Antimicrobial Resistance and Virulence Genes of Escherichia coli Isolates from Swine in Ontario

A total of 318 Escherichia coli isolates obtained from diarrheic and healthy pigs in Ontario from 2001 to 2003 were examined for their susceptibility to 19 antimicrobial agents. They were tested by PCR for the presence of resistance genes for tetracycline, streptomycin, sulfonamides, and apramycin and of 12 common virulence genes of porcine E. coli. Antimicrobial resistance frequency among E. coli isolates from swine in Ontario was moderate in comparison with other countries and was higher in isolates from pigs with diarrhea than in isolates from healthy finisher pigs. Resistance profiles suggest that cephamycinases may be produced by ≥8% of enterotoxigenic E. coli (ETEC). Resistance to quinolones was detected only in enterotoxigenic E. coli (≤3%). The presence of sul3 was demonstrated for the first time in Canada in porcine E. coli isolates. Associations were observed among tetA, sul1, aadA, and aac(3)IV and among tetB, sul2, and strA/strB, with a strong negative association between tetA and tetB. The paa and sepA genes were detected in 92% of porcine ETEC, and strong statistical associations due to colocation on a large plasmid were observed between tetA, estA, paa, and sepA. Due at least in part to gene linkages, the distribution of resistance genes was very different between ETEC isolates and other porcine E. coli isolates. This demonstrates that antimicrobial resistance epidemiology differs significantly between pathogenic and commensal E. coli isolates. These results may have important implications with regards to the spread and persistence of resistance and virulence genes in bacterial populations and to the prudent use of antimicrobial agents.     

Resistance to drugs and heavy metals, colicin production, and biochemical characteristics of selected bovine and porcine Escherichia coli strains.

A study was made of resistance to heavy metals and antibiotics, biochemical characteristics, and colicinogeny in selected strains of Escherichia coli of O serogroups 8, 9, 20, 64, 101, and X46. Of 42 strains that were investigated, 26 were porcine enterotoxigenic E. coli (ETEC), 8 were porcine non-enterotoxigenic E. coli (NETEC), and 8 were bovine ETEC. Multiple resistance to antimicrobial agents was common among the strains, and resistance to chloramphenicol and kanamycin was less common than resistance to other drugs, possibly reflecting the lower frequency of use of these agents in pigs and calves. Colicin production was a more common property of porcine ETEC (80.8%) than of porcine NETEC (25%), and all porcine ETEC of O serogroups 101 and 64 were colicinogenic. Equal numbers of bovine ETEC strains were colicinogenic as were non-colicinogenic. Resistance of bovine and porcine strains to sodium arsenate, mercury, and tellerium was 90, 16, and 5%, respectively. There was a close relationship between serogroup and biochemical reactions among the E. coli strains tested.     

Frequency and distribution of tetracycline resistance genes in genetically diverse, nonselected, and nonclinical Escherichia coli strains isolated from diverse human and animal sources

Nonselected and natural populations of Escherichia coli from 12 animal sources and humans were examined for the presence and types of 14 tetracycline resistance determinants. Of 1,263 unique E. coli isolates from humans, pigs, chickens, turkeys, sheep, cows, goats, cats, dogs, horses, geese, ducks, and deer, 31% were highly resistant to tetracycline. More than 78, 47, and 41% of the E. coli isolates from pigs, chickens, and turkeys were resistant or highly resistant to tetracycline, respectively. Tetracycline MICs for 61, 29, and 29% of E. coli isolates from pig, chickens, and turkeys, respectively, were >/=233 micro g/ml. Muliplex PCR analyses indicated that 97% of these strains contained at least 1 of 14 tetracycline resistance genes [tetA, tetB, tetC, tetD, tetE, tetG, tetK, tetL, tetM, tetO, tetS, tetA(P), tetQ, and tetX] examined. While the most common genes found in these isolates were tetB (63%) and tetA (35%), tetC, tetD, and tetM were also found. E. coli isolates from pigs and chickens were the only strains to have tetM. To our knowledge, this represents the first report of tetM in E. coli.     

Genome sequences and phylogenetic analysis of K88- and F18-positive porcine enterotoxigenic Escherichia coli

Porcine enterotoxigenic Escherichia coli (ETEC) continues to result in major morbidity and mortality in the swine industry via postweaning diarrhea. The key virulence factors of ETEC strains, their serotypes, and their fimbrial components have been well studied. However, most studies to date have focused on plasmid-encoded traits related to colonization and toxin production, and the chromosomal backgrounds of these strains have been largely understudied. Here, we generated the genomic sequences of K88-positive and F18-positive porcine ETEC strains and examined the phylogenetic distribution of clinical porcine ETEC strains and their plasmid-associated genetic content. The genomes of porcine ETEC strains UMNK88 and UMNF18 were both found to contain remarkable plasmid complements containing known virulence factors, potential novel virulence factors, and antimicrobial resistance-associated elements. The chromosomes of these strains also possessed several unique genomic islands containing hypothetical genes with similarity to classical virulence factors, although phage-associated genomic islands dominated the accessory genomes of these strains. Phylogenetic analysis of 78 clinical isolates associated with neonatal and porcine diarrhea revealed that a limited subset of porcine ETEC lineages exist that generally contain common toxin and fimbrial profiles, with many of the isolates belonging to the ST10, ST23, and ST169 multilocus sequencing types. These lineages were generally distinct from existing human ETEC database isolates. Overall, most porcine ETEC strains appear to have emerged from a limited subset of E. coli lineages that either have an increased propensity to carry plasmid-encoded virulence factors or have the appropriate ETEC core genome required for virulence.     

Characterization of an F18+ enterotoxigenic Escherichia coli strain from post weaning diarrhoea of swine, and of its conjugative virulence plasmid pTC

The enterotoxigenic Escherichia coli (ETEC) strain Ec2173, causing post weaning diarrhoea in swine, harbours six plasmids ranging from 13 to 200 kb in size. The heat stable toxin genes sta, stb and a tetracycline resistance gene were located on a self conjugative 120-kb plasmid, called pTC. In the cloned ColE1 type origin of replication of pTC a deletion was detected compared to other ColE1 replicons affecting the replication modulator gene rom. Epidemiological studies on ETEC isolates showed that pTC-like plasmids are widely distributed among porcine ETEC strains; thus representing an example of co-evolution of antibacterial resistance and virulence in pathogenic E. coli.     

产志贺样毒素和肠毒素大肠杆菌分子流行病学

[目的]人和动物腹泻的主要病原菌为大肠杆菌,本文主要研究贵州省致腹泻大肠杆菌毒力因子的分布类型.[方法]采用PCR技术对各毒力因子的基因分布进行研究.[结果]共分离到333株大肠杆菌,其中产肠毒素大肠杆菌(ETEC)在腹泻的人、猪、牛群中占优势,分别为:人群73(n=112),猪群82(n=106),牛群18(n=115).在ETEC菌株中检测到热敏肠毒素(lt)和不耐热肠毒素(st)基因,还存在lt/st并存现象.从人、猪、牛群中还检测到产志贺样毒素大肠杆菌(STEC),其中源自猪的STEC的检出率最高.大部分STEC同时携带lt、st或lt和st同时并存.编码F18菌毛的主亚基由fedA基因编码.对所分离大肠杆菌F18菌毛进行的研究结果表明,fedA基因主要与肠毒素基因共存,与stx基因并存的类型较少,25份猪源STEC菌株中仅有4份检测到fedA基因.[结论]贵州省人群、猪群和牛群致腹泻病原菌中以带F18菌毛的ETEC为主,STEC主要分布在腹泻的猪群中.     

Molecular characterization of multidrug-resistant Escherichia coli isolates from Irish cattle farms

This study describes the genotypic characteristics of a collection of 100 multidrug-resistant (MDR) Escherichia coli strains recovered from cattle and the farm environment in Ireland in 2007. The most prevalent antimicrobial resistance identified was to streptomycin (100%), followed by tetracycline (99%), sulfonamides (98%), ampicillin (82%), and neomycin (62%). Resistance was mediated predominantly by strA-strB (92%), tetA (67%), sul2 (90%), bla(TEM) (79%), and aphA1 (63%) gene markers, respectively. Twenty-seven isolates harbored a class 1 integrase (intI1), while qacEΔ1 and sul1 markers were identified in 25 and 26 isolates, respectively. The variable regions of these integrons contained aminoglycoside, trimethoprim, and β-lactam resistance determinants (aadA12, aadB-aadA1, bla(OXA-30)-aadA1, dfrA1-aadA1, dfrA7). Class 2 integrons were identified less frequently (4%) and contained the gene cassette array dfrA1-sat1-aadA1. Resistance to ampicillin, neomycin, streptomycin, sulfonamide, and tetracycline was associated with transferable high-molecular-weight plasmids, as demonstrated by conjugation assays. A panel of virulence markers was screened for by PCR, and genes identified included vt1, K5 in 2 isolates, papC in 10 isolates, and PAI IV(536) in 37 isolates. MDR commensal E. coli isolates from Irish cattle displayed considerable diversity with respect to the genes identified. Our findings highlight the importance of the commensal microflora of food-producing animals as a reservoir of transferable MDR.     

Comparative analysis of virulence genes, genetic diversity, and phylogeny of commensal and enterotoxigenic Escherichia coli isolates from weaned pigs

If the acquisition of virulence genes (VGs) for pathogenicity were not solely acquired through horizontal gene transfers of pathogenicity islands, transposons, and phages, then clonal clusters of enterotoxigenic Escherichia coli (ETEC) would contain few or even none of the VGs found in strains responsible for extraintestinal infections. To evaluate this possibility, 47 postweaning diarrhea (PWD) ETEC strains from different geographical origins and 158 commensal E. coli isolates from the gastrointestinal tracts of eight group-housed healthy pigs were screened for 36 extraintestinal and 18 enteric VGs using multiplex PCR assays. Of 36 extraintestinal VGs, only 8 were detected (fimH, traT, fyuA, hlyA, kpsMtII, k5, iha, and ompT) in the ETEC collection. Among these, hlyA (alpha-hemolysin) and iha (nonhemagglutinating adhesin) occurred significantly more frequently among the ETEC isolates than in the commensal isolates. Clustering analysis based on the VG profiles separated commensal and ETEC isolates and even differentiated serogroup O141 from O149. On the other hand, pulsed-field gel electrophoresis (PFGE) successfully clustered ETEC isolates according to both serotype and geographical origin. In contrast, the commensal isolates were heterogeneous with respect to both serotype and DNA fingerprint. This study has validated the use of VG profiling to examine pathogenic relationships between porcine ETEC isolates. The clonal relationships of these isolates can be further clarified by PFGE fingerprinting. The presence of extraintestinal VGs in porcine ETEC confirmed the hypothesis that individual virulence gene acquisitions can occur concurrently against a background of horizontal gene transfers of pathogenicity islands. Over time, this could enable specific clonotypes to respond to host selection pressure and to evolve into new strains with increased virulence.     

猪源大肠杆菌(ETEC、STEC、AEEC)毒力基因及其与O抗原型的关系

[目的]揭示从我国部分地区仔猪腹泻或水肿病病猪体内分离到的300个大肠杆菌分离株所属病原型(pathotype)、毒力基因及其与O血清型的关系.[方法]O血清型采用常规的凝集试验进行测定,毒力基因采用PCR方法检测.[结果]通过对这300个分离株的O血清型及其毒素、紧密素和黏附素基因进行鉴定,结果显示除50株未定型、17株自凝外,测定出233个分离株的血清型,这些分离株覆盖了45个血清型,其中以0149、0107、0139、093和091为主,共133株,占定型菌株的57.1%;拥有est Ⅰ、estⅡ、elt、stx2e和eae A基因的菌株分别为102(34.0%)、190(63.3%)、81(27.0%)、57(19.0%)和54(18.0%)株;分离株中有51株K88基因阳性(其中菌毛表达率为100%),75株F18基因阳性(其中菌毛表达率为50.7%),在K88菌株中,0149血清型与est Ⅰ或estⅡ elt密切相关,在F18菌株中,0107血清型与est Ⅰ或estⅡ、0139血清型与stx2e紧密相关.依其毒力特征可将这些分离株分为以下6种类型:ETEC、STEC、AEEC、ETEC/STEC、AEEC/ETEC和AEEC/ETEC/STEC,分别拥有190、24、36、32、17和1个菌株,占分离株的63.3%、8.0%、12.0%、10.7%、5.7%和0.3%.通过分析这些分离株的O血清型、毒素类型和黏附素型之间的相关性:猪源ETEC以0149、0107、093和098等血清型为主,0149:K88菌株主要与estⅡ或estⅡ elt肠毒素相关,0107:F18菌株主要与estⅡ相关,093和098血清型菌株主要与estⅡ肠毒素相关;STEC菌株以0139:F18血清型为主,拥有stx2e;AEEC菌株拥有紧密素,无明显优势血清型;ETEC/STEC菌株以0107:F18和0116:F18血清型为主,主要与est Ⅰ stx2e或estⅡ stx2e密切相关,ETEC/AEEC菌株以091和0107血清型为主,全部拥有肠毒素est Ⅰ和紧密素基因.[结论]我国至少存在6种病原型的猪肠道致病性大肠杆菌,其中ETEC为我国部分地区猪大肠杆菌病的主要病原,同时其病原型日益复杂.     

Antimicrobial resistance in generic Escherichia coli isolates from wild small mammals living in swine farm, residential, landfill, and natural environments in southern Ontario, Canada

To assess the impacts of different types of human activity on the development of resistant bacteria in the feces of wild small mammals, we compared the prevalences and patterns of antimicrobial resistance and resistance genes in generic Escherichia coli and Salmonella enterica isolates from fecal samples collected from wild small mammals living in four environments: swine farms, residential areas, landfills, and natural habitats. Resistance to antimicrobials was observed in E. coli isolates from animals in all environments: 25/52 (48%) animals trapped at swine farms, 6/69 (9%) animals trapped in residential areas, 3/20 (15%) animals trapped at landfills, and 1/22 (5%) animals trapped in natural habitats. Animals trapped on farms were significantly more likely to carry E. coli isolates with resistance to tetracycline, ampicillin, sulfisoxazole, and streptomycin than animals trapped in residential areas. The resistance genes sul2, aadA, and tet(A) were significantly more likely to be detected in E. coli isolates from animals trapped on farms than from those trapped in residential areas. Three S. enterica serotypes (Give, Typhimurium, and Newport) were recovered from the feces of 4/302 (1%) wild small mammals. All Salmonella isolates were pansusceptible. Our results show that swine farm origin is significantly associated with the presence of resistant bacteria and resistance genes in wild small mammals in southern Ontario, Canada. However, resistant fecal bacteria were found in small mammals living in all environments studied, indicating that environmental exposure to antimicrobials, antimicrobial residues, resistant bacteria, or resistance genes is widespread.     

Loss of virulence genes in Escherichia coli populations during manure storage on a commercial swine farm

Confined livestock production farms typically store their wastes prior to land application. Here, we employed three complementary approaches to evaluate changes in the population structure and stability of virulence genes in Escherichia coli during manure storage on a commercial farm that housed healthy swine. Isolates were genotyped by repetitive extragenic palindromic PCR using the BOXA1R primer and evaluated for the presence of selected virulence genes by PCR. Isolates obtained from the manure holding tank (n = 392) carried estB, fedA, stx(2e), astA, paa, aida-I, and sepA at lower frequencies than isolates obtained from fresh feces (n = 412). Fresh fecal material from the barn was added into diffusion chambers and immersed in the manure holding tank for 7 weeks. The fecal E. coli population was initially dominated by a single genotype, all isolates of which carried fedA and aida-I. After 7 weeks, a genotype that did not carry any virulence genes dominated the surviving population. In a second experiment, 48 fecal isolates of E. coli that varied in their genotypes and virulence gene complement were incubated in diffusion chambers in the manure holding tank for 3 weeks. Over 95% of the inoculum population carried at least one virulence gene, whereas after 3 weeks 90% of the recovered isolates carried no virulence genes. Taken together, these results indicate that during commercial manure storage, there was a significant reduction in the carriage of these virulence genes by E. coli. We propose that loss of virulence genes from enteric pathogens in the farm and in natural environments may, if generalized, contribute to the attenuation of a public health risk from contamination with agricultural wastes.     

Class 1 integrons, selected virulence genes, and antibiotic resistance in Escherichia coli isolates from the Minjiang River, Fujian Province, China

Widespread fecal pollution of surface waters in developing countries is a threat to public health and may represent a significant pathway for the global dissemination of antibiotic resistance. The Minjiang River drainage basin in Fujian Province is one of China's most intensive livestock and poultry production areas and is home to several million people. In the study reported here, Escherichia coli isolates (n = 2,788) were sampled (2007 and 2008) from seven surface water locations in the basin and evaluated by PCR for carriage of selected genes encoding virulence factors, primarily for swine disease. A subset of isolates (n = 500) were evaluated by PCR for the distribution and characteristics of class 1 integrons, and a subset of these (n = 200) were evaluated phenotypically for resistance to a range of antibiotics. A total of 666 (24%) E. coli isolates carried at least one of the virulence genes elt, fedA, astA, fasA, estA, stx(2e), paa, and sepA. Forty-one percent of the isolates harbored class 1 integrons, and these isolates had a significantly higher probability of resistance to tobramycin, cefoperazone, cefazolin, ciprofloxacin, norfloxacin, azitromycin, and rifampin than isolates with no class 1 integron detected. Frequencies of resistance to selected antibiotics were as high as or higher than those in fecal, wastewater, and clinical isolates in published surveys undertaken in China, North America, and Europe. Overall, E. coli in the Minjiang River drainage basin carry attributes with public health significance at very high frequency, and these data provide a powerful rationale for investment in source water protection strategies in this important agricultural and urban setting in China.     

Characterization of antimicrobial susceptibility and virulence genes of Salmonella serovars collected at a commercial turkey processing plant

Aims:  To determine the antimicrobial susceptibility profiles, distribution of class 1 integrons, virulence genes and genes encoding resistance to tetracycline ( tetA , tetC , tetD and tetE ) and streptomycin ( strA , strB and aadA1 ) in Salmonella recovered from turkeys.
Methods and Results:  The antimicrobial susceptibility of 80 isolates was determined using National Antimicrobial Resistance Monitoring System. The distribution of resistance genes, class 1 integrons and virulence genes was determined using PCR. Resistances to tetracycline (76·3%) and streptomycin (40%) were common. Sixty-two (77·5%) isolates displayed resistance against one or more antimicrobials and 33 were multi-drug resistant. tetA was detected in 72·5% of the isolates, while tetC , tetD and tetE were not detected. The strA and strB genes were detected in 73·8% of the isolates. Two isolates possessed class 1 integrons of 1 kb in size, containing the aadA1 gene conferring resistance to streptomycin and spectinomycin. Fourteen of the virulence genes were detected in over 80% of the isolates.
Conclusions:  This study shows that continuous use of tetracycline and streptomycin in poultry production selects for resistant strains. The Salmonella isolates recovered possess significant ability to cause human illness.
Significance and Impact of the Study:  Information from this study can be employed in guiding future strategies for the use of antimicrobials in poultry production.     

A high prevalence of antimicrobial resistant Escherichia coli isolated from pigs and a low prevalence of antimicrobial resistant E. coli from cattle and sheep in Great Britain at slaughter

The incidence of antimicrobial resistance and expressed and unexpressed resistance genes among commensal Escherichia coli isolated from healthy farm animals at slaughter in Great Britain was investigated. The prevalence of antimicrobial resistance among the isolates varied according to the animal species; of 836 isolates from cattle tested only 5.7% were resistant to one or more antimicrobials, while only 3.0% of 836 isolates from sheep were resistant to one or more agents. However, 92.1% of 2480 isolates from pigs were resistant to at least one antimicrobial. Among isolates from pigs, resistance to some antimicrobials such as tetracycline (78.7%), sulphonamide (66.9%) and streptomycin (37.5%) was found to be common, but relatively rare to other agents such as amikacin (0.1%), ceftazidime (0.1%) and coamoxiclav (0.2%). The isolates had a diverse range of resistance gene profiles, with tet(B), sul2 and strAB identified most frequently. Seven out of 615 isolates investigated carried unexpressed resistance genes. One trimethoprim-susceptible isolate carried a complete dfrA17 gene but lacked a promoter for it. However, in the remaining six streptomycin-susceptible isolates, one of which carried strAB while the others carried aadA, no mutations or deletions in gene or promoter sequences were identified to account for susceptibility. The data indicate that antimicrobial resistance in E. coli of animal origin is due to a broad range of acquired genes.     

Comparative Analysis of Virulence Genes, Genetic Diversity, and Phylogeny of Commensal and Enterotoxigenic Escherichia coli Isolates from Weaned Pigs

If the acquisition of virulence genes (VGs) for pathogenicity were not solely acquired through horizontal gene transfers of pathogenicity islands, transposons, and phages, then clonal clusters of enterotoxigenic Escherichia coli (ETEC) would contain few or even none of the VGs found in strains responsible for extraintestinal infections. To evaluate this possibility, 47 postweaning diarrhea (PWD) ETEC strains from different geographical origins and 158 commensal E. coli isolates from the gastrointestinal tracts of eight group-housed healthy pigs were screened for 36 extraintestinal and 18 enteric VGs using multiplex PCR assays. Of 36 extraintestinal VGs, only 8 were detected (fimH, traT, fyuA, hlyA, kpsMtII, k5, iha, and ompT) in the ETEC collection. Among these, hlyA (α-hemolysin) and iha (nonhemagglutinating adhesin) occurred significantly more frequently among the ETEC isolates than in the commensal isolates. Clustering analysis based on the VG profiles separated commensal and ETEC isolates and even differentiated serogroup O141 from O149. On the other hand, pulsed-field gel electrophoresis (PFGE) successfully clustered ETEC isolates according to both serotype and geographical origin. In contrast, the commensal isolates were heterogeneous with respect to both serotype and DNA fingerprint. This study has validated the use of VG profiling to examine pathogenic relationships between porcine ETEC isolates. The clonal relationships of these isolates can be further clarified by PFGE fingerprinting. The presence of extraintestinal VGs in porcine ETEC confirmed the hypothesis that individual virulence gene acquisitions can occur concurrently against a background of horizontal gene transfers of pathogenicity islands. Over time, this could enable specific clonotypes to respond to host selection pressure and to evolve into new strains with increased virulence.     

Genotypic and Phenotypic Profiles of Enterotoxigenic <Emphasis Type="Italic">Escherichia coli</Emphasis> Associated with Acute Diarrhea in Tunis,Tunisia

No past studies of acute diarrhea in Tunisia have examined the phenotypic and genotypic profiles of enterotoxigenic Escherichia coli (ETEC) isolates. We determined 65 ETEC isolates derived from a total of 327 E. coli isolates collected from a previous study (acute diarrheal and healthy persons, children and adults n = 214) and 32 E. coli isolates derived from an acute diarrheal outbreak in Kabaria-Ennour city, Tunis. All E. coli isolates were screened by polymerase chain reaction (PCR) for ETEC virulence genes: sta (heat-stable toxin gene) and elt (heat-labile toxin gene). Seventy-two percent (47 of 65) of ETEC strains expressed the sta gene only, 21.5% (14 of 65) expressed the elt gene and 6.1% (4 of 65) expressed both genes. For the outbreak isolates, the elt gene was predominant (10 isolates out of 14). Ganylioside GM1 enzyme-linked immunosorbent assay (GM1-ELISA) was used to validate the PCR results and this was confirmed by dot blot assay. The same results were obtained. The most common colonization factors (CFs) were CFA/I (44.6%) and coli surface antigen 6 (CS6) (11%), and 44.6% of the isolates showed no association with either CFAs. Resistance of ETEC isolates to tetracycline (38.5%), streptomycin (26%), and beta-lactam agents (ticarcillin 26%, amoxicillin 24.6%, cephalotin 21.5%) was common. Regarding serotypes, the majority of ETEC isolates serotyped as O86:H(-) (n = 16), O128:H2 (n = 11), and O127:H21 (n = 10). Other serotypes found were O111:H(-) (n = 6) and O126: H(-) (n = 5). DNA macrorestriction fragment analysis by pulsed-field gel electrophoresis (PFGE) using the XbaI enzyme was conducted to investigate the epidemiological clonal relationship among ETEC isolates. Major patterns were identified among which some of outbreak ETEC isolates belonged. These data suggest that a proportion of acute diarrhea in Tunis represents the confluence of small epidemics by clonality-related ETEC isolates that are transiently introduced or that persist in our community.     

Insights into the environmental resistance gene pool from the genome sequence of the multidrug-resistant environmental isolate Escherichia coli SMS-3-5

The increasing occurrence of multidrug-resistant pathogens of clinical and agricultural importance is a global public health concern. While antimicrobial use in human and veterinary medicine is known to contribute to the dissemination of antimicrobial resistance, the impact of microbial communities and mobile resistance genes from the environment in this process is not well understood. Isolated from an industrially polluted aquatic environment, Escherichia coli SMS-3-5 is resistant to a record number of antimicrobial compounds from all major classes, including two front-line fluoroquinolones (ciprofloxacin and moxifloxacin), and in many cases at record-high concentrations. To gain insights into antimicrobial resistance in environmental bacterial populations, the genome of E. coli SMS-3-5 was sequenced and compared to the genome sequences of other E. coli strains. In addition, selected genetic loci from E. coli SMS-3-5 predicted to be involved in antimicrobial resistance were phenotypically characterized. Using recombinant vector clones from shotgun sequencing libraries, resistance to tetracycline, streptomycin, and sulfonamide/trimethoprim was assigned to a single mosaic region on a 130-kb plasmid (pSMS35_130). The remaining plasmid backbone showed similarity to virulence plasmids from avian-pathogenic E. coli (APEC) strains. Individual resistance gene cassettes from pSMS35_130 are conserved among resistant bacterial isolates from multiple phylogenetic and geographic sources. Resistance to quinolones was assigned to several chromosomal loci, mostly encoding transport systems that are also present in susceptible E. coli isolates. Antimicrobial resistance in E. coli SMS-3-5 is therefore dependent both on determinants acquired from a mobile gene pool that is likely available to clinical and agricultural pathogens, as well, and on specifically adapted multidrug efflux systems. The association of antimicrobial resistance with APEC virulence genes on pSMS35_130 highlights the risk of promoting the spread of virulence through the extensive use of antibiotics.     

Studies on diarrheagenic Escherichia coli isolated from children with diarrhea in Myanmar

Escherichia coli isolates from 217 children in Myanmar with diarrhea were investigated for the presence of virulence genes related to diarrhea by colony hybridization and PCR. The genes examined were lt, stI, stII, stx1, stx2, eae, bfp, pCVD (which is the representative gene of plasmid of pCVD of EAEC), and ial (which is invasion-associated locus of the invasion plasmid found in EIEC). Isolates from 47 of 217 children (21.7%) possessed virulence genes characteristic of diarrheagenic E. coli. No instance was found of co-existence of different E. coli strains with different virulence genes in the same patient. Diarrheagenic E. coli are currently classified into five categories based on their virulence markers: ETEC, EHEC, EPEC, EAEC, and EIEC. Of the 47 isolates examined, 30 were EAEC, 12 were EPEC and 5 were ETEC. Susceptibility tests for antimicrobial agents showed that almost all diarrheagenic isolates were resistant to penicillin, tetracycline and streptomycin. However, the majority of strains were sensitive to cephalexin, nalidixic acid and norfloxacin. In particular, 42 of the 47 isolates were sensitive to norfloxacin, which is a fluoroquinolone. This study shows EAEC and EPEC are responsible for sporadic diarrhea in Myanmar and fluoroquinolones appear to be effective in the treatment of these patients.     

Characterization of antibiotic-resistant and potentially pathogenic Escherichia coli from soil fertilized with litter of broiler chickens fed antimicrobial-supplemented diets

The objective of this study was to characterize antimicrobial resistance and virulence determinants of Escherichia coli from soil amended with litter from 36-day-old broiler chickens ( Gallus gallus domesticus ) fed with diets supplemented with a variety of antimicrobial agents. Soil samples were collected from plots before and periodically after litter application in August to measure E. coli numbers. A total of 295 E.?coli were isolated from fertilized soil samples between August and March. Antibiotic susceptibility was determined by Sensititre, and polymerase chain reaction was performed to detect the presence of resistance and virulence genes. The results confirmed that E.?coli survived and could be quantified by direct plate count for at least 7 months in soil following litter application in August. The effects of feed supplementation were observed on E.?coli numbers in November and January. Among the 295 E.?coli, the highest antibiotic resistance level was observed against tetracycline and β-lactams associated mainly with the resistance genes tetB and bla(CMY-2), respectively. Significant treatment effects were observed for phylogenetic groups, antibiotic resistance profiles, and virulence gene frequencies. Serotyping, phylogenetic grouping, and pulsed-field gel electrophoresis confirmed that multiple-antibiotic-resistant and potentially pathogenic E.?coli can survive in soil fertilized with litter for several months regardless of antimicrobials used in the feed.     

Genes coding for Shiga-like toxin and heat-stabile enterotoxin in porcine strains of Escherichia coli

Besides diarrheagenic enterotoxigenic Escherichia coli (ETEC) that produce classical heat stable and/or heat labile enterotoxins (STs, LTs) and the class of Shiga-like toxin-producing entero-hemorrhagic E. coli (EHEC), a new category of E. coli is defined sharing similarities with ETEC and EHEC. DNA hybridization studies indicate that some E. coli serovars from porcine origin harbor genes encoding cytotonic ST and cytotoxic Shiga-like toxin. The presence of two potent toxins might contribute to the virulence of such strains and should be taken into consideration when bio-assays are performed.