Medical and economic impact of extraintestinal infections due to Escherichia coli: focus on an increasingly important endemic problem

Escherichia coli is probably the best-known bacterial species and one of the most frequently isolated organisms from clinical specimens. Despite this, underappreciation and misunderstandings exist among medical professionals and the lay public alike regarding E. coli as an extraintestinal pathogen. Underappreciated features include (i) the wide variety of extraintestinal infections E. coli can cause, (ii) the high incidence and associated morbidity, mortality, and costs of these diverse clinical syndromes, (iii) the pathogenic potential of different groups of E. coli strains for causing intestinal versus extraintestinal disease, and (iv) increasing antimicrobial resistance. In this era in which health news often sensationalizes uncommon infection syndromes or pathogens, the strains of E. coli that cause extraintestinal infection are an increasingly important endemic problem and underappreciated "killers". Billions of health care dollars, millions of work days, and hundreds of thousands of lives are lost each year to extraintestinal infections due to E. coli. New treatments and prevention measures will be needed for improved outcomes and a diminished disease burden.     

孕产妇产后感染病原菌的构成及耐药性分析

目的了解孕产妇产后感染病原菌的分布及耐药现状,为临床抗感染治疗提供依据。方法采用法国生物梅里埃VITEK-2全自动细菌鉴定分析仪进行鉴定和药敏试验,采用WHONET5.6软件进行数据统计分析。结果 2012年1月至2015年12月我院产科病房孕产妇共60 229例,产后感染336例,感染主要部位为宫腔、切口、血液和尿液。主要病原菌依次为大肠埃希菌、粪肠球菌、B型链球菌和金黄色葡萄球菌。大肠埃希菌对头孢他啶、头孢吡肟、妥布霉素、阿米卡星、头孢替坦、哌拉西林/他唑巴坦、头孢哌酮/舒巴坦、亚胺培南耐药率均15.00%;粪肠球菌和无乳链球菌对氨苄西林、青霉素耐药率为0.00%,耐甲氧西林金黄色葡萄球菌(MRSA)耐药率高达19.05%。结论我院产科病房感染主要部位为宫腔(43.18%),减少剖宫产率可以降低产后感染发生率,感染的主要病原菌为大肠埃希菌和粪肠球菌(63.23%),在预防用药的选择上应选择对大肠埃希菌和粪肠球菌均有效的抗生素以减少产后感染的发生率,同时了解孕产妇产后感染病原菌的构成和耐药性,有助于临床更快速有效地治疗产后感染。     

泌尿外科住院患者泌尿系感染病原菌分布及其耐药性分析

了解泌尿外科住院患者泌尿系感染病原菌的分布及其对常用抗菌药物的耐药情况。对泌尿外科泌尿系感染住院患者的消毒中段尿培养结果进行回顾性分析,尿培养菌株的鉴定、药敏分析和统计分析采用VITEK2全自动微生物仪。3 a中泌尿外科泌尿系感染住院患者共分离到细菌1 233株,其中革兰阴性杆菌772株,占62.61%,革兰阳性球菌353株,占28.63%,真菌82株,占6.65%。菌株数居前5位的细菌依次为:大肠埃希菌、肠球菌属、洋葱伯克霍尔德菌、假丝酵母菌属和变形杆菌。分离大肠埃希菌产ESBLs率为66.18%,粪肠球菌中未发现VRE菌株,屎肠球菌VRE为0.8%。未发现对美洛培南耐药的大肠埃希菌,对肠埃希菌耐药率在10%以下的有亚胺培南、阿米卡星、哌拉西林/他唑巴坦和呋喃妥因。洋葱伯克霍尔德菌和铜绿假单胞菌对所监测的抗菌药物均有不同程度的耐药。未发现对万古霉素、替考拉宁及氨苄西林耐药的粪肠球菌。泌尿系感染的病原菌以大肠埃希菌和肠球菌为主,非发酵革兰阴性杆菌及假丝酵母菌所占比例超过20%,不容忽视,病原菌的耐药率较高,临床医生应根据尿培养和药敏试验结果合理使用抗菌药物。     

Interaction of Escherichia coli K1 and K5 with Acanthamoeba castellanii trophozoites and cysts

The existence of symbiotic relationships between Acanthamoeba and a variety of bacteria is well-documented. However, the ability of Acanthamoeba interacting with host bacterial pathogens has gained particular attention. Here, to understand the interactions of Escherichia coli K1 and E. coli K5 strains with Acanthamoeba castellanii trophozoites and cysts, association assay, invasion assay, survival assay, and the measurement of bacterial numbers from cysts were performed, and nonpathogenic E. coli K12 was also applied. The association ratio of E. coli K1 with A. castellanii was 4.3 cfu per amoeba for 1 hr but E. coli K5 with A. castellanii was 1 cfu per amoeba for 1 hr. By invasion and survival assays, E. coli K5 was recovered less than E. coli K1 but still alive inside A. castellanii. E. coli K1 and K5 survived and multiplied intracellularly in A. castellanii. The survival assay was performed under a favourable condition for 22 hr and 43 hr with the encystment of A. castellanii. Under the favourable condition for the transformation of trophozoites into cysts, E. coli K5 multiplied significantly. Moreover, the pathogenic potential of E. coli K1 from A. castellanii cysts exhibited no changes as compared with E. coli K1 from A. castellanii trophozoites. E. coli K5 was multiplied in A. castellanii trophozoites and survived in A. castellanii cysts. Therefore, this study suggests that E. coli K5 can use A. castellanii as a reservoir host or a vector for the bacterial transmission.     

Comparison of virulence gene profiles of Escherichia coli strains isolated from healthy and diarrheic swine

A combination of uni- and multiplex PCR assays targeting 58 virulence genes (VGs) associated with Escherichia coli strains causing intestinal and extraintestinal disease in humans and other mammals was used to analyze the VG repertoire of 23 commensal E. coli isolates from healthy pigs and 52 clinical isolates associated with porcine neonatal diarrhea (ND) and postweaning diarrhea (PWD). The relationship between the presence and absence of VGs was interrogated using three statistical methods. According to the generalized linear model, 17 of 58 VGs were found to be significant (P < 0.05) in distinguishing between commensal and clinical isolates. Nine of the 17 genes represented by iha, hlyA, aidA, east1, aah, fimH, iroN(E. coli), traT, and saa have not been previously identified as important VGs in clinical porcine isolates in Australia. The remaining eight VGs code for fimbriae (F4, F5, F18, and F41) and toxins (STa, STb, LT, and Stx2), normally associated with porcine enterotoxigenic E. coli. Agglomerative hierarchical algorithm analysis grouped E. coli strains into subclusters based primarily on their serogroup. Multivariate analyses of clonal relationships based on the 17 VGs were collapsed into two-dimensional space by principal coordinate analysis. PWD clones were distributed in two quadrants, separated from ND and commensal clones, which tended to cluster within one quadrant. Clonal subclusters within quadrants were highly correlated with serogroups. These methods of analysis provide different perspectives in our attempts to understand how commensal and clinical porcine enterotoxigenic E. coli strains have evolved and are engaged in the dynamic process of losing or acquiring VGs within the pig population.     

Insights into a multidrug resistant Escherichia coli pathogen of the globally disseminated ST131 lineage: genome analysis and virulence mechanisms

Escherichia coli strains causing urinary tract infection (UTI) are increasingly recognized as belonging to specific clones. E. coli clone O25b:H4-ST131 has recently emerged globally as a leading multi-drug resistant pathogen causing urinary tract and bloodstream infections in hospitals and the community. While most molecular studies to date examine the mechanisms conferring multi-drug resistance in E. coli ST131, relatively little is known about their virulence potential. Here we examined E. coli ST131 clinical isolates from two geographically diverse collections, one representing the major pathogenic lineages causing UTI across the United Kingdom and a second representing UTI isolates from patients presenting at two large hospitals in Australia. We determined a draft genome sequence for one representative isolate, E. coli EC958, which produced CTX-M-15 extended-spectrum β-lactamase, CMY-23 type AmpC cephalosporinase and was resistant to ciprofloxacin. Comparative genome analysis indicated that EC958 encodes virulence genes commonly associated with uropathogenic E. coli (UPEC). The genome sequence of EC958 revealed a transposon insertion in the fimB gene encoding the activator of type 1 fimbriae, an important UPEC bladder colonization factor. We identified the same fimB transposon insertion in 59% of the ST131 UK isolates, as well as 71% of ST131 isolates from Australia, suggesting this mutation is common among E. coli ST131 strains. Insertional inactivation of fimB resulted in a phenotype resembling a slower off-to-on switching for type 1 fimbriae. Type 1 fimbriae expression could still be induced in fimB-null isolates; this correlated strongly with adherence to and invasion of human bladder cells and bladder colonisation in a mouse UTI model. We conclude that E. coli ST131 is a geographically widespread, antibiotic resistant clone that has the capacity to produce numerous virulence factors associated with UTI.     

Secretory monoclonal IgA class-switch variants against bacterial enteric pathogens in bile and intestinal secretions

Abstract In a previous study we analyzed the molecular forms of monoclonal IgA class-switch variants (moIgA variants) and their transport into murine respiratory secretions. The aim of the present study is to characterize the transport of moIgA variants into bile and intestinal secretions so that their applicability in a passive immunization model of the gut can be evaluated. Different moIgA variants were directly isolated from IgG1 and IgG2a producing hybridoma clones specific for the same surface determinants of bacterial enteric pathogens ( Salmonella typhimurium and Campylobacter jejuni ) as their respective parent IgG clones. Hepatobiliary transport experiments clearly revealed the selective transport of biologically active polymeric forms of the IgA variants into the murine and rat bile after intravenous injection. Biotinylation of polymeric IgA variants prior to intravenous injection resulted in the recovery of functional, labeled SIgA. Moreover biotin-labeled polymeric IgA variant was recovered in bile with an increased molecular weight, suggesting that the secretory component had been added during passage through the liver. When IgA variant and IgG parent clones were both used in a murine backpack tumor model for passive immunization, IgA variant was selectively transported into intestinal secretions in comparison to IgG. The experimental model described here is suitable for use in comparative studies on the role of IgA and IgG with identical specificity in invasive infections of the intestinal tract.     

Correlation between In Vivo Biofilm Formation and Virulence Gene Expression in Escherichia coli O104:H4

The emergence of novel pathogens poses a major public health threat causing widespread epidemics in susceptible populations. The Escherichia coli O104:H4 strain implicated in a 2011 outbreak in northern Germany caused the highest frequency of hemolytic uremic syndrome (HUS) and death ever recorded in a single E. coli outbreak. Therefore, it has been suggested that this strain is more virulent than other pathogenic E. coli (e.g., E. coli O157:H7). The E. coli O104:H4 outbreak strain possesses multiple virulence factors from both Shiga toxin (Stx)-producing E. coli (STEC) and enteroaggregative E. coli (EAEC), though the mechanism of pathogenesis is not known. Here, we demonstrate that E. coli O104:H4 produces a stable biofilm in vitro and that in vivo virulence gene expression is highest when E. coli O104:H4 overexpresses genes required for aggregation and exopolysaccharide production, a characteristic of bacterial cells residing within an established biofilm. Interrupting exopolysaccharide production and biofilm formation may therefore represent effective strategies for combating future E. coli O104:H4 infections.     

尿路感染相关病原菌的分布及耐药性分析

目的调查尿路感染病原菌的分布和耐药特点,为临床的抗感染治疗提供依据。方法收集2013年至2015年荆州市中心医院门诊和住院患者中,尿路感染患者送检的尿培养和血培养标本中检出的病原菌,采用Vitek2 Compact全自动微生物检测仪进行细菌鉴定,采用纸片扩散法和仪器法分别对革兰阴性杆菌和革兰阳性球菌进行药敏试验,药敏结果的判断依照CLSI M100-S24标准。数据分析采用WHONET5.6和SPSS 19.0软件,统计分析采用x~2检验。结果从尿路感染患者送检的标本中共检出各类非重复病原菌2 306株,其中门诊患者中检出19种100株,住院患者检出56种2 206株。导致尿路感染最多的两种病原菌为大肠埃希菌和粪肠球菌,分别检出1 241株和232株。导致尿脓毒血症最多的两种病原菌为大肠埃希菌和肺炎克雷伯菌,分别检出36株和10株。大肠埃希菌产ESBL.s率达67.9%,其对多种抗菌药物的耐药性均高于60.0%。粪肠球菌对大多数抗菌药物的耐药性均高于50.0%,仅对呋喃妥因和高浓度链霉素的耐药性较低,分别为12.0%和38.7%;未检出对万古霉素、利奈唑胺和替加环素耐药的粪肠球菌。结论导致尿路感染的病原菌种类繁多,大肠埃希菌和粪肠球菌是主要病原菌,其耐药情况严重;为保证治疗的有效性,临床医生应注重相关病原学和药敏检查结果。     

The dps gene of symbiotic "Candidatus Legionella jeonii" in Amoeba proteus responds to hydrogen peroxide and phagocytosis

To survive in host cells, intracellular pathogens or symbiotic bacteria require protective mechanisms to overcome the oxidative stress generated by phagocytic activities of the host. By genomic library tagging, we cloned a dps (stands for DNA-binding protein from starved cells) gene of the symbiotic "Candidatus Legionella jeonii" organism (called the X bacterium) (dps(X)) that grows in Amoeba proteus. The gene encodes a 17-kDa protein (pI 5.19) with 91% homology to Dps and DNA-binding ferritin-like proteins of other organisms. The cloned gene complemented the dps mutant of Escherichia coli and conferred resistance to hydrogen peroxide. Dps(X) proteins purified from E. coli transformed with the dps(X) gene were in oligomeric form, formed a complex with pBlueskript SKII DNA, and protected the DNA from DNase I digestion and H(2)O(2)-mediated damage. The expression of the dps(X) gene in "Candidatus Legionella jeonii" was enhanced when the host amoeba was treated with 2 mM H(2)O(2) and by phagocytic activities of the host cell. These results suggested that the Dps protein has a function protective of the bacterial DNA and that its gene expression responds to oxidative stress generated by phagocytic activities of the host cell. With regard to the fact that invasion of Legionella sp. into respiratory phagocytic cells causes pneumonia in mammals, further characterization of dps(X) expression in the Legionella sp. that multiplies in a protozoan host in the natural environment may provide valuable information toward understanding the protective mechanisms of intracellular pathogens.     

Tuning of expression level of the genes of interest located in the bacterial chromosome

The new method of construction of the set of E. coli clones, differing in the promoter strength upstream the gene of interest, has been developed and tested using native E. coli MG 1655 lacZ gene as the reporter. This method includes the construction of the promoter-carrying DNA fragment obtained by PCR with consensus P(tac) as a template and the primers that lead to randomization of 4 central nucleotides in the promoter "-35"-region, linking the obtained fragments with the selective marker (Cm(R)) followed by Red-driven integration of the resulted DNA fragments directly in E. coli MG1655 chromosome instead the native lacI-gene and promoter/operator region of lac-operon. Due to direct determination of LacZ-activity in the independently obtained clones-integrants, we have found 14 new promoters (from 44 = 256 possible variants) that differ in their strength up to 100 fold (LacZ-activity in the corresponding strains smoothly varies from 10(2) for the weakest tested promoter up to 10(4) Miller U detected for the initial P(tac)). Sequencing of obtained promoters revealed that randomization of three positions in the "-35"-region is sufficient to obtain representative promoter library that would decrease the total number of potential promoter variants from 256 up to 64. It seems probable that exploiting of the developed method leading to one-step construction the library of clones with varied expression of gene/operon of interest could be useful tool in the modem metabolic engineering for optimization of genes expression.     

Clonal relationships among Escherichia coli serogroup 06 isolates from human and animal infections

The clonal relationship of thirty E. coli strains of 0 antigen serotype 06 isolated from human, dog, pig or cow infections were investigated. Two main clones with serotypes 06 : H1 or 06 : H31, H- were identified. Isolates from humans, dogs, pigs and cows were found in both clones, indicating that animals are a possible source for human extraintestinal Escherichia coli strains. Two human ETEC (06 : H16) and two pig isolates (06 : H10) were not related to the 06 : H1 or 06 : H31, H- E. coli clones.     

Attaching and effacing pathogen-induced tight junction disruption in vivo

Diarrhoea is a hallmark of infections by the human attaching and effacing (A/E) pathogens, enterohaemorrhagic Escherichia coli (EHEC) and enteropathogenic E. coli (EPEC). Although the mechanisms underlying diarrhoea induced by these pathogens remain unknown, cell culture results have suggested that these pathogens may target tight junctions. Tight junctions in the colon function as physical intercellular barriers that separate and prevent mixing of the luminal contents with adlumenal regions of the epithelium. Consequently, it is thought that the disruption of intestinal epithelial tight junctions by A/E pathogens could result in a loss of barrier function in the alimentary tract; however, this remains unexamined. Here we demonstrate for the first time that A/E pathogen infection results in the morphological alteration of tight junctions during natural disease. Tight junction alteration, characterized by relocalization of the transmembrane tight junction proteins claudin 1, 3 and 5, is a functional disruption; molecular tracers, which do not normally penetrate uninfected epithelia, pass across pathogen-infected epithelia. Functional junction disruption occurs with a concomitant increase in colon luminal water content. The effects on tissue are dependent upon the bacterial type III effector EspF (E. coli secreted protein F), because bacteria lacking EspF, while able to colonize, are defective for junction disruption and result in decreased proportions of water in the colon compared with wild-type infection. These results suggest that the diarrhoea induced by A/E pathogens occurs as part of functional tight junction disruption.     

Autotransporter-encoding sequences are phylogenetically distributed among Escherichia coli clinical isolates and reference strains

Autotransporters are secreted bacterial proteins exhibiting diverse virulence functions. Various autotransporters have been identified among Escherichia coli associated with intestinal or extraintestinal infections; however, the specific distribution of autotransporter sequences among a diversity of E. coli strains has not been investigated. We have validated the use of a multiplex PCR assay to screen for the presence of autotransporter sequences. Herein, we determined the presence of 13 autotransporter sequences and five allelic variants of antigen 43 (Ag43) among 491 E. coli isolates from human urinary tract infections, diarrheagenic E. coli, and avian pathogenic E. coli (APEC) and E. coli reference strains belonging to the ECOR collection. Clinical isolates were also classified into established phylogenetic groups. The results indicated that Ag43 alleles were significantly associated with clinical isolates (93%) compared to commensal isolates (56%) and that agn43K12 was the most common and widely distributed allele. agn43 allelic variants were also phylogenetically distributed. Sequences encoding espC, espP, and sepA and agn43 alleles EDL933 and RS218 were significantly associated with diarrheagenic E. coli strains compared to other groups. tsh was highly associated with APEC strains, whereas sat was absent from APEC. vat, sat, and pic were associated with urinary tract isolates and were identified predominantly in isolates belonging to either group B2 or D of the phylogenetic groups based on the ECOR strain collection. Overall, the results indicate that specific autotransporter sequences are associated with the source and/or phylogenetic background of strains and suggest that, in some cases, autotransporter gene profiles may be useful for comparative analysis of E. coli strains from clinical, food, and environmental sources.     

Kinetics and strain specificity of rhizosphere and endophytic colonization by enteric bacteria on seedlings of Medicago sativa and Medicago truncatula

The presence of human-pathogenic, enteric bacteria on the surface and in the interior of raw produce is a significant health concern. Several aspects of the biology of the interaction between these bacteria and alfalfa (Medicago sativa) seedlings are addressed here. A collection of enteric bacteria associated with alfalfa sprout contaminations, along with Escherichia coli K-12, Salmonella enterica serotype Typhimurium strain ATCC 14028, and an endophyte of maize, Klebsiella pneumoniae 342, were labeled with green fluorescent protein, and their abilities to colonize the rhizosphere and the interior of the plant were compared. These strains differed widely in their endophytic colonization abilities, with K. pneumoniae 342 and E. coli K-12 being the best and worst colonizers, respectively. The abilities of the pathogens were between those of K. pneumoniae 342 and E. coli K-12. All Salmonella bacteria colonized the interiors of the seedlings in high numbers with an inoculum of 10(2) CFU, although infection characteristics were different for each strain. For most strains, a strong correlation between endophytic colonization and rhizosphere colonization was observed. These results show significant strain specificity for plant entry by these strains. Significant colonization of lateral root cracks was observed, suggesting that this may be the site of entry into the plant for these bacteria. At low inoculum levels, a symbiosis mutant of Medicago truncatula, dmi1, was colonized in higher numbers on the rhizosphere and in the interior by a Salmonella endophyte than was the wild-type host. Endophytic entry of M. truncatula appears to occur by a mechanism independent of the symbiotic infections by Sinorhizobium meliloti or mycorrhizal fungi.     

The structures of Escherichia coli O-polysaccharide antigens

Escherichia coli is usually a non-pathogenic member of the human colonic flora. However, certain strains have acquired virulence factors and may cause a variety of infections in humans and in animals. There are three clinical syndromes caused by E. coli : (i) sepsis/meningitis; (ii) urinary tract infection and (iii) diarrhoea. Furthermore the E. coli causing diarrhoea is divided into different 'pathotypes' depending on the type of disease, i.e. (i) enterotoxigenic; (ii) enteropathogenic; (iii) enteroinvasive; (iv) enterohaemorrhagic; (v) enteroaggregative and (vi) diffusely adherent. The serotyping of E. coli based on the somatic (O), flagellar (H) and capsular polysaccharide antigens (K) is used in epidemiology. The different antigens may be unique for a particular serogroup or antigenic determinants may be shared, resulting in cross-reactions with other serogroups of E. coli or even with other members of the family Enterobacteriacea . To establish the uniqueness of a particular serogroup or to identify the presence of common epitopes, a database of the structures of O-antigenic polysaccharides has been created. The E. coli database (ECODAB) contains structures, nuclear magnetic resonance chemical shifts and to some extent cross-reactivity relationships. All fields are searchable. A ranking is produced based on similarity, which facilitates rapid identification of strains that are difficult to serotype (if known) based on classical agglutinating methods. In addition, results pertinent to the biosynthesis of the repeating units of O-antigens are discussed. The ECODAB is accessible to the scientific community at http://www.casper.organ.su.se/ECODAB/ .     

Quinolone/fluoroquinolone susceptibility in Escherichia coli correlates with human polymicrobial bacteriuria and with in vitro interleukine-8 suppression

Urinary tract infections (UTIs) are frequently polymicrobial diseases mainly sustained by Escherichia coli in association with other opportunistic pathogens. Cystitis and pyelonephritis are usually accompanied by an inflammatory response, which includes neutrophil recruitment. Uropathogenic E. coli possess the ability to evade host defenses, modulating the innate immune response. The aim of this study was to determine whether particular E. coli strains correlate with polymicrobial bacteriuria and whether escape from the early host defenses and microbial synergy could lead to mixed UTIs. We evaluated 188 E. coli-positive urine samples and assessed the relationships among polymicrobism, neutrophil presence and several traits of E. coli isolates (virulence factors such as hlyA, fimA, papC and their relative products, i.e. hemolysin, type 1 and P fimbriae, and cnf1, their phylogenetic group) and their ability to suppress cytokine response in 5637 bladder epithelial cells. Escherichia coli susceptibility toward quinolones and fluoroquinolones, known to be linked to the pathogenicity of this species, was also considered. We found significant correlations among polymicrobial bacteriuria, absence of pyuria and quinolone/fluoroquinolone susceptibility of E. coli isolates and their enhanced capability to suppress interleukin-8 urothelial production when compared with the patterns induced by the resistant strains.     

Cloning and expression of a cDNA encoding a maize glutathione-S-transferase in E. coli.

The isolation and characterization of a family of maize glutathione-S-transferases (GST's) has been described previously. These enzymes are designated GSTs I, II and III based on size, substrate specificity and responsiveness to safeners. GST III has been shown to act on the herbicide alachlor as well as the commonly used substrate 1-chloro-2,4-dinitrobenzene (CDNB). Clones were isolated from a maize cDNA library in lambda gt10. Three clones contained the entire coding region for GST III. The sequences of these clones were consistent with the known amino terminal GST III protein sequence. Moreover, expression of one of these clones in E. coli resulted in a GST activity as measured with both CDNB and alachlor, proving that at least one of the clones encodes an active GST III species. With the enzyme expressed in E. coli it will become possible to study enzyme structure-function relationships ex planta. While a number of different GST proteins are present in maize tissue the GST III gene is present in single or low copy in the genome.     

Novel screening assay for in vivo selection of Klebsiella pneumoniae genes promoting gastrointestinal colonisation

ABSTRACT: BACKGROUND: Klebsiella pneumoniae is an important opportunistic pathogen causing pneumonia, sepsis and urinary tract infections. Colonisation of the gastrointestinal (GI) tract is a key step in the development of infections; yet the specific factors important for K. pneumoniae to colonize and reside in the GI tract of the host are largely unknown. To identify K. pneumoniae genes promoting GI colonisation, a novel genomic-library-based approach was employed. RESULTS: Screening of a K. pneumoniae C3091 genomic library, expressed in E. coli strain EPI100, in a mouse model of GI colonisation led to the positive selection of five clones containing genes promoting persistent colonisation of the mouse GI tract. These included genes encoding the global response regulator ArcA; GalET of the galactose operon; and a cluster of two putative membrane-associated proteins of unknown function. Both ArcA and GalET are known to be involved in metabolic pathways in Klebsiella but may have additional biological actions beneficial to the pathogen. In support of this, GalET was found to confer decreased bile salt sensitivity to EPI100. CONCLUSIONS: The present work establishes the use of genomic-library-based in vivo screening assays as a valuable tool for identification and characterization of virulence factors in K. pneumoniae and other bacterial pathogens.     

新生儿医院感染产超广谱β-内酰胺酶细菌流行状况及耐药性分析

目的了解新生儿医院感染中产超广谱β-内酰胺酶（ESBLs）细菌流行状况及耐药性,为预防和控制感染提供依据。方法对2011年1月至2013年12月间新生儿医院感染病原菌分布及耐药性进行回顾性分析;用VITEK-2 Compact微生物鉴定系统鉴定菌种和药敏试验。结果共检出病原菌192株,105株为肺炎克雷伯菌和大肠埃希菌,占54.7%;检出产ESBLs菌58株,全部来自肺炎克雷伯菌和大肠埃希菌;产ESBLs菌对青霉素类、头孢菌素类、单内酰环类抗菌药物高度耐药,耐药率〉80.0%,对头孢哌酮/舒巴坦、哌拉西林/他唑巴坦耐药率较低,耐药率〈20.0%,未检测到亚胺培南和美罗培南耐药株。结论产ESBLs肺炎克雷伯菌和大肠埃希菌是新生儿医院感染中主要的病原菌,且对常用抗菌药物耐药率较高,临床应加强病原菌的耐药性监测,合理使用抗菌药物。