA expression and its signaling transduction}; Cai S et al.; OBJECTIVE: To investigate the in vivo effects of Klebsiella pneumoniae endotoxin(LPS) on beta-defensin expression and the relevant signaling transduction pathway . METHODS: A LPS tolerant mouse C3H/HeJ with a point mutation at Toll-like receptor-4 (TLR4) gene and its wild type strain C3H/HeN were used in this study . C3H/HeJ and C3H/HeN were injected with 4 mg/kg of LPS intraperitoneally . The tracheas, lungs and kidneys of the C3H/HeJ and C3H/HeN were collected respectively at different LPS-treated time points, and the total RNA of each sample was extracted . The expression of mice beta-defensin-3 and/or beta-defensin-4 mRNA in these tissues was determined by reverse transcriptase-polymerase chain reaction (RT-PCR) . The sequence of cDNA amplified from the lung of C3H/HeN treated by LPS for 24 h was analyzed . By using western blot, p-I kappa B alpha (phosphorylated I kappa B alpha) and I kappa B alpha of in the lungs of C3H/HeJ and C3H/HeN were detected at different time points after treatment with LPS or without LPS . RESULTS: 1 . beta-defensin-4 mRNA was detected in the lungs of C3H/HeN after 24 h treatment with LPS . In contrast, no signal was determined in C3H/HeJ mice with LPS treatment and the C3H/HeN mice without LPS treatment . 2 . Compared with the control, increas of the p-I kappa B alpha was observed in the lungs of C3H/HeN at 4 h after treatment with LPS, while both the p-I kappa B alpha and I kappa B alpha contents showed a tendency to go down at 8 h after treatment and dramatically decreased at 24 h . But there were no changes in the of p-I kappa B alpha and I kappa B alpha content the lungs of C3H/HeJ under the same conditions . CONCLUSION: K . pneumoniea endotoxin could induce the expression of beta-defensin-4 mRNA in the lung of C3H/HeN, and TLR4-mediated NF-kappa B activation signaling pathway may be responsible for this event.

Infect Immun, 2003 Mar, 71(3), 1306 - 15
Intrapulmonary expression of macrophage inflammatory protein 1alpha (CCL3) induces neutrophil and NK cell accumulation and stimulates innate immunity in murine bacterial pneumonia; Zeng X et al.; Macrophage inflammatory protein 1alpha (MIP-1alpha) (CCL3) is an important mediator of leukocyte recruitment and activation in a variety of inflammatory states, including infection . A recombinant human type 5 adenovirus containing the murine MIP-1alpha cDNA (AdMIP-1alpha) was constructed to determine the effect of transient intrapulmonary expression of MIP-1alpha on leukocyte recruitment, activation, and bacterial clearance in a murine model of Klebsiella pneumoniae pneumonia . The intratracheal administration of AdMIP-1alpha resulted in both time- and dose-dependent expression of MIP-1alpha mRNA and protein within the lung . Importantly, the intrapulmonary overexpression of MIP-1alpha resulted in a maximal 35- and 100-fold reduction in lung and blood bacterial burden, respectively, in animals cochallenged with K . pneumoniae, which was associated with a significant increase in neutrophil and activated NK cell accumulation . Furthermore, the transgenic expression of MIP-1alpha during bacterial pneumonia resulted in enhanced expression of gamma interferon mRNA, compared to that observed in Klebsiella-challenged animals pretreated with control vector . These findings indicate an important role for MIP-1alpha in the recruitment and activation of selected leukocyte populations in vivo and identify this cytokine as a potential immunoadjuvant to be employed in the setting of localized bacterial infection.

Clin Infect Dis, 2003 Mar 1, 36(5), 575 - 9 Epub 2003 Feb 07.
Klebsiella infection in patients with thalassemia; Chung BH et al.; Klebsiella infection has previously been reported in a few patients with transfusion-dependent thalassemia . The incidence and clinical spectrum of this infection in our cohort of patients were reviewed retrospectively . Among 160 patients observed for 12 years, there were 15 episodes of Klebsiella infection that occurred in 12 patients (7.5%), resulting in an incidence of 0.78 infections per 100 patient-years . The clinical spectrum included sinusitis (4 cases), intracranial infection (5 cases), septicemia (4 cases), and abscesses of the liver, lung, kidney, and parotid gland (1 case each) . Three patients had recurrent infections involving different sites, 2 (16%) died of fulminant septicemia, and 3 (25%) had significant permanent neurological deficits . The antibiotic susceptibility pattern for the isolates was similar to the pattern for isolates recovered in the community . With regard to predisposing factors, iron overload and liver function derangement were found to be significant on univariate analysis (P=.046 and P=.049, respectively) but insignificant on multivariate analysis . Klebsiella infection was a serious and frequently encountered complication in our patients with transfusion-dependent thalassemia, resulting in high mortality and morbidity rates.

Enferm Infecc Microbiol Clin, 2003 Feb, 21(2), 72 - 6
{Risk factors associated with ceftazidime-resistant Klebsiella pneumoniae infection}; Bermejo J et al.; INTRODUCTION: Risk factors associated with ceftazidime-resistant Klebsiella pneumoniae (CAZ-R Kp) infection may vary among hospitals and in the same hospital at different time points . Knowledge of these factors is required to establish suitable infection control programs . METHODS: A case-control study was conducted to assess risk factors for CAZ-R Kp infection . Thirty-two cases were compared with 28 controls admitted to a 200-bed general hospital during 1999 and 2000 . RESULTS: In the univariate analysis Kp CAZ-R isolates were significantly associated with nosocomial acquisition (OR 5 17.40), prior antibiotic use (OR 5 14.94), particularly ciprofloxacin use (OR 5 5), and hospitalization stay of more than 6 days (OR 5 6.72) . Significantly associated variables in the logistic regression analysis included nosocomial acquisition (OR 5 9.29), prior antibiotic use (OR 5 6.21), and particularly, ciprofloxacin use (OR 5 10.84) . CONCLUSIONS: Efforts toward more rational overall antibiotic use and particularly ciprofloxacin use, combined with infection control measures are necessary to decrease the prevalence of CAZ-R Kp in our hospital.

FEMS Microbiol Lett, 2003 Jan 21, 218(1), 121 - 6
Identification and characterization of KvgAS, a two-component system in Klebsiella pneumoniae CG43; Lai YC et al.; A two-component system encoding gene cluster kvgAS that is present only in virulent Klebsiella pneumoniae CG43 was isolated and its sequence determined . RT-PCR and Southern analysis demonstrated that kvgAS is organized as an operon . No apparent effect of a kvgS deletion on bacterial virulence was observed in a mouse peritonitis model . In the presence of paraquat or 2,2-dipyridyl, the activity of kvgAS promoter in the kvgS mutant was found to be reduced to half of the level in the wild-type strain . The data suggest that the KvgAS system is autoregulated and plays a role in countering free radical stresses and sensing iron-limiting conditions.

Zhonghua Yi Xue Za Zhi (Taipei), 2002 Nov, 65(11), 534 - 9
Risk factors for endogenous endophthalmitis secondary to Klebsiella pneumoniae liver abscess; Sheu SJ et al.; BACKGROUND: To identify the risk factors for endogenous endophthalmitis secondary to Klebsiella pneumoniae (KP) liver abscess . METHODS: Retrospective chart review of 200 patients with KP liver abscess from 1990 to 2000 was performed . Data variables included age, sex, past history, systemic condition, initial and final visual acuity, slit lamp biomicroscopy, intraocular pressure, fundus, course and treatment . RESULTS: Extrahepatic metastasis developed in 44 patients, including 18 cases (23 eyes) of endophthalmitis . Seventeen eyes had final vision less than counting fingers . Diabetes had significant association with the development of extrahepatic metastasis (p = 0.045) and the poor visual outcome of endophthalmitis (p = 0.022), whereas, neither the initial vision nor vitrectomy was significantly related to the outcome of endophthalmitis . CONCLUSIONS: The visual prognosis of endogenous endophthalmitis secondary to KP liver abscess is generally poor, and diabetes might be a significant risk factor for the poor visual outcome . Even when the initial vision is as bad as light perception only, some eyes can be saved . Both internists and ophthalmologists should be alert enough to detect and treat these patients early.

Indian J Pathol Microbiol, 2000 Jan, 43(1), 35 - 40
Immunoglobulin-M estimation and C-reactive protein detection in neonatal septicemia; Krishna BV et al.; Immunoglobulin-M estimation, and C-Reactive Protein test and blood culture were performed on 57 neonates, clinically suspected to have septicemia . IgM level of > or = 20 mg/dl was found in 58.62% cases and CRP test was positive in 68.98% of culture proven sepsis . Blood culture was positive in 29 cases, with the predominant organism being Klebsiella . Of the 2 tests, CRP had a higher sensitivity (68.97%) while IgM estimation had a higher specificity (82.14%) . When the 2 tests were considered together the sensitivity and specificity further increased . Death rate among neonates with IgM levels < 20 mg/dl was 5 times higher than those with elevated IgM levels.

Eur J Biochem, 2003 Feb, 270(4), 635 - 45
Relation between domain evolution, specificity, and taxonomy of the alpha-amylase family members containing a C-terminal starch-binding domain; Janecek S et al.; The alpha-amylase family (glycoside hydrolase family 13; GH 13) contains enzymes with approximately 30 specificities . Six types of enzyme from the family can possess a C-terminal starch-binding domain (SBD): alpha-amylase, maltotetraohydrolase, maltopentaohydrolase, maltogenic alpha-amylase, acarviose transferase, and cyclodextrin glucanotransferase (CGTase) . Such enzymes are multidomain proteins and those that contain an SBD consist of four or five domains, the former enzymes being mainly hydrolases and the latter mainly transglycosidases . The individual domains are labelled A {the catalytic (beta/alpha)8-barrel}, B, C, D and E (SBD), but D is lacking from the four-domain enzymes . Evolutionary trees were constructed for domains A, B, C and E and compared with the 'complete-sequence tree' . The trees for domains A and B and the complete-sequence tree were very similar and contain two main groups of enzymes, an amylase group and a CGTase group . The tree for domain C changed substantially, the separation between the amylase and CGTase groups being shortened, and a new border line being suggested to include the Klebsiella and Nostoc CGTases (both four-domain proteins) with the four-domain amylases . In the 'SBD tree' the border between hydrolases (mainly alpha-amylases) and transglycosidases (principally CGTases) was not readily defined, because maltogenic alpha-amylase, acarviose transferase, and the archaeal CGTase clustered together at a distance from the main CGTase cluster . Moreover the four-domain CGTases were rooted in the amylase group, reflecting sequence relationships for the SBD . It appears that with respect to the SBD, evolution in GH 13 shows a transition in the segment of the proteins C-terminal to the catalytic (beta/alpha)8-barrel(domain A).

J Clin Microbiol, 2003 Feb, 41(2), 798 - 802
Use of multienzyme multiplex PCR amplified fragment length polymorphism typing in analysis of outbreaks of multiresistant Klebsiella pneumoniae in an intensive care unit; van der Zee A et al.; We developed and optimized a new modified amplified fragment length polymorphism (AFLP) typing method to obtain a multibanding fingerprint that can be separated by agarose gel electrophoresis . Both to maximize the discriminatory power and to facilitate the computer-assisted analysis, bacterial DNA was digested with four different restriction enzymes . After ligation of adaptors to the DNA fragments, PCR testing of various single primers was performed . Two single primers that gave optimal results with regard to band resolution and discriminatory power were selected and combined . The computer-assisted analysis of fingerprint patterns was performed with Pearson's product-moment correlation values of densitometric curves, without assigning bands to peaks . Thus, the analysis is not subject to human interpretation errors . With this method, we investigated two outbreaks of multiresistant Klebsiella pneumoniae in an intensive care unit and various sporadic isolates of K . pneumoniae and Klebsiella oxytoca . Cluster analysis of isolates analyzed in different experiments and on different gels showed that fingerprint patterns clustered correctly according to subspecies or to the outbreaks . Multienzyme multiplex PCR AFLP revealed that the first outbreak was caused by two different types of strains . Outbreak two was caused by yet another strain of K . pneumoniae . In conclusion, the typing method used here is easy to perform and highly reproducible, and due to generation of complex banding patterns, it has a higher discriminatory power . Furthermore, the multienzyme multiplex PCR fingerprints are easy to analyze, and a reliable database can be stored in the computer to facilitate comparison of future isolates of Klebsiella spp . The method can be performed in every clinical microbiology laboratory.

J Clin Microbiol, 2003 Feb, 41(2), 772 - 7
Occurrence of extended-spectrum and AmpC beta-lactamases in bloodstream isolates of Klebsiella pneumoniae: isolates harbor plasmid-mediated FOX-5 and ACT-1 AmpC beta-lactamases; Coudron PE et al.; We tested 190 Klebsiella pneumoniae bloodstream isolates recovered from 189 patients in 30 U.S . hospitals in 23 states to determine the occurrence of extended-spectrum beta-lactamase (ESBL) and AmpC beta-lactamase producers . Based on growth inhibition by clavulanic acid by disk and MIC test methods, 18 (9.5%) of the isolates produced ESBLs . Although the disk diffusion method with standard breakpoints identified 28 cefoxitin-nonsusceptible isolates, only 5 (18%) of these were confirmed as AmpC producers . Of two AmpC confirmatory tests, the three-dimensional extract test was easier to perform than was the double-disk approximation test using a novel inhibitor, Syn2190 . Three of the five AmpC producers carried the bla(FOX-5) gene, while the other two isolates harbored the bla(ACT-1) gene . All AmpC genes were transferable . In vitro susceptibility testing with standard inocula showed that all five AmpC-producing strains were susceptible to cefepime, imipenem, and ertapenem but that with a high inoculum, more of these strains were susceptible to the carbapenems than to cefepime . All but 1 of 14 screen-positive AmpC nonproducers (and ESBL nonproducers) were susceptible to ceftriaxone and cefepime at the standard inoculum as were 6 of 6 isolates that were randomly selected and tested with a high inoculum . These results indicate that (i) . a significant number of K . pneumoniae bloodstream isolates harbor ESBL or AmpC beta-lactamases, (ii) . confirmatory tests are necessary to identify true AmpC producers, and (iii) . in vitro, carbapenems are active against AmpC-producing strains of K . pneumoniae.

Diagn Microbiol Infect Dis, 2003 Jan, 45(1), 29 - 34
Comparison of BDPhoenix and VITEK2 automated antimicrobial susceptibility test systems for extended-spectrum beta-lactamase detection in Escherichia coli and Klebsiella species clinical isolates; Sturenburg E et al.; The present study compares the ability to detect extended-spectrum beta-lactamases (ESBL) among a collection of 34 ESBL producing clinical isolates belonging to Escherichia coli and Klebsiella species with two new rapid susceptibility and identification instruments-VITEK2 (bioMerieux, Marcy l'Etoile, France) vs . BDPhoenix (BD Biosciences, Sparks, MD) . ESBL content in these isolates was previously characterized on the basis of PCR amplification and sequencing results which were used as the reference method in our evaluation . BDPhoenix correctly determined the ESBL outcome for all strains tested (100% detection rate), whereas VITEK2 was not able to detect the ESBL status in 5 isolates (85% detection rate) . Detailed analysis revealed that the discrepancies were mainly observed with 'difficult-to-detect' strains . Misidentification was either due to low oximino cephalosporin MIC in these strains or was associated with pronounced 'cefotaximase' or 'ceftazidimase' phenotypes . Klebsiella oxytoca chromosomal beta-lactamase (K1) is phenotypically quite similar to ESBL enzymes . In order to evaluate whether the K1 and ESBL enzymes could be discriminated, we expanded our analysis by 8 clinical K . oxytoca strains with K1 phenotypes . VITEK2 gave excellent identification of these strains whereas 7 out of 8 were falsely labeled ESBL-positive by the BDPhoenix system.

Eur J Immunol, 2003 Feb, 33(2), 326 - 33
Outer membrane protein A renders dendritic cells and macrophages responsive to CCL21 and triggers dendritic cell migration to secondary lymphoid organs; Jeannin P et al.; Outer membrane protein A (OmpA) is a class of bacterial cell wall protein that is immunogenic without adjuvant . As specific immune responses are initiated in the lymph nodes (LN, we analyzed the effect of the OmpA from Klebsiella pneumoniae (KpOmpA) onchemokine/ chemokine receptor expression by APC and on cell migration to the LN . Upon contact with KpOmpA, human immature DC and macrophages acquire CCR7 expression and responsiveness to CCL21 . In parallel, CCR1 and CCR5 expression is down-regulated and CXCL8, CCL2, CCL3 and CCL5 production is up-regulated . Mice injected subcutaneously with KpOmpA present a transient inflammatory reaction at the site of injection accompanied by an enlargement of the draining LN with a higher proportion of DC and macrophages . Lastly, when exposed to KpOmpA prior injection, DC but not macrophages migrate to the draining LN . In conclusion, KpOmpA confers a migratory phenotype to DC and triggers their migration to the regional LN . This property contributes to explain how innate cells initiate adaptive immune response upon recognition of conserved bacterial components and also why OmpA is immunogenic in the absence of adjuvant.

Antimicrob Agents Chemother, 2003 Feb, 47(2), 755 - 8
Emergence in Klebsiella pneumoniae of a chromosome-encoded SHV beta-lactamase that compromises the efficacy of imipenem; Poirel L et al.; A Klebsiella pneumoniae isolate was identified that had reduced susceptibility to several expanded-spectrum cephalosporins and imipenem . That isolate produced a chromosome-encoded SHV-type beta-lactamase, SHV-38, that had an alanine to valine substitution in position Ambler 146 compared to beta-lactamase SHV-1 . The kinetic parameters for purified beta-lactamases SHV-38 and SHV-1 showed that the hydrolytic spectrum of SHV-38 included only ceftazidime and imipenem . This report is the first example of an SHV-type beta-lactamase capable of hydrolyzing imipenem.

Antimicrob Agents Chemother, 2003 Feb, 47(2), 559 - 62
Prevalence of plasmid-mediated quinolone resistance; Jacoby GA et al.; Quinolone resistance encoded by the qnr gene and mediated by plasmid pMG252 was discovered in a clinical strain of Klebsiella pneumoniae that was isolated in 1994 at the University of Alabama at Birmingham Medical Center . The gene codes for a protein that protects DNA gyrase from quinolone inhibition and that belongs to the pentapeptide repeat family of proteins . The prevalence of the gene has been investigated by using PCR with qnr-specific primers with a sample of more than 350 gram-negative strains that originated in 18 countries and 24 states in the United States and that included many strains with plasmid-mediated AmpC or extended spectrum beta-lactamase enzymes . qnr was found in isolates from the University of Alabama at Birmingham only during 6 months in 1994, despite the persistence of the gene for FOX-5 beta-lactamase, which is linked to qnr on pMG252 . Isolates from other locations were negative for qnr . The prevalence of mcbG in the same sample was also examined . mcbG encodes another member of the pentapeptide repeat family and is involved in immunity to microcin B17, which, like quinolones, targets DNA gyrase . A single clinical isolate contained mcbG on a transmissible R plasmid . This plasmid and one carrying the complete microcin B17 operon slightly decreased sparfloxacin susceptibility but had a much less protective effect than pMG252 . Plasmid-mediated quinolone resistance was thus rare in the sample examined.

Se Pu, 2000 Jul, 18(4), 328 - 31
{Research of indole-3-acetic acid biosynthetic pathway of Klebsiella oxytoca SG-11 by HPLC and GC-MS}; Lu ZX et al.; The plant growth promoting bacteria are closely associated to plant . The bacteria are used to adhering to plant rhizoplane, promoting plant growth by fixing nitrogen from atmosphere, secreting stimulating substances or producing antagonistic to plant pathogens . It was indicated that the biological nitrogen fixation played an important role in plant growth promoting function . In fact, it was verified recently by overall research that IAA does it . Therefore research of IAA production and biosynthetic pathway of plant growth-promoting bacteria is much more important . The various ways of IAA production indicated the strong or weak promoting function of bacterium to plants in general . The purpose of this paper is to determine whether IAA exists in cultured medium of Klebsiella axytoca SG-11 and biosynthetic pathway of IAA, in order to opimize cultural conditions for IAA production . Klebsiella axytoca SG-11 is a plant growth promoting bacterium, isolated from rice rhizoplane, which can fix nitrogen . The supernatant of SG-11 cultured medium determined by HPLC showed that 47.4 mg/L of IAA existed in LB medium and 1.2 mg/L of IAA, in basal medium . IAA in metabolite was identified by GC/MS as well . The intermediate determination of tryptamine, indole-3-acetamide, tryptophol and indole-3-acetonitrile indirectly indicated that IAA was biosynthesized in a pathway of indole-3-pyruvic acid . Meanwhile, tryptophol in metabolite of SG-11 was verified by GC/MS . The direct intermediates of indole-3-pyruvic acid and indole-3-acetaldehyde in the pathway can not be determined, because both are unstable under normal condition . As reversible conversion existed between indole-3 pyruvic aldehyde and tryptophol, the presence of tryptophol also proved the pathway of indole-3-pyruvic acid in the synthesis of IAA by Klebsiella axytoca SG-11 . The results laid basis for further research of plant growth-promoting function of the bacterium.

Lin Chuang Er Bi Yan Hou Ke Za Zhi, 2000 Apr, 14(4), 173 - 4
{HSP70 expression of middle ear mucosa in acute suppurative otitis media}; Bi AF et al.; OBJECTIVE: To study HSP70 expression of middle ear mucosa in acute suppurative otitis media . METHOD: An animal model of acute middle ear infection was established by inoculating Klebsiella Pneumoniae into middle ear of guinea pigs . Animals were sacrificed 1, 3, 5 and 7 days after inoculation . The expression of Hsp70 epitope related proteins in middle ear mucosa were analysed by modified western blot test . RESULT: Very light 70 kD bands were recognized by anti Hsp70 monoclonal antibody in the unstressed group . However, middle ear mucosa of the stressed group not only showed relatively strong 70 kD bands, but also expressed more intensely 31 kD and 17 kD bands . The infected mucosa expressed one 70 kD band at 1st day, the strongest band at 3rd day, falling at 5th day; 31 kD was recognized two bands at 3rd day, the strongest bands at 5th day, falling at 7th day; two bands of 17 kD were detected in all days, with the strongest bands at 7th day . CONCLUSION: The results suggested that middle ear mucosa expressed Hsp70 and Hsp70 epitope related proteins like 31 kD and 17 kD in acute otitis media.

J Inorg Biochem, 2003 Jan 1, 93(1-2), 18 - 32
Effects of substrates (methyl isocyanide, C2H2) and inhibitor (CO) on resting-state wild-type and NifV(-)Klebsiella pneumoniae MoFe proteins; McLean PA et al.; We report the use of electron nuclear double resonance (ENDOR) spectroscopy to examine how the metal sites in the FeMo-cofactor cluster of the resting nitrogenase MoFe protein respond to addition of the substrates acetylene and methyl isocyanide and the inhibitor carbon monoxide . 1H, 57Fe and 95Mo ENDOR measurements were performed on the wild-type and the NifV(-)proteins from Klebsiella pneumoniae . Among the molecules tested, only the addition of acetylene to either protein induced widespread changes in the 57Fe ENDOR spectra . Acetylene also induced increases in intensity from unresolved protons in the proton ENDOR spectra . Thus we conclude that acetylene may bind to the resting-state MoFe protein to perturb the FeMo-cofactor environment . On the other hand, the present results show that methyl isocyanide and carbon monoxide do not substantially alter the FeMo cofactor's geometric and electronic structures . We interpret this as lack of interaction between those two molecules and the FeMo cofactor in the resting state MoFe protein . Thus, although it is generally accepted that substrates or inhibitors bind to the FeMo-cofactor only under turnover condition, this work provides evidence that at least one substrate can perturb the active site of nitrogenase under non-catalytic conditions.

Ann Fr Anesth Reanim, 2002 Dec, 21(10), 807 - 11
{Discitis after spinal anesthesia for transurethral resection of the prostate}; Malinovsky JM et al.; We described a case of discitis and meningitis following spinal anaesthesia for transurethral resection of the prostate . The patient received antibiotics for a month before surgery, because of Klebsiella prostatitis . Spinal anaesthesia was performed in L3-L4 interspace by using 22G Quincke needle . Bacteriaemia occurred during the first postoperative hours . Ten days after spinal anaesthesia, patient suffered from lumbar pain, exacerbated by vertebral percussion, and motor weakness within lower limb, which was marked on right side . MRI examination showed L3-L4 discitis with psoas abcess in regard, and epiduritis marked around L3 right spinal root . CSF examination confirmed meningitis but no bacteria was found . Antibiotics were administered over a 6 weeks period, and then patient discharged from hospital without neurological sequellae . Infectious discitis related to disk puncture during spinal anaesthesia and postoperative bacteriaemia was likely in our patient.

J Bacteriol, 2003 Feb, 185(3), 788 - 800
RmpA2, an activator of capsule biosynthesis in Klebsiella pneumoniae CG43, regulates K2 cps gene expression at the transcriptional level; Lai YC et al.; The rmpA2 gene, which encodes an activator for capsular polysaccharide (CPS) synthesis, was isolated from a 200-kb virulence plasmid of Klebsiella pneumoniae CG43 . Based on the sequence homology with LuxR at the carboxyl-terminal DNA-binding motif, we hypothesized that RmpA2 exerts its effect by activating the expression of cps genes that are responsible for CPS biosynthesis . Two luxAB transcriptional fusions, each containing a putative promoter region of the K . pneumoniae K2 cps genes, were constructed and were found to be activated in the presence of multicopy rmpA2 . The activation is likely due to direct binding of RmpA2 to the cps gene promoter through its C-terminal DNA binding motif . Moreover, the loss of colony mucoidy in a K . pneumoniae strain deficient in RcsB, a regulator for cps gene expression, could be recovered by complementing the strain with a multicopy plasmid carrying rmpA2 . The CPS production in Lon protease-deficient K . pneumoniae significantly increased, and the effect was accompanied by an increase of RmpA2 stability . The expression of the rmpA2 gene was negatively autoregulated and could be activated when the organism was grown in M9 minimal medium . An IS3 element located upstream of the rmpA2 was required for the full activation of the rmpA2 promoter . In summary, our results suggest that the enhancement of K2 CPS synthesis in K . pneumoniae CG43 by RmpA2 can be attributed to its transcriptional activation of K2 cps genes, and the expression level of rmpA2 is autoregulated and under the control of Lon protease.

Chemosphere, 2003 Mar, 50(8), 1055 - 61
Biotransformation of cyanide to methane and ammonia by Klebsiella oxytoca; Kao CM et al.; Klebsiella oxytoca, isolated from cyanide-containing industrial wastewater, was shown to be able to biodegrade cyanide to non-toxic endproducts using cyanide as the sole nitrogen source . In this study, ammonia was one of the detected endproduct of cyanide biodegradation by the concentrated resting cells of K . oxytoca . Moreover, cyanide has been shown to be biotransformed to methane through the actions of concentrated resting cells . Biodegradation of cyanide by cell-free extracts was not observed, which might be due to the inactivation of nitrogenase (an oxygen-labial enzyme) caused by the oxygen exposure after cell disruption . Results show that the cyanide consumption by resting cells of K . oxytoca was induced when the pretreatment of these cells with cyanide was conducted . However, the cyanide-degrading capability of resting cells pretreated with ammonia was inhibited . The inhibition of cyanide degradation by resting cells of K . oxytoca was affected by the ammonia concentration . This might result from the suppression of nitrogenase activity of K . oxytoca by ammonia since nitrogenase was suggested to be the sole cyanide-degrading enzyme during the cyanide degradation process . Results from this study also show that the processes of cyanide biodegradation and ammonia production by resting cells occurred simultaneously . This suggests that the utilization of cyanide as nitrogen source by K . oxytoca might proceed using ammonia as an assimilatory substrate .

Vaccine, 2003 Jan 17, 21(5-6), 458 - 67
B- and T-cell responses to the mycobacterium surface antigen PstS-1 in the respiratory tract and adjacent tissues . Role of adjuvants and routes of immunization; Rodriguez A et al.; Induction of mucosal immunity in the respiratory tract is crucial for protection against respiratory infections . Here, we have investigated the effects of the routes of immunization as well as of three different adjuvants on the induction of mucosal immune responses . Mice were immunized using intranasal (i.n.) or intraperitoneal (i.p.) routes with the mycobacterium PstS-1 antigen . Cholera toxin (CT), detoxified pertussis toxin (detPT) and RU 41.740 from Klebsiella pneumoniae were compared as mucosal adjuvants . Our data showed that i.n . route of immunization induced the most favorable stimulation of mucosal antigen-specific IgA responses supported by mixed Th cells producing IL-4, IL-5, IFN-gamma . In contrast, i.p . immunizations elicited only enhancement of systemic responses, predominantly of the Th2 type . Furthermore, the use of CT as mucosal adjuvant resulted in the stimulation of a mixed Th cell response whereas detPT evoked mainly Th2 type of responses . Likewise CT, the RU 41.740 adjuvant elicited a mixed Th cell response, albeit supported by much lower numbers of CD4(+) T-cells . Thus, i.n . route of immunization favors the induction of mucosal and systemic immune responses, while the Th cell development at mucosal inductive site is influenced by the adjuvant used for immunizations.

Zh Mikrobiol Epidemiol Immunobiol, 2002 Sep-Oct, (5), 48 - 53
{Specific biological features of opportunistic bacteria determining dysbacteriosis in the composition of the large intestine normal microflora}; Levanova LA et al.; Dysbiotic manifestations in the gastrointestinal tract are widely spread . They are characterized by a prolonged, persistent course with the tendency to transition to the chronic form and poorly respond to corrective treatment . The occurrence of high concentrations is 70% for Klebsiella oxytoca, while for K . pneumoniae--within the limit of 30% . The most topical problem is colonization of the intestinal mucosa in children aged up to 1 year by Klebsiella, the seeded bacteria retaining their capacity for growth for up to 2-3 years despite the use of probiotics . As shown in this study, the occurrence of Klebsiella and the level of the antilysozyme and "antiinterferon" activity of these bacteria, their resistance to antimicrobial preparations increase, correlating with the level of dysbiotic disturbances.

Pharm Res, 2002 Dec, 19(12), 1801 - 7
Locally delivered polyclonal antibodies potentiate intravenous antibiotic efficacy against gram-negative infections; Barekzi NA et al.; PURPOSE: Comparison of the anti-microbial efficacy of locally delivered antibodies in tandem with conventional systemic administration of ceftazidime antibiotic therapy in two lethal gram-negative animal infection models . METHODS: Previously published lethal E . coli-induced closed peritonitis and Klebsiella-induced burn wound infections were generated in outbred female CF-1 mice cohorts . Pooled human polyclonal antibodies were injected locally into sites of infection in these mice simultaneously with intravenous infusions of the broad-spectrum antibiotic, ceftazidime . Mouse survival was compared in sham control cohorts vs . both ceftazidime-alone or antibody-alone systemically infused cohorts as well as local antibody-systemic ceftazidime combination therapy cohorts . Microbial burdens in blood and tissue samples (by agar plating), as well as interleukin-6 cytokine levels (using ELISA) correlated with sepsis, were monitored in sacrificed animals as a function of antimicrobial treatment regimen . RESULTS: Local delivery of human polyclonal antibodies to infection sites was shown to produce synergistic therapeutic efficacy in combination with systemic antibiotic administration in these lethal wound infection models in mice . Enhanced benefits of the unique combination therapy included host survival, bacterial burden both locally and systemically, and IL-6 levels in host serum . CONCLUSIONS: Commercial pooled human antibodies contain a broad spectrum of antimicrobial activity against gram-negative pathogens . Prevention of systemization of infection correlates with host survival in these models . Local control of infection using doses of local, high-titer polyclonal antibodies can enhance traditional approaches to curb systemic spread of infection using intravenous antibiotics . Antibodies provide antimicrobial efficacy independent of known pathogen resistance mechanisms.

Structure (Camb), 2003 Jan, 11(1), 109 - 19
Structure of glycerol dehydratase reactivase: a new type of molecular chaperone; Liao DI et al.; The function of glycerol dehydratase (GDH) reactivase is to remove damaged coenzyme B(12) from GDH that has suffered mechanism-based inactivation . The structure of GDH reactivase from Klebsiella pneumoniae was determined at 2.4 A resolution by the single isomorphous replacement with anomalous signal (SIR/AS) method . Each tetramer contains two elongated 63 kDa alpha subunits and two globular 14 kDa beta subunits . The alpha subunit contains structural features resembling both GroEL and Hsp70 groups of chaperones, and it appears chaperone like in its interactions with ATP . The fold of the beta subunit resembles that of the beta subunit of glycerol dehydratase, except that it lacks some coenzyme B(12) binding elements . A hypothesis for the reactivation mechanism of reactivase is proposed based on these structural features.

J Synchrotron Radiat, 2003 Jan 1, 10(Pt 1), 71 - 5 Epub 2002 Dec 24.
XAFS studies of nitrogenase: the MoFe and VFe proteins and the use of crystallographic coordinates in three-dimensional EXAFS data analysis; Strange RW et al.; This paper reports a three-dimensional EXAFS refinement of the Mo coordination sphere of the FeMoco cluster of the dithionite-reduced MoFe protein from Klebsiella pneumoniae nitrogenase (Kp1) using the 1.6 A-resolution crystallographic coordinates . At this resolution, the positions of the heavy (Fe and S) atoms of the cluster are well determined and there is excellent agreement between the crystallographic and EXAFS models . However, the lighter homocitrate and histidine ligands are poorly determined in the crystal structure, and it is shown that the application of EXAFS-derived distance restraints during the early stages of crystallographic refinement provides a means of substantially improving (by approximately 0.1 A) the final crystallographic model . The consistency of the EXAFS analysis with the crystallographic information in this case justifies applications of EXAFS to cases where protein crystal structures are absent . Thus, the VFe protein of V-nitrogenase has been shown by EXAFS to possess a V-atom site catalytically similar to the well characterized MoFe-nitrogenases, with V replacing Mo.

J Bacteriol, 2003 Jan, 185(2), 688 - 92
Isolation of a negative control mutant of the nitrogen assimilation control protein, NAC, in Klebsiella aerogenes; Janes BK et al.; A negative control mutant of the nitrogen assimilation control protein, NAC, has been isolated . Mutants with the leucine at position 111 changed to a nonhydrophobic residue activate transcription from hut and ure promoters, but fail to repress gdhA expression . This failure does not result from failure to bind to either of the two sites required for gdhA repression, but the binding at those sites is altered in the mutant . It appears that the NAC negative control mutants fail to form the complex structures (probably tetramers) formed by wild-type NAC at the gdhA promoter.

Zh Mikrobiol Epidemiol Immunobiol, 2002 Nov-Dec, (6), 13 - 8
{Effect of metal oxides on the growth, hemolytic and serologic properties of Klebsiella pneumoniae}; Aleksakhina NN et al.; Silicon, dysprosium, germanium, yttrium, iron, cobalt, samarium, lutecium oxides, as well as the mixture of 8 metal oxides, at a concentration of 20 g/l were found to produce a stimulating or inhibiting effect on the growth of K . pneumoniae strains 204 and K-9 . Silicon, dysprosium, germanium and yttrium oxides were shown to stimulate the growth of K . pneumoniae strain 204 . Iron, cobalt, samarium and lutecium oxides, as well as the mixtures of all oxides under study, inhibited the growth of this strain . Silicon, samarium and lutecium oxides produced no effect on the growth of K . pneumoniae strain K-9; at the same time germanium and yttrium oxides stimulated the growth of these bacteria, while dysprosium, iron, cobalt oxides, as well as the mixture of all oxides, inhibited their growth . The presence of metal oxides did not change the serological activity of the cultures of both strains growing old, i.e . by 24 hours of their growth . The addition of silicon, germanium and iron oxides to the culture medium increased the hemolytic activity of K . pneumoniae strain K-9 seven to ninefold in comparison with the control grown in a synthetic nutrient medium without metal oxides . The comparison of these two strains (K-9 and 204) revealed that K . pneumoniae strain K-9 possessed greater hemolytic activity.

J Biol Chem, 2003 Mar 7, 278(10), 7863 - 74 Epub 2002 Dec 28.
Kinetic and catalytic properties of dimeric KpnI DNA methyltransferase; Bheemanaik S et al.; KpnI DNA-(N(6)-adenine)-methyltransferase (KpnI MTase) is a member of a restriction-modification (R-M) system in Klebsiella pneumoniae and recognizes the sequence 5'-GGTACC-3' . It modifies the recognition sequence by transferring the methyl group from S-adenosyl-l-methionine (AdoMet) to the N(6) position of adenine residue . KpnI MTase occurs as a dimer in solution as shown by gel filtration and chemical cross-linking analysis . The nonlinear dependence of methylation activity on enzyme concentration indicates that the functionally active form of the enzyme is also a dimer . Product inhibition studies with KpnI MTase showed that S-adenosyl-l-homocysteine is a competitive inhibitor with respect to AdoMet and noncompetitive inhibitor with respect to DNA . The methylated DNA showed noncompetitive inhibition with respect to both DNA and AdoMet . A reduction in the rate of methylation was observed at high concentrations of duplex DNA . The kinetic analysis where AdoMet binds first followed by DNA, supports an ordered bi bi mechanism . After methyl transfer, methylated DNA dissociates followed by S-adenosyl-l-homocysteine . Isotope-partitioning analysis showed that KpnI MTase-AdoMet complex is catalytically active.

Acta Crystallogr D Biol Crystallogr, 2003 Jan, 59(Pt 1), 150 - 1 Epub 2002 Dec 19.
Expression, crystallization and preliminary X-ray analysis of isomaltulose synthase (PalI) from Klebsiella sp . LX3; Li N et al.; Isomaltulose synthase (PalI) catalyzes the hydrolysis of the alpha-1,2 bond between the glucose and fructose moieties of sucrose and the formation of alpha-1,6 and alpha-1,1 bonds between the two components to produce isomaltulose (alpha-D-glucosylpyranosyl-1,6-D-fructofranose) and trehalulose (alpha-D-glucosylpyranosyl-1,1-D-fructofranose), respectively . The PalI protein has been overexpressed, purified and crystallized at 295 K using the hanging-drop vapour-diffusion method . The crystals diffract to 2.2 A resolution using synchrotron radiation and belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 59.239, b = 94.153, c = 111.294 A.

Antimicrob Agents Chemother, 2003 Jan, 47(1), 342 - 9
In34, a complex In5 family class 1 integron containing orf513 and dfrA10; Partridge SR et al.; A complex class 1 integron, In34, found in a conjugative plasmid from a multidrug-resistant Klebsiella pneumoniae strain isolated in 1997 at a hospital in Sydney, Australia, was shown to have a backbone related to that of In2, which belongs to the In5 family . In In34, the aadB gene cassette replaces the aadA1a cassette in In2, and two additional resistance genes, dfrA10 and aphA1, that are not part of a gene cassette are present . The aphA1 gene is in a Tn4352-like transposon that is located in the tniA gene . The dfrA10 gene lies adjacent to a 2,154-bp DNA segment, known as the common region, that contains an open reading frame predicting a product of 513 amino acids (Orf513) . Orf513 is 66 and 55% identical to the products of two further open reading frames that, like the common region, are found adjacent to antibiotic resistance genes . A 27-bp conserved sequence was found at one end of each type of common region . The loss of dfrA10 due to homologous recombination between flanking direct repeats and incorporation of the excised circle by homologous recombination were demonstrated . Part of In34 is identical to the sequenced portion of In7, which is from a multidrug-resistant Escherichia coli strain that had been isolated 19 years earlier in the same hospital . In34 and In7 are in plasmids that contain the same six resistance genes conferring resistance to ampicillin, chloramphenicol, gentamicin, kanamycin, neomycin, tobramycin, trimethoprim, and sulfonamides, but the plasmid backbones appear to be unrelated, suggesting that translocation of a multiple-drug-resistance-determining region as well as horizontal transfer may have occurred.

Zhonghua Jie He He Hu Xi Za Zhi, 2002 Oct, 25(10), 607 - 9
{Drug resistance and genotyping of Klebsiella pneumoniae in lower respiratory tract infection}; Shi J et al.; OBJECTIVE: To understand the current drug resistance of Klebsiella pneumoniae in lower respiratory tract infection and the prevalence trend of extended spectrum beta-lactamase (ESBLs)-producing Klebsiella pneumoniae . METHODS: Drug resistance in 86 strains of Klebsiella pneumoniae and ESBLs-producing strains were genotyped by randomly amplified DNA polymorphisms (RAPD) . RESULTS: ESBLs-producing strains accounted for 20.9% of the 86 strains, and 83.3% of the ESBLs-producing strains were from respiratory ICU . The drug resistance rate of ESBLs positive bacteria was much higher than that of ESBLs negative bacteria . ESBLs positive bacteria were resistant to both the quinolones and a minoglylosides . Eighteen ESBLs-producing strains were divided into 12 types . 3 strains had the same profile (genotype A), and 5 strains had an identical profile (genotype B), but other strains had different profiles . CONCLUSIONS: Chromosomal DNA analysis proved that certain types of ESBLs-producing Klebsiella pneumoniae were epidemic in respiratory ICU . RAPD was an economic, simple but reliable method in epidemic study.

Chin Med J (Engl), 2002 Oct, 115(10), 1479 - 82
Epidemiological and antibiotic resistant study on extended-spectrum beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae in Zhejiang Province; Yu Y et al.; OBJECTIVE: To investigate the epidemiological status of extended-spectrum beta-lactamase (ESBL) producing Escherichia coli (E . coli) and Klebsiella pneumoniae (K . pneumoniae) and the drug resistance profiles of such organisms . METHODS: A total of 282 clinical isolates of E . coli and 180 of K . pneumoniae were collected from different districts of Zhejiang Province . Inhibitor potentiated broth dilution tests were performed for detecting extended-spectrum beta-lactamases . Etests were performed to detect the drug resistance of these strains against nine commonly used antibiotics . RESULTS: The prevalence of extended-spectrum beta-lactamases in E . coli and K . pneumoniae was 34.0% and 38.3%, respectively . The average prevalence of extended-spectrum beta-lactamases in E . coli and K . pneumoniae was 35.7% . The resistance prevalence of extended spectrum beta-lactamase producing strains to ceftazidime and cefotaxime was 40% and 26% respectively, so were those to cefepime, cefoxitin, piperacillin-tazobactam, cefoperazone-sulbactam, amikacin and ciprofloxacin . All these strains were sensitive to imipenem . CONCLUSION: The results in this study showed that the prevalence of extended-spectrum beta-lactamases was high, while extended-spectrum beta-lactamase producing strains were resistant to most antimicrobial agents except imipenem.

J Korean Med Sci, 2002 Dec, 17(6), 778 - 83
Glutamine on the luminal microbial environment after massive small bowel resection; Choi SH et al.; To evaluate the oral glutamine (GLN) on the luminal microbes and bacterial translocation (BT) in short bowel, 45 Wistar rats were utilized in three groups; A (control), and B and C (short bowel, 85% of small bowel resected) . The group A was fed with elemental diet (EmD), B with EmD+2% glycine, and C with EmD+2% GLN . The groups B and C were isocaloric and isonitrogenous . Wet weight, DNA, protein, and histomorphometry of the mucosa and parallel microbial culture from cecal contents, caval blood, and tissue blocks of the liver, spleen, and mesenteric lymph nodes were performed on the 5th, 10th, and 15th day . Mucosal growth was higher in group C than B . Colony forming units (CFU) from cecal contents increased more in group B than in C . BTs in A, B, and C were 7/15, 8/15, and 2/15, respectively . Total CFUs in blood and tissues were 5.8 X 10(4)/g, 5.5 X 10(6)/g, and 1.8 X 10(4)/g, respectively . As for BT, the most frequent organism was Klebsiella in A (79.3%), but E . coli in B and C (94.2% and 55.6%) . GLN seems to suppress luminal microbes, and reduces BT in short bowel due to enforced barrier function and proliferation of the mucosa.

Am J Trop Med Hyg, 2002 Nov, 67(5), 524 - 7
Hepatic hydatid cysts with superinfection in a non-endemic area in Taiwan; Chen YC et al.; Hepatic hydatid cysts are extremely rare in Taiwan . We report a case of complicated, multivesicular, hydatid cysts in the liver and a secondary infection with Klebsiella pneumoniae . The cysts were detected in an early stage by ultrasonography and computed tomography and treated successfully with radical resection.

Kaohsiung J Med Sci, 2002 Aug, 18(8), 386 - 92
Space infection of the head and neck; Wang LF et al.; Deep neck infection may be lethal, especially when life-threatening complications occur . We conducted a retrospective analysis of 184 patients with deep neck infection who were treated at Kaohsiung Medical University Hospital during the past 6 years . Factors such as age, sex, hospitalization days, clinical presentations, involved spaces, imaging studies, microbiology, and treatment method were analyzed . There were 122 men and 62 women with a mean age of 41.7 years . The average hospitalization was 8.4 days . The involved spaces, determined by physical examination and radiologic findings, were the peritonsillar space (59 patients), parapharyngeal space (77 patients), submandibular space (55 patients), and retropharyngeal space (20 patients) . Of the 49 patients for whom the origin of infection was identified, 29 were infected via the upper respiratory tract and 13 had infection of odontogenic origin . The most common isolated organism was Klebsiella pneumoniae . One hundred and thirty-seven patients (74.5%) underwent surgery, including repeated needle aspiration (87 patients) and surgical drainage (50 patients) . The remaining 47 patients recovered uneventfully with antibiotic therapy alone . Eighteen patients developed life-threatening complications, such as descending mediastinitis, sepsis, airway obstruction, and jugular vein thrombosis . Two patients died of septic shock . The combination of accurate diagnosis, effective antibiotic therapy, airway maintenance, and intensive surgical debridement for those who fail to respond to conservative treatment will lead to a good prognosis.

Sex Transm Infect, 2002 Dec, 78(6), 452 - 7
Donovanosis; O'Farrell N; Donovanosis, a chronic cause of genital ulceration, has recently been the subject of renewed interest after a long period of relative obscurity . The causative organism, Calymmatobacterium granulomatis, has been cultured for the first time in many years and a polymerase chain reaction diagnostic using a colorimetric detection system has been developed . Phylogenetic analysis confirms close similarities with the genus Klebsiella and a proposal made that C granulomatis be reclassified as Klebsiella granulomatis comb nov . Azithromycin has emerged as the drug of choice and should be used if the diagnosis is confirmed or suspected . In donovanosis endemic areas, syndromic management protocols for genital ulceration may need to be adapted locally . A significant donovanosis epidemic was reported in Durban from 1988-97 but the current status of this epidemic is unclear . The donovanosis elimination programme among Aboriginals in Australia appears successful and is a model that could be adopted in other donovanosis endemic areas . Overall, the incidence of donovanosis seems to be decreasing . Increased attention would undoubtedly be paid to donovanosis if policy makers recognised more readily the importance of genital ulcers in fuelling the HIV epidemic.

J Clin Microbiol, 2002 Dec, 40(12), 4666 - 9
Molecular epidemiology of extended-spectrum beta-lactamase-producing, fluoroquinolone-resistant isolates of Klebsiella pneumoniae in Taiwan; Yu WL et al.; Strains of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae (ESBL-KP) have emerged worldwide . Concomitant ciprofloxacin resistance with ESBL production in K . pneumoniae isolates would severely restrict treatment options . Among 39 (18.5%) of 211 ESBL-KP isolates resistant to ciprofloxacin (MIC, >/=4 micro g/ml), 37 (95%) were high level resistant (MIC, >/=16 micro g/ml) . These isolates were also cross resistant to the newer fluoroquinolones, including levofloxacin, gatifloxacin, gemifloxacin, and garenoxacin (BMS 284756) . Sitafloxacin was most active against these ciprofloxacin-resistant ESBL-KP isolates with MICs for 67% of the isolates being </=2 micro g/ml . The molecular epidemiology of these multiresistant isolates was investigated by automated ribotyping and pulsed-field gel electrophoresis (PFGE) . Ribotyping identified 18 different strains among the 39 ciprofloxacin-resistant ESBL-KP isolates . The majority (67%) of these isolates were contained in six ribogroups which were further confirmed by PFGE . The distribution of the six major strains of ciprofloxacin-resistant ESBL-KP within Taiwan included one (ribogroup 255.3-PFGE type E) with a nationwide distribution and several institution-specific strains . Interhospital cooperation appears necessary, with strict infection control practices coupled with restriction of fluoroquinolone and extended-spectrum beta-lactam use to control both the major epidemic strain and the more diverse strains observed within individual institutions.

Intensive Care Med, 2002 Dec, 28(12), 1718 - 23 Epub 2002 Oct 17.
Extended-spectrum beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae bloodstream infection: risk factors and clinical outcome; Du B et al.; OBJECTIVES: To study the risk factor for nosocomial bacteremia caused by Escherichia coli or Klebsiella pneumoniae producing extended-spectrum beta-lactamase (ESBL) and the influence on patient outcome . DESIGN: Retrospective, single-center study of consecutive bacteremic patients . SETTINGS: A university-affiliated teaching hospital . PATIENTS: A total of 85 patients with nosocomial bacteremia due to E . coli or K . pneumoniae were enrolled . INTERVENTION: None . MEASUREMENTS AND MAIN RESULTS: The demographic characteristics and clinical information including treatment were recorded upon review of patients' records . The primary end point was hospital mortality . Twenty-seven percent of isolates produced ESBLs . Previous treatment with 3rd-generation cephalosporins was the only independent risk factor for bacteremia due to ESBL-producing pathogens {odds ratio (OR) 4.146, P=0.008} . Antibiotic treatment was considered appropriate in 71 cases (83%), and failed in 23 patients (27%) . Twenty-one patients (25%) died in the hospital . Antibiotic treatment failure was the only independent risk factor for hospital mortality (OR 15.376, P=0.001) . Inappropriate antibiotic treatment might lead to significantly higher mortality rate (7/14 vs 14/71, P=0.016) . Patients treated with imipenem were more likely to survive while those receiving cephalosporin treatment tended to have a poorer outcome (1/19 vs 14/40, P=0.023) . CONCLUSIONS: More judicious use of cephalosporins, especially 3rd-generation cephalosporins, may decrease ESBL-producing E . coli or K . pneumoniae bacteremia, and also improve patient outcome.

J Bacteriol, 2002 Dec, 184(24), 6966 - 75
Repression of glutamate dehydrogenase formation in Klebsiella aerogenes requires two binding sites for the nitrogen assimilation control protein, NAC; Goss TJ et al.; In Klebsiella aerogenes, the gdhA gene codes for glutamate dehydrogenase, one of the enzymes responsible for assimilating ammonia into glutamate . Expression of a gdhAp-lacZ transcriptional fusion was strongly repressed by the nitrogen assimilation control protein, NAC . This strong repression (>50-fold under conditions of severe nitrogen limitation) required the presence of two separate NAC binding sites centered at -89 and +57 relative to the start of gdhA transcription . Mutants lacking either or both of these sites lost the strong repression . The distance between the two sites was less important than the face of the helix on which they lay . Insertion or deletion of 10 bp between the sites had little effect on the strong repression, but insertion of 5 bp or deletion of either 5 or 15 bp decreased the repression significantly . We propose that the strong repression of gdhAp-lacZ expression requires an interaction between the NAC molecules bound at the two sites . A weaker repression of gdhAp-lacZ expression (about threefold) required only the NAC site centered at -89 . This weaker repression appears to result from NAC's ability to prevent the action of a positive effector the target of which overlaps the NAC binding site centered at -89 . Point mutations and deletions of this region result in the same threefold reduction in gdhAp-lacZ expression as the presence of NAC at this site.

Clin Microbiol Infect, 2002 Nov, 8(11), 715 - 24
Comparison of screening methods for TEM- and SHV-derived extended-spectrum beta-lactamase detection; MacKenzie FM et al.; OBJECTIVE: To compare common extended-spectrum beta-lactamase (ESBL) screening methods and beta-lactams for their ability to detect TEM- and SHV-related ESBL enzymes . METHODS: This study compared disk diffusion testing by NCCLS methodology, the Jarlier double disk test, a disk-on-disk test, a modified three-dimensional test and the E test method for their sensitivity and specificity in detecting TEM- and SHV-related ESBL producers . Three negative and 22 positive controls were studied . These were two Klebsiella pneumoniae and 23 Escherichia coli transconjugants . Seventeen beta-lactam antibiotics were tested: cefamandole, cefotetan, cefoxitin, cefuroxime, cefixime, cefoperazone, cefotaxime, cefpodoxime, cefsulodin, ceftazidime, ceftibuten, ceftizoxime, ceftriaxone, moxalactam, cefepime, cefpirome and aztreonam . RESULTS: NCCLS disk diffusion was 14% sensitive with ceftriaxone, 36% with cefotaxime, 64% with aztreonam, 68% with cefpodoxime, and 73% with ceftazidime . Cefoperazone, cefamandole, cefpodoxime and cefpirome showed 91% sensitivity using the Jarlier test . Using the disk-on-disk test, cefsulodin showed 95% sensitivity, and cefoperazone, cefepime and cefamandole showed 91% sensitivity . With the modified three-dimensional test, cefoperazone, cefpodoxime and cefpirome showed 91% sensitivity . CONCLUSIONS: For practical reasons, we would recommend use of either the Jarlier test or the commercial cephalosporin disks containing clavulanic acid to screen for ESBL producers . Cefoperazone, cefamandole, cefpodoxime and cefpirome showed good sensitivity across the methods tested.

Arch Intern Med, 2002 Nov 25, 162(21), 2469 - 77
Risk factors for fluoroquinolone resistance in nosocomial Escherichia coli and Klebsiella pneumoniae infections; Lautenbach E et al.; BACKGROUND: The incidence of fluoroquinolone (FQ) resistance has increased markedly in recent years . Even in the common nosocomial pathogens Escherichia coli and Klebsiella pneumoniae, in which the emergence of FQ resistance was believed to be unlikely, increasing resistance to these agents has been noted . Risk factors for FQ resistance in these pathogens remain unknown . Although FQs are important components of the present antimicrobial arsenal, their continued usefulness is threatened by rising FQ resistance . OBJECTIVE: To identify risk factors for nosocomial FQ resistance . METHODS: A case-control study of hospitalized patients with infections due to FQ-resistant and FQ-susceptible E coli and K pneumoniae occurring between January 1, 1998, and June 30, 1999 . RESULTS: We included 123 patients with nosocomial FQ-resistant infections and 70 randomly selected patients with nosocomial FQ-susceptible infections . Independent risk factors (adjusted odds ratio {95% confidence interval}) for FQ resistance were (1) recent FQ use (5.25 {1.81-15.26}); (2) residence in a long-term care facility (3.65 {1.64-8.15}); (3) recent aminoglycoside use (8.86 {1.71-45.99}); and (4) older age (1.03 {1.01-1.06}) . CONCLUSIONS: Recent FQ use, residence in a long-term care facility, recent aminoglycoside use, and older age were all noted to be independent risk factors for FQ resistance among patients with nosocomial E coli and K pneumoniae infections . Efforts should be directed at recognition and modification of these risk factors to curb the rise in FQ resistance and preserve the utility of these agents in the treatment of common nosocomial gram-negative infections.

Antimicrob Agents Chemother, 2002 Dec, 46(12), 3984 - 6
AcrAB Efflux System: Expression and Contribution to Fluoroquinolone Resistance in Klebsiella spp; Mazzariol A et al.; Seven Klebsiella pneumoniae and four Klebsiella oxytoca clinical isolates with different levels of resistance to ciprofloxacin were studied . Mutations in the topoisomerase genes were found in almost all strains, but the contribution of a multidrug efflux system homologous to AcrAB in Escherichia coli was also observed . Overexpression of this efflux system was demonstrated by immunoblotting with antibodies against E . coli AcrA.

Antimicrob Agents Chemother, 2002 Dec, 46(12), 3926 - 32
Energy-dependent accumulation of norfloxacin and porin expression in clinical isolates of Klebsiella pneumoniae and relationship to extended-spectrum beta-lactamase production; Martinez-Martinez L et al.; The relationships between porin deficiency, active efflux of fluoroquinolones, and extended-spectrum beta-lactamase (ESBL) production were determined for 53 clinical isolates of Klebsiella pneumoniae . Thirty-two ESBL-positive strains (including 22 strains expressing porins and 10 strains lacking porins) and 21 ESBL-negative strains were evaluated . Active efflux of norfloxacin was defined as a >/=50% increase in the accumulation of norfloxacin in the presence of carbonyl cyanide m-chlorophenylhydrazone (CCCP) in comparison with the corresponding basal value in the absence of CCCP . The quinolone resistance-determining regions of both gyrA and parC from 13 strains, representing all isolates with different porin profiles and with or without active efflux, were determined . Porin loss was significantly more common among ESBL-positive strains (10 of 32 {31.2%}) than among ESBL-negative strains (0 of 2 {0%}) (P < 0.01) . Active efflux was observed in 7 of 10 (70%) strains lacking porins and in 4 of 43 (9.3%) strains producing porins (P < 0.001) . The 11 strains showing active efflux corresponded to 3 of 21 (14.3%) ESBL-negative strains and 8 of 32 (25.5%) ESBL-positive strains (P > 0.05) . Basal values of norfloxacin accumulation were higher in strains lacking active efflux than in those that had this mechanism (P < 0.05) . In the absence of topoisomerase changes, the contribution of either porin loss or active efflux to fluoroquinolone resistance in K . pneumoniae was negligible . It is concluded that among K . pneumoniae strains of clinical origin, porin loss was observed only in those producing ESBL, and that a significant number of porin-deficient strains also expressed active efflux of norfloxacin . In terms of fluoroquinolone resistance, both mechanisms are significant only in the presence of topoisomerase modifications.

Antimicrob Agents Chemother, 2002 Dec, 46(12), 3837 - 42
Occurrence of newer beta-lactamases in Klebsiella pneumoniae isolates from 24 U.S . hospitals; Moland ES et al.; Despite the discovery of novel beta-lactamases such as extended-spectrum beta-lactamases (ESBLs), imported AmpC, and carbapenem-hydrolyzing beta-lactamases at least a decade ago, there remains a low level of awareness of their importance and how to detect them . There is a need to increase the levels of awareness of clinical laboratories about the detection of newer beta-lactamases . Therefore, a study was conducted in 2000 to investigate the occurrence of these beta-lactamases in Klebsiella pneumoniae isolates at 24 U.S . medical centers . To enhance the likelihood of detecting imported AmpC and carbapenem-hydrolyzing beta-lactamases, participating laboratories were permitted to include archived strains (1996 to 2000) that were intermediate or resistant to either cefoxitin or imipenem . The beta-lactamase production of 408 isolates positive by screening of 1,123 isolates was investigated by ESBL phenotypic confirmation tests; and for AmpC and carbapenem-hydrolyzing beta-lactamases, three-dimensional tests, isoelectric focusing, beta-lactamase inhibitor studies, spectrophotometric assays, induction assays, and molecular tests were used . ESBL-producing isolates were detected at 18 of the 24 sites (75%), imported AmpC-producing isolates were detected at 10 sites (42%), inducible imported AmpC-producing isolates were detected at 3 sites (12.5%), and a molecular class A carbapenem-hydrolyzing enzyme was detected at 1 site (4%) . No class B or D carbapenem-hydrolyzing enzymes were detected . ESBLs and imported AmpC beta-lactamases were detected at a significant number of sites, indicating widespread penetration of these enzymes into U.S . medical institutions . Because these enzymes may significantly affect therapeutic outcomes, it is vital that clinical laboratories be aware of them and be able to detect their occurrence.

Antimicrob Agents Chemother, 2002 Dec, 46(12), 3829 - 36
Mechanisms of decreased susceptibility to cefpodoxime in Escherichia coli; Oliver A et al.; Cefpodoxime is one of five antimicrobial agents recommended by the National Committee for Clinical Laboratory Standards for screening isolates of Klebsiella spp . and Escherichia coli for extended-spectrum beta-lactamase (ESBL) production . In a prior study, we noted that among 131 E . coli isolates for which the MIC of at least one extended-spectrum cephalosporin (ESC) or aztreonam was > or =2 micro g/ml (suggesting the presence of ESBL production), there were 59 isolates (45.0%) for which the MIC of cefpodoxime was 2 to 4 micro g/ml (i.e., a positive ESBL screening test), but the MICs of ceftazidime, cefotaxime, and ceftriaxone were < or =1 micro g/ml (below the ESBL screening breakpoint) . Thus, the results appeared to be false-positive ESBL screening tests . These 59 isolates were divided into five phenotypic groups based on the susceptibility patterns of the organisms to a variety of beta-lactam agents and further characterized . The first group (32 isolates) all produced a TEM-1 beta-lactamase, and changes in the major outer membrane proteins were detected in representative strains . The second group (18 isolates) lacked bla(TEM) but showed a number of porin changes; some also showed a modest elevation in production of the AmpC chromosomal beta-lactamase . In the third phenotypic group (seven isolates) all expressed an OXA-30 beta-lactamase . Some also harbored altered porins . The two remaining phenotypes each had a distinct pattern of porin changes with or without beta-lactamase production . These data indicate that several factors are associated with decreased susceptibility to cefpodoxime in E . coli, but none of the mechanisms are related to ESBL production . Current screening methods produced false-positive ESBL results for these isolates . Such isolates should not be classified as containing ESBLs, nor should interpretations of ESCs or aztreonam susceptibility be changed to resistant on test reports for these isolates.

Planta, 2002 Nov, 216(1), 129 - 35 Epub 2002 Nov 12.
How the mutation glycine96 to alanine confers glyphosate insensitivity to 5-enolpyruvyl shikimate-3-phosphate synthase from Escherichia coli; Eschenburg S et al.; The enzyme 5-enolpyruvyl shikimate-3-phosphate (EPSP) synthase (EC 2.5.1.19) is essential for the biosynthesis of aromatic compounds in plants and microbes and is the unique target of the herbicide glyphosate . One of the first glyphosate-insensitive enzymes reported was a Gly96Ala mutant of EPSP synthase from Klebsiella pneumoniae . We have introduced this single-site mutation into the highly homologous EPSP synthase from Escherichia coli . The mutant enzyme is insensitive to glyphosate with unaltered affinity for its first substrate, shikimate-3-phosphate (S3P), but displays a 30-fold lower affinity for its second substrate, phosphoenolpyruvate (PEP) . Using X-ray crystallography, we solved the structure of Gly96Ala-EPSP synthase liganded with S3P to 0.17 nm resolution . The crystal structure shows that the additional methyl group from Ala96 protrudes into the active site of the enzyme . While the interactions between enzyme and S3P remain unaffected, the accessible volume for glyphosate binding is substantially reduced . Exploiting the crystallographic results for molecular modeling, we demonstrate that PEP but not glyphosate can be docked in the Gly96Ala-modified binding site . The predicted PEP binding site satisfies the earlier proposed interaction pattern for PEP with EPSP synthase and corroborates the assumption that glyphosate and PEP target the same binding site.

Indian Pediatr, 2002 Oct, 39(10), 952 - 7
Febrile episodes in childhood malignancies; Dubey AP et al.; The objectives of the present report were to study the site of infections and pathogenic organisms during febrile episodes in different childhood malignant conditions, to correlate febrile episode with Absolute Neutrophil Count (ANC) and to know the sensitivity pattern of bacteria to different antibiotics so as to know the most appropriate antibiotic regimen in these children . The study material comprised of forty two febrile episodes occurring in children aged lt 12 years with various malignancies . All the episodes were worked up in detail including complete history, physical examination and relevant hematological, microbiological and radiological investigations . Out of the 42 episodes, 15 (36%) occurred in children with acute leukemias, 20 (48%) in children with lymphomas and 7(17%) in children with solid tumors . 26 (62%) episodes were seen in children during chemotherapy, while 12% each in freshly diagnosed and remission and 14% in relapse cases . 12 (28%) episodes occurred in children with ANC < 500/mm3 . 36% were microbiologically confirmed . Klebsiella species was the commonest organism isolated followed by E . coli . Maximum sensitivity (75%) was seen with ciprofloxacin against both Klebsiella species and E.coli.

Curr Rheumatol Rep, 2002 Dec, 4(6), 458 - 65
Gastrointestinal issues in children with rheumatologic disease; Chehade M et al.; Distinctive arthritic patterns, some of which may parallel or even precede intestinal disease activity, are seen in inflammatory bowel disease . Some spondyloarthropathies are associated with transient ileocolic inflammation . Vasculitis frequently affects the gastrointestinal tract, predominantly manifesting with abdominal pain . In severe cases, intestinal ischemia and perforation may occur . Various arthritides are thought to be associated with other gastrointestinal diseases, such as celiac disease and hepatitis . The association between intestinal disease and arthritis is still being investigated . Interactions between the inflammatory intestinal cells and inflamed synovial cells have been demonstrated . Certain intestinal bacteria such as Klebsiella pneumoniae are suspected to play a role as triggers for the development of arthropathies . Genetic factors, especially human leukocyte antigen associations, are also being increasingly investigated for better characterization of the types of arthritis and possible prognostic implications . Various therapies, including nonsteroidal anti-inflammatory drugs, used to treat rheumatologic diseases have the potential to cause gastrointestinal complications.

Clin Microbiol Infect, 2002 Sep, 8(9), 579 - 88
Transferable plasmid mediating resistance to multiple antimicrobial agents in Klebsiella pneumoniae isolates in Greece; Galani I et al.; OBJECTIVE: To investigate the underlying resistance mechanisms in 10 Klebsiella pneumoniae isolates . METHODS: Ten K . pneumoniae strains according to distinct bacteriocin typing and REP-PCR, were examined for their plasmid content, their ability to transfer their resistance to aminoglycosides and third-generation cephalosporins, and their production of aminoglycoside-modifying enzymes and beta-lactamases . RESULTS: Transfer of resistance to the above-mentioned antibiotics as well as to co-trimoxazole and tetracycline in Escherichia coli strain RC 85 at a frequency of 5-106 was achieved for all strains by conjugation . Similar strains harbor a self-transferable multiresistant plasmid (80 kb) with similar EcoRI and HindIII restriction patterns . This plasmid encodes an extended-spectrum beta-lactamase which confers high-level resistance to third-generation cephalosporins and aztreonam . It produces SHV-5 beta-lactamase, as demonstrated by isoelectric focusing and DNA sequencing . Aminoglycoside resistance was co-transferred, and AAC(6')-I, mediating resistance to gentamicin, tobramycin, netilmicin and amikacin, and AAC(3)-I, mediating resistance to gentamicin and sisomycin, were encoded in all isolates and their transconjugants, while APH(3')-I, mediating resistance to kanamycin and neomycin, was encoded in seven strains . CONCLUSIONS: It appears that a multiresistant transferable plasmid encoding the SHV-5 beta-lactamase, causing unusually high resistance to ceftazidime and aztreonam, and the combination AAC(6')-I + AAC(3)-I of acetylating enzymes causing, also resistance to all clinically available aminoglycosides, is established in K . pneumoniae in Greece.

J Clin Microbiol, 2002 Nov, 40(11), 4030 - 6
Identification and broad dissemination of the CTX-M-14 beta-lactamase in different Escherichia coli strains in the northwest area of Spain; Bou G et al.; During the course of a molecular epidemiology study of mechanisms of antibiotic resistance in the area served by our hospital (516,000 inhabitants), we isolated the gene encoding CTX-M-14 beta-lactamase . Thirty clinical strains (27 Escherichia coli and 3 Klebsiella pneumoniae isolates) with a phenotype of extended-spectrum beta-lactamase were collected from January to October 2001 and studied for the presence of the CTX-M-14 beta- lactamase gene . By isoelectric point determination, PCR, and nucleotide sequencing, we detected the presence of this gene in 17 E . coli strains belonging to 15 different genotypes (REP-PCR) causing infections in 17 different patients . Epidemiological studies based on medical records did not suggest any relationship between the patients infected with these E . coli strains and, interestingly, 7 of 30 patients harboring strains with extended-spectrum beta-lactamases never had contact with the hospital environment before the clinical E . coli isolation . Conjugation experiments revealed that this gene was plasmid mediated in the 17 E . coli strains, and plasmid restriction fragment length polymorphisms showed 9 different patterns in the 17 E . coli strains . By PCR, the sequence of the tnpA transposase gene of the insert sequence ISEcp-1 was detected in all the plasmids harboring the CTX-M-14 gene . These results strongly suggest that plasmid dissemination between different E . coli strains in addition to a mobile element (transposon) around the beta-lactamase gene may be involved in the spreading of the CTX-M-14 gene . This study reinforces the hypothesis that the epidemiology of the prevalence of the beta-lactamase genes is changing and should alert the medical community to the increase in the emergence of the CTX-M beta-lactamases worldwide.

Appl Environ Microbiol, 2002 Nov, 68(11), 5336 - 41
Biodegradation of hexahydro-1,3,5-trinitro-1,3,5-triazine and its mononitroso derivative hexahydro-1-nitroso-3,5-dinitro-1,3,5-triazine by Klebsiella pneumoniae strain SCZ-1 isolated from an anaerobic sludge; Zhao JS et al.; In previous work, we found that an anaerobic sludge efficiently degraded hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX), but the role of isolates in the degradation process was unknown . Recently, we isolated a facultatively anaerobic bacterium, identified as Klebsiella pneumoniae strain SCZ-1, using MIDI and the 16S rRNA method from this sludge and employed it to degrade RDX . Strain SCZ-1 degraded RDX to formaldehyde (HCHO), methanol (CH3OH) (12% of total C), carbon dioxide (CO(2)) (72% of total C), and nitrous oxide (N2O) (60% of total N) through intermediary formation of methylenedinitramine (O(2)NNHCH(2)NHNO(2)) . Likewise, hexahydro-1-nitroso-3,5-dinitro-1,3,5-triazine (MNX) was degraded to HCHO, CH3OH, and N2O (16.5%) with a removal rate (0.39 micromol . h(-1) . g {dry weight} of cells(-1)) similar to that of RDX (0.41 micromol . h(-1) . g {dry weight} of cells(-1)) (biomass, 0.91 g {dry weight} of cells . liter(-1)) . These findings suggested the possible involvement of a common initial reaction, possibly denitration, followed by ring cleavage and decomposition in water . The trace amounts of MNX detected during RDX degradation and the trace amounts of hexahydro-1,3-dinitroso-5-nitro-1,3,5-triazine detected during MNX degradation suggested that another minor degradation pathway was also present that reduced -NO2 groups to the corresponding -NO groups.

Biochemistry, 2002 Nov 5, 41(44), 13253 - 63
Multiple inequivalent metal-nucleotide coordination environments in the presence of the VO2+-inhibited nitrogenase iron protein: pH-dependent structural rearrangements at the nucleotide binding site; Petersen J et al.; Nitrogenase naturally requires adenosine nucleoside triphosphates and divalent metal cations for catalytic activity . Their energy of hydrolysis controls several mechanistic functions, most probably via separate structural conformers of the nitrogenase Fe protein . To characterize the ligand environment of the divalent metal in the ternary complex, with ADP or ATP and the Fe protein from Klebsiella pneumoniae, the hyperfine structures have been investigated by electron paramagnetic resonance (EPR) spectroscopy by substituting naturally occurring diamagnetic Mg(2+) by paramagnetic oxovanadium . This metal replacement leads to inhibition of nitrogenase activity . Moreover, depending on pH, two distinctly different VO(2+) EPR spectra are detected . At pH 7.4 each of the vanadyl EPR hyperfine lines is further split into two . This indicates that several spectroscopically distinguishable metal coordination environments coexist for VO(2+)-nucleotide chelate complexes in the presence of the reduced Fe protein . Overall, a total of at least three distinct local metal coordination environments have been identified . We report the EPR parameters for each of the disparate metal coordinations measured at different pH values with ADP and ATP bound . EPR spectra have also been recorded for the oxidized Fe protein showing essentially similar spectra to that of the reduced protein . The EPR parameters of VO-nucleotides in the presence of the Fe protein are consistent, for all metal coordination environments, with direct metal ligation by nucleotide phosphate groups and the formation of mononucleotide complexes . The nucleotide binding environment with the highest ligand field strength is compatible with a metal coordination structure that is also found in various G-proteins with GTP bound . No significant EPR line width change is detected after exchange into D(2)O buffer solution for any of the pH forms although differences exist between the pH forms . The missing difference between the EPR parameters in the presence of ADP or ATP suggests that there is little or no conformational rearrangement between these two forms; this contrasts with behavior of G-proteins that undergo substantial conformational changes upon hydrolysis . This could be related to the inhibition of nitrogenase by VO(2+).

Nat Struct Biol, 2002 Dec, 9(12), 966 - 72
Modeling and experiment yields the structure of acireductone dioxygenase from Klebsiella pneumoniae; Pochapsky TC et al.; Here we report the structure of acireductone dioxygenase (ARD), the first determined for a new family of metalloenzymes . ARD represents a branch point in the methionine salvage pathway leading from methylthioadenosine to methionine and has been shown to catalyze different reactions depending on the type of metal ion bound in the active site . The solution structure of nickel-containing ARD (Ni-ARD) was determined using NMR methods . X-ray absorption spectroscopy, assignment of hyperfine shifted NMR resonances and conserved domain homology were used to model the metal-binding site because of the paramagnetism of the bound Ni2+ . Although there is no structure in the Protein Data Bank within 3 A r.m.s deviation of that of Ni-ARD, the enzyme active site is located in a conserved double-stranded b-helix domain . Furthermore, the proposed Ni-ARD active site shows significant post-facto structural homology to the active sites of several metalloenzymes in the cupin superfamily.

FEMS Microbiol Lett, 2002 Oct 8, 215(2), 255 - 9
Klebsiella pneumoniae KE-1 degrades endosulfan without formation of the toxic metabolite, endosulfan sulfate; Kwon GS et al.; For bioremediation of toxic endosulfan, endosulfan degradation bacteria, which do not form toxic endosulfan sulfate, were isolated from various soil samples using endosulfan as sole carbon and energy source . Among the 40 isolated bacteria, strain KE-1, which was identified as Klebsiella pneumoniae by physiological and 16S rDNA sequence analysis, showed superior endosulfan degradation activity . Analysis of culture pH, growth, free sulfate and endosulfan and its metabolites demonstrated that KE-1 biologically degrades 8.72 microg endosulfan ml(-1) day(-1) when incubated with 93.9 microg ml(-1) endosulfan for 10 days without formation of toxic endosulfan sulfate . Our results suggest that K . pneumoniae KE-1 degraded endosulfan by a non-oxidative pathway and that strain KE-1 has potential as a biocatalyst for endosulfan bioremediation.

J Hosp Infect, 2002 Oct, 52(2), 99 - 106
Clinical implications of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae bacteraemia; Kim BN et al.; To identify the clinical implications of extended-spectrum beta-lactamase (ESBL) production, 162 cases of Klebsiella pneumoniae bacteraemia in 154 adults were analysed . Of these cases, 44 (27.2%) were ESBL-producing (ESBLKP) . Common sources of ESBLKP bacteraemia included primary bacteraemia (34.1%) and biliary infection (29.5%) . The placement of a biliary drainage catheter, nosocomial acquisition, and prior antibiotic therapy were independently associated with ESBL production in multivariate analysis . More cases of ESBLKP than non-ESBLKP received inappropriate antibiotic therapy before culture results were reported (54.5 vs . 3.4%; P = 0.001) . In 19 cases of ESBLKP, no significant difference in mortality was observed between patients who received appropriate empiric antibiotic therapy and those who did not (26.3 vs . 20.8%; P = 0.67) . The mean length of hospital stay after the onset of bacteraemia was longer in the cases of ESBLKP than in the cases of non-ESBLKP (39.6 vs . 23.9 days; P = 0.008) . Directly related mortality was not significantly different between the cases of ESBLKP and the cases of non-ESBLKP (23.3 vs . 20.0%; P = 0.65) . None of the patients with biliary infection due to ESBLKP died (0/12; P = 0.03) . In conclusion, ESBL production was not significantly associated with death but it had a considerable impact on patients with K . pneumoniae bacteraemia .

Antimicrob Agents Chemother, 2002 Nov, 46(11), 3679 - 82
Expression of SHV-2 beta-lactamase and of reduced amounts of OmpK36 porin in Klebsiella pneumoniae results in increased resistance to cephalosporins and carbapenems; Crowley B et al.; A Klebsiella pneumoniae clinical isolate was resistant to cefoxitin, cefotaxime, ceftazidime, ceftazidime-clavulanate, piperacillin-tazobactam (MICs, >256 micro g/ml in all cases), and meropenem (MIC, 16 micro g/ml) and was intermediate to imipenem (MIC, 8 micro g/ml) . Decreased expression of the OmpK36 porin and expression of an SHV-2 beta-lactamase contributed to the observed resistance to these beta-lactam-containing agents.

Antimicrob Agents Chemother, 2002 Nov, 46(11), 3422 - 7
Complete nucleotide sequence of Klebsiella pneumoniae multiresistance plasmid pJHCMW1; Sarno R et al.; The multiresistance plasmid pJHCMW1, harbored by a clinical Klebsiella pneumoniae strain isolated from a neonate with meningitis, was sequenced . A circular sequence of 11,354 bp was generated, of which 7,993 bp make up Tn1331, a transposon including the antibiotic resistance genes aac(6')-Ib, aadA1, bla(OXA-9), and bla(TEM-1) . The gene aac(6')-Ib is included in a gene cassette, and both aadA1 and bla(OXA-9) are included in a single-gene cassette that may have arisen as a consequence of a recombination event involving two integrons . The pJHCMW1 plasmid replicates through a ColE1-like RNA-regulated mechanism, includes a functional oriT, and two loci with similarity to XerCD site-specific recombination target sites involved in plasmid stabilization by the resolution of multimers . One of these two loci, mwr, is active and has been the subject of previous studies, and the other, dxs, is not functional but binds the recombinase XerD with low affinity . Two additional open reading frames were identified, one with low similarity to two hypothetical membrane proteins from Mycobacterium tuberculosis and Mycobacterium leprae and the other with low similarity to psiB, a gene encoding a function that facilitates the establishment of the transferring plasmid in the recipient bacterial cell during the process of conjugation.

J Clin Neurosci, 2002 Sep, 9(5), 533 - 8
Klebsiella meningitis in adults: clinical features, prognostic factors and therapeutic outcomes; Lu CH et al.; Sixty adult Klebsiella meningitis patients have been identified at Kaohsiung Chang Gung Memorial Hospital in a period of 13 years . Most cases were associated with debilitating diseases, and devastating metastatic septic abscesses are common in diabetic patients with K . pneumoniae meningitis . Although the mortality rate has been significantly reduced in recent years, there has been an increase in nosocomial infections and the emergence of multi-antibiotic resistant strains . Significant prognostic factors include appropriate antibiotic therapy, the presence of septic shock, disseminated intravascular coagulation, and high cerebrospinal fluid protein levels and white blood cell counts . Initial empiric therapy with a third generation cephalosporin should be considered for community-acquired meningitis while antibiotics such as carbapenems should be considered as initial empiric therapy for patients with postneurosurgical meningitis . Early diagnosis and the use of appropriate antibiotics are of crucial importance.

Br J Pharmacol, 2002 Nov, 137(5), 621 - 8
Role of the platelet-activating factor (PAF) receptor during pulmonary infection with gram negative bacteria; Soares AC et al.; The lipid mediator PAF plays an important role in the phagocytosis of particles, including bacteria, and consequent production of pro-inflammatory cytokines, such as TNF-alpha and IL-8 . Using a PAF receptor antagonist (UK-74,505) and PAF receptor knock-out mice, we have investigated the relevance of PAF for the inflammatory changes and lethality after pulmonary infection with the gram-negative bacteria Klebsiella pneumoniae in mice . At an inoculum of 3 x 10(6) bacteria, there was marked pulmonary (bronchoalveolar lavage and lung) neutrophilia that started early (2.5 h after infection) and peaked at 48 h . All animals were dead by day 4 of infection . The chemokine KC and the pro-inflammatory cytokine TNF-alpha increased rapidly and persisted for 48 h in the lungs . Pretreatment with UK-74,505 (30 mg kg(-1) per day, p.o.) had no significant effects on the number of infiltrating neutrophils in BAL fluid or lung tissue, as assessed by histology and measuring myeloperoxidase, or on the concentrations of KC . In contrast, concentrations of TNF-alpha and the number of bacteria inside neutrophils were significantly diminished . In order to support a role for the PAF during K . pneumoniae infection, experiments were also carried out in PAFR-deficient mice . In the latter animals, lethality occurred earlier than in wild-type controls . This was associated with greater number of bacteria in lung tissue and diminished percentage of neutrophils containing bacteria in their cytoplasm . Our results suggest that PAF, acting on its receptor, plays a protective role during infection with K . pneumoniae in mice.

J Microbiol Immunol Infect, 2002 Sep, 35(3), 179 - 83
A pilot study of oral fleroxacin once daily compared with conventional therapy in patients with pyogenic liver abscess; Chen YW et al.; The object of this open-label, randomized, comparative study was to evaluate the efficacy and safety of fleroxacin versus conventional therapy (cefazolin plus gentamicin/cephalexin) in the treatment of patients with bacterial liver abscess . Thirty-one adult patients (26 men and 5 women) received fleroxacin 400 mg orally once daily for 3 weeks, and 30 adult patients (21 men and 9 women) received conventional therapy for 3 to 4 weeks . Patients was assessed on day 3 of treatment, thereafter every week during treatment, and at 7 to 14 days (compulsory follow-up) after treatment for assessment of bacteriologic, clinical, and safety parameters . A total of 20 patients in the fleroxacin group and 22 patients in the conventional therapy group were evaluated . Klebsiella pneumoniae was the predominant pathogen isolated in all evaluable cases . Bacteriologic cure was achieved in 14 (70%) of 20 patients on fleroxacin therapy compared with 18 (81.8%) of 22 patients on conventional therapy (p=0.48) . Clinical cure was achieved in 12 (60%) and 18 (81.8%) patients, and improvement in 2 (10%) and 1 (4.5%) patients in the fleroxacin and conventional therapy group, respectively . Most of adverse effects were of mild intensity . Oral fleroxacin once-daily administration is an effective, alternative treatment of bacterial liver abscess.

Infect Immun, 2002 Nov, 70(11), 6310 - 8
Divergent role of gamma interferon in a murine model of pulmonary versus systemic Klebsiella pneumoniae infection; Moore TA et al.; Klebsiella pneumoniae is a leading cause of both community-acquired and nosocomial gram-negative-bacterial pneumonia . A further clinical complication of pulmonary K . pneumoniae infection is dissemination of bacteria from the lung into the peripheral blood, resulting in bacteremia concurrent with the localized pulmonary infection . Here, we report studies detailing the divergent role of gamma interferon (IFN-gamma) in pulmonary versus systemic K . pneumoniae infection . Intratracheal inoculation of IFN-gamma knockout mice resulted in significantly increased mortality compared to that observed for wild-type infected animals . Increased mortality correlated with a 100-fold increase in pulmonary bacteria within 2 days postinfection and upregulation of lung-associated interleukin-10 (IL-10) mRNA . Interestingly, IFN-gamma knockout mice had a twofold reduction in plasma aminospartate transferase activity, indicating diminished liver injury following peripheral blood bacterial dissemination . To study the host response towards blood-borne bacteria in the absence of the ongoing pulmonary infection, intravenous inoculation studies were initiated . IFN-gamma knockout mice were no more susceptible to intravenous infection than their wild-type counterparts . The consistent observation in IFN-gamma knockout mice was for improved survival correlating with increased clearance of blood- and liver-associated bacteria . Intravenous inoculation resulted in a two- to threefold increase in hepatic IL-10 production 24 and 48 h postinfection . Liver injury was also significantly reduced in IFN-gamma knockout mice . These data indicate that IFN-gamma secretion is a critical mediator in the resolution of localized gram-negative pulmonary pneumonia . Surprisingly, host responses towards systemic infection with the same bacteria appear to be IFN-gamma independent.

Biochemistry, 2002 Oct 22, 41(42), 12607 - 17
Substrate-induced conformational change of a coenzyme B12-dependent enzyme: crystal structure of the substrate-free form of diol dehydratase; Shibata N et al.; Substrate binding triggers catalytic radical formation through the cobalt-carbon bond homolysis in coenzyme B12-dependent enzymes . We have determined the crystal structure of the substrate-free form of Klebsiella oxytoca diol dehydratase*cyanocobalamin complex at 1.85 A resolution . The structure contains two units of the heterotrimer consisting of alpha, beta, and gamma subunits . As compared with the structure of its substrate-bound form, the beta subunits are tilted by approximately 3 degrees and cobalamin is also tilted so that pyrrole rings A and D are significantly lifted up toward the substrate-binding site, whereas pyrrole rings B and C are only slightly lifted up . The structure revealed that the potassium ion in the substrate-binding site of the substrate-free enzyme is also heptacoordinated; that is, two oxygen atoms of two water molecules coordinate to it instead of the substrate hydroxyls . A modeling study in which the structures of both the cobalamin moiety and the adenine ring of the coenzyme were superimposed onto those of the enzyme-bound cyanocobalamin and the adenine ring-binding pocket, respectively, demonstrated that the distortions of the Co-C bond in the substrate-free form are already marked but slightly smaller than those in the substrate-bound form . It was thus strongly suggested that the Co-C bond becomes largely activated (labilized) when the coenzyme binds to the apoenzyme even in the absence of substrate and undergoes homolysis through the substrate-induced conformational changes of the enzyme . Kinetic coupling of Co-C bond homolysis with hydrogen abstraction from the substrate shifts the equilibrium to dissociation.

Int J Mol Med, 2002 Nov, 10(5), 627 - 30
Clinicopathological analysis of liver abscess in Japan; Okano H et al.; Currently, pyogenic liver abscess is not frequent, but it is a severe infectious disease . However a strategy for the effective treatment of liver abscess is not established . We analyzed 75 cases of liver abscess over an eight year period and evaluated their prognosis, any associated underlying disease, or the effect of percutaneous transhepatic abscess drainage (PTAD) . For all 75 cases, laboratory data were analyzed and imaging studies were performed . Next, PTAD and antibiotic administration were started on these cases as first choice treatments . These treatments were continued until the laboratory data of the patient were restored to within the normal range . Those cases that were PTAD non-effective or required operation for underlying diseases, underwent operations . Of the total 75 cases, 63 survived after treatment and 12 cases died . Bacteria were detected in 50 cases and Klebsiella pneumoniae was detected in 31 of these 50 cases, but 25 out of 75 cases were negative . The biliary system was the main route of infection . PTAD was effective, especially in cases that were complicated with disseminated intravascular coagulation (DIC) or acute renal failure (ARF) . PTAD is an effective treatment for liver abscess, it is especially useful in the restoration of severe general conditions as indicated by this study.

J Clin Microbiol, 2002 Oct, 40(10), 3703 - 11
Evaluation of the Etest ESBL and the BD Phoenix, VITEK 1, and VITEK 2 automated instruments for detection of extended-spectrum beta-lactamases in multiresistant Escherichia coli and Klebsiella spp; Leverstein-van Hall MA et al.; Seventy-four isolates of multiresistant Escherichia coli and Klebsiella spp . recovered during a 3-year period and 17 control strains with genotypically identified beta-lactamases were tested for the production of extended-spectrum beta-lactamases (ESBLs) by using the Etest and the VITEK 1, VITEK 2, and Phoenix automated instruments . The use of the Etest was evaluated by investigating its accuracy in detecting the ESBLs of the control strains and by comparing interpretation results of laboratory technicians and experts . The accuracy of the Etest was 94% . With the Etest as the reference for the clinical strains and the genotype as the reference for the control strains, the automated instruments detected the ESBLs with accuracies of 78% (VITEK 2), 83% (VITEK 1), and 89% (Phoenix) . No significant difference between the systems with regard to the control strains was detected . The VITEK 2 did, however, perform less well than the Phoenix (P = 0.03) on the collection of clinical isolates, mainly because of its high percentage of indeterminate test results (11%) . No significant difference between the performances of the VITEK 1 and either the VITEK 2 or the Phoenix was found . However, because of its associated BDXpert system the Phoenix showed the best performance . The Etest was found to be an accurate test but was limited by its indeterminate results (4%), its inability to differentiate between K1 hyperproduction and ESBLs, questionable guidelines concerning mutants inside the inhibition zones, and the inability of the technicians to recognize subtle zone deformations.

Shock, 2002 Sep, 18(3), 248 - 54
An essential role for lipopolysaccharide-binding protein in pulmonary innate immune responses; Fan MH et al.; Lipopolysaccharide (LPS)-binding protein (LBP) greatly facilitates LPS activation of monocytic cells through the CD14 receptor, triggering activation of innate immune responses . An acute phase protein, LBP is produced predominantly by the liver; however, we and others have shown that LBP is produced extrahepatically in multiple locations, including the lung . The importance of LBP in the lung has remained unclear . LBP may make the host more acutely sensitive to LPS and development of septic complications; alternatively, it may be protective, aiding in detection, opsonization, and killing of bacteria . Our objective was to determine the role LBP plays in local pulmonary immune defenses to bacterial challenge . LBP knockout mice and age-matched C57BL/6 wild-type controls were challenged with direct intratracheal inoculation of Klebsiella pneumoniae . We observed a significant increase in mortality, earlier onset of bacteremia, and greater pulmonary bacterial loads in LBP knockout mice compared with controls . Total lung myeloperoxidase (MPO) activity, neutrophil recruitment to the alveolar space, and levels of KC--a chemokine involved in neutrophil recruitment--in bronchoalveolar lavage (BAL) fluid and lung homogenates were found to be significantly diminished in knockout mice compared with controls . Together, our findings suggest that LBP is essential in local pulmonary innate immune responses against bacteria.

Nihon Kokyuki Gakkai Zasshi, 2002 Jun, 40(6), 530 - 5
{Four cases of Klebsiella pneumonia}; Tsukadaira A et al.; Typical Klebsiella pneumonia with mucous sputum is known as an opportunistic nosocomial infection . However, computed tomographic study of limiting in Klebsiella pneumonia is rare . We report three types of chest computed tomography (CT) findings for Klebsiella pneumonia . Case 1 shows typical lobar pneumonia (Friedlander pneumonia), Cases 2 and 3 show acute bronchopneumonia with subclinical aspiration, and Case 4, chronic Klebsiella pneumonia with typical cavitary lung abscesses . Of these four cases of Klebsiella pneumonia, three developed in the right lung, as determined radiologically, but esophagogastroduodenoscopy indicated that the lesions of Case 3 had developed in the left lingula and upper lobe.

J Int Med Res, 2002 Jul-Aug, 30(4), 445 - 8
Detection of antimicrobial resistance and extended-spectrum beta-lactamase production in Klebsiella pneumoniae strains from infected neonates; Aktas E et al.; The present study was designed to determine the antimicrobial resistance and extended-spectrum beta-lactamase (ESBL) activities of Klebsiella pneumoniae strains isolated from the neonatal intensive care unit of Ataturk University Hospital, Erzurum, Turkey . Antibiotic susceptibility of 40 isolates was detected by the standard disk diffusion method according to the National Committee for Clinical Laboratory Standards Guidelines . The double-disk synergy method was used to determine ESBL activity, which is associated with resistance to beta-lactam antibiotics . Twenty-four (60%) of 40 K . pneumoniae strains were found to produce ESBL . Of the antibiotics tested, meropenem was found to be the most effective (100%), and ampicillin the least effective (0%) . With the increasing incidence of antimicrobial resistance, which poses a clinically significant risk to vulnerable patients, it is important that clinical microbiology laboratories have accurate and timely information concerning the strains of bacteria present to enable them to predict which antibiotics are likely to be effective in treating the infections they may cause.

Nucleic Acids Res, 2002 Sep 15, 30(18), 3972 - 80
Secondary structure and DNA binding by the C-terminal domain of the transcriptional activator NifA from Klebsiella pneumoniae; Ray P et al.; The NifA protein of Klebsiella pneumoniae is required for transcriptional activation of all nitrogen fixation (nif) operons except the regulatory nifLA genes . At these operons, NifA binds to an upstream activator sequence (UAS), with the consensus TGT-N(10)-ACA, via a C-terminal DNA-binding domain (CTD) . Binding of the activator to this upstream enhancer-like sequence allows NifA to interact with RNA polymerase containing the alternative sigma factor, sigma(54) . The isolated NifA CTD is monomeric and binds specifically to DNA in vitro as shown by DNase I footprinting . Heteronuclear 3D NMR experiments have been used to assign the signals from the protein backbone . Three alpha-helices have been identified, based on secondary chemical shifts and medium range Halpha(i)-NH(i)( + 1), and NH(i)-NH(i)( + 1) NOEs . On addition of DNA containing a half-site UAS, several changes are observed in the NMR spectra, allowing the identification of residues that are most likely to interact with DNA . These occur in the final two helices of the protein, directly confirming that DNA binding is mediated by a helix-turn-helix motif.

Eur J Biochem, 2002 Sep, 269(18), 4484 - 94
The crystal structure of coenzyme B12-dependent glycerol dehydratase in complex with cobalamin and propane-1,2-diol; Yamanishi M et al.; Recombinant glycerol dehydratase of Klebsiella pneumoniae was purified to homogeneity . The subunit composition of the enzyme was most probably alpha 2 beta 2 gamma 2 . When (R)- and (S)-propane-1,2-diols were used independently as substrates, the rate with the (R)-enantiomer was 2.5 times faster than that with the (S)-isomer . In contrast to diol dehydratase, an isofunctional enzyme, the affinity of the enzyme for the (S)-isomer was essentially the same or only slightly higher than that for the (R)-isomer (Km(R)/Km(S) = 1.5) . The crystal structure of glycerol dehydratase in complex with cyanocobalamin and propane-1,2-diol was determined at 2.1 A resolution . The enzyme exists as a dimer of the alpha beta gamma heterotrimer . Cobalamin is bound at the interface between the alpha and beta subunits in the so-called 'base-on' mode with 5,6-dimethylbenzimidazole of the nucleotide moiety coordinating to the cobalt atom . The electron density of the cyano group was almost unobservable, suggesting that the cyanocobalamin was reduced to cob(II)alamin by X-ray irradiation . The active site is in a (beta/alpha)8 barrel that was formed by a central region of the alpha subunit . The substrate propane-1,2-diol and essential cofactor K+ are bound inside the (beta/alpha)8 barrel above the corrin ring of cobalamin . K+ is hepta-coordinated by the two hydroxyls of the substrate and five oxygen atoms from the active-site residues . These structural features are quite similar to those of diol dehydratase . A closer contact between the alpha and beta subunits in glycerol dehydratase may be reminiscent of the higher affinity of the enzyme for adenosylcobalamin than that of diol dehydratase . Although racemic propane-1,2-diol was used for crystallization, the substrate bound to glycerol dehydratase was assigned to the (R)-isomer . This is in clear contrast to diol dehydratase and accounts for the difference between the two enzymes in the susceptibility of suicide inactivation by glycerol.

Clin Infect Dis, 2002 Oct 1, 35(7), 834 - 41 Epub 2002 Sep 10.
Molecular epidemiology of a citywide outbreak of extended-spectrum beta-lactamase-producing Klebsiella pneumoniae infection; Quale JM et al.; Multidrug-resistant strains of Klebsiella pneumoniae are a problem in many hospitals . In 1999, the molecular epidemiology of K . pneumoniae with extended-spectrum beta-lactamases (ESBLs) was studied at 15 hospitals in Brooklyn . Of 824 unique patient isolates, 34% were presumptive ESBL producers . Of this subset, 34% were susceptible to cefoxitin, 42% to ciprofloxacin, 48% to ceftriaxone, 55% to piperacillin-tazobactam, 57% to amikacin, and 86% to cefepime . Ribotype analysis revealed 87 unique types . However, 2 clusters accounted for 35% of isolates and were present in most of the hospitals . One cluster was significantly more resistant to most antibiotics . Although there was a predominance of SHV-5, considerable heterogeneity of beta-lactamases was evident, even among isolates of the same cluster . A correlation was found between the use of cephalosporins and the prevalence of ESBL-producing strains of K . pneumoniae at each hospital . Our data suggest that there is an advanced outbreak of multidrug-resistant K . pneumonia infection that is affecting all Brooklyn hospitals.

Biosci Biotechnol Biochem, 2002 Jul, 66(7), 1524 - 30
Structural analysis of an extracellular polysaccharide bioflocculant of Klebsiella pneumoniae; Kobayashi T et al.; The glycoside composition and sequence of an extracellular polysaccharide flocculant of Klebsiella pneumoniae H12 was analyzed . GC and HPLC analysis of the acid-hydrolysate identified its constituent monosaccharides as D-Glc, D-Man, D-Gal, and D-GlcA in an approximate molar ratio of 3.9:1.0:2.3:3.6 . To analyze the glycoside sequence, the polysaccharide was partially hydrolyzed by acid and enzyme treatment . GC, HPLC, TLC, MALDI-TOF/MS, and 1H- and 13C- NMR spectroscopy characterized the obtained oligosaccharides . The results clarified the partial structure of H12 polysaccharide as a linear polymer of a unit of pentasaccharide with a side chain of one D-GlcA to D-Glc moiety (see below) . Although the existence of other sequences or other constituent glycosides could not be fully excluded, H12 polysaccharide must be a novel types as such a complicated unit for a polymer has not so far been reported . The partial structure of a H12 polysaccharide flocculant is also discussed in this report . {structure: see text}

J Microbiol Methods, 2002 Nov, 51(3), 407 - 10
Further modification of the Hodge test to screen AmpC beta-lactamase (CMY-1)-producing strains of Escherichia coli and Klebsiella pneumoniae; Yong D et al.; Cefoxitin-resistant Escherichia coli and Klebsiella pneumoniae are widespread in Korea . Significant proportions of them are considered to be CMY-1 producers . For effective screening of CMY-1 producers, the Hodge test was modified by using a cefoxitin disk and the performance was evaluated . The sensitivity and specifity of the test were 100% and 94.9%, respectively . The test was easier to perform than the three-dimensional extract test . This modified test should be suitable for screening CMY-1-producing strains of E . coli and K . pneumoniae .

Plasmid, 2002 Jul, 48(1), 73 - 6
Chromosomal sequences from Klebsiella pneumoniae flank the SHV-5 extended-spectrum beta-lactamase gene in pACM1; Preston KE et al.; The nucleotide sequence was determined for Anon 13, a 1250-bp SmaI fragment located approximately 2.8 kb downstream from bla(SHV-5) in pACM1 . Anon 13 is 99% identical to a segment of the unpublished sequence of the Klebsiella pneumoniae chromosome . Genes of the K . pneumoniae sequence are undefined, but conceptual amino acid translations of two ORFs in Anon 13 are homologous to L-fuculose-1-phosphate aldolase (FucA) and a conserved hypothetical protein present in the chromosomes of several species of bacteria . In addition, restriction mapping indicates that the region of homology between the K . pneumoniae chromosome and pACM1 is as least 7.9 kb and includes both Anon 13 and bla(SHV) . These observations demonstrate the chromosomal origin of the bla(SHV-5) on pACM1.

Indian J Chest Dis Allied Sci, 2002 Jul-Sep, 44(3), 173 - 6
Antibiotic resistance among gram-negative bacteria of lower respiratory tract secretions in hospitalized patients; Navaneeth BV et al.; Data on antibiotic resistance pattern of gram-negative bacterial isolates of lower respiratory tract secretions of hospitalized patients were fed into WHONET computer and analyzed for the year 1999 . Out of 860 samples, 269 (31.2%) were culture positive . Gram-negative bacteria (GNB) accounted for 238 (88.4%) positive samples . Non-fermenting gram-negative bacteria (NFGNB) were found in 34% samples, the other common ones being Klebsiella spp (29.8%) and Pseudomonas spp (17.2%) . GNB isolates from tracheal aspirates and sputum were 132 (55.4%) and 106 (44.5%) respectively . Adults (32.7%) and elderly patients (24.3%) recorded higher isolation of GNB as compared to pediatric patients (1.6%) . The highest mean resistance among predominant GNB in both tracheal aspirate (96.6%) and sputum (86.9%) was noted to ampicillin while the lowest mean resistance in tracheal aspirate (28%) and sputum (14.3%) was to amikacin . NFGNB of tracheal aspirates and sputum showed highest resistance of 50% and 32% to amikacin, respectively . Pseudomonas spp showed the highest variation in the resistance pattern between tracheal aspirates and sputum samples . Overall mean resistance was highest among tracheal aspirate isolates compared to sputum isolates.

Surgery, 2002 Aug, 132(2), 408 - 14
Gram-negative bacteria killed by complement are associated with more severe biliary infections and produce more tumor necrosis factor-alpha in sera; Stewart L et al.; BACKGROUND: We previously showed that gallstones contain bacteria and that illness severity correlates with bacterial presence . This study examined virulence differences of gram-negative biliary bacteria . METHODS: Gallstones and bile were cultured, and sera obtained, from 210 patients . Infection severity was staged as: none-no clinical infection; moderate-fever, leukocytosis; or severe-bacteremia, cholangitis, hypotension, abscess, or organ failure . Gram-negative biliary bacteria were tested against patient (and control) serum for complement-mediated bacterial killing and induction of tumor necrosis factor-alpha (TNFalpha) production (using cultured monocytes) with and without sera . These results were correlated with infection severity . RESULTS: A total of 98 (47%) patients had biliary bacteria . Infection severity distribution was none, 29%; moderate, 35%; and severe, 36% . Gram-negative organisms killed by complement were associated with more severe infections as follows: 13%, none; 60%, moderate; and 88%, severe infections (P =.024 and P <.0001, respectively vs none, chi-square test) . TNFalpha production in sera increased 182 pg/mL with complement resistant bacteria, but increased 546 pg/mL with bacteria killed by complement (P <.0001, killed vs not killed, Student's t test) . E coli and Klebsiella were the most virulent bacterial species . They were cultured from blood, usually killed by complement, and had the largest increase in TNFalpha production in sera . CONCLUSIONS: Gram-negative biliary bacteria killed by complement (as opposed to complement-resistant) were associated with more serious biliary infections including bacteremia and induced more TNFalpha production in sera . This suggests a potential role for complement activation and cytokine production in biliary sepsis.

J Immunol, 2002 Sep 15, 169(6), 3267 - 74
Surfactant protein D binds selectively to Klebsiella pneumoniae lipopolysaccharides containing mannose-rich O-antigens; Sahly H et al.; Surfactant protein D (SP-D) plays important roles in the regulation of innate immune responses in the lung . We have previously shown that SP-D can agglutinate and enhance the macrophage-dependent killing of specific unencapsulated phase variants of Klebsiella pneumoniae . In the present studies, we used 16 clinical isolates of Klebsiella representing four O-serotypes and examined the interaction of SP-D with their isolated LPSs . Although SP-D bound to the core oligosaccharide of rough LPS from all isolates, it selectively bound to smooth forms of LPS expressed by O-serotypes with mannose-rich repeating units in their O-polysaccharides . SP-D was more potent in agglutinating unencapsulated phase variants of O-serotypes expressing these SP-D "reactive" O-polysaccharides, and more effectively inhibited the adhesion of these serotypes to lung epithelial cells . This novel anti-adhesion activity required the multimerization of trimeric SP-D subunits (dodecamers) . Klebsiella serotypes expressing "nonreactive" LPS O-Ags were isolated at a significantly higher frequency from patients with K . pneumoniae . Our findings suggest that SP-D plays important roles in the clearance of opportunistic Gram-negative bacteria and contributes to known serotypic differences in the pathogenicity of Klebsiella through specific interactions with O-polysaccharides.

Chin Med J (Engl), 2002 Aug, 115(8), 1158 - 62
Klebsiella pneumoniae: epidemiology and analysis of risk factors for infections caused by resistant strains; Li J et al.; OBJECTIVE: To analyze the epidemiology of hospital and community-acquired infections caused by Klebsiella pneumoniae (K . pneumoniae) and risk factors for infections caused by resistant strains . METHODS: A retrospective observational study was performed to analyze the relationship between antimicrobial use and bacterial resistance . RESULTS: A K . pneumoniae infection was diagnosed in 0.47% of patients (169 of 36 179) admitted to the hospital between 1 March 1999 and 31 August 2000 . Of the 169 isolates, 166 (98.2%) were resistant to at least one antimicrobial and 91.1% (154/169) to two or more antibiotics . 98% were resistant to ampicillin, 77% to piperacillin, 64% to cephalothin, 60% to ampicillin/sulbactam, 59% to cefoperazone, 57% to cefazolin, 55% to cefuroxime, 51% to TMP-SMZ, 51% to tobramycin, 50% to gentamicin, 49% to aztreonam, cefetaxime and ceftriaxone respectively, 47% to ceftazidime, 47% to cefepime, 46% to ciprofloxacin, 46% to ticarcillin/clavulanate, 44% to amikacin, 38% to cefoxitin, 22% to piperacillin/tazobactam, while all strains were tested susceptible to imipenem . CONCLUSIONS: Prior receipt of amtimicrobial therapy was significantly associated with infection caused by a resistant organism and most strains were resistant to multiple antibiotics.

J Antimicrob Chemother, 2002 Sep, 50(3), 421 - 4
Significance of low-level resistance to ciprofloxacin in Klebsiella pneumoniae and the effect of increased dosage of ciprofloxacin in vivo using the rat granuloma pouch model; Fuursted K et al.; This study was designed to compare the killing effect of ciprofloxacin on strains of Klebsiella pneumoniae with different MICs of ciprofloxacin in vivo using the rat granuloma pouch infection model . Five different strains were used: one ciprofloxacin-susceptible strain (MIC 0.06 mg/L); one strain highly resistant to ciprofloxacin (MIC 8 mg/L); and three nalidixic acid-resistant strains with low-level resistance to ciprofloxacin (MIC 0.25-0.5 mg/L) . The efficacy of ciprofloxacin was evaluated 3 h after bacterial challenge (treating an acute infection) or after 3 days (treating a late infection) with a single intraperitoneal injection of ciprofloxacin (40 and 200 mg/kg) . Ciprofloxacin was bactericidal against both growing K . pneumoniae (acute infection model) and non-growing K . pneumoniae (late infection model), but the extent of killing was significantly higher on growing bacteria and against ciprofloxacin-susceptible K . pneumoniae . A peak concentration of ciprofloxacin, at the infection site, <3 x MIC was not sufficient for optimal bacterial elimination . However, it was possible to compensate for the lower killing in low-level ciprofloxacin-resistant K . pneumoniae by increasing the dosage of ciprofloxacin from 40 to 200 mg/kg, consistent with the higher MIC.

Clin Microbiol Infect, 2002 Jul, 8(7), 397 - 404
Klebsiella typing: pulsed-field gel electrophoresis (PFGE) in comparison with O:K-serotyping; Hansen DS et al.; OBJECTIVE: To compare pulsed-field gel electrophoresis (PFGE) typing and O:K-serotyping of Klebsiella in two different epidemiological settings . METHODS: One hundred and four bacteremia isolates without known epidemiological relation and 47 isolates from an outbreak in a neonatal intensive care unit (NICU) were K-typed by countercurrent immunoelectrophoresis (CCIE), O-typed by an inhibition enzyme-linked immunosorbent assay method, and typed by pulsed-field gel electrophoresis (PFGE) using the restriction enzyme XbaI . RESULTS: Typing data for the 104 bacteremia isolates were compared with regard to typability, number of types, maximum number of isolates per type, and the Discriminative Index (DI) . O-typing combined with K-typing (DI 0.98) as O:K-serotyping (DI 0.99) gave a very discriminative typing system, whereas O-typing alone was not very discriminative (DI 0.76) . PFGE (DI 1.00) was a more discriminative typing method than O:K-serotyping, as it could subdivide 13/22 O:K-serotypes into smaller groups . Isolates with the same PFGE-type had the same O:K-serotype, indicating that isolates with different O- and/or K-types could be expected to be of different PFGE-types . Typing of the 47 isolates from the outbreak in the NICU showed that 38 isolates belonged to a single clone, and that during an epidemic limited in time and space, differences in the electrophoretic patterns of up to five bands between a parental pattern type and a subtype may be found in the PFGE profiles . CONCLUSIONS: Both O:K-serotyping and PFGE typing are highly discriminative typing methods . PFGE is the most discriminative method and is excellent for typing outbreaks with few isolates . If large numbers of isolates are to be typed, a more convenient strategy might be first to K- or O:K-serotype isolates followed by PFGE typing of possible identical isolates . Since K- or O:K-serotyping is a definitive typing method, while PFGE typing is a comparative one, PFGE cannot, for the time being, replace O:K-serotyping for surveillance purposes.

Biochem Soc Trans, 2002 Aug, 30(4), 672 - 4
Mechanisms of mercury bioremediation; Essa AM et al.; Mercury is one of the most toxic heavy metals, and has significant industrial and agricultural uses . These uses have led to severe localized mercury pollution . Mercury volatilization after its reduction to the metallic form by mercury-resistant bacteria has been reported as a mechanism for mercury bioremediation {Brunke, Deckwer, Frischmuth, Horn, Lunsdorf, Rhode, Rohricht, Timmis and Weppen (1993) FEMS Microbiol . Rev . 11, 145-152; von Canstein, Timmis, Deckwer and Wagner-Dobler (1999) Appl . Environ . Microbiol . 65, 5279-5284} . The reduction/volatilization system requires to be studied further, in order to eliminate the escape of the metallic mercury into the environment . Recently we have demonstrated three different mechanisms for mercury detoxification in one organism, Klebsiella pneumoniae M426, which may increase the capture efficiency of mercury.

Ann Trop Med Parasitol, 2002 Jul, 96(5), 513 - 6
Causative agents of liver abscess in those with liver cirrhosis: a 10-year case review of hospitalized patients in Thailand; Wiwanitkit V et al.; Although patients with cirrhosis of the liver show relative immunosuppression and therefore have increased susceptibility to most infections, they rarely develop liver abscesses . In a retrospective case review, the pathogens causing the liver abscesses observed, between January 1992 and December 2001 at the King Chulalongkorn Memorial Hospital, Bangkok, Thailand, in 44 hospitalized patients diagnosed as cases of liver cirrhosis were investigated . The most common clinical symptoms and signs of the abscesses--abdominal pain (80%), fever and chills (73%), and abdominal tenderness (73%)--were similar to those seen in non-cirrhotic patients with abscesses . The frequency of liver abscess among the cirrhotic patients was low (0.46%) . Most (71%) of the abscesses were in the right lobe and most (71%) of those with abscesses only had a single abscess . Surprisingly, many of the abscesses (36%) were apparently caused by amoebae . Bacterial pathogens were identified in eight patients (18%) by blood culture and 15 (34%) patients by pus culture . Seven (16%) of the blood cultures and 13 (30%) of the pus contained Gram-negative aerobes, indicating that such pathogens, particularly Klebsiella pneumoniae (in six pus and six blood cultures) and Escherichia coli (in three pus cultures and one blood), were the most common causes of the bacterial abscesses . Pus culture appeared more successful than blood culture for bacterial abscesses, and amoebic abscesses could always be identified by direct microscopical examination of pus samples . Aspiration of liver abscesses, to obtain pus samples for culture and microscopy, is therefore recommended.

Pathology, 2002 Aug, 34(4), 362 - 4
Detection of extended spectrum beta-lactamases in the routine clinical microbiology laboratory; Midolo PD et al.; AIMS: To compare three methods of confirming the presence of an extended spectrum beta-lactamase (ESBL) enzyme with the initial detection (i.e., screening) by the Vitek AMS . METHODS: Gram-negative bacteria which flagged as ESBL-positive in the Vitek GNS card, or were suspected of harbouring an enzyme, were further tested by each of the following methods: (a) combination disc test using cefpodoxime, ceftazidime and cefotaxime with and without clavulanate; (b) cefotaxime ESBL Etest; and (c) Jarlier keyhole method with cefpodoxime (10 microg), cefotaxime (5 microg) and aztreonam (30 microg) placed 15mm away from an augmentin (30 microg) disc . RESULTS: A total of 52 isolates were investigated, representing an 18-month time period . Fifty of these were positive by Vitek . Twenty-eight (56%) were confirmed by other methods (true positives) . Of the 44% Vitek-positive/confirmatory test-negative (false positives), eight were Escherichia coli which was 53% of all E . coli tested . The majority of other false-positive isolates were Klebsiella oxytoca (24% overall) which were all Vitek- and Etest-positive but negative by the combination disc test . CONCLUSIONS: All ESBL-positive strains by Vitek should be confirmed by the combination disc test using all three antibiotics . This will enable differentiation of 'true' ESBLs from false-positive organisms, including K1 hyperbetalactamase-producing Klebsiella oxytoca and AmpC-producing organisms . The cefpodoxime combination discs gave the best differentiation in this study with only one ESBL organism being missed.

Infect Immun, 2002 Sep, 70(9), 4772 - 6
Role of the htrA gene in Klebsiella pneumoniae virulence; Cortes G et al.; We recently described the use of mini-Tn5 to generate complement-sensitive mutants derived from a complement-resistant Klebsiella pneumoniae clinical isolate deficient in the lipopolysaccharide O side chain . One mutant with a reduced capacity to survive in nonimmune human sera carried the transposon inserted in the htrA gene . We cloned and sequenced the gene and predicted from the deduced amino acid sequence that the putative HtrA homolog contains structural features similar to those of previously described HtrA proteins . To investigate the biological functions and the role of the htrA gene in the virulence of K . pneumoniae, we constructed an isogenic mutant by insertion-duplication mutagenesis . Characterization of the mutant showed that it had greater sensitivity to temperature (50 degrees C) and oxidative stress (H(2)O(2)) than the parent strain . Furthermore, the htrA mutant produced less capsule, bound more molecules of complement component C3, and was more sensitive to complement and whole-blood killing than was the parent strain . Finally, disruption of the htrA gene in a virulent K . pneumoniae strain caused a reduction of its virulence in a mice model . Our results indicate that the htrA gene plays an important role in the virulence of K . pneumoniae.

J Allergy Clin Immunol, 2002 Aug, 110(2), 262 - 8
Effects of the hindlimb-unloading model of spaceflight conditions on resistance of mice to infection with Klebsiella pneumoniae; Belay T et al.; BACKGROUND: It has been well documented in several studies that many immunologic parameters are altered in experimental animals and human subjects who have flown in space . However, it is not fully known whether these immunologic changes could result in increased susceptibility to infection . Hindlimb (antiorthostatic) unloading of rodents has been used successfully to simulate some of the effects of spaceflight on physiologic systems . OBJECTIVE: The objective of this study was to determine the effect of hindlimb unloading on the outcome of Klebsiella pneumoniae infection in mice . METHODS: Hindlimb-unloaded, hindlimb-restrained, and control mice were intraperitoneally infected with one 50% lethal dose of K pneumoniae 2 days after suspension . Mortality and bacterial load in several organs were compared among the groups . RESULTS: Unloaded mice showed significantly increased mortality and reduced mean time to death compared with that seen in the control groups . Kinetics of bacterial growth with smaller infective doses revealed that control mice were able to clear bacteria from the organs after 30 hours . In contrast, unloaded mice had continued bacterial growth at the same time point . CONCLUSION: The results of this study suggest that hindlimb unloading might enhance the dissemination of K pneumoniae, leading to increased mortality . The complex physiologic changes observed during hindlimb unloading, including stress, have a key role in the pathophysiology of this infection.

Malays J Pathol, 2001 Dec, 23(2), 73 - 8
Comparison of three different methods for the presumptive detection of ESBL production in ceftazidime-resistant strains of Klebsiella pneumoniae; Palasubramaniam S et al.; Twenty-eight (28) strains of ceftazidime-resistant Klebsiella pneumoniae were isolated from blood cultures of in-patients from University Hospital, Kuala Lumpur between March 1995 and August 1996 . Three methods were used to detect the production of ESBL enzymes by these strains . These three methods include the double-disc synergy test (DDST), inhibitor-potentiated disc-diffusion test (IPDD) and the E-test ESBL method . All strains could be identified as ESBL producers using the DDST method by a minimum of two beta-lactams and these included either a combination of ceftazidime and ceftriaxone with clavulanate respectively or cefotaxime and aztreonam with clavulanate respectively . Similarly using a combination of either cefotaxime and ceftriaxone with clavulanate or ceftriaxone and aztreonam with clavulanate respectively would have detected all strains as ESBL producers . The IPDD method could also detect for ESBL activity based on combinations of beta-lactam antibiotics with clavulanate respectively . All combinations of beta-lactam antibiotics could detect for ESBL activity in all the strains except a combination of either ceftazidime and aztreonam or cefotaxime and ceftriaxone with clavulanate respectively . The E-Test method using ceftazidime alone and in combination with clavulanate was found to be the most effective method in the presumptive identification of ESBL activity in all the strains.

Mol Cell Biochem, 2002 May-Jun, 234-235(1-2), 39 - 48
The role of nitric oxide in lung innate immunity: modulation by surfactant protein-A; O'Reilly P et al.; Surfactant protein A (SP-A) and alveolar macrophages are essential components of lung innate immunity . Alveolar macrophages phagocytose and kill pathogens by the production of reactive oxygen and nitrogen species . In particular, peroxynitrite, the reaction product of superoxide and nitric oxide, appears to have potent antimicrobial effects . SP-A stimulates alveolar macrophages to phagocytose and kill pathogens and is important in host defense . However, SP-A has diverse effects on both innate and adaptive immunity, and may stimulate or inhibit immune function . SP-A appears to mediate toxic or protective effects depending on the immune status of the lung . In contrast to mouse or rat cells, it has been difficult to demonstrate nitric oxide production by human macrophages . We have recently demonstrated that human macrophages produce nitric oxide and use it to kill Klebsiella pneumoniae . SP-A either stimulates or inhibits this process, depending on the activation state of the macrophage . Given its diverse effects on immune function, SP-A may prove to be an effective therapy for both infectious and inflammatory diseases of the lung.

Biotechnol Prog, 2002 Jul-Aug, 18(4), 694 - 9
Improving 1,3-propanediol production from glycerol in a metabolically engineered Escherichia coli by reducing accumulation of sn-glycerol-3-phosphate; Zhu MM et al.; High levels of glycerol significantly inhibit cell growth and 1,3-propanediol (1,3-PD) production in anaerobic glycerol fermentation by genetically engineered Escherichia coli (E . coli) strains expressing genes from the Klebsiella pneumoniae dha (K.pneumoniae) regulon . We have previously demonstrated that 1,3-PD production by the engineered E . coli can be improved by reducing the accumulation of methylglyoxal . This study focuses on investigation of another lesser-known metabolite in the pathways related to 1,3-PD production-glycerol-3-phosphate (G3P) . When grown anaerobically on glycerol in the absence of an exogenous acceptor, the engineered E . coli strains have intracellular G3P levels that are significantly higher than those in K . pneumoniae, a natural 1,3-PD producer . Furthermore, in the engineered E . coli strains, the G3P levels increase with increasing glycerol concentrations, whereas, in K . pneumoniae, the concentrations of G3P remain relatively constant . Addition of fumarate, which can stimulate activity of anaerobic G3P dehydrogenase, into the fermentation medium led to a greater than 30-fold increase in the specific activity of anaerobic G3P dehydrogenase and a significant decrease in concentrations of intracellular G3P and resulted in better cell growth and an improved production of 1,3-PD . This indicates that the low activity of G3P dehydrogenase in the absence of an exogenous electron acceptor is one of the reasons for G3P accumulation . In addition, spent media from E.coli Lin61, a glycerol kinase (responsible for conversion of glycerol to G3P) mutant, contains greatly decreased concentrations of G3P and shows improved production of 1,3-PD (by 2.5-fold), when compared to media from its parent strain E . coli K10 . This further suggests that G3P accumulation is one of the reasons for the inhibition of 1,3-PD production during anaerobic fermentation.

J Biol Chem, 2002 Sep 20, 277(38), 35263 - 6 Epub 2002 Jul 19.
Crystallographic analysis of the MoFe protein of nitrogenase from a nifV mutant of Klebsiella pneumoniae identifies citrate as a ligand to the molybdenum of iron molybdenum cofactor (FeMoco); Mayer SM et al.; The x-ray crystal structure of NifV(-) Klebsiella pneumoniae nitrogenase MoFe protein (NifV(-) Kp1) has been determined and refined to a resolution of 1.9 A . This is the first structure for a nitrogenase MoFe protein with an altered cofactor . Moreover, it is the first direct evidence that the organic acid citrate is not just present, but replaces homocitrate as a ligand to the molybdenum atom of the iron molybdenum cofactor (FeMoco) . Subsequent refinement of the structure revealed that the citrate was present at reduced occupancy.

Microb Pathog, 2002 Jul, 33(1), 1 - 6
Cloning of a gene encoding a unique haemolysin from Klebsiella pneumoniae and its potential use as a species-specific gene probe; Yin-Ching C et al.; A gene, designated khe, that encodes a haemolysin of Klebsiella pneumoniae CMC-1 has been cloned and sequenced . When expressed in Escherichia coli, a unique peptide of approximately 20kDa was identified . Nucleotide sequence analysis predicted a single open reading frame (ORF) of 486bp encoding a 162 amino acid polypeptide with an estimated pI of 6.77 . No extensive sequence homology could be identified between khe and any reported sequence at either the nucleotide or amino acid level . Furthermore, DNA hybridizations under high stringency conditions failed to show any cross hybridizations to several bacteria including K . oxytoca, K . planticola, K . terrigena and K . ornithinolytica . These data indicate that we have cloned a unique gene, which is highly conserved among tested K . pneumoniae isolates.

Antimicrob Agents Chemother, 2002 Aug, 46(8), 2613 - 8
Three-dimensional quantitative structure-activity relationship and comparative molecular field analysis of dipeptide hydroxamic acid Helicobacter pylori urease inhibitors; Mishra H et al.; A homology model of Helicobacter pylori urease was developed by using the crystal structure of urease from Klebsiella aerogenes (EC 3.5.1.5) as a template . The acetohydroxamic acid moiety was docked into the active pocket of the enzyme model, followed by relaxation of the complex by use of molecular dynamics . The resulting conformation was used as a template to construct 24 potential dipeptide hydroxamic acid inhibitors with which comparative molecular field analysis (CoMFA) was performed . The resulting model provided a cross-validation correlation coefficient (q(2)(L00)) of 0.610, a conventional r(2) value of 0.988, and an F (Fisher indication of statistical significance) value of 294.88 . We were able to validate the CoMFA model by using the 50% inhibitory concentrations of six compounds that were not included in the construction of the model . A very good structural correlation was observed between the amino acids in the model urease's active pocket and the contour maps derived from the CoMFA model . This correlation, accompanied by the validation supplied by use of the CoMFA data, illustrates that the model can aid in the prediction and design of novel H . pylori urease inhibitors.

Infection, 2002 Jun, 30(3), 125 - 31
Klebsiella bacteremia in children in southern Israel (1988-1997); Krontal S et al.; BACKGROUND: Klebsiella spp . have emerged in recent years as a major cause of gram-negative bacteremia in infants and children . We therefore aimed to document the epidemiology, antibiotic susceptibility pattern and outcome of both community-acquired and nosocomial Klebsiella spp . bacteremias in children . PATIENTS AND METHODS: From 1988-1997, 177 episodes of Klebsiella bacteremia, representing 15% of all gram-negative bacteremias, occurred at the Soroka Medical Center in 166 children aged 0-14 years . RESULTS: The overall incidence of Klebsiella bacteremia in southern Israel during the study period was 0.13/1,000, with an increase from 0.1 to 0.2/1,000 children from 1988-1992 to 1993-1997 (p = 0.02) . 113 and 64 episodes were recorded in Bedouin Arabs and Jewish children, respectively . The incidence of Klebsiella bacteremia was significantly higher in Bedouins compared to Jewish children (p < 0.001) . The incidence of Klebsiella bacteremia increased significantly among Jewish children from 1993-1997 compared to 1988-1992 . The incidence of Klebsiella bacteremia was 2/1,000 admissions, with an increase from 1.8 to 2.2/1,000 from 1993-1997 compared to 1988-1992 . The incidence of Klebsiella bacteremia was significantly higher among hospitalized Bedouin children compared to Jewish children (3.1 vs 1.4/1,000 admissions, p < 0.001) . There were 48 (27%), 24 (14%) and 98 (55%) Klebsiella bacteremia episodes at the pediatric departments, pediatric intensive care unit (PICU) and neonatal intensive care unit (NICU), respectively . 76% of Klebsiella bacteremia episodes were nosocomial; 66% occurred at NICU . 71% and 90% of Klebsiella bacteremia episodes occurring at NICU and PICU, respectively, were nosocomial . The overall incidence of nosocomial infections was 1.5/1,000 admissions, with an increase from 1.2 to 1.8/1,000 from 1993-1997 compared to 1988-1992 (p = 0.03) . The resistance rates of Klebsiella spp . to piperacillin, ceftriaxone, ceftazidime and gentamicin were 34%, 17%, 17% and 14%, respectively . A significant increase in the resistance rates to ceftriaxone and ceftazidime was observed from 1993-1997 compared to 1988-92 (21.9% vs 7.8%, p = 0.05 and 21.9% vs 5%, p = 0.03) . A significant increase in resistance to ceftriaxone was recorded at PICU and NICU (from 12% and 0%, respectively, from 1988-1992, to 61% and 16%, respectively, from 1993-1997, p = 0.02) . Overall mortality rate of Klebsiella bacteremia was 13% (21/167 cases, 12 and eight at PICU and NICU, respectively) . CONCLUSION: An increase in Klebsiella bacteremia was recorded in southern Israel during the 10 years of the study . A marked increase in the rate of nosocomial Klebsiella bacteremia occurred at all departments . Resistance to third-generation cephalosporins emerged frequently at PICU and NICU during the last period of the survey.

Eur J Clin Microbiol Infect Dis, 2002 Jun, 21(6), 471 - 3 Epub 2002 Jun 12.
Clinical impact of nosocomial Klebsiella bacteremia in critically ill patients; Blot SI et al.; In order to determine the clinical impact of Klebsiella bacteremia on critically ill patients, a matched cohort study was conducted between January 1992 and December 2000 . During the study period, all intensive care unit (ICU) patients with nosocomial Klebsiella bacteremia were defined as cases (n=52), but two of these patients were excluded from the matched cohort due to incomplete medical records . The remaining 50 patients were matched at a ratio of 1:2 with control patients (n=100) on the basis of the APACHE II severity of disease classification system . Patients with Klebsiella bacteremia experienced acute renal failure and hemodynamic instability more often than controls . They also had a longer ICU stay and longer ventilator dependence . In-hospital mortality rates for cases and controls were nearly equal (36% vs . 37%, respectively; P=0.905) . In conclusion, after adjusting accurately for severity of underlying disease and acute illness, no difference in mortality was found between ICU patients with Klebsiella bacteremia and their matched control subjects.

Eur J Clin Microbiol Infect Dis, 2002 Jun, 21(6), 419 - 26 Epub 2002 Jun 12.
Retrospective analysis of clinical and microbiological aspects of Klebsiella oxytoca bacteremia over a 10-year period; Kim BN et al.; From 1991 to 2000, 125 sporadic cases of Klebsiella oxytoca bacteremia were analyzed retrospectively to review clinical features and to identify the risk factors associated with resistance to extended-spectrum cephalosporins and fatal outcome . Bacteremia was acquired nosocomially in 52% of the patients . Almost all patients (97%) had an underlying disease, with biliary and pancreatic disease occurring most frequently (55%) . The biliary tract was the most common site of infection (44%) . Resistance to extended-spectrum cephalosporins was identified in 22 of the 125 (18%) Klebsiella oxytoca blood isolates and resistance to ciprofloxacin in 9 (7%) . Only previous antibiotic therapy was strongly associated with resistance to extended-spectrum cephalosporins in patients with Klebsiella oxytoca bacteremia ( P=0.009) . The mortality rate was 24% and was higher in patients infected with isolates resistant to extended-spectrum cephalosporins (41% vs . 20%; P=0.04) . In multivariate analysis, fatal outcome was independently associated with septic shock, deteriorated mental status, polymicrobial bacteremia, and solid tumor . Surgical therapy had a protective effect (OR, 0.06; 95% CI, 0.005-0.7; P=0.03) . In conclusion, Klebsiella oxytoca bacteremia was most commonly associated with biliary tract infection . Previous antibiotic therapy was strongly associated with resistance to extended-spectrum cephalosporins in patients with Klebsiella oxytoca bacteremia.

J Microbiol Immunol Infect, 2002 Jun, 35(2), 85 - 8
Klebsiella pneumoniae liver abscess in Taiwan is not caused by a clonal spread strain; Cheng HP et al.; In Taiwan, the incidence of pyogenic liver abscess caused by Klebsiella pneumoniae has been increasing over the past 2 decades . Although most of the patients have no concurrent biliary tract disease, diabetes mellitus is thought to be an important risk factor for the disease . The incidence of metastatic infections in K . pneumoniae liver abscess, such as endogenous endophthalmitis and other extrahepatic infections, is also higher than that in liver abscess caused by other microbes . Furthermore, the incidence of metastatic infections in K . pneumoniae liver abscess in Taiwan is higher than Western countries . The reasons why K . pneumoniae liver abscess is so common in Taiwan and why diabetes mellitus is a risk factor for the disease are not clear . In this study, blood isolates from 40 patients with K . pneumoniae liver abscess treated at the Taipei Veterans General Hospital from 1995 through 2000 were randomly selected for study . Pulsed-field gel electrophoresis, ribotyping, and serotyping were used for cluster analysis . A total of 15 strains were of serotype K1 and 25 strains were of a serotype other than K1 . No major cluster or a closely related strain of K . pneumoniae was found . In conclusion, the results obtained from pulse-field gel electrophoresis and ribotyping of K . pneumoniae isolates do not suggest that liver abscess in Taiwan is primarily caused by a single genetically related strain.

Biosci Biotechnol Biochem, 2002 May, 66(5), 996 - 1001
Isolation and culture conditions of a Klebsiella pneumoniae strain that can utilize ammonium and nitrate ions simultaneously with controlled iron and molybdate ion concentrations; Kim YJ et al.; A bacterial strain F-5-2, isolated from soil and identified as Klebsiella pneumoniae, removed NH4+ completely in 24 h of aerobic cultivation in a medium containing 1 mg/ml of NH4NO3 . However, 70% of the NO3- originally provided remained . When 100 microM Fe2+ was added to the medium, both NH4+ and NO3- were removed simultaneously and completely from the culture within 6 h of incubation . In addition, the amount of MoO4- in the medium markedly affected the bacterial cell growth and utilization of NH4+ and NO3- . The bacterium could remove 4 mg/ml of NH4NO3 completely in 48 h of aerobic cultivation in a medium containing 100 microM Fe2+ and 0.8 pM MoO4(2-) . The total nitrogen in the culture containing its cells was decreased to 14% of that in the NH4NO3 originally provided . GC-MS analysis demonstrated that N2 was generated from the nitrogen atoms of both NH4+ and NO3-.

Appl Environ Microbiol, 2002 Jul, 68(7), 3462 - 6
Klebsiella pneumoniae produces no histamine: Raoultella planticola and Raoultella ornithinolytica strains are histamine producers; Kanki M et al.; Histamine fish poisoning is caused by histamine-producing bacteria (HPB) . Klebsiella pneumoniae and Klebsiella oxytoca are the best-known HPB in fish . However, 22 strains of HPB from fish first identified as K . pneumoniae or K . oxytoca by commercialized systems were later correctly identified as Raoultella planticola (formerly Klebsiella planticola) by additional tests . Similarly, five strains of Raoultella ornithinolytica (formerly Klebsiella ornithinolytica) were isolated from fish as new HPB . R . planticola and R . ornithinolytica strains were equal in their histamine-producing capabilities and were determined to possess the hdc genes, encoding histidine decarboxylase . On the other hand, a collection of 61 strains of K . pneumoniae and 18 strains of K . oxytoca produced no histamine.

Indian Pediatr, 2002 Jun, 39(6), 529 - 37
Respiratory distress in neonates with special reference to pneumonia; Mathur NB et al.; OBJECTIVE: (i) To find causes of respiratory distress in neonates brought to a referral neonatal unit with symptoms suggestive of respiratory disorder; (ii) to evaluate clinical signs for diagnosis of neonatal pneumonia; (iii) To determine bacterial etiology of neonatal pneumonia; and (iv) To determine indicators of fatality in neonatal pneumonia . DESIGN: Prospective descriptive . SETTING: Referral neonatal unit of a teaching hospital . SUBJECTS: 150 neonates admitted with respiratory symptoms consecutively . METHODS: All neonates presenting with respiratory symptoms were included in the study . The diagnosis of the cause of respiratory distress was based on guidelines recommended by the National Neonatology Forum . Clinical features, FiO2 requirement, sepsis screen, X-ray chest, blood culture and antibiotic sensitivity, arterial blood gases and other relevant investigations were documented in a structured proforma . The neonates were regularly followed up for outcome . Multivariate unweighted logistic regression was done to find out the indicators of fatality in neonatal pneumonia for those variables which were significantly associated with outcome on univariate analysis . RESULTS: Pneumonia was found to be the most common cause (68.6%) of respiratory distress in neonates . Other conditions included hydline membrane disease (HMD), transient tachypneia of new born (TTNB), birth asphyxia with hypoxic ischemic encephalopathy (HIE) and meconium aspiration syndrome (MAS) . Clinical signs and symptoms were non specific and did not differentiate between pneumonia and other causes of respiratory distress . Respiratory rate was less than 60 per minute in 11.6% neonates with pneumonia . The most common organism responsible for neonatal pneumonia was Klebsiella pneumoniae . Chest X-ray was clear in 15% of neonates with pneumonia . On univariate analysis weight < 2000 g, gestation age < 34 weeks, age at presentation < 72 hours, lethargy, absent neonatal reflexes, shock, positive C-reactive protein, positive ventilatory support, blood culture positivity, Silverman Score >3, FiO2 >40%, pH < 7.2, alveolar arterial gradient (AaDO2) > 250 mmHg and arterial alveolar tension ratio (a/A) of < 0.25 were significantly associated with mortality in neonates with pneumonia . However, on multivariate analysis, only AaDO2 of > 250 mmHg was found to be independent predictor of fatality in neonatal pneumonia . CONCLUSION: Pneumonia was the most common cause of respiratory distress in neonates . Clinical features and X-ray chest missed the diagnosis of pneumonia in 15 cases and had to be corroborated with sepsis screen and blood culture . AaDO2 > 250 mmHg was an independent predictor of fatality in neonatal pneumonia

Acta Crystallogr D Biol Crystallogr, 2002 Jul, 58(Pt 7), 1237 - 9 Epub 2002 Jun 20.
Crystallization of the FAD-independent acetolactate synthase of Klebsiella pneumoniae; Pang SS et al.; Leucine and valine are formed in a common pathway from pyruvate in which the first intermediate is 2-acetolactate . In some bacteria, this compound also has a catabolic fate as the starting point for the butanediol fermentation . The enzyme (EC 4.1.3.18) that forms 2-acetolactate is known as either acetohydroxyacid synthase (AHAS) or acetolactate synthase (ALS), with the latter name preferred for the catabolic enzyme . A significant difference between AHAS and ALS is that the former requires FAD for catalytic activity, although the reason for this requirement is not well understood . Both enzymes require the cofactor thiamine diphosphate . Here, the crystallization and preliminary X-ray diffraction analysis of the Klebsiella pneumoniae ALS is reported . Data to 2.6 A resolution have been collected at 100 K using a rotating-anode generator and an R-AXIS IV++ detector . Crystals have unit-cell parameters a = 137.4, b = 143.9, c = 134.4 A, alpha = 90, beta = 108.4, gamma = 90 degrees and belong to space group C2 . Preliminary analysis indicates that there are four monomers located in each asymmetric unit.

Eur J Biochem, 2002 Jun, 269(12), 2997 - 3004
Role of conserved residues within helices IV and VIII of the oxaloacetate decarboxylase beta subunit in the energy coupling mechanism of the Na+ pump; Schmid M et al.; The membrane-bound beta subunit of the oxaloacetate decarboxylase Na+ pump of Klebsiella pneumoniae catalyzes the decarboxylation of enzyme-bound biotin . This event is coupled to the transport of 2 Na+ ions into the periplasm and consumes a periplasmically derived proton . The connecting fragment IIIa and transmembrane helices IV and VIII of the beta subunit are highly conserved, harboring residues D203, Y229, N373, G377, S382, and R389 that play a profound role in catalysis . We report here detailed kinetic analyses of the wild-type enzyme and the beta subunit mutants N373D, N373L, S382A, S382D, S382T, R389A, and R389D . In these studies, pH profiles, Na+ binding affinities, Hill coefficients, Vmax values and inhibition by Na+ was determined . A prominent result is the complete lack of oxaloacetate decarboxylase activity of the S382A mutant at Na+ concentrations up to 20 mm and recovery of significant activities at elevated Na+ concentrations (KNa approximately 400 mm at pH 6.0), where the wild-type enzyme is almost completely inhibited . These results indicate impaired Na+ binding to the S382 including site in the S382A mutant . Oxaloacetate decarboxylation by the S382A mutant at high Na+ concentrations is uncoupled from the vectorial events of Na+ or H+ translocation across the membrane . Based on all data with the mutant enzymes we propose a coupling mechanism, which includes Na+ binding to center I contributed by D203 (region IIIa) and N373 (helix VIII) and center II contributed by Y229 (helix IV) and S382 (helix VIII) . These centers are exposed to the cytoplasmic surface in the carboxybiotin-bound state of the beta subunit and become exposed to the periplasmic surface after decarboxylation of this compound . During the countertransport of 2 Na+ and 1 H+ Y229 of center II switches between the protonated and deprotonated Na+-bound state.

Tunis Med, 2002 Jan, 80(1), 26 - 8
{Multidrug resistance in Klebsiella pneumoniae: multicenter study}; Boutiba-Ben Boubaker I et al.; The extensive use of broad spectrum antibiotics, especially the third generation cephalosporins (C3G), was followed by the emergence of newer plasmid mediated betalactamases called extended spectrum betalactamases (ESBLs) . To assess the impact of K . pneumoniae resistant to 3GC in Tunisia, this study was conducted in 3 teaching hospitals . A total of 1110 strains of K pneumoniae was collected . The antibiotics susceptibilities were tested by diffusion method using Mueller-Hinton agar . The quality control was regularly performed . I ESBLs producing solates were detected using the double-disc synergy test . Data analysis was done using the Whonet 4 software . 23.6% K . pneumoniae isolates showed phenotype pattern of ESBLs producers . The double-disc synergy test was positive in 75% of the cases . These isolates were recovered from hospitalized patients in different wards but mainly from pediatrics (23.6%), medicine (23.2%), surgery (22.9%), intensive care units (11%) and neonatology (11%) . 54% were isolated from urines, 22% from blood cultures . These isolates remained susceptible to imipenem (100%) and most of them to cefoxitin (96.4%) but all had associated resistance to aminoglycosides, quinolones and trimethoprim-sulfamethoxazole . The prevalence of multidrug resistant K . pneumoniae is high . This resistance can be minimized by the implementation of infection control measures including handwashing and isolation procedures.

Antimicrob Agents Chemother, 2002 Jul, 46(7), 2262 - 5
Novel plasmid-encoded class C beta-lactamase (MOX-2) in Klebsiella pneumoniae from Greece; Raskine L et al.; Klebsiella pneumoniae KOL, a clinical strain resistant to various beta-lactams, was isolated from the stools of a patient from Greece . This strain harbored a new pI 9.1 plasmid-mediated AmpC beta-lactamase with unusually high levels of hydrolytic activity for cefoxitin and cefotetan that we named MOX-2 . Sequencing of bla(MOX-2) revealed 93.2, 92.9, 92.7, and 73.1% identities with the deduced amino acid sequences of CMY-8, MOX-1, CMY-1, and the AmpC beta-lactamase of Aeromonas sobria, respectively.

Immunopharmacol Immunotoxicol, 2002 May, 24(2), 245 - 54
Suppression of IgE antibody response in mice by a polysaccharide, AZ9, produced by Klebsiella oxytoca strain TNM3; Sugihara R et al.; A soil bacterium, Klebsiella oxytoca TNM3 was found to produce a polysaccharide named AZ9 that shows suppressive effects on IgE antibody response in mice . When mice were administered with 50 to approximately 100 mg/kg AZ9 subcutaneousely for 4 consecutive days after immunization with trinitrophenyl (TNP)-keyhole limpet hemocyanin, anti-TNP IgE production was significantly suppressed, while the level of anti-TNP IgM was affected marginally . In AZ9-administered mice, IL-4 secretion from splenic cells was reduced to approximately 30% of the untreated control . Thus, AZ9 suppression of IgE production may be due to attenuating effects on the Th2-type response . Although oral administration of AZ9 alone had no effects on IgE production, ovalbumin (OVA)-induced oral tolerance of anti-TNP IgE response to TNP-OVA was markedly augmented when a suboptimal dose of OVA was administered orally in combination with AZ9 . Collectively, our data suggest that AZ9 has beneficial suppressive effects on IgE-dependent allergic responses.

Eur J Pharmacol, 2002 Jun 7, 445(1-2), 115 - 23
Effect of endotoxin on doxorubicin transport across blood-brain barrier and P-glycoprotein function in mice; Zhao YL et al.; The aim of this study was to investigate whether Klebsiella pneumoniae endotoxin modifies transport of doxorubicin, a P-glycoprotein substrate, across the blood-brain barrier and P-glycoprotein function in mice . Doxorubicin (30 mg/kg) was administered into the tail vein or fluorescein isothiocyanate-labeled dextran (FD-4) was infused (20 microg/min) into the right jugular vein of mice intravenously injected with endotoxin (10 mg/kg) 6 or 24 h earlier . Blood and brain samples were collected 4 h after injection of doxorubicin or 1 h after infusion of FD-4 . We examined using Western blotting the influence of endotoxin on the expression of P-glycoprotein in brains obtained 6, 12 and 24 h after injection . Endotoxin did not change the plasma and brain concentrations and brain-to-plasma concentration ratio (K(p) value) of FD-4 . No histopathological changes in brain capillaries were observed . These results suggest that endotoxin does not cause damage to brain capillaries . Plasma and brain concentrations of doxorubicin in mice treated 6 h earlier with endotoxin were significantly higher than those in control and mice treated 24 h earlier . However, endotoxin did not significantly change the K(p) value of doxorubicin . The protein level of P-glycoprotein was significantly, but slightly down-regulated 6 h after endotoxin treatment . However, the levels remained almost unchanged after 12 and 24 h . The present results suggest that Klebsiella pneumoniae endotoxin has no effect on the brain capillary integrity and doxorubicin transport across the blood-brain barrier in mice . It is likely that P-glycoprotein function might be sufficient to transport doxorubicin in spite of decreased levels of P-glycoprotein in the brain.

Arch Pediatr, 2002 May, 9(5), 463 - 8
{Genotypic exploration of a hospital neonatal outbreak due to Klebsiella pneumoniae producing extended-spectrum-betalactamase}; Boukadida J et al.; BACKGROUND: The aim of the study was to explore nosocomial neonatal outbreak of Klebsiella pneumoniae producing extended-spectrum-betalactamase by macrorestriction genotyping . PATIENTS AND METHODS: Over a 25 days period, a hospital neonatal outbreak due to Klebsiella pneumoniae producing extended-spectrum-betalactamase affected 14 newborn infants admitted to a university hospital in Sousse (Tunisia) . We collected 21 strains of Klebsiella pneumoniae producing extended-spectrum-betalactamase . Susceptibility testing to 17 antibiotics was determined . Macrorestriction genotyping of strains was determined by pulsed-field-electrophoresis . Neonatal intensive care unit survey was undertaken . RESULTS: A macrorestriction genotyping subdivided 21 strains into 3 clonally groups . Only cefoxitin, colistin, imipenem, amikacin and quinolons were active on the whole of strains . All infected babies died . The hygiene insufficiency and contamination of transfusion products at the time of their dividing in neonatal intensive care unit were incriminated . Handholding due to work overcharge was the main cause of bacterial diffusion . CONCLUSION: Multiclonal outbreak of Klebsiella pneumoniae producing extended-spectrum-betalactamase appeared following hygiene insufficiency related to work overcharge.

Appl Environ Microbiol, 2002 Jun, 68(6), 2676 - 82
Isomaltulose synthase from Klebsiella sp . strain LX3: gene cloning and characterization and engineering of thermostability; Zhang D et al.; The gene (palI) encoding isomaltulose synthase (PalI) from a soil bacterial isolate, Klebsiella sp . strain LX3, was cloned and characterized . PalI converts sucrose into isomaltulose, trehalulose, and trace amounts of glucose and fructose . Sequence domain analysis showed that PalI contains an alpha-amylase domain and (beta/alpha)(8)-barrel structures, suggesting that it belongs to the alpha-amylase family . Sequence alignment indicated that the five amino acid residues of catalytic importance in alpha-amylases and glucosyltransferases (Asp(241), Glu(295), Asp(369), His(145), and His(368)) are conserved in PalI . Purified recombinant PalI displayed high catalytic efficiency, with a Km of 54.6 +/- 1.7 mM for sucrose, and maximum activity (approximately 328.0 +/- 2.5 U/mg) at pH 6.0 and 35 degrees C . PalI activity was strongly inhibited by Fe3+ and Hg2+ and was enhanced by Mn2+ and Mg2+ . The half-life of PalI was 1.8 min at 50 degrees C . Replacement of selected amino acid residues by proline significantly increased the thermostability of PalI . Simultaneous replacement of Glu(498) and Arg(310) with proline resulted in an 11-fold increase in the half-life of PalI at 50 degrees C.

Arch Microbiol, 2002 Jun, 177(6), 500 - 6 Epub 2002 Apr 11.
Characterization of the citrate/acetate antiporter CitW of Klebsiella pneumoniae; Kastner CN et al.; The genome of Klebsiella pneumoniae contains at least three different genes encoding citrate transporters . Recently, a third and hitherto unknown gene encoding a citrate transport system ( citW) was identified . Escherichia coli transformed with a plasmid expressing citW was able to grow on citrate as sole carbon and energy source, identifying CitW as a citrate carrier . In this report, we provide evidence that further specifies CitW as a Na(+)-independent citrate/citrate and citrate/acetate exchanger . Kinetic analysis of citrate uptake at different pH values identified Hcitrate(2-) as the transported citrate species, with a K(m) of 25 microM . Since citW is expressed under anoxic conditions and acetate is the main end-product of citrate fermentation in K . pneumoniae, citrate/acetate exchange might be its in vivo function . Sequence similarity searches identified CitW (454 amino acids, 48.15 kDa) as a member of the 2-hydroxycarboxylate transporter family (TC 2.A.24) . The substrate specificity seems to partially contradict this phylogenetic classification, but appears logical with respect to the putative functional role of CitW in the citrate fermentation pathway of K . pneumoniae.

Biochemistry, 2002 May 28, 41(21), 6761 - 9
XAS investigation of the structure and function of Ni in acireductone dioxygenase; Al-Mjeni F et al.; Acireductone dioxygenases (ARDs) are enzymes involved in the methionine recycle pathway, which regulates aspects of the cell cycle . Klebsiella pneumoniae produces two enzymes that share a common polypeptide sequence and differ only in the metal ion present . Reaction of acireductone (1,2-dihydroxy-3-keto-5-methylthiopentene) with Fe-ARD and dioxygen produces formate and 2-keto-4-methylthiobutanoic acid, the alpha-ketoacid precursor of methionine . Ni-ARD reacts with acireductone and dioxygen to produce methylthiopropionate, CO, and formate and does not lie on the methionine recycle pathway . An X-ray absorption spectroscopy (XAS) study of the structure of the catalytic Ni center in resting Ni-ARD enzyme and the enzyme-substrate complex is reported . This study establishes the structure of the Ni site in resting Ni-ARD as containing a six coordinate Ni site composed of O/N-donor ligands including 3-4 histidine residues, demonstrates that the substrate binds to the Ni center in a bidentate fashion by displacing two ligands, at least one of which is a histidine ligand, and provides insight into the mechanism of catalysis employed by a Ni-containing dioxygenase . Efficiently relaxed and hyperfine-shifted resonances are observed in the (1)H nuclear magnetic resonance spectrum of Ni-ARD that can be attributed to the His imidazoles ligating the paramagnetic Ni ion and are consistent with the XAS results regarding His ligation . These resonances show significant perturbation in the presence of substrate, confirming that the metal ion interacts directly with the substrate.

Protist, 2002 Mar, 153(1), 39 - 45
Purine composition of the crystalline cytoplasmic inclusions of Paramecium tetraurelia; Creutz CE et al.; Crystalline cytoplasmic inclusions were isolated by differential centrifugation from mass cultures of Paramecium tetraurelia feeding on Klebsiella pneumonia . Physical and chemical measurements of intact and solubilized crystals determined that they consist primarily of guanine and hypoxanthine with traces of xanthine . Crystals from the mutant sombre consist primarily of xanthine, suggesting there is a disorder of purine metabolism in this mutant.

Antimicrob Agents Chemother, 2002 Jun, 46(6), 2024 - 8
Cloning of a cation efflux pump gene associated with chlorhexidine resistance in Klebsiella pneumoniae; Fang CT et al.; Expression libraries of a chlorhexidine-resistant Klebsiella pneumoniae strain were constructed and transformed into Escherichia coli XLOLR . Twenty chlorhexidine-resistant transformants were obtained after selection . All clones contained a novel 903-nucleotide locus . Its sequences were compatible with a cation efflux pump, and the locus was thus designated as cepA . Retransformation using cepA-containing plasmids conferred chlorhexidine resistance to both XLOLR and a chlorhexidine-sensitive K . pneumoniae strain . Therefore, CepA is associated with chlorhexidine resistance and may act as a cation efflux pump.

Antimicrob Agents Chemother, 2002 Jun, 46(6), 2000 - 3
TEM-71, a novel plasmid-encoded, extended-spectrum beta-lactamase produced by a clinical isolate of Klebsiella pneumoniae; Rasheed JK et al.; TEM-71, a novel extended-spectrum beta-lactamase from a Klebsiella pneumoniae clinical isolate, had an isoelectric point of 6.0 and a substrate profile showing preferential hydrolysis of cefotaxime over ceftazidime . It differed from TEM-1 by two substitutions, Gly238Ser and Glu240Lys, and was under the control of the strong P4 promoter.

Appl Biochem Biotechnol, 2002 Spring, 98-100, 687 - 98
Production of 1,3-propanediol by Klebsiella pneumoniae; Huang H et al.; 1,3-Propanediol (1,3-PD) has numerous applications from polymers to cosmetics, foods, lubricants, and medicines . Recently, there are strong industrial interests in a new kind of polyester, polytrimethylene terephthalate, with 1,3-PD as a monomer . This new polyester shows significant promise for use in carpeting and textiles . In this article we introduce a mild aerobic fermentation process using a strain screened from Klebsiella pneumoniae ATCC 25955, which is insensitive to oxygen, to produce 1,3-PD . We also describe a two-step fermentation process starting with glucose that was converted into glycerol with a glycerol-producing yeast, followed by K . pneumoniae that converts glycerol into 1,3-PD without intermediate isolation and purification of glycerol.

Appl Biochem Biotechnol, 2002 Spring, 98-100, 327 - 40
Reduction of furfural to furfuryl alcohol by ethanologenic strains of bacteria and its effect on ethanol production from xylose; Gutierrez T et al.; The ethanologenic bacteria Escherichia coli strains KO11 and LYO1, and Klebsiella oxytoca strain P2, were investigated for their ability to metabolize furfural . Using high performance liquid chromatography and 13C-nuclear magnetic resonance spectroscopy, furfural was found to be completely biotransformed into furfuryl alcohol by each of the three strains with tryptone and yeast extract as sole carbon sources . This reduction appears to be constitutive with NAD(P)H acting as electron donor . Glucose was shown to be an effective source of reducing power . Succinate inhibited furfural reduction, indicating that flavins are unlikely participants in this process . Furfural at concentrations >10 mM decreased the rate of ethanol formation but did not affect the final yield . Insight into the biochemical nature of this furfural reduction process may help efforts to mitigate furfural toxicity during ethanol production by ethanologenic bacteria.

J Chemother, 2002 Apr, 14(2), 140 - 6
Surveillance of antimicrobial resistance in gram-negative isolates from intensive care units in Turkey: analysis of data from the last 5 years; Leblebicioglu H et al.; A multicenter antimicrobial surveillance program was established in Turkey in 1995 to monitor the predominant Gram-negative pathogens from intensive care units (ICUs) and antimicrobial resistance patterns of these isolates . Sixteen hospitals participated in the study and a total of 1479 isolates from 1,100 patients were collected . The isolates were tested for their susceptibility against 13 antibiotics by E-test method . Minimum inhibitory concentrations (MICs) for each isolate were determined for imipenem, ceftazidime, ceftazidime-clavulanate, cefoperazone-sulbactam, ceftriaxone, cefepime, cefuroxime, piperacillin-tazobactam, ticarcillin-clavulanate, gentamicin, amikacin and ciprofloxacin . The most common isolates were Pseudomonas spp . (28.2%), Escherichia coli (19.2%) and Klebsiella spp . (19.1%) . We found very high resistance rates to all major antibiotics that are used to treat serious infections . Although imipenem is the most active agent, it had an overall susceptibility rate of 68% . Half of the tested Klebsiella spp . strains were found to produce ESBL . This is a very high rate when compared with the literature . Cross-resistance among species was also investigated . 52% of ciprofloxacin-resistant strains were also resistant to imipenem, 80% to ceftazidime, 97% to ceftriaxone, 86% to amikacin and 19% of imipenem-resistant strains were susceptible to ceftazidime and 18% to amikacin . When susceptibilities of the years 1995 and 1999 were compared, the most interesting finding was the decrease in resistance to 3rd generation cephalosporins . In conclusion, this national clinical isolate database shows that resistance rates are high, the change over years is not predictable and continuous surveillance is necessary to monitor antimicrobial resistance and to guide antibacterial therapy.

J Clin Invest, 2002 May, 109(10), 1311 - 9
Myeloperoxidase produces nitrating oxidants in vivo; Gaut JP et al.; Despite intense interest in pathways that generate reactive nitrogen species, the physiologically relevant mechanisms for inflammatory tissue injury remain poorly understood . One possible mediator is myeloperoxidase, a major constituent of neutrophils, monocytes, and some populations of macrophages . The enzyme uses hydrogen peroxide and nitrite to generate 3-nitrotyrosine in vitro . To determine whether myeloperoxidase produces nitrating intermediates in vivo, we used isotope dilution gas chromatography/mass spectrometry to quantify 3-nitrotyrosine in two models of peritoneal inflammation: mice infected with Klebsiella pneumoniae and mice subjected to cecal ligation and puncture . Both models developed an intense neutrophil inflammatory response, and the inflammatory fluid contained markedly elevated levels of 3-chlorotyrosine, a marker of myeloperoxidase action . In striking contrast, 3-nitrotyrosine levels rose only in the mice infected with K . pneumoniae . Levels of total nitrite and nitrate were 20-fold higher in mice injected with K . pneumoniae than in mice subjected to cecal ligation and puncture . Levels of 3-nitrotyrosine failed to increase in mice infected with K . pneumoniae that lacked functional myeloperoxidase . Our observations provide strong evidence that myeloperoxidase generates reactive nitrogen species in vivo and that it operates in this fashion only when nitrite and nitrate become available . This article was published online in advance of the print edition . The date of publication is available from the JCI website, http://www.jci.org.

J Hosp Infect, 2002 Apr, 50(4), 309 - 11
The higher disinfectant resistance of nosocomial isolates of Klebsiella oxytoca: how reliable are indicator organisms in disinfectant testing?
Gebel J, Sonntag HG, Werner HP, Vacata V, Exner M, Kistemann T.
The Children's Clinic in Giessen, Germany recently reported several severe infections with Klebsiella oxytoca resulting in deaths of two neonates . The putative source of the infections was a contaminated infusion solution . The resistance to disinfectant of the K . oxytoca isolates was investigated in three independent laboratories and was indeed found to be significantly increased . Comparative tests with standard strains of K . oxytoca and other recommended bacterial surrogates showed the disinfection procedures used were fully effective . The higher resistance of the nosocomial isolates may have developed due to improper handling and storage of the cleaning utensils . This report describes the events and draws conclusions concerning the use of disinfectants, the treatment of cleaning utensils, the reliability of procedures for testing disinfectants, and suggests additional measures .

Pharmacotherapy, 2002 May, 22(5), 578 - 85
Effect of antibiotics on polymorphonuclear neutrophil apoptosis; Healy DP et al.; STUDY OBJECTIVE: To evaluate the effects of various antibiotics-direct and indirect as a result of bacterial killing-on polymorphonuclear neutrophil (PMN) apoptosis . DESIGN: In vitro analysis . SETTING: Research laboratory . INTERVENTION: Whole blood collected from healthy human subjects was incubated with and without Klebsiella pneumoniae (1.0 x 10(5) colony-forming units {cfu}/ml) plus ceftazidime 50 microg/ml, gentamicin, ciprofloxacin, trovafloxacin, tetracycline, doxycycline, erythromycin, azithromycin (each 5 microg/ml), or lipopolysaccharide 10 microg/ml . After staining with fluorescein-conjugated annexin V, red blood cells were lysed, and the remaining white blood cells were assessed by flow cytometry with gating on PMNs . MEASUREMENTS AND MAIN RESULTS: In the absence of K . pneumoniae infection, antibiotic exposure directly decreased PMN apoptosis by 17.8% (range -25.0% to -13.9%, p=0.008) compared with untreated cells . In the presence of K . pneumoniae, all antibiotic treatments, even those with poor in vitro activity for the bacterial isolate, decreased PMN apoptosis by 26.2% (range -38.0% to -17.8%, p<0.001) compared with untreated control cells and by 36.6% compared with untreated (no antibiotic) K . pneumoniae-stimulated cells (range -46.2% to -28.0%, p<0.001) . CONCLUSIONS: All tested antibiotics in clinically relevant concentrations resulted in modest yet consistent decreases in PMN apoptosis . The magnitude of this change increased slightly in the presence of K . pneumoniae infection . In vivo studies are needed to determine whether antibiotic-associated prolongation of PMN survival improves host response to infection.

Pediatr Infect Dis J, 2002 Mar, 21(3), 260 - 2
Characterization of Klebsiella pneumoniae and Escherichia coli strains that produce CTX-M-2-type broad spectrum beta-lactamase isolated from a child with leukemia; Ohkawa T et al.; An 8-year-old girl with acute leukemia had bacteremia caused by Klebsiella pneumoniae producing CTX-M-2-type broad spectrum beta-lactamase . K . pneumoniae and Escherichia coli strains producing the same enzyme and harboring identical conjugative plasmids were recovered from stoor culture . Patients with frequent episodes of neutropenia and prophylactic administration of beta-lactams are at risk of harboring colonizing strains that produce broad spectrum beta-lactamases.

Arch Pediatr, 2002 Apr, 9(4), 385 - 7
{Acute non-traumatic mediastinitis}; Chat L et al.; Acute mediastinitis is uncommon . When it occurs, it usually follows an esophageal perforation or thoracic surgery . CASE REPORT: We report on a case of a ten-year-old girl with non traumatic mediastinitis secondary to a pleuro-pulmonary infection due to Klebsiella pneumoniae.

Emerg Infect Dis, 2002 May, 8(5), 522 - 4
Cefepime MIC as a predictor of the extended-spectrum beta-lactamase type in Klebsiella pneumoniae, Taiwan; Yu WL et al.; To guide selection of carbapenems or fourth-generation cephalosporins as therapy, 110 Klebsiella pneumoniae isolates with extended-spectrum beta-lactamases from Taiwan were characterized by phenotypic (MICs), molecular, and chemical methods . MIC patterns of ceftazidime and cefepime clearly differentiate strains treatable by cefepime and those capable of efficiently hydrolyzing available cephalosporins (CTX-M series and SHV-types) . Continued use of cefepime appears to be a treatment option in cases for which MIC results are available and interpreted by the criteria presented.

Arch Intern Med, 2002 May 13, 162(9), 1021 - 7
Characteristics of bacteremia between community-acquired and nosocomial Klebsiella pneumoniae infection: risk factor for mortality and the impact of capsular serotypes as a herald for community-acquired infection; Tsay RW et al.; BACKGROUND: Although several epidemiological surveys of Klebsiella clinical isolates have been performed, few studies have correlated the clinical isolate with disease . OBJECTIVE: To compare the clinical and bacteriological characteristics of Klebsiella pneumoniae bacteremia acquired as community or nosocomial infections . METHODS: We prospectively enrolled 158 consecutively hospitalized patients with K pneumoniae bacteremia . Clinical data were reviewed . Antimicrobial susceptibility testing and capsular serotyping were performed . We used the chi(2) test, the Fisher exact test, or the t test for statistic analysis . RESULTS: Underlying diabetes mellitus was more common in community-acquired than in nosocomial infection (46/94 {49%} vs . 8/64 {12%}; P<.001) . On the other hand, neoplastic disease (34/64 {53%} vs . 13/94 {14%}; P<.001) and antibiotic resistance (P<.01) were more frequent in patients with nosocomial compared with community-acquired infections . Klebsiella pneumoniae liver abscesses, which were all community acquired, accounted for the source of 22 (23%) of 94 community-acquired K pneumoniae infections . No attributable source of infection was found for 37 (58%) of the 64 nosocomial infections vs . 15 (16%) of the 94 community-acquired infections . Only 58 isolates (36.7%) could be serotyped; of these, capsular serotypes K1, K2, and K28 accounted for 37 (23.4%), 8 (5.1%), and 6 (3.8%), respectively, of all strains . However, typeable isolates were significantly more common among community-acquired than nosocomial isolates (42/94 {45%} vs . 16/64 {25%}; P =.01), especially for serotype K1 (28/94 {30%} vs . 9/64 {14%}; P =.02) . Significant risk factors for mortality included nosocomial infection, lung infection, thrombocytopenia, leukopenia, ceftazidime resistance, inappropriate antimicrobial therapy, and septic shock . CONCLUSIONS: Significant differences were identified between community-acquired and nosocomial K pneumoniae bacteremia . Ceftazidime resistance in nosocomial K pneumoniae bacteremia carried a high risk for mortality, and serotype K1 in K pneumoniae was more prevalent in community-acquired infection, suggesting more virulence.

J Biol Chem, 2002 Jul 12, 277(28), 25070 - 81 Epub 2002 May 01.
Structures of lipopolysaccharides from Klebsiella pneumoniae . Eluicidation of the structure of the linkage region between core and polysaccharide O chain and identification of the residues at the non-reducing termini of the O chains; Vinogradov E et al.; Deamination of LPSs from Klebsiella pneumoniae released O-chain polysaccharides together with a fragment of the core oligosaccharide . The structures of the products from serotypes O1, O2a, O2a,c, O3, O4, O5, and O12 were determined by NMR spectroscopy and chemical methods, identifying the linkage region between the O antigens and the core as well as novel residues at the non-reducing ends of the polysaccharides . All serotypes had an identical linkage between the O chain and core.

Pneumologia, 2001 Jul-Sep, 50(3), 154 - 8
{The study of the frequency and features of bacteria isolated in patients with non-tuberculosis respiratory disorders admitted in "M . Nasta" Institute in the first trimester 2001}; Diaconescu C et al.; The aim of this study was to determine the spectrum of bacteria isolated in patients with non-TB respiratory disorders, in order to find the frequency of germs isolated globally and in each of the 6 clinical syndromes: suppurations, chronic obstructive diseases exacerbations (COPD, bronchial asthma), pneumopathies, post-TB syndromes, tumors and interstitial lung diseases . We found the greatest global frequency in anaerobes (35%) . This is in concordance with the high frequency of broncho-pulmonary suppurations (51% of the cases studied) . In second place comes Ps . aeruginosa (18%), followed by Str . Pneumoniae (16%), H . influenzae (11%) and Klebsiella pn . (10%) . Considering the germs identified in the various clinical syndromes, we found a high frequency of anaerobes associated to suppurations (51%), interstitial lung diseases (43%) and tumors (37%), while Ps . aeruginosa is first in post-TB syndromes (50%) and COPD exacerbations (21%), equal to H . influenzae . In pneumonias, Str . Pneumoniae was most frequently isolated (38%) followed by H . influenzae (25%) . The susceptibility testing of strains of Klebsiella and Ps . aeruginosa revealed the increasing tendency to resistance to broad spectrum antibiotics, especially for Ps . aeruginosa, with consecutive difficulties in finding the appropriate treatment.

Intensive Care Med, 2002 Apr, 28(4), 438 - 42 Epub 2002 Mar 15.
Activity of antibacterial impregnated central venous catheters against Klebsiella pneumoniae; Yorganci K et al.; OBJECTIVE: Antibiotically coated or impregnated catheters are effective in eliminating gram-positive bacteria from their surfaces . However, their activity against gram-negative bacteria is not well known . The aim of this study was to evaluate and compare the adherence, persistence and colonization of Klebsiella pneumoniae on catheter surfaces and also to assess bacteriostatic and bactericidal levels . DESIGN: Randomized, controlled, laboratory study . SETTING: University surgical microbiology laboratory . SUBJECTIVE: Silver sulfadiazine-chlorhexidine impregnated (SSC), minocycline and rifampin bonded (M+R), silver, platinum and carbon incorporated (SP+C) and non-antiseptic central venous catheter segments . INTERVENTIONS: Catheter segments were immersed in 1 ml of phosphate buffered saline (0.01 mol/l) with 0.25% dextrose (PBSD) and incubated at 37 degrees C . The PBSD was replaced daily . Effluents were frozen at -70 degrees C for subsequent determination of bacteriostatic and bactericidal activity . On days 1,3,7,14 and 21 after initial immersion, 1 ml standardized inoculum of Klebsiella pneumoniae was added to 90 tubes for a period of 30 min . The inoculum was then replaced with PBSD . One third of the samples were immediately sonicated and plated for the determination of bacterial adherence . The remaining segments were incubated for 4 and 24 h, followed by the same procedure to determine bacterial persistence and colonization with time . All plates were read after 24 h of incubation . MEASUREMENTS AND RESULTS: There was a significant reduction in initial bacterial adherence for SP+C catheters on all days ( p<0.05) . SSC catheters prevented initial bacterial adherence for the first 7 days only ( p<0.05) . SSC and SP+C catheters prevented bacterial persistence and further colonization on all days . However M+R catheters prevented bacterial colonization for 3 days only . Effluent studies indicated that the impregnated agents in catheter SSC were bactericidal compared to catheter M+R, which were bacteriostatic to K . pneumoniae . No antibacterial activity was detected in the effluents from catheter SP+C . CONCLUSIONS: SSC and SP+C catheters are effective in eliminating K . pneumoniae from their surfaces for at least 21 days . M+R catheters are less effective in eliminating bacterial adherence and colonization may be due to their bacteriostatic property.

Antimicrob Agents Chemother, 2002 May, 46(5), 1481 - 91
Bloodstream infections by extended-spectrum beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae in children: epidemiology and clinical outcome; Kim YK et al.; To determine the epidemiologic features and clinical outcomes of bloodstream infections caused by extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli and Klebsiella pneumoniae isolates, cases of bacteremia caused by these organisms in children were analyzed retrospectively . Among the 157 blood isolates recovered from 1993 to 1998 at the Seoul National University Children's Hospital, the prevalence of ESBL production was 17.9% among the E . coli isolates and 52.9% among the K . pneumoniae isolates . The commonest ESBLs were SHV-2a and TEM-52 . A novel ESBL, TEM-88, was identified . Pulsed-field gel electrophoresis analysis of the ESBL-producing organisms showed extensive diversity in clonality . The medical records of 142 episodes were reviewed . The risk factors for bloodstream infection with ESBL-producing organisms were prior hospitalization, prior use of oxyimino-cephalosporins, and admission to an intensive care unit within the previous month . There was no difference in clinical severity between patients infected with ESBL-producing strains (the ESBL group) and those infected with ESBL-nonproducing strains (the non-ESBL group) at the time of presentation . However, the overall fatality rate for the ESBL group was significantly higher than that for the non-ESBL group: 12 of 45 (26.7%) versus 5 of 87 (5.7%) (P = 0.001) . In a subset analysis of patients treated with extended-spectrum cephalosporins with or without an aminoglycoside, favorable response rates were significantly higher in the non-ESBL group at the 3rd day (6 of 17 versus 33 of 51; P = 0.035), the 5th day (6 of 17 versus 36 of 50; P < 0.05), and the end of therapy (9 of 17 versus 47 of 50; P < 0.001) . In conclusion, the ESBL production of the infecting organisms has a significant impact on the clinical course and survival of pediatric patients with bacteremia caused by E . coli and K . pneumoniae.

Antimicrob Agents Chemother, 2002 May, 46(5), 1212 - 7
ColE1-like plasmid pIP843 of Klebsiella pneumoniae encoding extended-spectrum beta-lactamase CTX-M-17; Cao V et al.; The resistance of Klebsiella pneumoniae BM4493, isolated in Ho Chi Minh City, Vietnam, to cefotaxime and aztreonam was due to production of a novel beta-lactamase, CTX-M-17 . The bla(CTX-M-17) gene was borne by 7,086-bp plasmid pIP843, which was entirely sequenced and which was found to belong to the ColE1 family . The 876-bp bla(CTX-M-17) gene differed from bla(CTX-M-14) by 2 nucleotides, which led to the single amino acid substitution Glu289-->Lys . bla(CTX-M-17) was flanked upstream by an ISEcp1-like element and downstream by an insertion sequence (IS) IS903 variant designated IS903-C . The transcriptional start site of bla(CTX-M-17) was located 109 nucleotides upstream from the initiation codon in the ISEcp1-like element, which also provided the promoter sequences . Plasmid pIP843, which was non-self-transferable and nonmobilizable, contained five open reading frames transcribed in the same orientation . Regions homologous to sequences coding for putative RNA II and RNA I transcripts, a rom gene, which is involved in initiation of replication, and a cer-like gene, which is responsible for the stability of ColE1-like plasmids, were identified . Consensus sequences for putative replication (oriV) and transfer (oriT) origins were present . Results of primer extension experiments indicated that ISEcp1 provides the promoter for expression of bla(CTX-M-17) and may contribute to dissemination of this gene.

FEMS Microbiol Lett, 2002 Mar 5, 208(2), 179 - 85
Isolation and characterization of dibenzofuran-degrading bacteria; Fukuda K et al.; Two bacterial strains capable of utilizing dibenzofuran (DF) as a sole carbon source were isolated from soil samples of reclaimed land . The strains designated HL1 and HL7 were identified as Klebsiella sp . and Sphingomonas sp., respectively, on the basis of biochemical characteristics and the sequences of the 16S ribosomal DNA . Sphingomonas sp . strain HL7 degraded non-, mono- and also dichlorinated DF and dibenzo-p-dioxin (DD) . Klebsiella sp . strain HL1 was able to degrade non- and monochlorinated DFs and DDs, but not dichlorinated ones . The metabolites formed from DF by strains HL1 and HL7 were similar to those by dioxin-degrading bacteria Sphingomonas sp . strain RW1 except for salicylic acid and catechol . Strain HL7 had a gene homologous to that encoding the dioxin dioxygenase alpha-subunit (dxnA1) gene of Sphingomonas sp . strain RW1 . However, Southern hybridization analysis showed that the size of an EcoRV-digested genomic fragment involving the dioxin dioxygenase gene of strain HL7 was smaller than that of strain RW1, and that strain HL1 did not have the homologous gene . Strains HL1 and HL7 provided useful information regarding the dioxygenase genes.

FEMS Microbiol Lett, 2002 Feb 5, 207(2), 159 - 64
Polyphosphate produced in recombinant Escherichia coli confers mercury resistance; Pan-Hou H et al.; An Escherichia coli strain was generated by fusion of a merA-deleted broad-spectrum mer operon from Pseudomonas K-62 with a bacterial polyphosphate kinase gene (ppk) from Klebsiella aerogenes in vector pUC119 . A large amount of the ppk-specified polyphosphate was identified in the mercury-induced bacterium with the fusion plasmid designated pMKB18 but not in the cells without mercury induction . These results suggest that the synthesis of polyphosphate as well as the expression of the mer genes is mercury-inducible and regulated by merR . The E . coli strain with pMKB18 was more resistant to both Hg2+ and C6H5Hg+ than its isogenic strain with cloning vector pUC119 . The recombinant strain accumulated more mercury from Hg2+- and C6H5Hg+-contaminated medium . Hg2+ transported into the cytoplasm appeared to be bound by chelation with the polyphosphate produced by the recombinant cells . The transported phenylmercury was degraded to Hg2+ before the chelation since polyphosphate did not directly chelate with C6H5Hg+ . These results indicate that polyphosphate is capable of reducing the cytotoxicity of the transported Hg2+ probably via chelation between polyphosphate and Hg2+.

Infect Immun, 2002 May, 70(5), 2583 - 90
Molecular analysis of the contribution of the capsular polysaccharide and the lipopolysaccharide O side chain to the virulence of Klebsiella pneumoniae in a murine model of pneumonia; Cortes G et al.; Klebsiella pneumoniae is a common cause of gram-negative bacterial nosocomial pneumonia . Two surface polysaccharides, lipopolysaccharide (LPS) O side chain and capsular polysaccharide (CPS), are critical for the microorganism in causing sepsis, but little is known about their role in pneumonia . To investigate their contribution in the pathogenesis of K . pneumoniae pneumonia, we characterized the host response to bacterial challenge with a highly virulent clinical isolate or with isogenic insertion-duplication mutants deficient in CPS or LPS O side chain in a murine model of pneumonia . Animals challenged intratracheally with the wild-type or LPS O side chain-deficient strain developed pneumonia and became bacteremic before death . Extensive lung lesions as well as pleuritis, vasculitis, and edema were observed by histopathological examination, and polymorphonuclear infiltration was also demonstrated . In contrast, none of the animals challenged with the unencapsulated strain developed pneumonia or bacteremia . Examination of tissue from this group did not identify lung lesions, and none of the infected animals died . Analysis of the early host defense mechanisms that contributed to the clearance of the unencapsulated mutant showed that the levels of C3 deposited on the unencapsulated mutant surface were threefold higher than those for the wild-type and LPS O side chain-deficient strains . Furthermore, phagocytosis of the unencapsulated mutant by human alveolar macrophages (AM) was more efficient than that of the wild-type and LPS O side chain-deficient strains . We conclude that CPS, but not LPS O side chain, is essential for Klebsiella pneumonia because it modulates the deposition of C3 and protects the microorganisms against human AM phagocytosis.

Zhonghua Yi Xue Za Zhi, 2002 Feb 25, 82(4), 279 - 83
{Molecular characteristics of beta-lactamase from Klebsiella pneumoniae}; Zhang Y et al.; OBJECTIVE: To study the molecular characteristics of beta-lactamase from Klebsiella pneumoniae . METHODS: Beta-lactamase was prepared from strains of Klebsiella pneumoniae isolated clinically for which the minimal inhibitory concentration (MIC) of cefoperazone had been >or= 8 mg/L and then was reduced by 50% by the addition of sulbatam . The isoelectric points of different beta -lactamases were examined by LKB2117 Multiphor II Electrophoresis System . The molecular weights were examined by polyacrylamide gel electrophoresis . PCR was used to detect the TEM-type genes . The PCR products were sequenced by chemiluminescence . RESULTS: All four strains of Klebsiella pneumoniae produced more than two kinds of beta-lactamase . TEM-type genes were amplified from all strains . 97% - 98% of the DNA sequence and amino acid sequence of the TEM-type enzymes from 2 strains of Klebsiella pneumoniae were identical with those of TEM-1 enzyme . The isoelectric points of the beta-lactamases ranged from 5.4 to 9.30 and the molecular weights were between 23.0 and 43.0 KD . The beta-lactamases from Klebsiella pneumoniae 99592 and K . pneumoniae 99607 hydrolysed ceftazidine, cefotaxime, and ceftriaxone . The beta-lactamase from K . pneumoniae 99595 had stronger activity against ceftazidime, ceftaxime, and ceftriaxone than that from K . pneumoniae 99607 . CONCLUSION: Klebsiella pneumoniae produces TEM-type extended spectrum beta -lactamase against ceftazidine, ceftaxime, and ceftriaxone.

J Bacteriol, 2002 May, 184(9), 2439 - 46
Identification of a gene cluster in Klebsiella pneumoniae which includes citX, a gene required for biosynthesis of the citrate lyase prosthetic group; Schneider K et al.; The biosynthesis of the 2'-(5"-phosphoribosyl)-3'-dephospho-coenzyme A (CoA) prosthetic group of citrate lyase (EC 4.1.3.6), a key enzyme of citrate fermentation, proceeds via the initial formation of the precursor 2'-(5"-triphosphoribosyl)-3'-dephospho-CoA and subsequent transfer to apo-citrate lyase with removal of pyrophosphate . In Escherichia coli, the two steps are catalyzed by CitG and CitX, respectively, and the corresponding genes are part of the citrate lyase gene cluster, citCDEFXG . In the homologous citCDEFG operon of Klebsiella pneumoniae, citX is missing . A search for K . pneumoniae citX led to the identification of a second genome region involved in citrate fermentation which comprised the citWX genes and the divergent citYZ genes . The citX gene was confirmed to encode holo-citrate lyase synthase, whereas citW was shown to encode a citrate carrier, the third one identified in this species . The citYZ genes were found to encode a two-component system consisting of the sensor kinase CitY and the response regulator CitZ . Remarkably, both proteins showed >or=40% sequence identity to the citrate-sensing CitA-CitB two-component system, which is essential for the induction of the citrate fermentation genes in K . pneumoniae . A citZ insertion mutant was able to grow anaerobically with citrate, indicating that CitZ is not essential for expression of citrate fermentation genes . CitX synthesis was induced to a basal level under anaerobic conditions, independent of citrate, CitB, and CitZ, and to maximal levels during anaerobic growth with citrate as the sole carbon source . Similar to the other citrate fermentation enzymes, CitX synthesis was apparently subject to catabolite repression.

Curr Med Chem, 2002 Feb, 9(4), 437 - 42
In vitro and in vivo activities of ciprofloxacin and levofloxacin against an SHV-5 extended-spectrum beta-lactamase-producing Klebsiella pneumoniae strain; Szabo D et al.; Activities of ciprofloxacin and levofloxacin against an SHV-5 extended-spectrum beta-lactamase (ESBL)-producing Klebsiella pneumoniae strain were studied in vitro and in vivo in septic mice using a high inoculum . Susceptibility to ciprofloxacin and levofloxacin was independent of the inoculum size . In killing curve studies, after 24 hours the initial 7.69 log10 CFU/ml increased in the control to 9.34, while it was reduced to 4.83 by ciprofloxacin and to 4.25 by levofloxacin . Mice were infected with 10(7) CFU/g of K . pneumoniae intraperitoneally . Treatment started 2 hours later, when the mean blood bacterial counts were 7.33 log10 CFU/ml, and lasted for 26 hours from the time of infection . Blood bacterial count was reduced from 7.33 log10 CFU/ml to 4.08 log10 CFU/ml by ciprofloxacin (20 mg/kg/6 hours), and to 3.60 log10 CFU/ml by levofloxacin (50 mg/kg/6 hours) 8 hours after the infection, which differed significantly from the infected untreated group . Ciprofloxacin and levofloxacin prolonged significantly the survival of mice compared with the infected untreated group (p<0.001 for both groups) . There were not significant differences either in the survival (p=1.0) or in the blood bacterial counts (p=0.216 after 8 hours) between ciprofloxacin and levofloxacin group . Based on these results both ciprofloxacin and levofloxacin could be alternative therapeutic agents for the infection caused by ESBL-producing Klebsiella strains.

Am J Physiol Lung Cell Mol Physiol, 2002 May, 282(5), L944 - 56
Killing of Klebsiella pneumoniae by human alveolar macrophages; Hickman-Davis JM et al.; We investigated putative mechanisms by which human surfactant protein A (SP-A) effects killing of Klebsiella pneumoniae by human alveolar macrophages (AMs) isolated from bronchoalveolar lavagates of patients with transplanted lungs . Coincubation of AMs with human SP-A (25 microg/ml) and Klebsiella resulted in a 68% decrease in total colony forming units by 120 min compared with AMs infected with Klebsiella in the absence of SP-A, and this SP-A-mediated effect was abolished by preincubation with N(G)-monomethyl-L-arginine . Incubation of transplant AMs with SP-A increased intracellular Ca(2+) concentration ({Ca(2+)}(i)) by 70% and nitrite and nitrate (NO(x)) production by 45% (from 0.24 +/- 0.02 to 1.3 +/- 0.21 nmol small middle dot 10(6) AMs(-1).h(-1)) . Preincubation with 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester inhibited the increase in {Ca(2+)}(i) and abrogated the SP-A-mediated Klebsiella phagocytosis and killing . In contrast, incubation of AMs from normal volunteers with SP-A decreased both {Ca(2+)}(i) and NO(x) production and did not result in killing of Klebsiella . Significant killing of Klebsiella was also seen in a cell-free system by sustained production of peroxynitrite (>1 microM/min) at pH 5 but not at pH 7.4 . These findings indicate that SP-A mediates pathogen killing by AMs from transplant lungs by stimulating phagocytosis and production of reactive oxygen-nitrogen intermediates.

J Immunol, 2002 Apr 15, 168(8), 4018 - 24
Leptin-deficient mice exhibit impaired host defense in Gram-negative pneumonia; Mancuso P et al.; Leptin is an adipocyte-derived hormone that is secreted in correlation with total body lipid stores . Serum leptin levels are lowered by the loss of body fat mass that would accompany starvation and malnutrition . Recently, leptin has been shown to modulate innate immune responses such as macrophage phagocytosis and cytokine synthesis in vitro . To determine whether leptin plays a role in the innate host response against Gram-negative pneumonia in vivo, we compared the responses of leptin-deficient and wild-type mice following an intratracheal challenge of Klebsiella pneumoniae . Following K . pneumoniae administration, we observed increased leptin levels in serum, bronchoalveolar lavage fluid, and whole lung homogenates . In a survival study, leptin-deficient mice, as compared with wild-type mice, exhibited increased mortality following K . pneumoniae administration . The increased susceptibility to K . pneumoniae in the leptin-deficient mice was associated with reduced bacterial clearance and defective alveolar macrophage phagocytosis in vitro . The exogenous addition of very high levels of leptin (500 ng/ml) restored the defect in alveolar macrophage phagocytosis of K . pneumoniae in vitro . While there were no differences between wild-type and leptin-deficient mice in lung homogenate cytokines TNF-alpha, IL-12, or macrophage-inflammatory protein-2 after K . pneumoniae administration, leukotriene synthesis in lung macrophages from leptin-deficient mice was reduced . Leukotriene production was restored by the addition of exogenous leptin (500 ng/ml) to macrophages in vitro . This study demonstrates for the first time that leptin-deficient mice display impaired host defense in bacterial pneumonia that may be due to a defect in alveolar macrophage phagocytosis and leukotriene synthesis.

J Mol Microbiol Biotechnol, 2002 May, 4(3), 235 - 42
Regulation of nitrogen fixation in Klebsiella pneumoniae and Azotobacter vinelandii: NifL, transducing two environmental signals to the nif transcriptional activator NifA; Schmitz RA et al.; The enzymatic reduction of molecular nitrogen to ammonia requires high amounts of energy, and the presence of oxygen causes the catalyzing nitrogenase complex to be irreversible inactivated . Thus nitrogen-fixing microorganisms tightly control both the synthesis and activity of nitrogenase to avoid the unnecessary consumption of energy . In the free-living diazotrophs Klebsiella pneumoniae and Azotobacter vinelandii, products of the nitrogen fixation nifLA operon regulate transcription of the other nifoperons . NifA activates transcription of nif genes by the alternative form of RNA-polymerase, sigma54-holoenzyme; NifL modulates the activity of the transcriptional activator NifA in response to the presence of combined nitrogen and molecular oxygen . The translationally-coupled synthesis of the two regulatory proteins, in addition to evidence from studies of NifL/NifA complex formation, imply that the inhibition of NifA activity by NifL occurs via direct protein-protein interaction in vivo . The inhibitory function of the negative regulator NifL appears to lie in the C-terminal domain, whereas the N-terminal domain binds FAD as a redox-sensitive cofactor, which is required for signal transduction of the internal oxygen status . Recently it was shown, that NifL acts as a redox-sensitive regulatory protein, which modulates NifA activity in response to the redox-state of its FAD cofactor, and allows NifA activity only in the absence of oxygen . In K . pneumoniae, the primary oxygen sensor appears to be Fnr (fumarate nitrate reduction regulator), which is presumed to transduce the signal of anaerobiosis towards NifL by activating the transcription of gene(s) whose product(s) function to relieve NifL inhibition through reduction of the FAD cofactor . In contrast, the reduction of A . vinelandii-NifL appears to occur unspecifically in response to the availability of reducing equivalents in the cell . Nitrogen status of the cells is transduced towards the NifL/NifA regulatory system by the GlnK protein, a paralogue PII-protein, which appears to interact with the NifL/NifA regulatory system via direct protein-protein interaction . It is not currently known whether GlnK interacts with NifL alone or affects the NifL/NifA-complex; moreover the effects appear to be the opposite in K . pneumoniae and A . vinelandii . In addition to these environmental signals, adenine nucleotides also affect the inhibitory function of NifL; in the presence of ATP or ADP the inhibitory effect on NifA activity in vitro is increased . The NifL proteins from the two organisms differ, however, in that stimulation of K . pneumoniae-NifL occurs only when synthesized under nitrogen excess, and is correlated with the ability to hydrolyze ATP . In general, transduction of environmental signals to the nif regulatory system appears to involve a conformational change of NifL or the NifL/NifA complex . However, experimental data suggest that K . pneumoniae and A . vinelandii employ significantly different species-specific mechanisms of signal transduction.

Hepatol Res, 2002 Apr, 22(4), 307 - 312
Septic endophthalmitis and meningitis associated with Klebsiella pneumoniae liver abscess; Ohmori S et al.; We report a female case of septic endophthalmitis and meningitis associated with Klebsiella pneumoniae liver abscess which was thought to be caused by duodeno-biliary reflux related to choledochoduodenostomy . We treated this patient by ultrasonography-guided percutaneous abscess drainage and intravenous administration of third generation antibiotics . However, the visual function of her left eye was eventually lost . Reports of liver abscess with metastatic lesions are rare, and our experience suggests that more physicians should be alert to septic metastatic lesions such as K . pneumoniae liver abscess or bacteremia with complaints of ocular or central nervous system symptoms.

Cent Afr J Med, 2001 May, 47(5), 119 - 23
Prevalence of causative organisms in corneal ulcers seen at Sekuru Kaguvi Eye Unit, Harare, Zimbabwe; Wani MG et al.; OBJECTIVE: To investigate the prevalence of organisms causing corneal ulcers, determine frequency of ocular risk factors and assess the value of Gram stain in initial identification of pathogens . DESIGN: A descriptive cross sectional study . SETTING: Sekuru Kaguvi Eye Unit at Parirenyatwa Group of Hospitals in Harare, Zimbabwe . MATERIALS AND METHODS: We enrolled 43 patients with corneal ulcers for the study . Demographic data was obtained, measurement of visual acuity was done by Snellen's chart and a Topcon Slit Lamp was used to determine the characteristics of ulcers for all the patients . Corneal scrapings were obtained and examined by microscopy and culture methods to determine etiologic organisms . Positivity rates between Gram stain and culture methods were compared . MAIN OUTCOME MEASURES: Prevalence of different bacterial isolates in corneal ulcers, risk factors to infection and laboratory methods used for detection . RESULTS: Gram-positive cocci were the most prevalent organisms found (54%) followed by E . coli (18%) . Pseudomonas, Klebsiella and Narcodia constituted about 9% each . Trauma, use of traditional eye medicine and previous viral disease were the common risk factors . CONCLUSION: Bacteria were the only organisms identified and Gram-positive cocci were the most common isolates . Gram stain of corneal smears was not reliable in initial recognition of offending pathogens.

Arch Microbiol, 2002 Mar, 177(3), 223 - 34 Epub 2001 Dec 12.
Membrane association of Klebsiella pneumoniae NifL is affected by molecular oxygen and combined nitrogen; Klopprogge K et al.; In the diazotroph Klebsiella pneumoniae, NifL and NifA regulate transcription of the nitrogen fixation genes in response to molecular oxygen and combined nitrogen . We recently showed that Fnr is the primary oxygen sensor . Fnr transduces the oxygen signal towards the negative regulator NifL by activating genes whose products reduce the FAD moiety of NifL under anoxic conditions . These Fnr-dependent gene products could be membrane-bound components of the anaerobic electron transport chain . Consequently, in this study we examined the localization of NifL within the cell under various growth conditions . In K . pneumoniae grown under oxygen- and nitrogen-limited conditions, approximately 55% of the total NifL protein was found in the membrane fraction . However, when the cells were grown aerobically or shifted to nitrogen sufficiency, less than 10% of total NifL was membrane-associated . In contrast to NifL, NifA was located in the cytoplasm under all growth conditions tested . Further studies using K . pneumoniae mutant strains showed that, under derepressing conditions but in the absence of either the primary oxygen sensor Fnr or the primary nitrogen sensor GlnK and the ammonium transporter AmtB, NifL was located in the cytoplasm and inhibited NifA activity . These findings suggest that under nitrogen- and oxygen-limitation, a significantly higher membrane affinity of NifL might create a spatial gap between NifL and its cytoplasmic target protein NifA, thereby impairing inhibition of NifA by NifL . Localization of GlnK further showed that, under nitrogen-limited conditions but independent of the presence of oxygen, 15-20% of the total GlnK is membrane-associated.

ANZ J Surg, 2001 Dec, 71(12), 744 - 6
Pyogenic liver abscess complicated by endogenous endophthalmitis; Tan YM et al.; BACKGROUND: Endogenous endophthalmitis is an inflammation of ocular tissues that can lead to deterioration of and loss of vision . Rarely, this can complicate the course of a patient with pyogenic liver abscess . METHODS: Over an 18-month period, 68 patients were treated for pyogenic liver abscesses . Three patients, all of whom were male and with diabetes, were diagnosed with a Klebsiella pneumoniae liver abscess complicated by endogenous endophthalmitis . Open surgical or percutaneous drainage of the liver abscess was undertaken and the symptomology and outcome of the endophthalmitis reviewed . RESULTS: There was no mortality in our series . Two patients presented with simultaneous abdominal and ocular symptoms and one patient had ocular symptoms 3 days after surgical drainage of the liver abscess . Despite aggressive treatment, all patients had permanent deterioration of visual function with one patient becoming blind and requiring evisceration of the infected eye . CONCLUSION: Ocular symptoms in patients treated for pyogenic abscesses must be dealt with urgently with an ophthalmologic consultation . Increased awareness of this complication and a high index of suspicion are paramount for salvage of visual function.

Biochim Biophys Acta, 2002 Feb 11, 1594(2), 243 - 54
The inhibitory form of NifL from Klebsiella pneumoniae exhibits ATP hydrolyzing activity only when synthesized under nitrogen sufficiency; Klopprogge K et al.; The inhibitory function of Klebsiella pneumoniae NifL on NifA transcriptional activity in vitro is stimulated by ATP and ADP when NifL is synthesized under nitrogen sufficiency (NifL(NH4)) . Further characterizations showed that NifL(NH4) binds and hydrolyzes ATP (2500 mU/mg) . Analyzing fusions between MalE and different portions of NifL, we localized both the ATP binding site and ATP hydrolysis activity to the N-terminal domain of NifL . In contrast, NifL synthesized under nitrogen limitation is not affected by adenine nucleotides and exhibits no ATP hydrolyzing activity . These major differences indicate that the stimulation of the inhibitory function of NifL and the ability to hydrolyze ATP depend on a specific NifL conformation induced by ammonium . We hypothesize that the presence of ammonium alters the conformation of NifL, enabling it to use the energy of ATP hydrolysis to increase the efficiency of NifL-NifA complex formation.

Mol Microbiol, 1997 Jul, 25(1), 3 - 10
Enzymic and genetic basis for bacterial growth on malonate; Dimroth P et al.; Various bacteria are able to grow aerobically or anaerobically on malonate as sole source of carbon and energy . Independent of the mechanism for energy conservation, the decarboxylation of malonate is the key reaction in the decomposition of this compound . To achieve malonate decarboxylation under physiological conditions, the substrate must be converted into an activated (thioester) derivative . We report here on the malonate decarboxylases of Malonomonas rubra and Klebsiella pneumoniae . These enzymes perform an interesting substrate activation mechanism by generating a malonyl thioester with the enzyme . Formation of the malonyl-S-enzyme involves an 'activation module' that comprises the acetylation of a specific thiol group of an acyl carrier protein (ACP) and the transfer of the ACP moiety to malonate, yielding malonyl-S-ACP and acetate . The malonyl-S-ACP is subsequently decarboxylated with regeneration of the acetyl-ACP . The malonate activation mechanism is related to the activation of citrate by citrate lyase . The relationship extends to the identical 2'-(5''-phosphoribosyl)-3'-dephospho-CoA thiol cofactor that is bound covalently to the corresponding ACP subunit . In Klebsiella pneumoniae, malonate is decarboxylated by a water-soluble enzyme complex . In the anaerobic bacterium Malonomonas rubra, malonate decarboxylation is catalysed by a set of water-soluble as well as membrane-bound enzymes that function together in converting the free energy of the decarboxylation reaction into delta muNa+ . Therefore, this malonate decarboxylase includes a biotin carrier protein that accepts the CO2 moiety from malonyl-S-ACP and delivers it to a membrane-bound decarboxylase acting as a Na+ pump . Genes encoding the individual protein components that perform the decarboxylation of malonate in K . pneumoniae or M . rubra have been identified within the mdc and mad gene clusters respectively . The function of most of the derived proteins could be envisaged from sequence similarities with proteins of known functions . The genetic evidence firmly supports the idea that malonate decarboxylation is carried out by the two different decarboxylases, as deduced from the biochemical studies of the enzymes.

Chemotherapy, 2002 Mar, 48(1), 10 - 4
Selection of Klebsiella pneumoniae mutants with high-level cefotaxime resistance during growth in serum containing therapeutic concentrations of cefotaxime; Bedenic B; BACKGROUND: In a previous investigation of the genetic characterization of extended-spectrum beta-lactamases (ESBLs) in Klebsiella pneumoniae from Zagreb, Croatia, 20 strains were found to produce SHV-2 beta-lactamase . Those strains displayed varying degrees of beta-lactam resistance and a wide range of beta-lactamase activity . We concluded that more resistant isolates were hyperproducers of SHV-2 beta-lactamase . METHODS: In this investigation, we tried to develop hyperproducing variants from 8 low-level SHV-2 beta-lactamase-producing Klebsiella strains by subculturing them in serum containing therapeutic concentrations of cefotaxime (CTX) . RESULTS: In most cases, there was a moderate increase in CTX resistance (twofold to threefold), except in one strain which displayed a 16-fold increase in the minimum inhibitory concentration (MIC) of CTX after incubation in the serum . That strain showed a marked increase in enzyme activity as well . The strains with a moderate increase in CTX MIC did not produce more enzyme after exposure to the serum, except for one strain which had a threefold rise in beta-lactamase activity after exposure to serum . CONCLUSIONS: In this investigation, it was established that the mutants with high-level CTX resistance developed very quickly in the biological fluids containing therapeutic concentrations of CTX . It is reasonable to expect that a similar process occurs in patients infected with an ESBL-producing K . pneumoniae strain during antibiotic treatment . Since most of the high-level CTX-resistant mutants did not have a marked rise in beta-lactamase activity after exposure to serum, it is possible that the elevated resistance was due to some other mechanism, such as reduced penicillin-binding protein affinity, changes in outer membrane proteins or efflux by multidrug efflux pumps .

Gig Sanit, 2002 Jan-Feb, (1), 54 - 6
{Effects of toxic substances on opportunistic microorganisms}; Boiko OV et al.; Anti-immunoglobulin activity of immunoglobulins, rarely receiving attention in literature, occupies a special place among the special bacterial substances released by bacteria exocellularly . We studied the effects of some pollutants (phenol, diethanolamine, nitric and hydrochloric acids) on this pathogenicity factor . Phenol and hydrochloric acid added to tap water caused statistically significant changes in anti-immunoglobulin activity of Klebsiella strains.

Antimicrob Agents Chemother, 2002 Apr, 46(4), 1098 - 100
First description of Klebsiella pneumoniae harboring CTX-M beta-lactamases (CTX-M-14 and CTX-M-3) in Taiwan; Yu WL et al.; Klebsiella pneumoniae isolates from Taiwan medical centers (50 strains; 1998 to 2000) with a CTX-M resistance phenotype (ceftazidime susceptible and ceftriaxone or cefotaxime nonsusceptible) were selected for initial isoelectric focusing analysis . beta-Lactamases with pIs of 7.9 (n = 22) and 8.4 (n = 28) in addition to 5.4 and/or 7.6 were detected . DNA gene sequencing identified the beta-lactamases with pIs of 7.9 and 8.4 as CTX-M-14 and CTX-M-3, respectively . Molecular typing suggested inter- and intrahospital clonal dissemination of these Taiwanese CTX-M-producing Klebsiella strains.

Emerg Infect Dis, 2002 Feb, 8(2), 160 - 6
Community-acquired Klebsiella pneumoniae bacteremia: global differences in clinical patterns; Ko WC et al.; We initiated a worldwide collaborative study, including 455 episodes of bacteremia, to elucidate the clinical patterns of Klebsiella pneumoniae . Historically, community-acquired pneumonia has been consistently associated with K . pneumoniae . Only four cases of community-acquired bacteremic K . pneumoniae pneumonia were seen in the 2-year study period in the United States, Argentina, Europe, or Australia; none were in alcoholics . In contrast, 53 cases of bacteremic K . pneumoniae pneumonia were observed in South Africa and Taiwan, where an association with alcoholism persisted (p=0.007) . Twenty-five cases of a distinctive syndrome consisting of K . pneumoniae bacteremia in conjunction with community-acquired liver abscess, meningitis, or endophthalmitis were observed . A distinctive form of K . pneumoniae infection, often causing liver abscess, was identified, almost exclusively in Taiwan.

EMBO J, 1983, 2(1), 39 - 44
Nitrogen control of the nif regulon in Klebsiella pneumoniae: involvement of the ntrA gene and analogies between ntrC and nifA; Merrick MJ; The ntrC and nifA gene products of Klebsiella pneumoniae are transcriptional activators involved in general nitrogen control and nif-specific regulation, respectively . Multicopy plasmids expressing either ntrC or nifA from a foreign promoter were used to study the relationship between these two genes and ntrA . The nifA product substituted for ntrC product in activation of a number of genes including nifLA, hutUH and genes for arginine and proline utilisation . NtrC could not substitute for nifA in transcriptional activation of the nifHDKY operon . In ntrA- strains, neither the ntrC nor the nifA product functioned to activate transcription of nif promoters . In vitro transcription/translation studies with plasmid clones demonstrated similar levels of expression of ntrC and nifA in ntr+ and ntrA- S-30 extracts . Hence, lack of activator function in an ntrA mutant indicates that both the ntrC and nifA products require a functional ntrA for activity . When expressed from foreign promoters, both the ntrC and nifA products were active in conditions which would normally repress nif expression . Hence, the ntrA product was apparently not limiting in these conditions.

Med Sci Monit, 2002 Mar, 8(3), BR100 - 4
Resistance of Klebsiella pneumoniae strains producing and not producing ESBL (extended-spectrum beta-lactamase) type enzymes to selected non-beta-lactam antibiotics; Sekowska A et al.; BACKGROUND: Bacteria of the Klebsiella genus may cause numerous infections in human, which are often treated with beta-lactam antibiotics . The fundamental mechanism of Klebsiella resistance to penicillins or cephalosporins involves the production of enzymes--extended-spectrum beta-lactamases (ESBLs) . Because of resistance of many Klebsiella spp . strains to beta-lactams, alternative antibiotic therapy can make use of aminoglycosides and quinolones . MATERIAL/METHODS: The study analyzed the prevalence of ESBL -type enzymes among 256 Klebsiella pneumoniae strains isolated from various clinical materials collected from patients hospitalized between 1997 and 2000 . ESBLs were detected by double-disk synergy test (DDST) . The prevalence of strains resistant to selected aminoglycosides (gentamicin, amikacin, netilmicin) and quinolones (ciprofloxacin, norfloxacin, nalidixic acid) in the particular years was analyzed . Drug sensitivity was determined by disk-diffusion method according to the recommendations of the National Reference Center for Microbial Drug Sensitivity . RESULTS: During the analyzed time interval, a significant increase of the number of K . pneumoniae ESBL(+) strains was noted: in 1997 - 16.5% (14/85) and in 2000 - 40.4% (22/54) (p<0.001) . Among the ESBL(+) strains, an increase of the number of strains resistant to the tested antibiotics, except for nalidixic acid, was demonstrated CONCLUSIONS: A statistically significant increase of multidrug-resistant K . pneumoniae strains, including strains producing ESBLs, was demonstrated in the analyzed material.

Microbiology, 2002 Mar, 148(Pt 3), 843 - 52
Metabolism of sucrose and its five isomers by Fusobacterium mortiferum; Pikis A et al.; Fusobacterium mortiferum utilizes sucrose {glucose-fructose in alpha(1-->2) linkage} and its five isomeric alpha-D-glucosyl-D-fructoses as energy sources for growth . Sucrose-grown cells are induced for both sucrose-6-phosphate hydrolase (S6PH) and fructokinase (FK), but the two enzymes are not expressed above constitutive levels during growth on the isomeric compounds . Extracts of cells grown previously on the sucrose isomers trehalulose alpha(1-->1), turanose alpha(1-->3), maltulose alpha(1-->4), leucrose alpha(1-->5) and palatinose alpha(1-->6) contained high levels of an NAD+ plus metal-dependent phospho-alpha-glucosidase (MalH) . The latter enzyme was not induced during growth on sucrose . MalH catalysed the hydrolysis of the 6'-phosphorylated derivatives of the five isomers to yield glucose 6-phosphate and fructose, but sucrose 6-phosphate itself was not a substrate . Unexpectedly, MalH hydrolysed both alpha- and beta-linked stereomers of the chromogenic analogue p-nitrophenyl glucoside 6-phosphate . The gene malH is adjacent to malB and malR, which encode an EII(CB) component of the phosphoenolpyruvate-dependent sugar:phosphotransferase system and a putative regulatory protein, respectively . The authors suggest that for F . mortiferum, the products of malB and malH catalyse the phosphorylative translocation and intracellular hydrolysis of the five isomers of sucrose and of related alpha-linked glucosides . Genes homologous to malB and malH are present in both Klebsiella pneumoniae and the enterohaemorrhagic strain Escherichia coli O157:H7 . Both these organisms grew well on sucrose, but only K . pneumoniae exhibited growth on the isomeric compounds.

Proc Natl Acad Sci U S A, 2002 Mar 5, 99(5), 2696 - 701
Microcin E492, a channel-forming bacteriocin from Klebsiella pneumoniae, induces apoptosis in some human cell lines; Hetz C et al.; The cytotoxic effect of microcin E492, a low-molecular-mass channel-forming bacteriocin (7,887 Da) produced by a strain of Klebsiella pneumoniae, was characterized in HeLa cells . At low (5 microg/ml) and intermediate (10 microg/ml) concentrations, microcin E492 induced biochemical and morphological changes typical of apoptosis, such as cell shrinkage, DNA fragmentation, extracellular exposure of phosphatidylserine, caspase activation, and loss of mitochondrial membrane potential . Treatment with zVAD-fmk, a general caspase inhibitor, completely blocked the cytotoxic effect of this bacteriocin . At higher microcin concentrations (>20 microg/ml) a necrotic phenotype was observed . Induction of apoptosis by microcin E492 was associated with the release of calcium from intracellular stores, probably after microcin-triggered ion channel formation . Microcin E492 also presented a cytotoxic effect on Jurkat and RJ2.25 cells, but had no effect on KG-1 cells nor on a primary culture of human tonsil endothelial cells, suggesting that there is a specific interaction of the bacteriocin with components of the target cell surface . This report describes a bacteriocin that has the capacity to induce apoptosis in human cell lines.

J Clin Microbiol, 2002 Mar, 40(3), 799 - 804
Extended-spectrum beta-lactamase-producing Klebsiella pneumoniae in a neonatal intensive care unit in the high-prevalence area of Athens, Greece; Lebessi E et al.; Extended-spectrum beta-lactamase-producing Klebsiella pneumoniae (EPKP) strains are frequently implicated in outbreaks in neonatal intensive care units (NICUs) . During the period from 1997 to 1998, 21 infections and 23 colonizations with EPKP were recorded in the NICU of a children's hospital in Athens, Greece . Seventeen of the infected and 12 of the colonized neonates had been referred from other hospitals . The remaining infections and colonizations occurred during the current hospitalization . Pulsed-field gel electrophoresis typing showed that the latter cases were due to an outbreak strain that persisted in the unit, while the repeated introduction of EPKP carriers was mostly due to clonal outbreaks in two maternity hospitals.

Zh Mikrobiol Epidemiol Immunobiol, 2001 Sep-Oct, (5), 46 - 9
{Use of prodigiozan and tactivin for treatment of patients with chronic Klebsiella infections}; Titov LP et al.; Two-stage immunocorrection in patients with chronic Klebsiella infections by consecutive administration of prodigiozan and tactivin, has been shown to facilitate the restoration of the balance of immunoregulating cells, to enhance proliferative response to T- and B-mitogens, thus producing favorable clinical dynamics . The possibility of chronic Klebsiella infections therapy, based on the use of immunocorrecting preparations (prodigiozan and tactivin) and antibiotics capable of intracellular penetration (sumamed and cyprofloxacin) is discussed.

Zhonghua Bing Li Xue Za Zhi, 2000 Dec, 29(6), 421 - 3
{An etiological and pathologic study of Rhinoscleroma}; Zhang S et al.; OBJECTIVE: To investigate the clinicopathologic characteristics and mechanism of tissue injury caused by Klebsiella rhinoscleromatis (KR) infection . METHODS: Thirty-three cases of rhinoscleromas in the nasal and pharyngeal region seen in this hospital from Jan . 1980 to Mar . 2000 were studied retrospectively . Warthin-Starry (W-S) stain was used to study all cases . Five cases were studied further with transmission electron microscope . RESULTS: W-S stain revealed a great number of KR in Mikulicz cells within the granuloma . Electron microscopy demonstrated a number of phagosomes in the cytoplasm of Mikulicz cells where many KR were found . A small number of other organelles such as endoplasmic reticulums and lysozymes were squeezed to the side of cells . There were many granular substances on the surface of intracellular bacteria, which were not found on the extracellular KR . CONCLUSION: KR is a facultative intracellular bacteria that is able to resist the digestion of macrophages and to proliferate in them . Formation of granulomas and fibrosis in KR infiltrated regions is the major cause of tissue injury.

Clin Microbiol Infect, 1998, 4(10), 570 - 576
Bactericidal activity of three beta-lactams alone or in combination with a beta-lactamase inhibitor and two aminoglycosides against Klebsiella pneumoniae harboring extended-spectrum beta-lactamases; Roussel-Delvallez M et al.; OBJECTIVE: To study the bactericidal activity of beta-lactam antibiotics (imipenem, cefepime, cefpirome) alone or in combination with a beta-lactamase inhibitor (sulbactam) in the presence or absence of aminoglycoside (amikacin or isepamicin) against Klebsiella pneumoniae strains producing extended-spectrum beta-lactamases (ESBLs) . METHODS: We characterized 10 strains by means of analytic isoelectric focusing and pulsed-field gel electrophoresis . The ESBLs produced by these strains were derived from either TEM (TEM-1, TEM-2) or SHV-1 . The killing-curve method was used for this bacterial investigation . Bacteria (final inoculum 5x105 CFU/mL) were incubated with antibiotics at clinical concentrations obtained in vivo . RESULTS: All the combinations with cefepime or cefpirome + sulbactam were bactericidal, with a 4 log10 decrease being obtained within 6 h without regrowth at 24 h, whereas imipenem alone, and combinations, gave a bactericidal effect within 6 h . The two cephalosporins alone decreased the inoculum of 4 log10 at 6 h but regrowth was observed at 24 h . When the aminoglycoside was added, this bactericidal effect was obtained within 3 h with amikacin and within 1 h with isepamicin . CONCLUSIONS: Cefepime + sulbactam or cefpirome + sulbactam may be an alternative to imipenem for the treatment of patients with ESBL-producing K . pneumoniae . Aminoglycosides are often associated in nosocomial infections due to ESBL-producing K . pneumoniae: isepamicin acted faster than amikacin, but both worked well . To conclude, it may be prudent to avoid extended-spectrum cephalosporins as single agent when treating serious infections due to ESBL-producing K . pneumoniae . Addition of a beta-lactamase inhibitor such as sulbactam +/- aminoglycoside is advisable to avoid failure of treatment.

Clin Microbiol Infect, 1997, 3(5), 549 - 554
A simple and reliable method to screen isolates of Escherichia coli and Klebsiella pneumoniae for the production of TEM- and SHV-derived extended-spectrum beta-lactamases; Thomson KS et al.; OBJECTIVE: To evaluate which of 24 beta-lactams used in susceptibility tests best discriminated between strains of Klebsiella pneumoniae and Escherichia coli that produce extended spectrum beta-lactamases (ESBLs) from strains that produce older, more familiar, plasmid-mediated beta-lactamases such as TEM-1 and SHV-1 . METHODS: Susceptibility to the 24 beta-lactam agents was determined by agar dilution and disk diffusion methodologies, using 27 strains of K . pneumoniae and E . coli that produced 22 different older plasmid-mediated beta-lactamases and 28 strains that produced 17 different ESBLs . RESULTS: In general, strains that produced ESBLs were intermediate or resistant to cefpodoxime, whereas those that produced other beta-lactamases were susceptible to this agent . The agar dilution test exhibited 96% sensitivity and 100% specificity in discriminating these two groups of organisms . The disk diffusion test exhibited 100% sensitivity and 96% specificity . All other beta-lactam agents tested were inferior discriminators between the two groups of organisms . CONCLUSIONS: Agar dilution and disk diffusion tests with cefpodoxime can be used to discriminate strains of K . pneumoniae and E . coli that produce ESBLs from those that produce older, plasmid-mediated beta-lactamases.

Carbohydr Res, 2002 Mar 1, 337(5), 383 - 90
Synthesis of alpha-Manp-(1-->2)-alpha-Manp-(1-->3)-alpha-Manp-(1-->3)-Manp, the tetrasaccharide repeating unit of Escherichia coli O9a, and alpha-Manp-(1-->2)-alpha-Manp-(1-->2)-alpha-Manp-(1-->3)-alpha-Manp-(1-->3)-Manp, the pentasaccharide repeating unit of E . coli O9 and Klebsiella O3; Chen L et al.; The tetrasaccharide repeating unit of Escherichia coli O9a, alpha-D-Manp-(1-->2)-alpha-D-Manp-(1-->3)-alpha-D-Manp-(1-->3)-D-Manp, and the pentasaccharide repeating unit of E . coli O9 and Klebsiella O3, alpha-D-Manp-(1-->2)-alpha-D-Manp-(1-->2)-alpha-D-Manp-(1-->3)-alpha-D-Manp-(1-->3)-D-Manp, were synthesized as their methyl glycosides . Thus, selective 3-O-allylation of p-methoxyphenyl alpha-D-mannopyranoside via a dibutyltin intermediate gave p-methoxyphenyl 3-O-allyl-alpha-D-mannopyranoside (2) in good yield . Benzoylation (-->3), then removal of 1-O-methoxyphenyl (right arrow4), and subsequent trichloroacetimidation afforded the 3-O-allyl-2,4,6-tri-O-benzoyl-alpha-D-mannopyranosyl trichloroacetimidate (5) . Condensation of 5 with methyl 4,6-O-benzylidene-alpha-D-mannopyranoside (6) selectively afforded the (1-->3)-linked disaccharide 7 . Benzoylation of 7, debenzylidenation, benzoylation, and deallylation gave methyl 2,4,6-tri-O-benzoyl-alpha-D-mannopyranosyl-(1-->3)-2,4,6-tri-O-benzoyl-alpha-D-mannopyranoside (11) as the disaccharide acceptor . Coupling of 11 with (1-->2)-linked mannose disaccharide donor 17 or trisaccharide donor 21, followed by deacylation, furnished the target tetrasaccharide and pentasaccharide, respectively.

Arch Pathol Lab Med, 2002 Mar, 126(3), 372 - 4
Malakoplakia of liver diagnosed by a needle core biopsy: a case report and review of the literature; Hartman G et al.; Although malakoplakia of the genitourinary tract and colon is reported frequently in the literature, malakoplakia that occurs primarily in the liver is rare, and only 4 cases have been described thus far . To our knowledge, this is the first case of malakoplakia of the liver diagnosed by a needle core biopsy . This case occurred in a 19-year-old man with small bowel ileus following Klebsiella pneumonia.

Clin Microbiol Infect, 1999 Apr, 5(4), 184 - 189
Epidemiologic typing of international collections of Klebsiella spp.: computerized biochemical fingerprinting compared with serotyping, phage typing and pulsed-field gel electrophoresis; Tullus K et al.; OBJECTIVE: The Phene Plate (PhP) biochemical fingerprinting system is based on measurements of the kinetics of selected biochemical reactions performed in microtiter plates, and computerized data-processing . This study compared the performance of the PhP system as an epidemiologic tool with other commonly used typing systems . METHODS: PhP typing was applied to 107 nosocomial Klebsiella spp . isolates from 10 collections, mostly representing outbreaks . The results were compared with those obtained by capsular (K) serotyping, phage typing and, for a subset of isolates (n=33), pulsed-field gel electrophoresis (PFGE) . RESULTS: Clusters of identical or closely related isolates based on serotype, phage type, PhP type and PFGE type were found in most collections . The typeability was 100%, 95%, 94% and 68% for PhP, K, PFGE and phage typing, respectively . The agreement between the typing methods was high (88-96%) . The discriminatory power was high for PhP and PFGE (diversity index 0.95 and 0.97, respectively), but lower for phage typing (diversity index 0.91) and K typing (diversity index 0.87) . CONCLUSIONS: Like serotyping and PFGE, PhP typing is useful in studies of the nosocomial epidemiology of Klebsiella spp . Combining PhP typing with PFGE or K typing rarely yielded additional information when comparing isolates within each collection, but PFGE sometimes discriminated between isolates of similar PhP type derived from different collections.

Infect Immun, 2002 Mar, 70(3), 1075 - 80
Role of lung epithelial cells in defense against Klebsiella pneumoniae pneumonia; Cortes G et al.; The airway epithelium represents a primary site for the entry of pathogenic bacteria into the lungs . It has been suggested for many respiratory pathogens, including Klebsiella pneumoniae, that adhesion and invasion of the lung epithelial cells is an early stage of the pneumonia process . We observed that poorly encapsulated K . pneumoniae clinical isolates and an isogenic unencapsulated mutant invaded lung epithelial cells more efficiently than highly encapsulated strains independent of the K type . By contrast, the unencapsulated mutant was completely avirulent in a mouse model of pneumonia, unlike the wild-type strain, which produced pneumonia and systemic infection . Furthermore, the unencapsulated mutant bound more epithelially produced complement component C3 than the wild-type strain . Our results show that lung epithelial cells play a key role as a host defense mechanism against K . pneumoniae pneumonia, using two different strategies: (i) ingestion and control of the microorganisms and (ii) opsonization of the microorganisms . Capsular polysaccharide avoids both mechanisms and enhances the virulence of K . pneumoniae.

Biologicals, 2001 Sep-Dec, 29(3-4), 293 - 8
Stability and CTL-activity of P40/ELA melanoma vaccine candidate; Beck A et al.; The decapeptide ELA (ELAGIGILTV), a Melan-A/MART-1 antigen immunodominant peptide analogue, is an interesting melanoma vaccine candidate alone or in combination with other tumour antigens . P40, the recombinant outer membrane protein A of Klebsiella pneumoniae (kpOmpA), was recently shown to target dendritic cells and to induce peptide-specific CTLs . Here we investigated the adjuvant role of P40 mixed or chemically conjugated to ELA . This compound is an N-terminal glutamic acid-containing peptide . However, it has been reported that the amino group and the gamma-carboxylic group of glutamic acids easily condense to form pyroglutamic derivatives . Usually, to overcome this stability problem, peptides of pharmaceutical interest were developed with a pyroglutamic acid instead of N-terminal glutamic acid, without loss of pharmacological properties . Unfortunately, the pyroglutamic acid derivative (PyrELA) as well as the N-terminal acetyl capped derivative (AcELA) failed to elicit CTL activity when mixed with P40 adjuvant protein . Despite the apparent minor modifications introduced by PyrELA and AcELA, these two derivatives have probably lower affinity than ELA for the class I Major Histocompatibility Complex . Furthermore, this stability problem is worse in the case of clinical grade ELA, produced as an acetate salt, like most of the pharmaceutical grade peptides . We report here that the hydrochloride shows a higher stability than the acetate and may be suitable for use in man .

Antimicrob Agents Chemother, 2002 Mar, 46(3), 659 - 64
Identification and minisequencing-based discrimination of SHV beta-lactamases in nosocomial infection-associated Klebsiella pneumoniae in Brisbane, Australia; Howard C et al.; Extended-spectrum beta-lactamases (ESBLs) are active against oxyimino cephalosporins and monobactams . Twenty-one Klebsiella pneumoniae isolates obtained between 1991 and 1995 at the Princess Alexandra Hospital in Brisbane, Australia, were subject to amplification and sequencing of the SHV beta-lactamase-encoding genes . Thirteen strains were phenotypically ESBL positive . Of these, six strains carried the blaSHV-2a gene and seven strains carried the blaSHV-12 gene . Eight strains were phenotypically ESBL negative . Of these, seven strains carried the non-ESBL blaSHV-11 gene and one strain carried the non-ESBL blaSHV-1 gene . There was complete correspondence between the ESBL phenotype and the presence or absence of an ESBL-encoding gene(s) . In addition, it was determined that of the 13 ESBL-positive strains, at least 4 carried copies of a non-ESBL-encoding gene in addition to the blaSHV-2a or blaSHV12 gene . A minisequencing-based assay was developed to discriminate the different SHV classes . This technique, termed "first-nucleotide change," involves the identification of the base added to a primer in a single-nucleotide extension reaction . The assay targeted polymorphisms at the first bases of codons 238 and 240 and reliably discriminated ESBL-positive strains from ESBL-negative strains and also distinguished strains carrying blaSHV-2a from strains carrying blaSHV-12 . In addition, this method was used to demonstrate an association between the relative copy numbers of blaSHV genes in individual strains and the levels of antibiotic resistance.

Infect Control Hosp Epidemiol, 2001 Nov, 22(11), 725 - 8
Outbreak of nosocomial sepsis and pneumonia in a newborn intensive care unit by multiresistant extended-spectrum beta-lactamase-producing Klebsiella pneumoniae: high impact on mortality; Martinez-Aguilar G et al.; We describe a case-control study of a small outbreak of nosocomial sepsis and pneumonia with high mortality due to clonal dissemination of a multiresistant Klebsiella pneumoniae in the neonatal intensive care unit of a Mexican institution . Our study helped to change nosocomial infection control policy in this hospital.

Infect Control Hosp Epidemiol, 2001 Nov, 22(11), 723 - 5
Multiresistant extended-spectrum beta-lactamase-producing Klebsiella pneumoniae causing an outbreak of nosocomial bloodstream infection; Gonzalez-Vertiz A et al.; This article describes an outbreak of bloodstream infection due to clonal dissemination of multiresistant Klebsiella pneumoniae in a neonatal area, during August 1999, in Mexico City General Hospital . The intestinal tract was the likely reservoir, and intensification of Contact Precaution measures contained the outbreak.

Pediatr Infect Dis J, 2002 Feb, 21(2), 169 - 70
Trichosporon asahii: an unusual cause of invasive infection in neonates; Panagopoulou P et al.; Trichosporon asahii causes white piedra, an infection of hair shafts and onychomycosis in immunocompetent patients, as well as various localized or disseminated invasive infections in immunodeficient hosts . We describe a 26-week gestation 890-g vaginally delivered female neonate who had severe respiratory distress syndrome and on the sixth day of life developed Klebsiella pneumoniae sepsis . At the same time two blood cultures were positive for T . asahii . The neonate was also colonized with T . asahii in the pharynx and perineum . The infant was successfully treated with conventional amphotericin B.

Gut, 2002 Mar, 50(3), 420 - 4
A global emerging disease of Klebsiella pneumoniae liver abscess: is serotype K1 an important factor for complicated endophthalmitis?
Fung CP, Chang FY, Lee SC, Hu BS, Kuo BI, Liu CY, Ho M, Siu LK.
BACKGROUND AND AIMS: Over the past two decades in Taiwan, pyogenic liver abscess has usually been caused by a single microorganism, Klebsiella pneumoniae, and is frequently associated with the serious complication of endophthalmitis, especially in diabetic patients . However, the relationship between the clinical presentation and bacterial factors remains unclear . The aim of this study was to investigate the clinical features of patients and the serotype and ribotype of K pneumoniae liver abscess . METHODS: From July 1991 to June 1998, a total of 134 cases of K pneumoniae liver abscess with 248 K pneumoniae isolates from the same patients were collected from two large medical centres in northern Taiwan . Clinical data were collected from medical records . Serotyping and ribotyping were performed using the countercurrent immunoelectrophoresis method and automated Riboprinter . RESULTS: Serotyping revealed that the most common serotypes were K1 (63.4%) and K2 (14.2%) . K1 isolates occurred at a significantly higher frequency (p<0.01) than all other serotypes . Among 134 patients, 105 (78.4%) had suffered from diabetes mellitus for 3-15 years . Fourteen patients (10.4%) had metastatic infection to the eye causing septic endophthalmitis . Liver aspirates, and blood and vitreous pus cultures yielded the same serotype of K pneumoniae in all patients . Among patients with septic endophthalmitis, 92.3% (13/14) were diabetic, and 85.7% (12/14) of the isolates belonged to serotype K1 . For molecular typing, different degrees of genetic polymorphism among isolates with the same K1 serotype suggested no particular prevalence of any one strain in K pneumoniae liver abscess . CONCLUSION: K pneumoniae serotype K1 was significantly associated with liver abscess and the complication of endophthalmitis, especially in diabetic patients . Physicians should request an immediate report of serotyping and susceptibility test results simultaneously if a diagnosis of pyogenic liver abscess has been made so that early and appropriate management for possible complications will not be delayed . The use of ceftriaxone because of its higher concentration in the aqueous humor is suggested to decrease the chance of septic endophthalmitis.

Biosci Biotechnol Biochem, 2001 Dec, 65(12), 2604 - 12
Induction of 4-hydroxycinnamate decarboxylase in Klebsiella oxytoca cells exposed to substrates and non-substrate 4-hydroxycinnamate analogs; Hashidoko Y et al.; The 4-hydroxycinnamate decarboxylase (4-HCD)-inducing activity of several substrate analogs toward Klebsiella oxytoca was investigated . Four E-cinnamate-class compounds, E-4-hydroxycinnamic acid (1), caffeic acid (2), ferulic acid (3) and E-2,4-dihydroxycinnamic acid (4), all of which were accepted as substrates, all of which were accepted as substrates of 4-HCD, enable K . oxytoca cells to induce the decarboxylase at a 2.0 mM concentration, while five non-substrate compounds of the E-cinnamate class so far tested were completely inactive . However, 6-hydroxy-2-naphthoic acid (11) and 7-hydroxycoumarin 3-carboxylic acid (14), both of which are non-cinnamate-class analogs of the substrate, acted as strong 4-HCD inducers, even at a 0.5 mM concentration . The 4-HCD-inducing activities of compounds 11 and 14 at 0.5 mM were 10-12-fold higher than that of substrate 1 . Compound 11 maintained its 4-HCD-inducing activity toward cultured cells through the late-log and stationary phases, unlike 1 that induced 4-HCD only in the early log phase . SDS-PAGE electrophoresis of protein mixtures from the cultured cells exposed to any 4-HCD inducer indicated that the 21.5 kDa protein was always present.

Chang Gung Med J, 2001 Nov, 24(11), 688 - 96
Klebsiella pneumoniae bacteremia: community-acquired vs . nosocomial infections; Yang PY et al.; BACKGROUND: This study attempted to determine the clinical manifestations and influential factors affecting the prognosis of patients with community-acquired and nosocomial bacteremia of Klebsiella pneumoniae (K . pneumoniae) . METHODS: We retrospectively reviewed the medical records of 211 patients who had a clinically significant episode of K . pneumoniae bacteremia from January 1997 until December 1999 . RESULTS: Most reports describe K . pneumoniae bacteremia as typically nosocomial, but in our study approximately 3 of 4 episodes were community-acquired . Without including "unknown origin", the most common infectious site for both community-acquired and nosocomial bacteremia was the hepatobiliary tract . The overall mortality for all 211 patients with K . pneumoniae bacteremia was 25.1% . Significantly higher mortality rates occurred in patients who were elderly (> 65 years), had a nosocomial infection, for whom the respiratory tract was the portal of entry, and ultimately fatal conditions or acute complications were due to shock or renal insufficiency . CONCLUSIONS: Patients with community-acquired and nosocomial bacteremia had different types of underlying diseases . Isolates from nosocomial infections were significantly more frequently resistant to aminoglycosides, antipseudomonal penicillin, and all three generations of cephalosporins . In this regard, an aggressive empirical therapeutic approach to infections of K . pneumoniae is suggested.

Antonie Van Leeuwenhoek, 2001 Sep, 79(3-4), 319 - 26
Cloning, sequencing and characterization of Fnr from Klebsiella pneumoniae; Grabbe R et al.; The transcription factor Fnr (fumarate nitrate reductase regulator) globally regulates gene expression in response to oxygen deprivation in Escherichia coli . We report here the cloning and sequencing of the fnr gene from the facultative anaerobic bacterium Klebsiella pneumoniae M5al, another member of the enteric bacteria . The deduced amino acid sequence of K . pneumoniae fnr showed very high similarity (98% amino acid identity) to the Fnr protein from E . coli and contained the four essential cysteine residues which are presumed to build the oxygen-sensing {4Fe4S}+2 center . Transfer of the K . pneumoniae gene to a fnr mutant of E . coli complemented the mutation and permitted synthesis of nitrate reductase and fumarate reductase during anaerobic growth . A gene fusion between K . pneumoniae fnr and glutathione S-transferase was constructed and expressed in E . coli under anaerobic conditions in order to make the protein available in preparative amounts . The overproduced protein was purified by glutathione-Sepharose 4B affinity chromatography in the absence of oxygen, and biochemically characterized.

J Antimicrob Chemother, 2002 Feb, 49(2), 261 - 7
Relationship between blaSHV-12 and blaSHV-2a in Korea; Kim J et al.; In contrast to the USA and Europe, where SHV-2, SHV-4 and SHV-5 are the prevalent extended-spectrum SHV enzymes, in Korea SHV-2a and SHV-12 are the most frequently identified extended-spectrum SHV enzymes . A 6.6 kb BamHI fragment containing the bla(SHV-12) gene of strain K7746 isolated from one university hospital in Korea was cloned into the pCRScriptCAM vector . Sequencing of the constructed recombinant plasmid pK7746-C1 revealed that the immediate upstream sequence of the bla(SHV-12) gene showed little similarity to the part of the prototype bla(SHV-1) gene due to the insertion of an IS26 element next to the -10 region . Instead, the upstream sequences of bla(SHV-12) retained 100% DNA identity with the part of plasmid pMPA2a from Klebsiella pneumoniae KPZU-3 carrying bla(SHV-2a) . The restriction map of the inserted 6.6 kb DNA fragment of plasmid pK7746-C1 was also homologous to that of plasmid pMPA2a, suggesting a common lineage of bla(SHV-12) and bla(SHV-2a) . We also studied, using PCR, the upstream non-coding region of several SHV beta-lactamase genes for the presence of IS26 sequence . The flanking IS26 sequence in the immediate upstream region of the bla(SHV) gene was not detected in five standard strains producing SHV-1, SHV-2, SHV-3, SHV-4 or SHV-5 . However, IS26 was detected in all 69 clinical strains producing SHV-2a or SHV-12 isolated from three university hospitals in Korea during 1993-1999 . The above findings suggest a direct evolution of SHV-12 from SHV-2a, not from SHV-2 to -5, and it is considered to be one of the reasons for the absolute predominance of SHV-2a and SHV-12 in Korea.

J Infect Chemother, 1999 Jun, 5(2), 91 - 96
Two sporadic cases of infections due to Klebsiella pneumoniae resistant to expanded-spectrum cephalosporins in Japan; Hirakata Y et al.; TEM- or SHV-type extended-spectrum beta-lactamases (ESBLs) are of clinical concern in Europe and the United States, whereas bacterial strains producing such types of ESBLs have not been reported in Japan . We report here two cases of infection due to Klebsiella pneumoniae resistant to extended-spectrum cephalosporins in Japan . A ceftadizime-resistant K . pneumoniae strain (minimum inhibitory concentration; 32 &mgr;g/ml) was isolated transiently from the sputum of an 87-year-old woman with acute myocardial infarction and pneumonia (patient 1) . Ceftadizime-susceptible and -resistant (minimum inhibitory concentration; >/=8 &mgr;g/ml) K . pneumoniae strains were isolated over a month from the blood, ascites, and feces of a 44-year-old man after bone marrow transplantation for acute lymphoblastic leukemia (patient 2); this patient died of K . pneumoniae sepsis and peritonitis followed by multiple organ failure . These isolates produced penicillinase, which was inhibited by clavulanic acid . A polymerase chain reaction (PCR) study showed that both isolates carried the SHV or LEN genes, but not the TEM, Toho-1, and IMP-1 genes . The pulsed-field gel electrophoresis profile of the strain isolated from patient 1 was genetically distinguishable from the profiles of the strains isolated from patient 2 . It appeared that mutation of the beta-lactamase gene may have occurred in the body of patient 2, since the genotypes of the ceftadizime-susceptible and -resistant isolates from this patient were identical . Another 12 strains of K . pneumoniae, isolated from other patients in the same wards during the period in which the K . pneumoniae strains were isolated from patients 1 and 2, did not produce ESBLs and showed different genotypes . The results suggest that these isolates of resistant K . pneumoniae did not spread by cross transmission in the hospital and that the two cases were sporadic . Surveillance of these types of resistant bacteria is necessary, since they may well be present in other hospitals in Japan . Although the organisms are suspected to produce SHV-type ESBLs or LEN-1 variant beta-lactamases, further studies are necessary to specify the resistance genes.

Acta Crystallogr D Biol Crystallogr, 2002 Feb, 58(Pt 2), 374 - 6 Epub 2002 Jan 24.
Crystallization and preliminary X-ray structure determination of jack bean urease with a bound antibody fragment; Sheridan L et al.; Urease allows organisms to use exogenous and internally generated urea as a nitrogen source, by catalyzing the hydrolysis of urea to form ammonia and carbon dioxide . Urease may also participate in the systemic nitrogen-transport pathways and possibly acts as a toxic defence protein . Jack bean urease (JBU) was the first nickel-metalloenzyme identified and was crystallized as early as 1926 . Despite this, the structure has not yet been determined . An antibody fragment, Fv, that has a high affinity for JBU has been used to aid crystallization . The complex, which retains full enzyme activity, forms very small crystals that diffract weakly to 3.3 A . The crystals belong to the rhombohedral space group R32, with unit-cell parameters a = b = 228.6, c = 130.9 A . The structure of the urease molecule has been solved by molecular replacement using the structure of homogenous enzyme from Klebsiella aerogenes as a search model.

J Bacteriol, 2002 Feb, 184(4), 1028 - 40
Characterization of GlnK1 from Methanosarcina mazei strain Gö1: complementation of an Escherichia coli glnK mutant strain by GlnK1; Ehlers C et al.; Trimeric PII-like signal proteins are known to be involved in bacterial regulation of ammonium assimilation and nitrogen fixation . We report here the first biochemical characterization of an archaeal GlnK protein from the diazotrophic methanogenic archaeon Methanosarcina mazei strain Go1 and show that M . mazei GlnK1 is able to functionally complement an Escherichia coli glnK mutant for growth on arginine . This indicates that the archaeal GlnK protein substitutes for the regulatory function of E . coli GlnK . M . mazei GlnK1 is encoded in the glnK1-amtB1 operon, which is transcriptionally regulated by the availability of combined nitrogen and is only transcribed in the absence of ammonium . The deduced amino acid sequence of the archaeal glnK1 shows 44% identity to the E . coli GlnK and contains the conserved tyrosine residue (Tyr-51) in the T-loop structure . M . mazei glnK1 was cloned and overexpressed in E . coli, and GlnK1 was purified to apparent homogeneity . A molecular mass of 42 kDa was observed under native conditions, indicating that its native form is a trimer . GlnK1-specific antibodies were raised and used to confirm the in vivo trimeric form by Western analysis . In vivo ammonium upshift experiments and analysis of purified GlnK1 indicated significant differences compared to E . coli GlnK . First, GlnK1 from M . mazei is not covalently modified by uridylylation under nitrogen limitation . Second, heterotrimers between M . mazei GlnK1 and Klebsiella pneumoniae GlnK are not formed . Because M . mazei GlnK1 was able to complement growth of an E . coli glnK mutant with arginine as the sole nitrogen source, it is likely that uridylylation is not required for its regulatory function.

Refuat Hapeh Vehashinayim, 2001 Oct, 18(3-4), 35 - 9, 109-10
{Parapharyngeal and peritonsillar infection following mandibular third molar extraction}; Abu el-Naaj I et al.; Numerous complications following impacted third molar extractions have been described previously . Among these are swelling, infection, subdermal hematomas, nerve injuries, injuries to adjacent teeth and mandibular fractures . The parapharyngeal space is a funnel-shaped space with its base located at the base of the skull and its apex near the hyoid bone . It is bounded medially by the superior constrictor muscle and the tonsillar fossa and laterally by the medial pterygoid muscle, the mandibular ramus, the deep lobe of the parotid gland and the posterior belly of the digastric muscle . The junction of the buccinator and superior constrictor muscles at the pterygomandibular raphe forms the anterior border . The posterior border is formed by the vertebral column and the prevertebral muscles . Infratemporal and peritonsillar space infections following third molar extractions are relatively rare because of anatomical barriers that exist in that area, but complications of such infections are considered to be highly severe and sometimes even life threatening . These complications include septic thrombophlebitis of the internal jugular vein, septic aneurysms of the internal carotid artery and mediastinitis . This article presents a parapharyngeal and peritonsillar space abscess resulting from a third molar extraction . Several articles reviewed in this paper revealed similar signs and symptoms in parapharyngeal abscesses . The main radiological finding was soft tissue swelling in the prevertebral area . Cultures have demonstrated Klebsiella pneumoniae as the dominant microorganism in these infections . Early identification and correct diagnosis of parapharyngeal and peritonsillar abscesses are necessary to avoid life-threatening complications that may accompany such infections . Airway control should receive top priority in treatment, followed by extensive surgical drainage and administration of high dose organism-specific antibiotics as well as removal of the source of infection.

Biochemistry, 2002 Jan 29, 41(4), 1285 - 92
Subunit gamma of the oxaloacetate decarboxylase Na(+) pump: interaction with other subunits/domains of the complex and binding site for the Zn(2+) metal ion; Schmid M et al.; The oxaloacetate decarboxylase Na(+) pump of Klebsiella pneumoniae is an enzyme complex composed of the peripheral alpha subunit and the two integral membrane-bound subunits beta and gamma . The alpha subunit consists of the N-terminal carboxyltransferase domain and the C-terminal biotin domain, which are connected by a flexible proline/alanine-rich linker peptide . To probe interactions between the two domains of the alpha subunit and between alpha-subunit domains and the gamma subunit, the relevant polypeptides were synthesized in Escherichia coli and subjected to copurification studies . The two alpha-subunit domains had no distinct affinity toward each other and could, therefore, not be purified as a unit on avidin-sepharose . The two domains reacted together catalytically, however, performing the carboxyl transfer from oxaloacetate to protein-bound biotin . This reaction was enhanced up to 6-fold in the presence of the Zn(2+)-containing gamma subunit . On the basis of copurification with different tagged proteins, the C-terminal biotin domain but not the N-terminal carboxyltransferase domain of the alpha subunit formed a strong complex with the gamma subunit . Upon the mutation of gamma H78 to alanine, the binding affinity to subunit alpha was lost, indicating that this amino acid may be essential for formation of the oxaloacetate decarboxylase enzyme complex . The binding residues for the Zn(2+) metal ion were identified by site-directed and deletion mutagenesis . In the gamma D62A or gamma H77A mutant, the Zn(2+) content of the decarboxylase decreased to 35% or 10% of the wild-type enzyme, respectively . Less than 5% of the Zn(2+) present in the wild-type enzyme was found if the two C-terminal gamma-subunit residues H82 and P83 were deleted . Corresponding with the reduced Zn(2+) contents in these mutants, the oxaloacetate decarboxylase activities were diminished . These results indicate that aspartate 62, histidine 77, and histidine 82 of the gamma subunit are ligands for the catalytically important Zn(2+) metal ion.

Dig Surg, 2001, 18(6), 459 - 65; discussion 465-6
Pyogenic liver abscess: an audit of 10 years' experience and analysis of risk factors; Lee KT et al.; BACKGROUND/AIMS: Despite continuous improvement in image modalities, availability of potent antibiotics and advancement in the knowledge and treatment of pyogenic liver abscess, mortality remains high . The high mortality rate has underlined the important role of prognostic factors and prompts a number of studies to identify the risk factors . The present study aims to audit our experience in managing patients with pyogenic hepatic abscess during the period of 1989-1999, and to document changes in etiology, bacteriology and outcome, and to identify any risk factor associated with mortality . METHODS: One hundred and thirty-three patients with pyogenic hepatic abscess were studied to determine the demographic characteristics, clinical features, laboratory, bacteriological findings, methods of treatment, final outcome and risk factor analysis . All patients were treated with parenteral antibiotics . One hundred and twelve patients were subjected to ultrasound-guided percutaneous aspiration of the abscess . A percutaneous drainage catheter was inserted after aspiration in all patients . Laparotomy was done in 21 patients . RESULTS: The overall hospital mortality rate was 6% (8/133) . Biliary tract disease was the most frequently identified cause . Leukocytosis, hypoalbuminemia and hyperbilirubinemia were common laboratory findings . The most common microorganism cultured was Klebsiella pneumoniae . The most common concomitant disease was diabetes mellitus . On univariate analysis, large abscess, diabetes mellitus and sepsis were significantly associated with hospital mortality . On multivariate logistic regression analysis, the presence of sepsis (p = 0.0031) was found to be an independent risk factor . CONCLUSIONS: In addition to early diagnosis and prompt treatment, making every effort to treat patients with adverse prognostic factors and systemic complications, the hospital mortality rate will be decreased significantly .

Antimicrob Agents Chemother, 2002 Feb, 46(2), 500 - 10
Genes encoding TEM-4, SHV-2, and CTX-M-10 extended-spectrum beta-lactamases are carried by multiple Klebsiella pneumoniae clones in a single hospital (Madrid, 1989 to 2000); Coque TM et al.; Over a 12-year period (1989 to 2000), 159 Klebsiella pneumoniae isolates harboring extended-spectrum beta-lactamases (ESBLs) (4.8% of the total number of K . pneumoniae isolates obtained) were recovered from 58 patients, who were mainly hospitalized in intensive care and surgery units . For 62 representative isolates from 58 patients, 31 clonal types harboring TEM-4 (n = 5), SHV-2 (n = 7), SHV2a (n = 4), SHV-5 (n = 1), CTX-M-10 (n = 13), or CTX-M-9 (n = 1) beta-lactamases were identified by pulsed-field gel electrophoresis . This is the first report to document the presence of the CTX-M-10 or the CTX-M-9 beta-lactamase in K . pneumoniae . These beta-lactamases were previously identified in Escherichia coli isolates from Spain . Only two of five K . pneumoniae TEM-4 clones caused more than a single case of infection, with one of them spreading for 9 months . A single plasmid was detected among these TEM-4 clones . Only two of seven K . pneumoniae clones containing SHV-2 and three of four strains harboring SHV-2a were detected in more than one case of infection . Plasmids encoding SHV-2 or SHV-2a were unrelated . Four of 13 K . pneumoniae CTX-M-10 clones were found in more than one patient, with two of them recovered 2 and 5 years apart . As in the case of the SHV-2 isolates, we were unable to document a common transmissible genetic element that could explain the polyclonal structure of our isolates . Nevertheless, the spread of a single gene may be suggested by the presence of a conserved set of noncoding polymorphisms in the sequences . Most ESBL-producing K . pneumoniae clones were ephemeral, being poorly selected and maintained in the hospital setting, but the genes encoding ESBL persisted successfully over the years that the strains were recovered, probably as a minority gene population in the hospital metagenome.

Arch Pharm Res, 2001 Dec, 24(6), 590 - 6
A novel plasmid-mediated beta-lactamase that hydrolyzes broad-spectrum cephalosporins in a clinical isolate of Klebsiella pneumoniae; Kwak JH et al.; A new extended-spectrum beta-lactamase with an isoelectric point (pI) of 6.2 was detected in Klebsiella pneumoniae F161 that was isolated from a patient with infection . This strain was highly resistant to the third or fourth generation cephalosporins such as ceftazidime, ceftriaxone, cefoperazone, and cefpirome . Analysis of this strain by the double disk diffusion test showed synergies between amoxicillin-clavulanate (AMX-CA) and cefotaxime, and AMX-CA and aztreonam, which suggested that this strain produced a extended-spectrum beta-lactamase (ESBL) . Genetic analysis revealed that the resistance was due to the presence of a 9.4-kb plasmid, designated as pKP161, encoding for new beta-lactamase gene (bla) . Sequence analysis showed that a new bla gene of pKP161 differed from bla(TEM-1) by three mutations leading to the following amino acid substitutions: Val84 --> Ile, Ala184 --> Val, and Gly238 --> Ser . These mutations have not been reported previously in the TEM type beta-lactamases produced by clinical strains . The novel beta-lactamase was overexpressed in E . coli and purified by ion exchange chromatography on Q-Sepharose and CM-Sepharose, and then further purified by gel filtration on Sehadex G-200 . The catalytic activity of the purified beta-lactamase was confirmed by the nitrocefin disk.

J Bacteriol, 2002 Feb, 184(3), 812 - 20
Role of GlnK in NifL-mediated regulation of NifA activity in Azotobacter vinelandii; Rudnick P et al.; In several diazotrophic species of Proteobacteria, P(II) signal transduction proteins have been implicated in the regulation of nitrogen fixation in response to NH(4)(+) by several mechanisms . In Azotobacter vinelandii, expression of nifA, encoding the nif-specific activator, is constitutive, and thus, regulation of NifA activity by the flavoprotein NifL appears to be the primary level of nitrogen control . In vitro and genetic evidence suggests that the nitrogen response involves the P(II)-like GlnK protein and GlnD (uridylyltransferase/uridylyl-removing enzyme), which reversibly uridylylates GlnK in response to nitrogen limitation . Here, the roles of GlnK and GlnK-UMP in A . vinelandii were studied to determine whether the Nif (-) phenotype of glnD strains was due to an inability to modify GlnK, an effort previously hampered because glnK is an essential gene in this organism . A glnKY51F mutation, encoding an unuridylylatable form of the protein, was stable only in a strain in which glutamine synthetase activity is not inhibited by NH(4)(+), suggesting that GlnK-UMP is required to signal adenylyltransferase/adenylyl-removing enzyme-mediated deadenylylation . glnKY51F strains were significantly impaired for diazotrophic growth and expression of a nifH-lacZ fusion . NifL interacted with GlnK and GlnKY51F in a yeast two-hybrid system . Together, these data are consistent with those obtained from in vitro experiments (Little et al., EMBO J., 19:6041-6050, 2000) and support a model for regulation of NifA activity in which unmodified GlnK stimulates NifL inhibition and uridylylation of GlnK in response to nitrogen limitation prevents this function . This model is distinct from one proposed for the related bacterium Klebsiella pneumoniae, in which unmodified GlnK relieves NifL inhibition instead of stimulating it.

Mikrobiol Z, 2001 May-Jun, 63(3), 13 - 21
Cellular and lipopolysaccharide fatty acid composition of the type strains of Klebsiella pneumoniae, Klebsiella oxytoca, and Klebsiella nonpathogenic species; Vasyurenko ZP et al.; The cellular and lipopolysaccharide (LPS) fatty acid compositions of the type strains of Klebsiella pneumoniae, K . oxytoca, K . terrigena, K . planticola, and "K . trevisanii" were studied . The cellular fatty acids of klebsiellae were presented by straight-chain saturated and monounsaturated, cyclopropane, and hydroxy fatty acids . Hexadecanoic, methylenehexadecanoic, octadecenoic and hexadecenoic acids prevailed . The K . pneumoniae strain mainly differed from the strains of other species by two and more times lower level of dodecanoic acid in cells . Variations of cyclopropane and unsaturated fatty acid contents in cells were observed . LPS fatty acids profiles of klebsiellae mainly consisted of straight-chain saturated and hydroxy fatty acids with predominance of tetradecanoic and 3-hydroxytetradecanoic acids . LPS fatty acids profiles of K . oxytoca, K . terrigena, K . planticola, and "K . trevisanii" strains were very similar and differed from that of the K . pneumoniae strain by higher levels of dodecanoic acid (approximately 5-6 times) and absence of 2-hydroxytetradecanoic acid . The obtained data indicated more close relatedness of K . oxytoca, K . terrigena, and K . planticola and some their remoteness from K . pneumoniae.

Eur Cytokine Netw, 2001 Oct-Dec, 12(4), 581 - 6
Reduced adrenal response and increased mortality after systemic Klebsiella pneumoniae infection in interleukin-6-deficient mice; van Enckevort FH et al.; During bacterial infections, both the immune system and the hypothalamus-pituitary-adrenal (HPA) axis are activated . The role of IL-6 in the activation of the HPA axis during bacterial sepsis is not fully understood . The aim of the present study was to investigate the role of endogenous IL-6 in a potentially lethal infection with Klebsiella pneumoniae and the concomitant activation of the HPA axis . We examined the mortality of IL-6-/- and IL-6+/+ mice after intravenous (i.v.) infection with K . pneumoniae as well as the bacterial outgrowth in several organs . Subsequently, the influence of endogenous IL-6 on the effect of i.v . administration of K . pneumoniae on the plasma levels of corticosterone and the pro-inflammatory cytokines TNF-alpha and IL-1alpha was investigated in these mice . The present study demonstrates that IL-6-/- mice are more susceptible than IL-6+/+ mice to a systemic Gram-negative infection with K . pneumoniae, leading to increased outgrowth of microorganisms in the organs of the mice . Moreover, this infection is associated with a reduced adrenal response in IL-6-/- mice . We conclude that IL-6-/- mice are more susceptible to Gram-negative bacterial infections, which is mainly due to an impaired recruitment of granulocytes to the site of infection in the absence of IL-6 . Furthermore, the reduced adrenal response may be an explanation for the strong inflammatory response with higher TNF-alpha plasma levels in IL-6-/- mice.

Zhonghua Jie He He Hu Xi Za Zhi, 1999 Dec, 22(12), 739 - 42
{The study on COPD rat model produced by bacterial infection}; Xu H et al.; OBJECTIVE: To observe the role of bacterial infection in pathogenesis of COPD . METHODS: The COPD animal model was developed by intranasal repeated injecting Klebsiella pneumoniae(K) or pneumococcal pneumoniae(P) into rat respiratory tract . Histomorphyological changes were observed, PaO2, PaCO2 and right ventricular systolic pressure (RVSP) were analysed . RESULTS: 1 week after injecting K and 4 week after injecting P, the epithelia of bronchioles showed obvious injury . From the 4th week there was severe chronic inflammatory process of bronchioles in 2 experimental groups including thickened wall, narrowed lumen and developed emphysema . In addition, the walls of arterioles accompanying bronchioles were also thickened obviously . Right ventricular systolic pressure raised in 2 experimental groups (P < 0.01) . From the 16th week, PaO2 dropped and PaCO2 raised in K group . CONCLUSIONS: Repeated injecting intranasally of the proper amount of klebsiella pneumoniae or pneumococcal pneumoniae into rats' lungs can induce rat small airway inflammation and emphysema . Combining with PaO2, PaCO2 and RVSP analysis, we suggest the model established shows main features of COPD.

Zhonghua Bing Li Xue Za Zhi, 2001 Oct, 30(5), 353 - 6
{Study on the rat airway epithelial cell injury induced by bacterial infection and its pathogenesis}; Xiong M et al.; OBJECTIVE: To explore the rat airway, especially small airway epithelium injury induced by Klebsiella pneumoniae infection and its pathogenesis . METHODS: A rat airway inflammation and COPD model was induced by repeated intranasal infection with Klebsiella pneumoniae . The injury changes were dynamically observed under SEM, TEM and LM . Actin, TNF-alpha, Fos, Jun protein and their mRNA expression levels in small airway epithelium were analyzed by immunohistochemistry and in situ hybridization . The changes of lung TNF-alpha content were examined by radioimmunoassay . RESULTS: 1 week after Klebsiella pneumoniae infection the epithelial cilia cohered, fell down, partially fell off and the tight intercellular connection destroyed . 2 and 4 weeks after the infection, the injury of bronchus epithelia worsened with pronounced chronic inflammation in all bronchi . The bronchioles walls became thickened, lumen narrowed and emphysema was noted . Compared with control group, TNF-alpha mRNA expression was higher (P < 0.01) in the 2nd and 4th week while TNF-alpha protein was higher at 4 weeks (P < 0.01) . From 1st week after the infection, Fos protein and mRNA expression began to elevate (P < 0.01) . TNF-alpha content of lung began to increase (P < 0.01) and kept at a higher level until the 8th week . CONCLUSIONS: Repeated intranasal injection of a certain amount of Klebsiella pneumoniae can induce airway inflammation and emphysema . The main features of early airway epithelia injury are damage of cilia and intercellular junctions, the cause of which may be associated with changes in microfilament arrangement of epithelia . High expression of TNF-alpha protein in the airway epithelia and lung parallels with epithelium injury . Fos and Jun proteins may also play important roles in up-regulating the TNF-alpha protein.

Zhonghua Yi Xue Za Zhi (Taipei), 2001 Sep, 64(9), 551 - 4
Klebsiella pneumoniae orbital cellulitis; Lin CT et al.; We present a case of orbital cellulitis due to Klebsiellapneumoniae infection . The computed tomography revealed cellulitis over the right cheek with abscess formation in the right eyelid and ipsilateral chronic obstructive sinusitis . Incision and drainage of orbital abscess and functional endoscopic sinus surgery were performed sequentially . Klebsiella pneumoniae was confirmed via culture from these two lesions . The patient achieved complete remission without sequalae after surgical intervention and systemic antibiotics . We describe a successfully treated case of Klebsiella pneumoniae orbital cellulitis, which has never been reported before.

Biol Pharm Bull, 2001 Dec, 24(12), 1423 - 6
Evaluation of ppk-specified polyphosphate as a mercury remedial tool; Pan-Hou H et al.; To evaluate the utility of polyphosphate kinase gene (ppk)-specified polyphosphate in mercury remediation, a fusion plasmid, pMK27, with ppk from Klebsiella aerogenes and mercury transport genes, merT and merP, from Pseudomonas K-62, was constructed . The transcription and translation of ppk, merT and merP were found to be mercury-inducible . The ppk-specified polyphosphate was identified in cells preinduced by Hg2+, but not in cells without mercury induction, suggesting that the synthesis of polyphosphate is regulated by merR . The hypersensitive phenotype to Hg2+, shown by bacteria with pMRD141, which contains merT and merP, was almost completely restored to its original levels when the ppk was introduced into the plasmid, suggesting that the Hg2+-toxicity was reduced by the polyphosphate, probably via chelation formation . Bacteria with pMK27 accumulated approximately 6-fold more mercury than the bacteria with cloning vector, pUC119 . These results clearly demonstrate that the polyphosphate is capable of retaining mercury in the cells without taxing the cells . Based on the results obtained in the present study, the fusion plasmid pMK27 may serve as a strategy for mercury remediation.

Acta Gastroenterol Latinoam, 2001 Oct, 31(4), 307 - 12
{Spontaneous bacterial peritonitis in hepatic cirrhosis with ascites: incidence, bacteriology and mortality in Uruguay}; Rubinstein P et al.; Spontaneous bacterial peritonitis (SBP) is a frequent and severe complication of cirrhotic patients with ascites . In order to analyze the incidence, bacteriology and in-hospital mortality, we studied 64 consecutive patients with cirrhosis and ascites (47 males, 17 females average age 59 years) hospitalized in a general adults 3rd level hospital (Pasteur hospital, Montevideo, Uruguay), between September 1998 and May 2000 . The diagnostic criteria was more than 250 polymorphonuclear cells/cu.mm . in ascitic fluid and/or a positive culture . We found 17 SBP in 17 patients (10 males 24-81 years) which means an incidence of 26.56% . 15 alcoholic cirrhosis and 2 autoimmune disease . 12% (2/17) were asymptomatic; 8/17 were SBP culture positive (5 E . Coli, 2 St . Pneumoniae, 1 Klebsiella sp.), and 9 were culture negative . The mortality rate associated with SBP was 47% (8/17), greater than the cirrhotic group without SBP (12.7% p < 0.01).

Emerg Infect Dis, 2002 Jan, 8(1), 100 - 2
Primary liver abscess caused by one clone of Klebsiella pneumoniae with two colonial morphotypes and resistotypes; Hsueh PR et al.; Two diabetic patients with primary liver abscess, who initially responded unsatisfactorily to intravenous ceftriaxone or cefoxitin treatment and had abscess drainage, were found to be infected with a single clone of Klebsiella pneumoniae with two different colonial morphotypes and resistotypes . Primary liver abscess caused by second-generation cephalosporin-resistant K . pneumoniae strains may be an emerging problem in Taiwan.

J Bacteriol, 2002 Jan, 184(1), 200 - 6
Domain interactions on the ntr signal transduction pathway: two-hybrid analysis of mutant and truncated derivatives of histidine kinase NtrB; Martinez-Argudo I et al.; We have used the yeast two-hybrid system to analyze protein-protein interactions mediated by domains of regulatory proteins of the ntr signal transduction system, including interactions among NtrB derivatives and their interactions with NtrC and PII from Klebsiella pneumoniae . Interactions took place only between proteins or protein domains belonging to the ntr signal transduction system and not between proteins or domains from noncognate regulators . NtrB and its transmitter domain, but not NtrC, CheA, or the cytoplasmic C terminus of EnvZ, interacted with PII . In addition, interaction of NtrB with NtrC, but not with PII, depended on the histidine phosphotransfer domain . Point mutation A129T, diminishing the NtrC phosphatase activity of NtrB, affected the strength of the signals between NtrC and the transmitter module of NtrB but had no impact on PII signals, suggesting that A129T prevents the conformational change needed by NtrB to function as a phosphatase for NtrC, rather than disturbing binding to PII.

Clin Microbiol Infect, 2001 Nov, 7(11), 626 - 35
Effect of inoculum size on the antibacterial activity of cefpirome and cefepime against Klebsiella pneumoniae strains producing SHV extended-spectrum beta-lactamases; Bedenic B et al.; OBJECTIVE: To determine the effects of varying inoculum size on in vitro susceptibility of SHV extended-spectrum beta-lactamase (ESBL)-producing Klebsiella pneumoniae isolates to cefepime and cefpirome compared to previously established cephalosporins and aztreonam . METHODS: Antibiotic susceptibilities were determined by disk diffusion test, the MIC broth microdilution method, and time-kill studies with two different inocula of 10(5) and 10(7) CFU/mL . The strains were classified into four groups according to the type of beta-lactamase they produce: SHV-2, SHV-5, SHV-12, and ESBL-negative klebsiellae . RESULTS: The antibacterial activities of cefpirome and cefepime were comparable to that of cefotaxime, but were significantly greater than those of ceftazidime and aztreonam . An inoculum effect was detected for all broad-spectrum cephalosporins, but it was more pronounced with cefpirome and cefepime compared to older cephalosporins . The disk diffusion test proved to be not sensitive enough for the detection of an inoculum effect, particularly for cefepime . CONCLUSIONS: The present study found that most SHV-producing klebsiellae have MICs of cefpirome that imply susceptibility at the moderate inoculum size, in spite of the fact that, according to the NCCLS, all ESBL producers should be considered resistant to all cephalosporins, independent of MIC values . With a high inoculum, most of the strains seemed to be resistant to both antibiotics . Furthermore, the bactericidal activities of cefpirome and cefepime against isogenic Escherichia coli strains producing SHV-2, SHV-4 and SHV-5 beta-lactamases, respectively, were also inoculum dependent . Bactericidal activity against SHV-4 and SHV-5 beta-lactamase producers was obtained only at the moderate inoculum, whereas the SHV-2 beta-lactamase producer was efficiently killed with both antibiotics, regardless of the inoculum size.

Curr Microbiol, 2002 Jan, 44(1), 31 - 7
4-Hydroxybenzoate uptake in Klebsiella planticola strain DSZ1 is driven by DeltapH; Allende JL et al.; Klebsiella planticola strain DSZ1 has the ability to degrade different aromatic compounds such as benzoate and organochlorinated as propachlor and alachlor . DSZ1 strain cells mineralised 4-hydroxybenzoate (4HBA) through a meta-cleavage pathway, yielding protocatechuate as dihydroxylated intermediate, with a specific rate of CO2 formation 0.12 x 10-6 (cpm/OD) h-1, and a rate of 4-HBA utilisation of 0.75 mmol h-1 . Aerobically the 4HBA transport system is driven by gradient of protons (DeltapH), but is not ATP-driven . Under anaerobic conditions, the system can use the nitrate reduction as a final electron acceptor in respiration . A kinetic analysis of the 4HBA transport system revealed a Kt value of 16 microM with a Vmax value of 25 nmol/min.mg at pH 7.

J Clin Microbiol, 2001 Dec, 39(12), 4433 - 9
Outbreak of infection with multidrug-resistant Klebsiella pneumoniae carrying bla(IMP-8) in a university medical center in Taiwan; Yan JJ et al.; Klebsiella pneumoniae strains with the transferable carbapenem-hydrolyzing metallo-beta-lactamases, which include IMP- and VIM-type enzymes, remain extremely rare . To investigate whether IMP- or VIM-producing K . pneumoniae isolates had spread at a university medical center in Taiwan, a total of 3,458 clinical isolates of K . pneumoniae consecutively collected in 1999 and 2000 were tested by the agar diffusion method, colony hybridization, PCR, and nucleotide sequencing . A total of 40 isolates (1.2%), or 17 nonrepetitive isolates, from 16 patients were found to carry bla(IMP-8), a metallo-beta-lactamase gene recently identified from a K . pneumoniae strain in Taiwan . Carriage of bla(VIM) or other bla(IMP) genes was detected in none of the remaining isolates . Of the 17 nonrepetitive bla(IMP-8)-positive isolates, 15 isolates (88.2%) appeared susceptible to imipenem (MICs, <or=4 microg/ml) and meropenem (MICs, <or=1 microg/ml), indicating the difficulty in detecting bla(IMP-8) in K . pneumoniae by routine susceptibility tests; 14 isolates (82.4%) produced SHV-12 as well; and 14 isolates (82.4%) were also resistant to fluoroquinolones . The organisms caused wound infections in eight patients and bloodstream infections in three patients . They were not directly associated with the death of nine patients . Before the recovery of the bla(IMP-8)-positive isolates, all 16 patients had undergone various surgical procedures, and 15 patients had been admitted to the surgical intensive care unit, suggesting a nosocomial outbreak . Two major patterns were observed by pulsed-field gel electrophoresis for 14 of the 17 nonrepetitive isolates, indicating that the clonal spread was mainly responsible for the outbreak.

Z Naturforsch {C}, 2001 Sep-Oct, 56(9-10), 787 - 91
Ammonium phosphate as a sole nutritional supplement for the fermentative production of 2,3-butanediol from sugar cane juice; Berbert-Molina MA et al.; The production of 2,3-butanediol by Klebsiella pneumoniae from sugar cane juice supplemented with different salts was studied . This microorganism is able to degrade sucrose present in sugar cane juice containing ammonium phosphate as the sole nutritional supplement . With a sugar cane juice-based medium containing approximately 180 g sucrose/l and 8.0 g (NH4)2HPO4/l, over 70 g 2,3-butanediol plus acetoin/l were formed . This result is comparable to that achieved with a sugar cane juice-based medium containing several nutrients, although the kinetic profiles of these runs presented significant differences . With the ammonium phosphate-enriched medium, cell growth was initially favoured by both the strong oxygen supply and the higher water activity due to the lower concentration of nutrients . After 14 h, the limitation in some nutrients led to the interruption of cell growth, and decreasing rates for product formation and substrate consumption were observed . During the stationary phase of this run, sucrose was preferentially converted to product, and the substrate was completely depleted after 35 h of the process . With the complete medium, the substrate was totally consumed after 36 h of run . In this case, the higher initial concentration of nutrients reduced the overall process rate but sustained the cell growth for 27 h . Conversion yields of 0.40 g product/g sucrose and productivities close to 2.0 g/l x h were obtained under both conditions.

Antimicrob Agents Chemother, 2001 Dec, 45(12), 3462 - 7
Effect of endotoxin on P-glycoprotein-mediated biliary and renal excretion of rhodamine-123 in rats; Ando H et al.; The effects of Klebsiella pneumoniae endotoxin on the biliary excretion and renal handling of rhodamine-123 were investigated in rats at different times after intraperitoneal injection (1 mg/kg of body weight) . The typical substrates for P glycoprotein, i.e., cyclosporine, colchicine, and erythromycin, inhibited the biliary clearance of rhodamine-123, whereas a substrate for organic cation transporter, cimetidine, did not inhibit clearance, suggesting that rhodamine-123 is transported mainly by P glycoprotein . The biliary, renal, and tubular secretory clearances of rhodamine-123 and the glomerular filtration rate significantly decreased 6 h after injection of endotoxin but returned to control levels by 24 h . These results suggest that endotoxin-induced decreases in P-glycoprotein-mediated biliary excretion and renal handling of rhodamine-123 were probably due to impairment of P-glycoprotein-mediated transport ability . Pretreatment with pentoxifylline (50 mg/kg) significantly inhibited endotoxin-induced increases in tumor necrosis factor alpha (TNF-alpha) levels in plasma, which ameliorated the endotoxin-induced reduction of the biliary excretion of rhodamine-123 . It is likely that endotoxin-induced impairment of the transport of rhodamine-123 is caused, in part, by overproduction of TNF-alpha . The effect of endotoxin on the expression of P-glycoprotein mRNA in liver and kidneys of rats was investigated by using a reverse transcriptase PCR . The expression of Mdr1a mRNA in both liver and kidney decreased 6 h after endotoxin injection and returned to control levels after 24 h, whereas the expression of Mdr1b mRNA in liver increased at both times and that in kidney decreased at 24 h . These findings suggest that K . pneumoniae endotoxin dramatically decreases P-glycoprotein-mediated biliary and renal excretion of rhodamine-123 probably by decreasing the expression of Mdr1a, which is likely due to increased plasma TNF-alpha levels.

Antimicrob Agents Chemother, 2001 Dec, 45(12), 3355 - 61
CTX-M-type extended-spectrum beta-lactamase that hydrolyzes ceftazidime through a single amino acid substitution in the omega loop; Poirel L et al.; Escherichia coli ILT-1, Klebsiella pneumoniae ILT-2, and K . pneumoniae ILT-3 were isolated in May 1999 in Paris, France, from a rectal swab of a hospitalized 5-month-old girl . These isolates had a clavulanic acid-inhibited substrate profile that included expanded-spectrum cephalosporins . The MICs of cefotaxime were higher for E . coli ILT-1 and K . pneumoniae ILT-2 than for K . pneumoniae ILT-3, while the opposite was found for the MICs of ceftazidime . Genetic and biochemical analyses revealed that E . coli ILT-1 and K . pneumoniae ILT-2 produced the CTX-M-18 beta-lactamase, while K . pneumoniae ILT-3 produced the CTX-M-19 beta-lactamase . The amino acid sequence of the CTX-M-18 beta-lactamase differed from that of the CTX-M-9 beta-lactamase by an Ala-to-Val change at position 231, while CTX-M-19 possessed an additional Pro-to-Ser change at position 167 in the omega loop of Ambler class A enzymes . The latter amino acid substitution may explain the CTX-M-19-mediated hydrolysis of ceftazidime, which has not been reported for other CTX-M-type enzymes . The bla(CTX-M-18) and bla(CTX-M-19) genes were located on transferable plasmids that varied in size (ca . 60 and 50 kb, respectively) but that showed similar restriction patterns.

Carbohydr Res, 2001 Nov 21, 336(3), 229 - 35
Synthesis of an L-rhamnose tetrasaccharide, the common and major structure of the repeating unit of the O-antigenic polysaccharide of a strain of Klebsiella pneumoniae and Pseudomonas holci; Zhang J et al.; A tetrasaccharide, alpha-L-Rhap-(1-->3)-alpha-L-Rhap-(1-->2)-alpha-L-Rhap-(1-->2)-L-Rhap, the common and major structure of the repeating unit of the O-antigenic polysaccharide of a strain of Klebsiella pneumoniae and Pseudomonas holci was synthesized as its methyl and octyl glycosides . Selective 3-O-glycosylation of allyl alpha-L-rhamnopyranoside with 2,3,4-tri-O-acetyl-alpha-L-rhamnopyranosyl trichloroacetimidate gave allyl 2,3,4-tri-O-acetyl-alpha-L-rhamnopyranosyl-(1-->3)-alpha-L-rhamnopyranoside (3) . Benzoylation, deallylation, and trichloroacetimidation afforded 2,3,4-tri-O-acetyl-alpha-L-rhamnopyranosyl-(1-->3)-2,4-di-O-benzoyl-alpha-L-rhamnopyranosyl trichloroacetimidate (6) . Self condensation of 3,4-di-O-benzoyl-beta-L-rhamnopyranose 1,2-methyl orthoester or 1,2-octyl orthoester gave methyl or octyl 2-O-acetyl-3,4-di-O-benzoyl-alpha-L-rhamnopyranosyl-(1-->2)-3,4-di-O-benzoyl-alpha-L-rhamnopyranoside (16 or 17), and subsequent selective deacetylation gave the disaccharide acceptor (18 or 19) . Coupling of 6 with 18 (or 19), followed by deacylation in ammonia-saturated methanol, produced the target tetrasacharide.

J Med Microbiol, 2001 Nov, 50(11), 959 - 64
Protection against pulmonary infection with Klebsiella pneumoniae in mice by interferon-gamma through activation of phagocytic cells and stimulation of production of other cytokines; Yoshida K et al.; The study was designed to determine the role of interferon (IFN)-gamma in inflammatory responses against experimentally induced pneumonia caused by Klebsiella pneumoniae . The host immunological responses in IFN-gamma gene knockout (IFN-gamma(-/-)) mice and immunocompetent control mice were compared . K . pneumoniae strain T-113 was inoculated intranasally into anaesthetised mice to induce pneumonia . Infected control mice survived significantly longer than infected IFN-gamma(-/-) mice . Viable bacterial counts in lungs and blood abruptly increased in IFN-gamma(-/-) mice; in contrast, a gradual decrease in the number of bacteria was noted in control mice . During the early stages of infection, the concentrations of interleukin (IL)-1beta and IL-6 in broncho-alveolar lavage fluid and IL-1beta in serum of IFN-gamma(-/-) mice were significantly lower than in control mice . During the late stage of infection, serum IL-6 level in IFN-gamma(-/-) mice was significantly higher than in control mice . These results suggest that the defective immunological host response, including inflammatory cytokine production caused by deficiency of IFN-gamma, is one of the mechanisms that allow the progression of pulmonary infection to systemic septicaemia.

Curr Microbiol, 2001 Nov, 43(5), 322 - 7
Volatilization of mercury by immobilized bacteria (Klebsiella pneumoniae) in different support by using fluidized bed bioreactor; Zeroual Y et al.; Klebsiella pneumoniae, a mercury-resistant bacterial strain able to reduce ionic mercury to metallic mercury, was isolated from wastewater of Casablanca . This strain exhibits high minimal inhibition concentrations for heavy metals such as mercury 2400 microM, lead 8000 microM, silver 2400 microM, and cadmium 1000 microM . This bacterium was immobilized in alginate, polyacrylamide, vermiculite, and cooper beech and was used for removing mercury from a synthetic water polluted by mercury by using a fluidized bead bioreactor . Immobilized bacterial cells of Klebsiella pneumoniae could effectively volatilize mercury and detoxify mercury compounds . Moreover, the efficiency of mercury volatilization was much greater than with the native cells . The highest cleanup and volatilization rates were obtained when Klebsiella pneumoniae was entrapped in alginate beads, with a cleanup rate of 100% and a volatilization rate of 89% . Immobilized cells in alginate continuously volatilized mercury even after 10 days without loss of activity.

Mol Microbiol, 2001 Oct, 42(1), 229 - 43
Structure, organization and characterization of the gene cluster involved in the production of microcin E492, a channel-forming bacteriocin; Lagos R et al.; Microcin E492 is a low-molecular-weight, channel-forming bacteriocin produced and excreted by Klebsiella pneumoniae RYC492 . A 13 kb chromosomal DNA fragment from K . pneumoniae RYC492 was sequenced, and it was demonstrated by random Tn5 mutagenesis that most of this segment, which has at least 10 cistrons, is needed for the production of active microcin and its immunity protein . Genes mceG and mceH correspond to an ABC exporter and its accessory protein, respectively, and they are closely related to the colicin V ABC export system . The microcin E492 system also requires the product of gene mceF as an additional factor for export . Despite the fact that this bacteriocin lacks post-translational modifications, genes mceC, mceI and mceJ are needed for the production of active microcin . Genes mceC and mceI are homologous to a glycosyl transferase and acyltransferase, respectively, whereas mceJ has no known homologue . Mutants in these three genes secrete an inactive form of microcin, able to form ion channels in a phospholipidic bilayer, indicating that the mutation of these microcin genes does not alter the process of membrane insertion . On the other hand, microcin isolated from mutants in genes mceC and mceJ has a lethal effect when incubated with spheroplasts of sensitive cells, indicating that the microcin defects in these mutants are likely to alter receptor recognition at the outer membrane . A model for synthesis and export is proposed as well as a novel maturation pathway that would involve conformational changes to explain the production of active microcin E492.

J Bacteriol, 2001 Nov, 183(22), 6607 - 19
Roles of glutamate synthase, gltBD, and gltF in nitrogen metabolism of Escherichia coli and Klebsiella aerogenes; Goss TJ et al.; Mutants of Escherichia coli and Klebsiella aerogenes that are deficient in glutamate synthase (glutamate-oxoglutarate amidotransferase {GOGAT}) activity have difficulty growing with nitrogen sources other than ammonia . Two models have been proposed to account for this inability to grow . One model postulated an imbalance between glutamine synthesis and glutamine degradation that led to a repression of the Ntr system and the subsequent failure to activate transcription of genes required for the use of alternative nitrogen sources . The other model postulated that mutations in gltB or gltD (which encode the subunits of GOGAT) were polar on a downstream gene, gltF, which is necessary for proper activation of gene expression by the Ntr system . The data reported here show that the gltF model is incorrect for three reasons: first, a nonpolar gltB and a polar gltD mutation of K . aerogenes both show the same phenotype; second, K . aerogenes and several other enteric bacteria lack a gene homologous to gltF; and third, mutants of E . coli whose gltF gene has been deleted show no defect in nitrogen metabolism . The argument that accumulated glutamine represses the Ntr system in gltB or gltD mutants is also incorrect, because these mutants can derepress the Ntr system normally so long as sufficient glutamate is supplied . Thus, we conclude that gltB or gltD mutants grow slowly on many poor nitrogen sources because they are starved for glutamate . Much of the glutamate formed by catabolism of alternative nitrogen sources is converted to glutamine, which cannot be efficiently converted to glutamate in the absence of GOGAT activity . Finally, GOGAT-deficient E . coli cells growing with glutamine as the sole nitrogen source increase their synthesis of the other glutamate-forming enzyme, glutamate dehydrogenase, severalfold, but this is still insufficient to allow rapid growth under these conditions.

J Org Chem, 1998 Oct 2, 63(20), 6797 - 6801
Stereoselective Synthesis of Tilivalline(1); Nagasaka T et al.; Tilivalline 1, a metabolite from Klebsiella pneumoniae var . ocytoca, was easily synthesized in five steps from (S)-proline and 3-(benzyloxy)isatoic anhydride 4g . This synthesis is based on modified Curtius reactions of 3-substituted phthalic anhydrides 11 to 3-substituted isatoic anhydrides 4, conversion of lactams 6 to the acyliminium precursors 7 and stereoselective introduction of indole from the less hindered side of 7.

Inorg Chem, 1996 Jun 19, 35(13), 3792 - 3803
Dinuclear Nickel(II) Complexes as Models for the Active Site of Urease; Volkmer D et al.; Dinuclear nickel(II) complexes of the ligands 2,6-bis{bis((2-benzimidazolylmethyl)amino)methyl}-p-cresol (bbapOH), N,N,N',N'-tetrakis(2-benzimidazolylmethyl)-2-hydroxy-1,3-diaminopropane (tbpOH), N-methyl-N,N',N'-tris(2-benzimidazolylmethyl)-2-hydroxy-1,3-diaminopropane (m-tbpOH) and 1-{N,N-bis(2-benzimidazolylmethyl)amino}-3-{2-(3,5-dimethyl-1H-pyrazol-1-yl)ethoxy}-2-hydroxypropane (bpepOH) were prepared in order to model the active site of urease . The novel asymmetric structures of the dinuclear complexes were characterized by X-ray structure analysis . The complex {Ni(2)(bbapO)(ClO(4))(H(2)O)(MeOH)}(ClO(4))(2).Et(2)O, 1, crystallizes in the monoclinic space group P2(1)/c, with a = 10.258(2) A, b = 19.876(3) A, c = 25.592(4) A, and beta = 97.12(2) degrees . The nickel ions in 1 are bridged by the phenoxy donor of the ligand and a perchlorate anion . The complexes {Ni(2)(tbpO)(MeCOO)(H(2)O)}(ClO(4))(2).H(2)O.Et(2)O, 2, {Ni(2)(m-tbpO)(PhCOO)(EtOH)(2)}(ClO(4))(2).EtOH, 3, and {Ni(2)(bpepO)(MeCOO)(H(2)O)(2)}(ClO(4))(2).H(2)O.Et(2)O.2EtOH, 4, also crystallize in the monoclinic crystal system with the following unit cell parameters: 2, C2/c, a = 35.360(13) A, b = 10.958(3) A, c = 24.821(10) A, beta = 103.55(3) degrees; 3, Cc, a = 14.663(5) A, b = 32.630(13) A, c = 9.839(3) A, beta = 92.49(2) degrees; 4, C2/c, a = 27.689(13) A, b = 12.187(5) A, c = 31.513(14) A, beta = 115.01(3) degrees . The dinuclear centers of all these complexes are bridged by the alkoxy donor of the ligand and a carboxylate function . Compounds 2 and 3 have one of the nickel ions in a five-coordinated, trigonal bipyramidal coordination environment and thus show a high structural similarity to the dinuclear active site of urease from Klebsiella aerogenes . Furthermore, their magnetic and spectroscopic properties were determined and related to those of the urease enzymes . Activity toward hydrolysis of test substrates (4-nitrophenyl)urea, 4-nitroacetanilide, 4-nitrophenyl phosphate or bis(4-nitrophenyl) phosphate by the dinuclear complexes were examined by UV spectroscopic measurements.

Antimicrob Agents Chemother, 2001 Nov, 45(11), 3189 - 94
Cloning and biochemical characterization of FOX-5, an AmpC-type plasmid-encoded beta-lactamase from a New York City Klebsiella pneumoniae clinical isolate; Queenan AM et al.; Klebsiella pneumoniae 5064, isolated in New York, carried plasmid-mediated resistance to multiple beta-lactams and was unresponsive to clavulanic acid . The beta-lactamase gene responsible for cephalosporin resistance encoded FOX-5, with 96 to 97% amino acid identities to other members of the FOX family of beta-lactamases . The bla(FOX-5) coding region was located next to a transposase gene from the Aeromonas salmonicida insertion element ISAS2.

Gastroenterol Clin Biol, 2001 Aug-Sep, 25(8-9), 814 - 6
{Diclofenac-induced colitis complicated by Klebsiella oxytoca infection}; Soussi F et al.; A 25-year-old man presented with abdominal pain and bloody diarrhea . Colonoscopy showed hemorrhagic proctocolitis with superficial erosions . Histology was consistent with the diagnosis of ulcerative colitis and biopsy cultures were negative . Despite treatment with prednisolone (40 mg/day), his clinical condition deteriorated and he was referred to our institution . On repeated questioning, the patient reported self-medication with diclofenac (200 mg/day) for 6 weeks to treat tendinitis prior to the beginning of digestive symptoms . Rectosigmoidoscopy confirmed bleeding colitis and repeated biopsy cultures showed Klebsiella oxytoca . Corticosteroids were stopped and ofloxacin (400 mg/day) was prescribed for 14 days . Diarrhea quickly resolved . Colonoscopy 8 weeks later showed only patchy erythematous mucosa without bleeding or erosions . Two years later, the patient remains asymptomatic with normal total colonoscopy . The definitive diagnosis was de novo NSAID-induced colitis subsequently complicated by Klebsiella oxytoca infection.

Infect Immun, 2001 Nov, 69(11), 7140 - 5
Identification of genes induced in vivo during Klebsiella pneumoniae CG43 infection; Lai YC et al.; A novel in vivo expression technology (IVET) was performed to identify Klebsiella pneumoniae CG43 genes that are specifically expressed during infection of BALB/c mice . The IVET employed a UDP glucose pyrophosphorylase (galU)-deficient mutant of K . pneumoniae which is incapable of utilizing galactose and synthesizing capsular polysaccharide, as demonstrated by its low virulence to BALB/c mice and a white nonmucoid colony morphology on MacConkey-galactose agar . By using a functional galU gene as the reporter, an IVE promoter could render the galU mutant virulent while maintaining the white nonmucoid colony phenotype . A total of 20 distinct sequences were obtained through the in vivo selection . Five of them have been identified previously as virulence-associated genes in other pathogens, while another five with characterized functions are involved in regulation and transportation of nutrient uptake, biosynthesis of isoprenoids, and protein folding . No known functions have been attributed to the other 10 sequences . We have also demonstrated that 2 of the 20 IVE genes turn on under iron deprivation, whereas the expression of another five genes was found to be activated in the presence of paraquat, a superoxide generator.

Carbohydr Res, 2001 Oct 15, 335(4), 291 - 6
Structural analysis of the core region of the lipopolysaccharides from eight serotypes of Klebsiella pneumoniae; Vinogradov E et al.; The core regions of the lipopolysaccharides (LPS) from Klebsiella pneumoniae serotypes O1, O2a, O2a,c, O3, O4, O5, O8, and O12 were analysed using NMR spectroscopy, ESI-MS spectroscopy, and chemical methods . All the LPSs had similar core structures, as shown below, differing only in the number and position of beta-D-galacturonic acid substituents: {carbohydrate structure: see text} where P is H or alpha-Hep, J, K is H or beta-GalA . LPS from all serotypes contained varying proportions of structures having additional or missing phosphate substituents . The core from serotype O1 contained a minor amount of a previously described variant with alpha-DD-Hep-(1-->2)-alpha-DD-Hep-(1-->6)-alpha-GlcN-(1--> replacing the alpha-Hep-(1-->4)-alpha-Kdo-(2-->6)-alpha-GlcN-(1--> component.

J Biol Chem, 2001 Dec 28, 276(52), 49359 - 64 Epub 2001 Oct 08.
Crystal structure of Klebsiella aerogenes UreE, a nickel-binding metallochaperone for urease activation; Song HK et al.; UreE is proposed to be a metallochaperone that delivers nickel ions to urease during activation of this bacterial virulence factor . Wild-type Klebsiella aerogenes UreE binds approximately six nickel ions per homodimer, whereas H144*UreE (a functional C-terminal truncated variant) was previously reported to bind two . We determined the structure of H144*UreE by multi-wavelength anomalous diffraction and refined it to 1.5 A resolution . The present structure reveals an Hsp40-like peptide-binding domain, an Atx1-like metal-binding domain, and a flexible C terminus . Three metal-binding sites per dimer, defined by structural analysis of Cu-H144*UreE, are on the opposite face of the Atx1-like domain than observed in the copper metallochaperone . One metal bridges the two subunits via the pair of His-96 residues, whereas the other two sites involve metal coordination by His-110 and His-112 within each subunit . In contrast to the copper metallochaperone mechanism involving thiol ligand exchanges between structurally similar chaperones and target proteins, we propose that the Hsp40-like module interacts with urease apoprotein and/or other urease accessory proteins, while the Atx1-like domain delivers histidyl-bound nickel to the urease active site.

Hong Kong Med J, 2001 Sep, 7(3), 303 - 6
Septic metastatic endophthalmitis complicating Klebsiella pneumoniae liver abscess in a non-diabetic Chinese man; Wong TY et al.; Septic metastatic endophthalmitis is a rare but serious disease . Endophthalmitis arising from Klebsiella pneumoniae liver abscess has been reported with diabetes mellitus as a major associated condition, but is rarely seen in patients without diabetes . A non-diabetic patient with liver abscess complained of right eye discomfort and floaters 3 days after admission . Both blood and liver aspirate cultured Klebsiella pneumoniae . The patient was treated initially with systemic and subconjunctival antibiotics followed by intravitreal antibiotics with successful visual salvation . Previous reports from the literature showed poor visual outcome despite treatment and delayed recognition was often the cause . Clinicians should be alert to endophthalmitis whenever a patient with Klebsiella pneumoniae liver abscess complains of ocular symptoms . Urgent ophthalmological assessment should be sought.

Am J Respir Cell Mol Biol, 2001 Sep, 25(3), 335 - 40
Interleukin-17 and lung host defense against Klebsiella pneumoniae infection; Ye P et al.; Bacterial pneumonia remains an important cause of morbidity and mortality worldwide, especially in immune-compromised patients . Cytokines and chemokines are critical molecules expressed in response to invading pathogens and are necessary for normal lung bacterial host defenses . Here we show that interleukin (IL)-17, a novel cytokine produced largely by CD4+ T cells, is produced in a compartmentalized fashion in the lung after challenge with Klebsiella pneumoniae . Moreover, overexpression of IL-17 in the pulmonary compartment using a recombinant adenovirus encoding murine IL-17 (AdIL-17) resulted in the local induction of tumor necrosis factor-alpha, IL-1beta, macrophage inflammatory protein-2, and granulocyte colony-stimulating factor (G-CSF); augmented polymorphonuclear leukocyte recruitment; and enhanced bacterial clearance and survival after challenge with K . pneumoniae . However, simultaneous treatment with AdIL-17 provided no survival benefit after intranasal K . pneumoniae challenge . These data show that IL-17 may have a role in priming for enhanced chemokine and G-CSF production in the context of lung infection and that optimally timed gene therapy with IL-17 may augment host defense against bacterial pneumonia.

J Antimicrob Chemother, 2001 Oct, 48(4), 493 - 500
A procedure for evaluation and documentation of susceptibility test methods using the susceptibility of Klebsiella pneumoniae to ciprofloxacin as a model; Schumacher H et al.; A new procedure for the evaluation and documentation of susceptibility test methods is described . To illustrate the procedure, four basically different susceptibility test methods were examined in a routine laboratory . The test parameter detection of decreased susceptibility to ciprofloxacin (breakpoint MIC 0.25 mg/L) among 94 selected isolates of Klebsiella pneumoniae was used . In addition to comparison of frequency histograms and regression analysis, the accuracies of the susceptibility test methods were determined using the receiver operating characteristic procedure . For each of the methods, the sensitivity (SN), specificity (SP), positive predictive value (PV+) and negative predictive value (PV-) for detection of decreased susceptibility to ciprofloxacin were calculated and plotted against a range of ciprofloxacin inhibition zones determined by the various susceptibility test methods or MICs determined by the Etest (Etest MICs) . The results illustrate the accuracy and the robustness of the methods, which can be used to expose the need for training and instruction of laboratory staff . It becomes possible to optimize and justify the choice of inhibition zone breakpoints or Etest MIC breakpoints according to the SN and SP of the method employed . Furthermore, the consequences of adjustments of these breakpoints on the PV+ and PV- can be analysed and related to different clinical and epidemiological situations . We believe that our approach can be used as a model for the evaluation and documentation of susceptibility test methods in general.

J Clin Microbiol, 2001 Oct, 39(10), 3696 - 9
Evaluation of efficiency of screening extended-spectrum beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae in hospitals where the bacteria are increasingly prevalent; Lee K et al.; The disk screening methods for extended-spectrum beta-lactamase-producing strains were evaluated . The confirmatory work is reduced significantly in settings such as those in this study, by changing the cefpodoxime breakpoint to < or =20 mm and by not testing cefoxitin-resistant isolates . Cefotaxime and ceftazidime disk screening is reliable, and the laboratory-prepared cefotaxime- and ceftazidime-clavulanic acid disks are stable at -20 degrees C for 12 weeks.

Clin Infect Dis, 2001 Oct 15, 33(8), 1288 - 94 Epub 2001 Sep 14.
Epidemiological investigation of fluoroquinolone resistance in infections due to extended-spectrum beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae; Lautenbach E et al.; The incidence of infections due to extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli and Klebsiella pneumoniae (ESBL-EK) has increased markedly in recent years . Treatment is difficult because of frequent multidrug resistance . Although fluoroquinolones offer effective therapy for ESBL-EK infections, their usefulness is threatened by increasing fluoroquinolone resistance . To identify risk factors for fluoroquinolone resistance in ESBL-EK infections, a case-control study of all patients with ESBL-EK infections from 1 June 1997 through 30 September 1998 was conducted . Of 77 ESBL-EK infections, 43 (55.8%) were resistant to fluoroquinolones . Independent risk factors for fluoroquinolone resistance were fluoroquinolone use (odds ratio {OR}, 11.20; 95% confidence interval {CI}, 1.99-63.19), aminoglycoside use (OR, 5.83; 95% CI, 1.12-30.43), and long-term care facility residence (OR, 3.39; 95% CI, 1.06-10.83) . The genotypes of fluoroquinolone-resistant ESBL-EK isolates were closely related . Efforts should be directed at modification of these risk factors to preserve the utility of fluoroquinolones in the treatment of ESBL-EK infections.

Aliment Pharmacol Ther, 2001 Oct, 15(10), 1647 - 53
Immune sensitization to food, yeast and bacteria in Crohn's disease; van den Bogaerde J et al.; BACKGROUND: Complex food proteins and enteric flora may act as antigenic stimuli in Crohn's disease . This study assessed the prevalence and magnitude of lymphocyte priming to these antigens in Crohn's disease . METHODS: A total of 31 Crohn's disease patients (median age 42 years, range 25-72 years) and 22 healthy controls (median 29 years, 23-43 years) were studied . Peripheral blood lymphocytes were collected and incubated with antigens in hanging drop culture for 4 days . The antigens tested were cow's milk, cereals, cabbage group, citrus group, peanut group, Saccharomyces (yeast), Bacteroides, E . coli and Klebsiella . On the 4th day 3H-thymidine incorporation was measured after a 4-h pulse . Responses to antigens were considered positive if mean proliferative values were above the 99% confidence interval for background proliferation . RESULTS: The mean background and mitogen-stimulated proliferation did not differ between patients and controls . The mean proliferation to antigens was not above background in controls, but in Crohn's patients proliferative responses to all food and bacterial antigens were significantly higher than background values . Twenty-three out of 31 Crohn's patients and five out of 22 controls (P=0.0003) responded to one or more antigens . Sixteen Crohn's patients and two controls responded to four or more antigens (P=0.001, Fisher's exact test) . CONCLUSION: The reactivity of peripheral lymphocytes to food, yeast and bacterial antigens, especially multiple antigens, is common in Crohn's disease . These sensitized lymphocytes may contribute to the inflammatory process.

Eur J Clin Microbiol Infect Dis, 2001 Jul, 20(7), 505 - 8
Molecular epidemiology of extended-spectrum beta-lactamases from Klebsiella pneumoniae strains isolated in Zagreb, Croatia; Bedenic B et al.; In order to assess the molecular epidemiology of 40 previously identified extended-spectrum beta-lactamase (ESBL)-producing clinical isolates of Klebsiella pneumoniae, gene sequencing was performed . While the previous examination of these isolates revealed one TEM producer, the sequencing procedure performed in this study identified 13 additional TEM producers, and all of the sequenced genes reflected production of nonESBL TEM-1 . All 38 suspected SHV producers were confirmed to be carriers of blaSHV-ESBL genes using the PCR/Nhel test and sequencing . Among them, types SHV-2, SHV-5, and SHV-12 were found in 20, 10, and 7 isolates, respectively, and SHV-2a was identified in 1 . SHV-5 and SHV-12 conferred higher resistance to ceftazidime and cefepime, while SHV-2 and SHV-2a raised the minimum inhibitory concentrations of cefotaxime and cefpirome . Fourth-generation cephalosporins were found to be more active against the isolates than third-generation cephalosporins.

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