Coliforms from hides and meat.

Coliform tests were performed on 85 hide and 75 meat samples. IMViC reactions were determined on isolates from positive confirmed and fecal tests, and strains other than Escherichia coli were identified. Strains typed as Aerobacter aerogenes types I and II were identified as Enterobacter cloacae (51.4%), Klebsiella pneumoniae (21.5%), Enterobacter aerogenes (15%), and Enterobacter liquefaciens, Serratia, and unidentified coliforms (12.1%). K. pneumoniae appeared to be responsible for less than 1% positive fecal tests.     

Coliforms from hides and meat.

Coliform tests were performed on 85 hide and 75 meat samples. IMViC reactions were determined on isolates from positive confirmed and fecal tests, and strains other than Escherichia coli were identified. Strains typed as Aerobacter aerogenes types I and II were identified as Enterobacter cloacae (51.4%), Klebsiella pneumoniae (21.5%), Enterobacter aerogenes (15%), and Enterobacter liquefaciens, Serratia, and unidentified coliforms (12.1%). K. pneumoniae appeared to be responsible for less than 1% positive fecal tests.     

A specific property of bacteria in the genus Klebsiella--a color reaction with 5-aminosalicylic acid in the nutrient medium

For the first time bacteria of the genus Klebsiella have been found to possess a specific property, characteristic of this genus only, i.e. the capacity of giving color reaction with 5-aminosalicylic acid. This reaction can be observed in all Klebsiella species under study: K. pneumoniae (94.4 +/- 2.3%), K. ozaenae (93.3 +/- 4.5%), K. rhinoscleromatis (100%), K. oxytoca (88.0 +/- 4.9%), K. mobilis (92-5 +/- 4.3% of the strains). In all other bacteria under study (40 species, 22 genera and 7 families) the reaction is negative. The test for the color reaction with 5-aminosalicylic acid confirms the belonging of K. mobilis (Enterobacter aerogenes) to the genus Klebsiella, thus making it possible to simplify and accelerate the identification of Klebsiella.     

Phylogenetical relationship based on groE genes among phenotypically related Enterobacter,Pantoea, Klebsiella,Serratia and Erwinia species

In an attempt to define the phylogenetical relationship among 17 phenotypically related species of genera Enterobacter, Pantoea, Serratia, Klebsiella and Erwinia, we determined almost all of their groE operon sequences using the polymerase chain reaction direct sequencing method. The number of nucleotide substitutions per site was 0.12+/-0.030. The value was 3.6-fold higher than that of 16S rDNA. As a result, we were successful in constructing molecular phylogenetic trees which had a finer resolution than that based on the 16S rDNA sequences. The phylogenetic trees based on the nucleotide sequences and deduced amino acid sequences of groE operons indicated that the members of genera Enterobacter, Pantoea and Klebsiella were closely related to each other, while Serratia and Erwinia species except Erwinia carotovora, made distinct clades. The close relationship between Enterobacter aerogenes and Klebsiella pneumoniae, that had been suggested by biochemical tests and DNA hybridization, was also supported by our molecular phylogenetic trees.     

Homology among bacterial catalase genes

Catalase activities in crude extracts of exponential and stationary phase cultures of various bacteria were visualized following gel electrophoresis for comparison with the enzymes from Escherichia coli. Citrobacter freundii, Edwardsiella tarda, Enterobacter aerogenes, Klebsiella pneumoniae, and Salmonella typhimurium exhibited patterns of catalase activity similar to E. coli, including bifunctional HPI-like bands and a monofunctional HPII-like band. Proteus mirabilis, Erwinia carotovora, and Serratia marcescens contained a single band of monofunctional catalase with a mobility intermediate between the HPI-like and HPII-like bands. The cloned genes for catalases HPI (katG) and HPII (katE) from E. coli were used as probes in Southern hybridization analyses for homologous sequences in genomic DNA of the same bacteria. katG was found to hybridize with fragments from C. freudii, Ent. aerogenes, Sal. typhimurium, and K. pneumoniae but not at all with Ed. tarda, P. mirabilis, S. marcesens, or Er. carotovora. katE hybridized with C. freundii and K. pneumoniae DNAs and not with the other bacterial DNAs.     

Prevalences of zoonotic bacteria among seabirds in rehabilitation centers along the Pacific Coast of California and Washington, USA

Many seabirds are rehabilitated annually by wildlife rehabilitation centers along the Pacific Coast, USA. Although various strains of zoonotic bacteria have been isolated from seabirds, risks to rehabilitators at these centers have not been well documented. From November 2001 through January 2003, we determined the prevalence of detectable enteric fauna by isolation and characterization of Gram-negative bacteria from cloacal swabs taken from 26 common murres (Uria aalge), 49 gulls (Larus spp.), and 14 other seabirds treated by rehabilitators in California and Washington (USA). At least 25 bacterial species were identified, including multiple strains of Escherichia coli, as well as Enterobacter cloacae, Citrobacter freundii, and Klebsiella pneumoniae. Antibiotic resistance was found in 13 of 19 bacterial isolates tested, including E. coli, K. pneumoniae, Acinetobacter baumanii, and Pseudomonas aeruginosa. Potential transfer of these bacteria poses a risk to wildlife rehabilitators and to seabirds in these centers, as well as to free-ranging birds.     

Potential prophylactic value of bovine colostrum in necrotizing enterocolitis in neonates: an in vitro study on bacterial attachment, antibody levels and cytokine production

Necrotizing enterocolitis (NEC) is an important disease of low birth-weight neonates. The immaturity of the gut mucosa may result in close contact between the host epithelium and microorganisms which are normally confined to the gut lumen. Damage of the mucosa due to endotoxin, cytokine production or other factors is believed to then occur. The aim of this study was to determine whether spray-dried bovine colostrum demonstrated potential in vitro as a prophylactic for NEC. Antiadherence was measured using a tissue culture assay and antibody levels against Enterobacteriaceae were determined by ELISA. The effect of bovine colostrum on the production of cytokines implicated in NEC was determined by a multiplex bead assay. Enterobacter cloacae, Klebsiella oxytoca, Escherichia coli, Serratia marcescens and Klebsiella pneumoniae ssp. pneumoniae were common in both NEC positive and NEC negative infants and IgA and IgG1 antibodies to these species were present in the bovine colostrum. Pretreatment with bovine colostrum produced a significant decrease (P<0.001) in attachment of bacteria to HT-29 cells. Bovine colostrum significantly increased the production of IL-8 in HT-29 cells and IL-8, IL-6 and TNF-alpha in THP-1 cells (P<0.001). The potential of bovine colostrum to increase the production of inflammatory mediators could limit its usefulness.     

Isolation of some pathogenic bacteria from the great spruce bark beetle, <Emphasis Type="Italic">Dendroctonus micans</Emphasis> and its specific predator, <Emphasis Type="Italic">Rhizophagus grandis</Emphasis>

Some bacteria were isolated from Dendroctonus micans and its specific predator, Rhizophagus grandis. Six bacteria from D. micans were identified as Bacillus pumilus, Enterobacter intermedius, Citrobacter freundii, Cellulomonas flavigena, Microbacterium liquefaciens and Enterobacter amnigenus, three bacteria from R. grandis as Klebsiella pneumoniae, Pantoea agglomerans and Serratia grimesii, on the basis of fatty acid methyl ester analysis and carbon utilization profile by using Microbial Identification and Biolog Microplate Systems. Their insecticidal effects were tested on larvae and adults of D. micans.     

Klebsiella, Enterobacter, and Serratia: Biochemical Differentiation and Susceptibility to Ampicillin and Three Cephalosporin Derivatives

Three hundred twenty-nine strains of the tribe Klebsielleae were compared by several biochemical tests and by susceptibility to selected antibiotics. Biochemical tests included urease, amino acid decarboxylase, and hydrogen sulfide production; fermentation of lactose and dextrose; motility; and tests in the IMViC (indole, methyl red, Voges-Proskauer, citrate) series. The isolates were: Klebsiella species, 67.5%; Enterobacter species, 28%, and Serratia species, 4.5%. Minimal inhibitory concentrations of cephaloridine, cephalothin, and a new cephalosporin, cephalexin, and of ampicillin were determined by the agar dilution procedure. Cephalosporins at 20 mug/ml or less inhibited 90% of the Klebsiella strains but only 15% of the Enterobacter strains. Ampicillin inhibited 27% of Enterobacter strains and 17% of Klebsiella strains. Serratia isolates were insensitive to the cephalosporins and ampicillin. The results suggest that precise identification of this group to the generic level can be accomplished readily in the clinical laboratory and that such information is helpful in the preliminary selection of an antibiotic for treatment of clinical infections.     

Evolution of chemotactic-signal transducers in enteric bacteria.

The methyl-accepting chemotactic-signal transducers of the enteric bacteria are transmembrane proteins that consist of a periplasmic receptor domain and a cytoplasmic signaling domain. To study their evolution, transducer genes from Enterobacter aerogenes and Klebsiella pneumoniae were compared with transducer genes from Escherichia coli and Salmonella typhimurium. There are at least two functional transducer genes in the nonmotile species K. pneumoniae, one of which complements the defect in serine taxis of an E. coli tsr mutant. The tse (taxis to serine) gene of E. aerogenes also complements an E. coli tsr mutant; the tas (taxis to aspartate) gene of E. aerogenes complements the defect in aspartate taxis, but not the defect in maltose taxis, of an E. coli tar mutant. The sequence was determined for 5 kilobases of E. aerogenes DNA containing a 3' fragment of the cheA gene, cheW, tse, tas, and a 5' fragment of the cheR gene. The tse and tas genes are in one operon, unlike tsr and tar. The cytoplasmic domains of Tse and Tas are very similar to those of E. coli and S. typhimurium transducers. The periplasmic domain of Tse is homologous to that of Tsr, but Tas and Tar are much less similar in this region. However, several short sequences are conserved in the periplasmic domains of Tsr, Tar, Tse, and Tas but not of Tap and Trg, transducers that do not bind amino acids. These conserved regions include residues implicated in amino-acid binding.     

Influence of the addition of mineral salts on the inhibitory activity of strains of the Arthrinium genus

The influence which the addition of certain mineral salts has on the inhibiting activity of strains belonging to the Arthrinium genus was studied. The salts employed were AgNO3, FeCl2, MnCl2 and CuSO4, in concentrations of 10, 50, 100, 150 and 200 mg/l, in the presence of Bacillus subtilis, Enterobacter cloacae, Staphylococcus aureus, Escherichia coli, Serratia marcescens, Klebsiella pneumoniae, Candida albicans and Aspergillus niger. For B. subtilis and S. aureus greater inhibition was observed after the addition of FeCl2 (200 mg/l) and MnCl2 (10 mg/l). E. coli showed a greater sensibility in the presence of AgNO3 (200 mg/l) and CuSO4 (150 mg/l). Inhibition before Enterobacter cloacae and Serratia marcescens was not increased by the addition of any of the salts. The addition of MnCl2 at 10 mg/l concentration to the basal medium showed a considerable increase of inhibitory activity over Klebsiella pneumoniae and Candida albicans. Activity for Aspergillus niger decreased when any of the salts were added to the medium.     

Enterobacteriaceae islated from honey bees,Apis mellifera,treated with 2,4-D and antibiotics

Klebsiella pneumoniae, Shigella sp., Enterobacter cloacae, Escherichia coli, Enterobacter hafniae, Arizona sp., Enterobacter aerogenes, Serratia liquefaciens, and Citrobacter sp. were isolated from the intestinal contents of honey bees, Apis mellifera, which were obtained either from untreated colonies, from colonies fed the herbicide 2,4-D, or from colonies fed a combination of oxytetracycline and fumagillin. Antibiotics depressed the growth of Enterobacteriaceae; 2,4-D had little effect on the enteric microflora of bees.     

A molecular phylogeny of enteric bacteria and implications for a bacterial species concept

A molecular phylogeny for seven taxa of enteric bacteria (Citrobacter freundii, Enterobacter cloacae, Escherichia coli, Hafnia alvei, Klebsiella oxytoca, Klebsiella pneumoniae, and Serratia plymuthica) was made from multiple isolates per taxa taken from a collection of environmental enteric bacteria. Sequences from five housekeeping genes (gapA, groEL, gyrA, ompA, and pgi) and the 16S rRNA gene were used to infer individual gene trees and were concatenated to infer a composite molecular phylogeny for the species. The isolates from each taxa formed tight species clusters in the individual gene trees, suggesting the existence of 'genotypic' clusters that correspond to traditional species designations. These sequence data and the resulting gene trees and consensus tree provide the first data set with which to assess the utility of the recently proposed core genome hypothesis (CGH). The CGH provides a genetically based approach to applying the biological species concept to bacteria.     

Fermentation of polysaccharides by Klebsielleae and other facultative bacilli.

Fermentations of 10 polysaccharides by species of the family Enterobacteriaceae were examined. Algin, guar, karaya, xanthan, and xylan were not fermented by any of the strains tested. Most of the activity was found in the tribe Klebsielleae. Klebsiella oxytoca fermented amylopectin (97% of the strains studied), carrageenan (100%), inulin (68%), polypectate (100%), and tragacanth (100%). Klebsiella pneumoniae fermented amylopectin (91%), carrageenan (100%), and tragacanth (86%). Carrageenan was also fermented by Enterobacter aerogenes (100%), Enterobacter agglomerans (63%), Enterobacter cloacae (95%), and Pectobacterium (38%). Pectobacterium shared polypectate fermentation (100%) with K. oxytoca. With one exception, Serratia strains were negative on all polysaccharides. These results, along with other evidence, indicate that (i) the genus Klebsiella is biochemically the most versatile genus of the tribe, (ii) because of its distinct characteristics, K. oxytoca warrants species designation separate from K. pneumoniae, and (iii) some food additives generally considered indigestible can be metabolized by a few species of facultative bacilli, whereas others appear to be resistant.     

Serotypes and pyocin types of Pseudomonas aeruginosa isolated from natural waters

The susceptibility of strains of Enterobacter cloacae, Klebsiella pneumoniae, Serratia marcescens, Pseudomonas aeruginosa, Proteus mirabilis and Escheriehia coli to six aminoglycosides was tested in media of different osmolarity and ionic content. We observed that increasing osmolarity decreased susceptibility of these Gram-negative bacteria to all antibiotics used. On the other hand, raising of ionic strength increased the susceptibility to tobramycin, neomycin and gentarnicin in all bacteria tested.     

Glutathione transferase in bacteria: subunit composition and antigenic characterization

The presence of glutathione transferase (GST; EC 2.5.1.18) in Escherichia coli ATCC 25922, E. coli ATCC 25422, Proteus vulgaris ATCC 8427, Pseudomonas aeruginosa ATCC 27853, Klebsiella oxytoca CIP 666, K. oxytoca AF 101, Enterobacter cloacae CIP 6085, Serratia marcescens CIP 6755, and Proteus mirabilis AF 2924 was investigated. Using 1-chloro-2,4-dinitrobenzene as substrate, GST activity was found in the glutathione-(GSH-)affinity-purified fraction of all strains tested. SDS-PAGE analysis of GSH-affinity-purified enzyme indicated that the GSTs of all these bacteria are dimers of two identical subunits of Mr about 22,500. Rabbit antiserum directed against the major isoenzyme present in Proteus mirabilis AF 2924, Pm-GST-6.0, was used to investigate the antigenic properties of bacterial GSTs. Western blot analysis indicated that a GST antigenically identical to Pm-GST-6.0 is present in Enterobacter cloacae CIP 6085, Escherichia coli ATCC 25422 and Proteus vulgaris ATCC 8427, but absent in Escherichia coli ATCC 25922, Klebsiella oxytoca CIP 666, K. oxytoca AF 101 and Serratia marcescens CIP 6755. The presence of Pm-GST-6.0, but not mammalian GST, increased the MIC values of amikacin, ampicillin, cefotaxime, cephalothin and nalidixic acid for E. coli ATCC 25922. It is suggested that bacterial GST may represent a defense against the effects of antibiotics.     

Gram-negative bacteria resistant to antibiotics in foods

From 154 food samples, including vegetables (lettuce), milk and meals served at school it was possible to isolate and identify 400 Gram negative bacilli distributed among 339 enteric bacteria (Escherichia, Shigella, Citrobacter, Klebsiella, Enterobacter, Serratia and Proteus) and other 61 non enteric bacilli (Acinetobacter, Flavobacterium, Aeromonas and Pseudomonas). Submitting this cultures to the drugs sulfadiazine (Su), streptomycin (Sm), tetracycline (Tc), chloramphenicol (Cm), kanamycin (Km), ampicillin (Ap), nalidixic acid (Nal) and gentamycin (Gm) it was observed only six stocks susceptible to all drugs and total sensibility to Gm. Among enteric bacteria the profiles Su (27,6%) and Su-Ap (39,6%) predominated, while for the non enteric bacilli percentages of 18.0 for Ap and 9.8 for Su-Ap were detected. Aiming to better characterization of resistance, experiments of conjugation were made with standard strains of Escherichia coli K 12. Great concern was raised by the recognition of these cultures due to the elevated R+ taxes for the enteric bacilli that were close to 90% (milk and food at school) and about 70% in relation to lettuce.     

深圳市儿童社区获得性肺炎细菌病原学及其耐药性

目的研究儿童社区获得性肺炎细菌病原学及其耐药性特征,指导临床合理应用抗菌药物。方法对2006年2月至2007年3月1年期间住院的5岁及5岁以下社区获得性肺炎病人,进行深部呼吸道吸引物细菌培养,并且检测分离菌株对常用抗菌药物的耐药性。结果1441例病人中,722例检出细菌共761株,分离阳性率为50．1％,分离菌依次为肺炎克雷伯菌170株（22．3％）、大肠埃希菌130株（17．1％）、肺炎链球菌89株（11．7％）、金黄色葡萄球菌63株（8．3％）及流感和副流感嗜血杆菌60株（7．9％）。耐甲氧西林金黄色葡萄球菌（MRSA）检出率为15．9％;对青霉素不敏感的肺炎链球菌（包括PISP和PRSP）检出率为84．3％;肺炎克雷伯菌、大肠埃希菌、粘质沙雷菌和阴沟肠杆菌产ESBLs的检出率分别为31．2％、46．2％、94．8％和16．8％;流感嗜血杆菌和副流感嗜血杆菌对氨苄西林的耐药率为36％和40％;铜绿假单胞菌和鲍曼复合不动杆菌对亚胺培南的耐药率分别为10．7％和13．2％。结论在深圳儿童社区获得性肺炎的分离菌中,革兰阴性菌明显多于革兰阳性菌,分离菌依次为肺炎克雷伯菌、大肠埃希菌、肺炎链球菌、金黄色葡萄球菌及流感和副流感嗜血杆菌。分离细菌对常用抗菌药物的耐药性较为严重。     

Evolution of the ferric enterobactin receptor in gram-negative bacteria.

Using sodium dodecyl sulfate-polyacrylamide gel electrophoresis of iron-deficient and replete cell envelopes, 59Fe-siderophore uptake studies, and Western immunoblots and cytofluorimetric analyses with monoclonal antibodies (MAbs), we surveyed a panel of gram-negative bacteria to identify outer membrane proteins that are structurally related to the Escherichia coli K-12 ferric enterobactin receptor, FepA. Antibodies within the panel identified FepA epitopes that are conserved among the majority of the bacteria tested, as well as epitopes present in only a few of the strains. In general, epitopes of FepA that are buried in the outer membrane bilayer were more conserved among gram-negative bacteria than epitopes that are exposed on the bacterial cell surface. The surface topology and tertiary structure of FepA are quite similar in E. coli and Shigella flexneri but differ in Salmonella typhimurium. Of the 18 different genera tested, 94% of the bacteria transported ferric enterobactin, including members of the previously unrecognized genera Citrobacter, Edwardsiella, Enterobacter, Haemophilus, Hafnia, Morganella, Neisseria, Proteus, Providencia, Serratia, and Yersinia. The ferric enterobactin receptor contains at least one buried epitope, recognized by MAb 2 (C. K. Murphy, V. I. Kalve, and P. E. Klebba, J. Bacteriol. 172:2736-2746, 1990), that is conserved within the structure of an iron-regulated cell envelope protein in all the bacteria that we have surveyed. With MAb 2, we identified and determined the Mr of cell envelope antigens that are immunologically related to E. coli FepA in all the gram-negative bacteria tested. Collectively, the library of anti-FepA MAbs showed unique patterns of reactivity with the different bacteria, allowing identification and discrimination of species within the following gram-negative genera: Aeromonas, Citrobacter, Edwardsiella, Enterobacter, Escherichia, Haemophilus, Hafnia, Klebsiella, Morganella, Neisseria, Proteus, Providencia, Pseudomonas, Salmonella, Serratia, Shigella, Vibrio, and Yersinia.