Sewage samples from seven locations in the United States were analyzed for Escherichia coli isolates which were resistant to trimethoprim-sulfamethoxazole (SXT). The prevalence rate of SXT resistant organisms varied between the different geographical locales. The majority of th...
Morcatti Coura, Fernanda; Diniz, Soraia de Araújo; Silva, Marcos Xavier; Mussi, Jamili Maria Suhet; Barbosa, Silvia Minharro; Lage, Andrey Pereira; Heinemann, Marcos Bryan
This study analyzes the occurrence and distribution of phylogenetic groups of 391 strains of Escherichia coli isolated from poultry, cattle, and water buffalo. The frequency of the phylogroups was A = 19%, B1 = 57%, B2 = 2.3%, C = 4.6%, D = 2.8%, E = 11%, and F = 3.3%. Phylogroups A (P < 0.001) and F (P = 0.018) were associated with E. coli strains isolated from poultry, phylogroups B1 (P < 0.001) and E (P = 0.002) were associated with E. coli isolated from cattle, and phylogroups B2 (P = 0.003) and D (P = 0.017) were associated with E. coli isolated from water buffalo. This report demonstrated that some phylogroups are associated with the host analyzed and the results provide knowledge of the phylogenetic composition of E. coli from domestic animals. PMID:26421310
The increase in resistance rates to trimehtoprim-sulfamethoxazole (TMP/SMX) in isolates of Escherichia coli has become a matter of increasing concern. This has been particularly true in reference to community acquired urinary tract infections (UTI). This study utilized sewage i...
Martínez, Aida Juliana; Bossio, Carolina Paba; Durango, Adriana Coral; Vanegas, Maria Consuelo
The aim of this study was to characterize Shiga toxigenic Escherichia coli (STEC) by PCR using strains isolated from ham, beef, and cattle in Colombia. A total of 189 E. coli strains were tested for the presence of the uidA, stx1, and stx2 genes, and identification was confirmed by the automated PCR BAX system for E. coli O157:H7. Genes encoding Shiga-like toxins (stx) were found in eight (6.06%) of 132 strains previously isolated from minced beef; four (50%) of these strains yielded amplification products for both toxin genes (stx1 and stx2), and four (50%) yielded products only for the stx2 toxin. None of the strains analyzed were positive by PCR for the presence of the single base-pair mutation in the uidA gene from E. coli O157:H7; these results were confirmed by the BAX system analysis. A multiplex PCR assay was standardized for the three genes. Results from this study confirmed previous data about the low prevalence of E. coli O157:H7 and Shiga-like toxins in Colombia and is the first known report of the prevalence of non-O157 enterohemorrhagic E. coli in this country.
Abduzaimovic, Amila; Aljicevic, Mufida; Rebic, Velma; Vranic, Sabina Mahmutovic; Abduzaimovic, Kadrija; Sestic, Sabina
Objectives: The aim of this study was to examine the presence of antimicrobial resistance / susceptibility strains of Escherichia coli in inpatients and outpatients. Materials and methods: It is a retrospective study carried out at the Department of Microbiology, Parasitology and Virology Faculty of Medicine, University of Sarajevo. In cooperation with the Microbiological laboratory of the Cantonal Hospital Zenica and the Microbiological laboratory of the General Hospital Tesanj, 3863 urine samples were processed in the period from March 1st to March 31st 2016. Results: Our study showed that E. coli had the highest antimicrobial resistance to trimethoprim / sulfamethoxazole (38.61%), followed by amoxicillin / clavulanic acid (19.62%), ciprofloxacin (9.49%), gentamicin (8.86%), cephalexin (8.23%), nitrofurantoin (8.23%), cefuroxime (7.52%), ceftazidime (6.33%), cefuroxime (89.87%), amikacin (4.43%). Conclusions: The isolated strains of E. coli showed the highest resistance to trimethoprim / sulfamethoxazole and amoxicillin / clavulanic acid. The isolated strains of E. coli showed the greatest susceptibility to amikacin and ceftazidime. Gender distribution of positive E. coli isolates showed statistically significant differences in favor of females. PMID:28144190
Hasvold, J; Bradford, L; Nelson, C; Harrison, C; Attar, M; Stillwell, T
Neonatal sepsis is a significant cause of morbidity and mortality among term and preterm infants. Ampicillin and gentamicin are standard empiric therapy for early onset sepsis. Four cases of neonatal sepsis secondary to Escherichia coli (E. coli) found to be gentamicin resistant occurred within a five week period in one neonatal intensive care unit (NICU). To determine whether these cases could be tied to a single vector of transmission, and to more broadly evaluate the incidence of gentamicin resistant strains of E. coli in the neonatal population at our institution compared to other centers, we reviewed the charts of the four neonates (Infants A through D) and their mothers. The E. coli isolates were sent for Pulse Field Gel Electrophoresis (PFGE) to evaluate for genetic similarity between strains. We also reviewed all positive E. coli cultures from one NICU over a two year period. Infants A and B had genetically indistinguishable strains which matched that of urine and placental cultures of Infant B's mother. Infant C had a genetically distinct organism. Infant D, the identical twin of Infant C, did not have typing performed. Review of all cultures positive for E. coli at our institution showed a 12.9 percent incidence of gentamicin-resistance. A review of other studies showed that rates of resistance vary considerably by institution. We conclude that gentamicin-resistant E. coli is a relatively uncommon cause of neonatal sepsis, but should remain a consideration in patients who deteriorate despite initiation of empiric antibiotics.
Dou, Xinhong; Gong, Jiansen; Han, Xiangan; Xu, Ming; Shen, Haiyu; Zhang, Di; Zhuang, Linlin; Liu, Jiasheng; Zou, Jianmin
In order to investigate the biological characteristics of avian pathogenic Escherichia coli (APEC) isolated in eastern China, a total of 243 isolates were isolated from diseased poultry on different farms during the period from 2007 to 2014. These isolates were characterized for serogroups (polymerase chain reaction and agglutination), the presence of virulence-associated genes (fimC, iss, ompA, fyuA, stx2f, iroC, iucD, hlyE, tsh, cvaC, irp2, and papC) and class I integrons (polymerase chain reaction), drug susceptibilities (disk diffusion method) and the biofilm-forming abilities (semi-quantitative method). The results showed that the most predominant serogroups were O78 (87 isolates, 35.8%) and O2 (35 isolates, 14.4%). Gene profiling found that fimC and ompA were frequently distributed among the isolates and that 77.4% of the isolates were positive for class 1 integrons. Overall, isolates displayed resistance to tetracycline (97.5%), nalidixic acid (82.3%), ampicillin (81.1%), sulphafurazole (80.7%), streptomycin (79.0%), trimethoprim (78.2%) and cotrimoxazole (78.2%). Multiple-drug resistance was exhibited in 80.3% of the isolates, and the presence of class 1 integrons is associated with multidrug resistance. Finally, 151 isolates had the ability to form biofilms in vitro, and drug resistance seemed relative to biofilm-forming abilities.
USA Abstract Introduction: One approach to control enterotoxigenic Escherichia coli (ETEC) infections has been to develop vaccines focused on...results show a lack of clonality among Egypt CS6 E. coli isolates and supports the use and the further research on vaccines targeting this cell surface...has received considerable attention as a target for vaccine development [11-14]. CS6 is immunogenic in humans both after natural infection and
Maslow, J N; Whittam, T S; Gilks, C F; Wilson, R A; Mulligan, M E; Adams, K S; Arbeit, R D
The clonal relationships among 187 bloodstream isolates of Escherichia coli from 179 patients at Boston, Mass., Long Beach, Calif., and Nairobi, Kenya, were determined by multilocus enzyme electrophoresis (MLEE), analysis of polymorphisms associated with the ribosomal operon (ribotyping), and serotyping. MLEE based on 20 enzymes resolved 101 electrophoretic types (ETs), forming five clusters; ribotyping resolved 56 distinct patterns concordant with the analysis by MLEE. The isolates at each study site formed a genetically diverse group and demonstrated similar clonal structures, with the same small subset of lineages accounting for the majority of isolates at each site. Moreover, two ribotypes accounted for approximately 30% of the isolates at each study site. One cluster contained the majority (65%) of isolates and, by direct comparison of the ETs and ribotypes of individual isolates, was genetically indistinguishable from the largest cluster for each of two other collections of E. coli causing pyelonephritis and neonatal meningitis (R. K. Selander, T. K. Korhonen, V. Väisänen-Rhen, P. H. Williams, P. E. Pattison, and D. A. Caugent, Infect. Immun. 52:213-222, 1986; M. Arthur, C. E. Johnson, R. H. Rubin, R. D. Arbeit, C. Campanelli, C. Kim, S. Steinbach, M. Agarwal, R. Wilkinson, and R. Goldstein, Infect. Immun. 57:303-313, 1989), thus defining a virulent set of lineages. The isolates within these virulent lineages typically carried DNA homologous to the adhesin operon pap or sfa and the hemolysin operon hly and expressed O1, O2, O4, O6, O18, O25, or O75 antigens. DNA homologous to pap was distributed among isolates of each major cluster, whereas hly was restricted to isolates of two clusters, typically detected in pap-positive strains, and sfa was restricted to isolates of one cluster, typically detected in pap- and hly-positive strains. The occurrence of pap-positive isolates in the same geographically and genetically divergent lineages suggests that this
Goodsell, David S.
Diverse biological data may be used to create illustrations of molecules in their cellular context. I describe the scientific results that support a recent textbook illustration of an "Escherichia coli cell". The image magnifies a portion of the bacterium at one million times, showing the location and form of individual macromolecules. Results…
Investigations were carried out into the susceptibility of a total of 223 strains of Escherichia coli to therapeutic agents with the employment of the disk diffusion method. The organisms were isolated from internal organs and bone marrow of birds died of coli septicaemia. The serologic classification of the strains was defined with the use of 88 anti-group OK-agglutinating sera obtained through hyperimmunization of rabbits with the following Escherichia coli serotypes: 01-063, 068, 071, 073, 075, 078, 086, 0101, 0103, 0111-0114, 0119, 0124, 0129, 0135-0141, 0146, 0147, and 0149. It was found that serologically the strains referred as follows: 01-41 strains, 02-70 strains, 04-2 strains, 08-3 strains, 026-1 strain, 078-70 strains, 0111-2 strains, 0103-1 strain, 0141-1 strain. The number of untypable strains amounted to 32. Highest number of strains proved sensitive to colistin--96.06%, the remaining drugs following in a descending order: flumequine--95.65%, apramycin - 95.5%, gentamycin--93.72%, amoxicillin--93,8%, amikacin--88.57%, carbenicillin--86.88%, furazolidone--83,13%, and kanamycin--79.36%. High was the percent of strains resistant to tetracycline--66.17%, spectinomycin--61.67%, ampicillin--51.12%, chloramphenicol--50.23%, and streptomycin--44.84%.
Oteo, Jesús; Lázaro, Edurne; Cuevas, Óscar; García-Cobos, Silvia; Pérez-Vázquez, María; de Abajo, F. J.
To determine the evolution and trends of amoxicillin–clavulanic acid resistance among Escherichia coli isolates in Spain, we tested 9,090 blood isolates from 42 Spanish hospitals and compared resistance with trends in outpatient consumption. These isolates were collected by Spanish hospitals that participated in the European Antimicrobial Resistance Surveillance System network from April 2003 through December 2006. PMID:18680650
Bidet, Philippe; Mariani-Kurkdjian, Patricia; Grimont, Francine; Brahimi, Naïma; Courroux, Céline; Grimont, Patrick; Bingen, Edouard
Forty-seven non-epidemic Escherichia coli O157 : H7 isolates causing haemolytic uraemic syndrome in France were characterized. The isolates clustered into 36 clones using PFGE typing. All the isolates harboured eae and one or more copies of stx2 and belonged to phylogenetic group D. Nine per cent were resistant to amoxicillin.
Oteo, Jesús; Campos, José; Lázaro, Edurne; Cuevas, Oscar; García-Cobos, Silvia; Pérez-Vázquez, María; de Abajo, F J
To determine the evolution and trends of amoxicillin-clavulanic acid resistance among Escherichia coli isolates in Spain, we tested 9,090 blood isolates from 42 Spanish hospitals and compared resistance with trends in outpatient consumption. These isolates were collected by Spanish hospitals that participated in the European Antimicrobial Resistance Surveillance System network from April 2003 through December 2006.
Medon, P P; Lanser, J A; Monckton, P R; Li, P; Symons, R H
Commercially available kits containing alkaline phosphatase-labeled oligonucleotide probes for Escherichia coli heat-stable enterotoxins (STI-H, STI-P, and STII) and the heat-labile enterotoxin were compared with bioassays and radiolabeled recombinant DNA probes to identify enterotoxigenic E. coli from 100 clinical isolates. There was very good agreement between the three methods. PMID:3053766
Ikedo, M; Komatsu, O; Hara-Kudo, Y; Yamamoto, S; Kumagai, S
Agar media for isolation of enterohaemorrhagic Escherichia coli (EHEC) have been developed primarily for E. coli O157, because this bacterium has most frequently caused EHEC infection. However, there have been few studies for isolation of other serotypes of EHEC, and media appropriate for isolation of such organisms, especially from food samples, are not yet available. Among such serotypes, E. coli O26 has often been isolated from clinical specimens from patients and animals, but not from food samples in outbreaks, because of lack of an appropriate method for isolation. In this study, we tried to develop a new chromogenic agar medium for selective isolation of E. coli O26 using the characteristics of E. coli O26. Fifteen strains of E. coli O26, 11 strains of E. coli O157 and 36 strains of other sero-types E. coli were tested for fermentation of rhamnose, cellobiose, dulcitol, salicin, raffinose, sorbitol, sucrose, lactose, mannitol, arabinose, maltose, xylose and glucose. Rhamnose was fermented by all E. coli strains except for E. coli O26. The other substrates were not effective for differentiating E. coli O26 from the other strains of E. coli. Thus the medium containing rhamnose and 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside, which is a substrate of beta-galactosidase specific to coliforms, produced a color of E. coli O26 colonies different from colors of the other bacteria. Furthermore, cefixime and sodium tellulite were added to the composition of the medium for gaining higher selectivity.
Cunha, Francisco Afrânio; Maia, Kamila Rocha; Mallman, Eduardo José Jucá; Cunha, Maria da Conceição Dos Santos Oliveira; Maciel, Antonio Auberson Martins; Souza, Ieda Pereira de; Menezes, Everardo Albuquerque; Fechine, Pierre Basílio Almeida
Nanotechnology can be a valuable ally in the treatment of infections. Silver nanoparticles (AgNPs) are structures that have antimicrobial activity. The aim of this study was to produce AgNPs by green methods, characterize these structures, and assess their antimicrobial activity against Escherichia coli associated with the antibiotic ciprofloxacin. AgNPs were characterized by spectroscopic and microscopic techniques. Antimicrobial activity was evaluated by the disk diffusion method against 10 strains of E. coli. The synthesized AgNPs showed a spherical shape and a size of 85.07 ± 12.86 nm (mean ± SD). AgNPs increased the activity of ciprofloxacin by 40% and may represent a new therapeutic option for the treatment of bacterial infections.
CUNHA, Francisco Afrânio; MAIA, Kamila Rocha; MALLMAN, Eduardo José Jucá; CUNHA, Maria da Conceição dos Santos Oliveira; MACIEL, Antonio Auberson Martins; de SOUZA, Ieda Pereira; MENEZES, Everardo Albuquerque; FECHINE, Pierre Basílio Almeida
SUMMARY Nanotechnology can be a valuable ally in the treatment of infections. Silver nanoparticles (AgNPs) are structures that have antimicrobial activity. The aim of this study was to produce AgNPs by green methods, characterize these structures, and assess their antimicrobial activity against Escherichia coli associated with the antibiotic ciprofloxacin. AgNPs were characterized by spectroscopic and microscopic techniques. Antimicrobial activity was evaluated by the disk diffusion method against 10 strains of E. coli. The synthesized AgNPs showed a spherical shape and a size of 85.07 ± 12.86 nm (mean ± SD). AgNPs increased the activity of ciprofloxacin by 40% and may represent a new therapeutic option for the treatment of bacterial infections. PMID:27680178
Kikuchi, Y; Kojima, H; Tanaka, T; Takatsuka, Y; Kamio, Y
We report here on the existence of a new gene for lysine decarboxylase in Escherichia coli K-12. The hybridization experiments with a cadA probe at low stringency showed that the homologous region of cadA was located in lambda Kohara phage clone 6F5 at 4.7 min on the E. coli chromosome. We cloned the 5.0-kb HindIII fragment of this phage clone and sequenced the homologous region of cadA. This region contained a 2,139-nucleotide open reading frame encoding a 713-amino-acid protein with a calculated molecular weight of 80,589. Overexpression of the protein and determination of its N-terminal amino acid sequence defined the translational start site of this gene. The deduced amino acid sequence showed 69.4% identity to that of lysine decarboxylase encoded by cadA at 93.7 min on the E. coli chromosome. In addition, the level of lysine decarboxylase activity increased in strains carrying multiple copies of the gene. Therefore, the gene encoding this lysine decarboxylase was designated Idc. Analysis of the lysine decarboxylase activity of strains containing cadA, ldc, or cadA ldc mutations indicated that ldc was weakly expressed under various conditions but is a functional gene in E. coli. PMID:9226257
Cheong, Youjin; Jin Kim, Young; Kang, Heeyoon; Choi, Samjin; Joo Lee, Hee
We introduce a label-free spectroscopic method to classify subtypes of quinolone-nonsusceptible Escherichia coli (E. coli) isolates obtained from human blood cultures. Raman spectroscopy with a 30-nm gold-deposited, surface-enhanced Raman scattering (SERS) substrate was used to evaluate three multilocus sequencing typing (MLST)-predefined groups including E. coli ATCC25922, E. coli ST131:O75, and E. coli ST1193:O25b. Although there was a coffee-ring effect, the ring zone was selected at the ideal position to screen E. coli isolates. Strong Raman peaks were present at 1001-1004 cm(-1) (CC aromatic ring breathing stretching vibrational mode of phenylalanine), 1447-1448 cm(-1) (CH2 scissoring deformation vibrational mode), and 1667 cm(-1) (amide I α-helix). Although the three MLST-predefined E. coli isolates had similar Raman spectral patterns, a support vector machine (SVM) learning algorithm-assisted principal component analysis (PCA) analysis had superior performance in detecting the presence of quinolone-nonsusceptible E. coli isolates as well as classifying similar microbes, such as quinolone-nonsusceptible E. coli ST131:O75 and E. coli ST1193:O25b isolates. Therefore, this label-free and nondestructive technique is likely to be useful for clinically diagnosing quinolone-nonsusceptible E. coli isolates with the MLST method.
Lafolie, Jeremy; Nicolas-Chanoine, Marie-Hélène; Grenouillet, Frédéric; Hocquet, Didier; Bertrand, Xavier
The prevalence of Escherichia coli sequence type 131 (ST131) and its subclone H30 was assessed among a collection of 490 E. coli isolated in 2013 in a French university hospital. The prevalence of ST131 was 4% among bloodstream isolates (regardless of antimicrobial resistance) and 17.2% among extended-spectrum β-lactamase (ESBL)-producing isolates. Although a much lower prevalence of ST131 was found among bloodstream E. coli isolates compared with other countries, a large predominance of H30 subclone within ST131 was confirmed. It was also confirmed that, among ESBL-producing E. coli, ST131 isolates were more frequently resistant to amoxicillin/clavulanic acid, ceftazidime, fluoroquinolones and aminoglycosides than non-ST131 isolates.
Amézquita-Montes, Zorangel; Tamborski, Maria; Kopsombut, Usa G; Zhang, Chengxian; Arzuza, Octavio S; Gómez-Duarte, Oscar G
Foodborne pathogens are a leading cause of mild-to-severe gastrointestinal illnesses worldwide. Escherichia coli pathotypes have been known to cause gastrointestinal illnesses in children less than 5 years old in Colombia. However, insufficient information is available on the prevalence of E. coli contamination of food products and the kind of E. coli food product reservoirs. The two objectives of this study were designed to address this issue. The first objective was to ascertain coliform, E. coli, and pathogenic E. coli contamination of food products readily available for human consumption in Cartagena, Colombia. The second objective was to evaluate the relationship between pathogenic E. coli isolated from food products and those isolated from cases of diarrhea in children. Food product samples consisting of pasteurized milk, unpasteurized fruit juice, ground beef, cheese, and vegetables were obtained at four retail stores. The food samples were cultured in liquid media and tested for the presence of coliforms and E. coli. E. coli isolates were tested by polymerase chain reaction for the presence of pathogenic E. coli. Coliforms, E. coli, and E. coli intestinal pathotypes contamination were detected in 88.4%, 53%, and 2.1% of food product samples, respectively. Ground beef and cheese were the only food samples contaminated with E. coli intestinal pathotypes including enteropathogenic (EPEC), Shiga toxin-producing (STEC), and enterotoxigenic E. coli (ETEC). Closed multilocus sequencing typing relationships between diarrheagenic E. coli isolates from food products and from individuals with diarrhea suggest that food products readily available at public markets in Cartagena can transmit ETEC and possibly EPEC and STEC. We demonstrated that a high proportion of food products for human consumption available at public markets in Cartagena are contaminated with coliforms, E. coli, and E. coli intestinal pathogens. Furthermore, food products containing E. coli intestinal
Amézquita-Montes, Zorangel; Tamborski, Maria; Kopsombut, Usa G.; Zhang, Chengxian; Arzuza, Octavio S.
Abstract Foodborne pathogens are a leading cause of mild-to-severe gastrointestinal illnesses worldwide. Escherichia coli pathotypes have been known to cause gastrointestinal illnesses in children less than 5 years old in Colombia. However, insufficient information is available on the prevalence of E. coli contamination of food products and the kind of E. coli food product reservoirs. The two objectives of this study were designed to address this issue. The first objective was to ascertain coliform, E. coli, and pathogenic E. coli contamination of food products readily available for human consumption in Cartagena, Colombia. The second objective was to evaluate the relationship between pathogenic E. coli isolated from food products and those isolated from cases of diarrhea in children. Food product samples consisting of pasteurized milk, unpasteurized fruit juice, ground beef, cheese, and vegetables were obtained at four retail stores. The food samples were cultured in liquid media and tested for the presence of coliforms and E. coli. E. coli isolates were tested by polymerase chain reaction for the presence of pathogenic E. coli. Coliforms, E. coli, and E. coli intestinal pathotypes contamination were detected in 88.4%, 53%, and 2.1% of food product samples, respectively. Ground beef and cheese were the only food samples contaminated with E. coli intestinal pathotypes including enteropathogenic (EPEC), Shiga toxin–producing (STEC), and enterotoxigenic E. coli (ETEC). Closed multilocus sequencing typing relationships between diarrheagenic E. coli isolates from food products and from individuals with diarrhea suggest that food products readily available at public markets in Cartagena can transmit ETEC and possibly EPEC and STEC. We demonstrated that a high proportion of food products for human consumption available at public markets in Cartagena are contaminated with coliforms, E. coli, and E. coli intestinal pathogens. Furthermore, food products containing E. coli
Arivett, Brock A; Ream, Dave C; Fiester, Steven E; Kidane, Destaalem; Actis, Luis A
Members of the Escherichia coli bacterial family have been grouped as ESKAPE (Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species) pathogens because of their extensive drug resistance phenotypes and increasing threat to human health. The genomes of six extended-spectrum β-lactamase (ESBL)-producing E. coli strains isolated from wounded military personnel were sequenced and annotated.
Nayme, Kaotar; Barguigua, Abouddihaj; Bouchrif, Brahim; Karraouan, Bouchra; El Otmani, Fatima; Elmdaghri, Naima; Zerouali, Khalid; Timinouni, Mohammed
This study was conducted to assess the retail food as a possible vehicle for antimicrobial resistant, particularly quinolones resistant and pathogenic Escherichia coli. We determined the prevalence and characteristics of nalidixic acid (Nal) resistant E. coli isolates from diverse retail food samples. In all, 70 (28%) of 250 E. coli isolates studied were Nal-resistant E. coli and 91% of these were multi-drug resistant. Plasmid mediated quinolone resistance genes were identified in 32 isolates, including aac(6')-Ib-cr (n = 16), qnrS1 (n = 11) and qnrB19 (n = 7). Mutations in gyr A and par C genes were detected among 80% of the isolates, and the isolates showed substitution Ser83-Leu and Asp87-Asn in gyrA and Ser80-Ile in parC. In addition, three different gene cassettes were identified (aadA1, aadA7, aac(3)-Id) in 18%. Virulence-associated genes stx1, eae, sfa, hlyA and stx2 were found in six (8%), three (4%), two (3%), three (4%) and three (4%) isolates, respectively. E. coli isolates of phylogenetic group A were dominant (64%, 45/70). Pulsed field gel electrophoresis revealed none epidemiological relationship between these isolates. The results of this work report the higher frequency of Nal-resistant E. coli isolates from Moroccan retail food samples including MDR and pathogenic isolates.
Stannarius, C; Bürgi, E; Regula, G; Zychowska, M A; Zweifel, C; Stephan, R
Based on Directive (EC) No 99/2003, monitoring programs on the development of antimicrobial resistance in bacteria from livestock are implemented in many European countries. The aim of the present study was (i) to establish comprehensive baseline data on the antimicrobial resistance situation in Escherichia coli isolates obtained from healthy pigs (pooled fecal samples) originating from 60 Swiss pig-breeding farms, and (ii) to analyze differences in the resistance frequency between Escherichia coli isolates from weaned pigs and sows. Susceptibility testing (disc diffusion method) was performed on 429 isolates from weaned pigs and 431 isolates from sows. Overall, 17.7% of the isolates from weaned pigs and 22.5% of the Escherichia coli isolates from sows were susceptible to all antibiotics tested. Low resistance prevalence was found for amoxicillin, amoxicillin/clavulanic acid, ampicillin, cefquinome, ciprofloxacin, colistin, florfenicol, and gentamicin. The most frequently found resistances were against streptomycin (60.6% of the isolates from weaners and 64.3% of the isolates from sows), sulfonamide (51.5% and 26.9%), tetracycline (35.2% and 22.0%), and trimethoprim (27.5% and 11.1%). With exception of colistin, most resistances were found for those antibiotics commonly used on the farms. Except for ciprofloxacin and streptomycin, isolates from weaned pigs showed higher resistance prevalence than those from sows. This difference was significant for cefquinome, florfenicol, sulfonamide, tetracycline, and trimethoprim (p<0.05).
Senkel, I Arthur; Jolbitado, Beverly; Zhang, Yifan; White, David G; Ayers, Sherry; Meng, Jianghong
Contaminated apple cider has been implicated in several Escherichia coli O157:H7 outbreaks. In an attempt to investigate sources and modes of entry of E. coli into apple cider, samples of fresh apple, pomace, and cider and equipment and mill floor swabs were analyzed for standard plate counts (SPC), total coliforms (TC), fecal coliforms (FC), and E. coli. E. coli was isolated from 14 (33%) of 42 samples of bottled fresh cider, from food equipment in 6 (67%) of 9 mills, and from apples, pomace, or cider in 7 (78%) of 9 mills. Seventy-five E. coli isolates were further characterized for Shiga toxin-producing E. coli (STEC)-associated virulence factors, antimicrobial susceptibility, and pulsed-field gel electrophoresis (PFGE) type. No E. coli O157:H7 or other STEC was identified. Serotyping and PFGE revealed 64 distinct profiles, suggesting that recovered E. coli arose from multiple independent sources. However, on one occasion, E. coli isolated from the source apple sample was closely related to the E. coli identified in the finished cider sample. E. coli isolates were further tested for antimicrobial susceptibility to 17 antimicrobial agents of human and veterinary importance. Fourteen (19%) of the 75 isolates were resistant to at least one of the antimicrobial agents tested, and 9 (12%) were resistant to at least two of these agents. Of the resistant isolates recovered, 64% were resistant to tetracycline and 57% were resistant to streptomycin. Overall, the level of E. coli contamination in source apple samples did not differ significantly from those in samples of pomace, cider at the press, and cider entering the bottling tank; therefore, source apples cannot be dismissed as a potential contributor of E. coli to the cider-making process.
In recent years, non-O157 Shiga toxin-producing Escherichia coli(STEC) have become an emerging problem. Efforts have been devoted to facilitating and speeding their detection, however, their isolation from high background microbiota foods remains problematic. To solve this problem, immunomagnetic se...
Keyes, Kathleen; Hudson, Charlene; Maurer, John J.; Thayer, Stephan; White, David G.; Lee, Margie D.
Florfenicol is an antibiotic approved for veterinary use in cattle in the United States in 1996. Although this drug is not used in poultry, we have detected resistance to florfenicol in clinical isolates of avian Escherichia coli. Molecular typing demonstrated that the florfenicol resistance gene, flo, was independently acquired and is plasmid encoded. PMID:10639375
Yousfi, Massilia; Touati, Abdelaziz; Mairi, Assia; Brasme, Lucien; Gharout-Sait, Alima; Guillard, Thomas; De Champs, Christophe
The emergence and worldwide spread of carbapenemase-producing Enterobacteriaceae is of great concern to public health. The aim of this study was to investigate the occurrence of carbapenemase-producing Escherichia coli in companion animals in Algeria. Two hundred fecal samples were obtained from healthy and diseased dogs and cats in one veterinary office and private owners in Bejaia city, Algeria, during November 2014 to March 2015. Isolates were screened by polymerase chain reaction for the presence of carbapenemase, acquired plasmidic AmpC (pAmpC) and extended-spectrum beta-lactamase genes. Five carbapenemase-producing E. coli isolates were detected including four OXA-48-producing isolates and one isolate producing NDM-5. Coexpression of ESBL and pAmpC genes was observed in these isolates. Phylogenetic grouping revealed that these isolates belonged to A and D phylogroups. The results of this study show that carbapenemase-producing E. coli spread to the companion animals in Algeria.
CHANG, Shao-Kuang; LO, Dan-Yuan; WEI, Hen-Wei; KUO, Hung-Chih
This study determined the antimicrobial resistance profiles of Escherichia coli isolates from dogs with a presumptive diagnosis of urinary tract infection (UTI). Urine samples from 201 dogs with UTI diagnosed through clinical examination and urinalysis were processed for isolation of Escherichia coli. Colonies from pure cultures were identified by biochemical reactions (n=114) and were tested for susceptibility to 18 antimicrobials. The two most frequent antimicrobials showing resistance in Urinary E. coli isolates were oxytetracycline and ampicillin. Among the resistant isolates, 17 resistance patterns were observed, with 12 patterns involving multidrug resistance (MDR). Of the 69 tetracycline-resistant E. coli isolates, tet(B) was the predominant resistance determinant and was detected in 50.9% of the isolates, whereas the remaining 25.5% isolates carried the tet(A) determinant. Most ampicillin and/or amoxicillin-resistant E. coli isolates carried blaTEM-1 genes. Class 1 integrons were prevalent (28.9%) and contained previously described gene cassettes that are implicated primarily in resistance to aminoglycosides and trimethoprim (dfrA1, dfrA17-aadA5). Of the 44 quinolone-resistant E. coli isolates, 38 were resistant to nalidixic acid, and 6 were resistant to nalidixic acid, ciprofloxacin and enrofloxacin. Chromosomal point mutations were found in the GyrA (Ser83Leu) and ParC (Ser80Ile) genes. Furthermore, the aminoglycoside resistance gene aacC2, the chloramphenicol resistant gene cmlA and the florfenicol resistant gene floR were also identified. This study revealed an alarming rate of antimicrobial resistance among E. coli isolates from dogs with UTIs. PMID:25720807
de Cleber Jacob Silva Paula; Marin, José Moacir
From January to December 2006, 92 Escherichia coli isolates from 25 diarrheic dogs were analyzed by screening for the presence of adhesin-encoding genes (pap, sfa, afa), hemolysin and aerobactin genes. Virulence gene frequencies detected in those isolates were: 12% pap, 1% sfa, 10% hemolysin and 6.5% aerobactin. Ten isolates were characterized as extraintestinal pathogenic E. coli (ExPEC) strains; all showed a multidrug resistance phenotype that may represent a reason for concern due the risk of dissemination of antimicrobial resistant genes to the microbiota of human beings. PMID:24031253
Lutterodt, G; Foppen, J W A; Uhlenbrook, S
We present a new methodology to scale up bacteria transport experiments carried out in the laboratory to practical field situations. The key component of the methodology is to characterize bacteria transport not by a constant sticking efficiency, but by a range of sticking efficiency values determined from laboratory column experiments. In this study, initially, we harvested six Escherichia coli strains from springs in Kampala, the capital of Uganda, and then we carried out a number of experiments with 1.5m high columns of quartz sand with various sampling ports in order to determine the fraction of bacteria as a function of sticking efficiency. Furthermore, we developed a simple mathematical formulation, based on the steady-state analytical solution for the transport of mass in the subsurface, to arrive at bacteria concentrations as a function of transport distance. The results of the quartz sand column experiments indicated that the fractional bacteria mass and sticking efficiency of most of the strains we harvested could be adequately described by a power law. When applying the power distributions to the field situation in Kampala, we found that the transport distance required to reduce bacteria concentrations with five log units ranged from 1.5 to 23m, and this was up to three times more than when using a constant sticking efficiency. The methodology we describe is simple, can be carried out in a spreadsheet, and in addition to parameters describing transport, like pore water flow velocity and dispersion, only two constants are required, which define the relation between sticking efficiency and percentage of bacteria mass.
Jensen, Stina Rikke; Mirsepasi-Lauridsen, Hengameh Chloé; Thysen, Anna Hammerich; Brynskov, Jørn; Krogfelt, Karen A; Petersen, Andreas Munk; Pedersen, Anders Elm; Brix, Susanne
Escherichia coli (E. coli) may be implicated in the pathogenesis of inflammatory bowel disease (IBD), as implied from a higher prevalence of mucosa-associated E. coli in the gut of IBD-affected individuals. However, it is unclear whether different non-diarrheagenic E. coli spp. segregate from each other in their ability to promote intestinal inflammation. Herein we compared the inflammation-inducing properties of non-diarrheagenic LF82, 691-04A, E. coli Nissle 1917 (ECN) and eleven new intestinal isolates from different locations in five IBD patients and one healthy control. Viable E. coli were cultured with human monocyte-derived dendritic cells (moDCs) and monolayers of intestinal epithelial cells (IECs), followed by analysis of secreted cytokines, intracellular levels of reactive oxygen species and cellular death. The IBD-associated E. coli LF82 induced the same dose-dependent inflammatory cytokine profile as ECN and ten of the new E. coli isolates displayed as high level IL-12p70, IL-1β, IL-23 and TNF-α from moDCs irrespective of their site of isolation (ileum/colon/faeces), disease origin (diseased/non-diseased) or known virulence factors. Contrarily, 691-04A and one new IBD E. coli isolate induced a different cellular phenotype with enhanced killing of moDCs and IECs, coupled to elevated IL-18. The cytopathic nature of 691-04A and one other IBD E. coli isolate suggests that colonization with specific non-diarrheagenic E. coli could promote intestinal barrier leakage and profound intestinal inflammation, while LF82, ECN and the remaining non-diarrheagenic E. coli isolates hold notorious pro-inflammatory characteristics that can progress inflammation in case of intestinal barrier leakage.
Mohammed Hamzah, Aseel; Mohammed Hussein, Aseel; Mahmoud Khalef, Jenan
The isolation and characterization of Escherichia coli O157:H7 strains from 22 out of 174 fecal samples from petting zoo animals representing twenty-two different species (camel, lion, goats, zebra, bear, baboon monkey, Siberian monkey, deer, elk, llama, pony, horses, fox, kangaroo, wolf, porcupine, chickens, tiger, ostrich, hyena, dogs, and wildcats) were investigated. One petting Al-Zawraa zoological society of Baghdad was investigated for E. coli O157:H7 over a 16-month period that spanned two summer and two autumn seasons. Variation in the occurrence of E. coli O157:H7-positive petting zoo animals was observed, with animals being culture positive only in the summer months but not in the spring, autumn, or winter. E. coli O157:H7 isolates were distinguished by agglutination with E. coli O157:H7 latex reagent (Oxoid), identified among the isolates, which showed that multiple E. coli strains were isolated from one petting zoo animal, in which a single animal simultaneously shed multiple E. coli strains; E. coli O157:H7 was isolated only by selective enrichment culture of 2 g of petting zoo animal feces. In contrast, strains other than O157:H7 were cultured from feces of petting zoo animals without enrichment. PMID:24489514
Gopee, N V; Adesiyun, A A; Caesar, K
A longitudinal study was conducted of the prevalence and characteristics of Escherichia coli in mammals, birds, and reptiles housed at the Emperor Valley Zoo, Trinidad. During a 6-mo study period, swabs were obtained from fecal samples that were randomly collected from the enclosures of animals from these three taxonomic groups every 3 wk. With snakes, both cloacal and fecal swabs were obtained. Fecal and cloacal swabs were cultured for E. coli on eosin methylene blue agar. The production of mucoid colonies and hemolytic colonies and non-sorbitol fermenter status were identified. The occurrence of O157 strains was determined amongst E. coli isolates that were non-sorbitol fermenters, and the disc diffusion method was used to determine the antibiograms of isolates. The frequency of E. coli isolation was significantly higher in mammals compared with birds and reptiles. Overall, the frequencies of isolation of E. coli from omnivores. herbivores, and carnivores, 87.2%, 70.0%, and 57.3%, respectively, regardless of animal class, were significantly different. Most (99.6%) of the E. coli isolates tested for antibiotic sensitivity exhibited resistance to one or more of the eight antimicrobial agents used. The possession of phenotypic virulence markers by the E. coli isolates studied and the generally high resistance to antimicrobial agents may have health implications for the zoological collection.
Mitchell, Natalie M.; Johnson, James R.; Johnston, Brian; Curtiss, Roy
Chicken products are suspected as a source of extraintestinal pathogenic Escherichia coli (ExPEC), which causes diseases in humans. The zoonotic risk to humans from chicken-source E. coli is not fully elucidated. To clarify the zoonotic risk posed by ExPEC in chicken products and to fill existing knowledge gaps regarding ExPEC zoonosis, we evaluated the prevalence of ExPEC on shell eggs and compared virulence-associated phenotypes between ExPEC and non-ExPEC isolates from both chicken meat and eggs. The prevalence of ExPEC among egg-source isolates was low, i.e., 5/108 (4.7%). Based on combined genotypic and phenotypic screening results, multiple human and avian pathotypes were represented among the chicken-source ExPEC isolates, including avian-pathogenic E. coli (APEC), uropathogenic E. coli (UPEC), neonatal meningitis E. coli (NMEC), and sepsis-associated E. coli (SEPEC), as well as an undefined ExPEC group, which included isolates with fewer virulence factors than the APEC, UPEC, and NMEC isolates. These findings document a substantial prevalence of human-pathogenic ExPEC-associated genes and phenotypes among E. coli isolates from retail chicken products and identify key virulence traits that could be used for screening. PMID:25480753
Sáez-López, Emma; Guiral, Elisabet; Fernández-Orth, Dietmar; Villanueva, Sonia; Goncé, Anna; López, Marta; Teixidó, Irene; Pericot, Anna; Figueras, Francesc; Palacio, Montse; Cobo, Teresa; Bosch, Jordi; Soto, Sara M.
Vaginal Escherichia coli colonization is related to obstetric infections and the consequent development of infections in newborns. Ampicillin resistance among E. coli strains is increasing, which is the main choice for treating empirically many obstetric and neonatal infections. Vaginal E. coli strains are very similar to extraintestinal pathogenic E. coli with regards to the virulence factors and the belonging to phylogroup B2. We studied the antimicrobial resistance and the genetic virulence profile of 82 E. coli isolates from 638 vaginal samples and 63 isolated from endometrial aspirate, placental and amniotic fluid samples from pregnant women with obstetric infections. The prevalence of E. coli in the vaginal samples was 13%, which was significant among women with associated risk factors during pregnancy, especially premature preterm rupture of membranes (p<0.0001). Sixty-five percent of the strains were ampicillin-resistant. The E. coli isolates causing obstetric infections showed higher resistance levels than vaginal isolates, particularly for gentamicin (p = 0.001). The most prevalent virulence factor genes were those related to the iron uptake systems revealing clear targets for interventions. More than 50% of the isolates belonged to the virulent B2 group possessing the highest number of virulence factor genes. The ampicillin-resistant isolates had high number of virulence factors primarily related to pathogenicity islands, and the remarkable gentamicin resistance in E. coli isolates from women presenting obstetric infections, the choice of the most appropriate empiric treatment and clinical management of pregnant women and neonates should be carefully made. Taking into account host-susceptibility, the heterogeneity of E. coli due to evolution over time and the geographical area, characterization of E. coli isolates colonizing the vagina and causing obstetric infections in different regions may help to develop interventions and avoid the aetiological link
Muto, T; Arai, K; Miyai, M
To compare the isolating efficiency of diarrhea-causing Escherichia coli between Fluorocult agar plates, which reveal the beta-glucuronidase activity of E. coli, and a combination of SS and DHL agar plates, a total of 330 fecal specimens collected from outpatients were examined. Diarrhea-causing E. coli, identified by serological and toxigenic characters, were demonstrated in 52 samples. Among these specimens, 35 samples tested were positive on the Fluorocult agar plates, and 26 samples on the combination of SS and DHL agar plates. However, only 10 samples were positive on both the Fluorocult agar plates and the combination of SS and DHL agar plates. Thus, using Fluorocult agar plates for the isolation of diarrhea-causing E. coli in addition to the conventional SS and DHL agar plates will improve isolating efficacy.
Vogeleer, Philippe; Tremblay, Yannick D. N.; Jubelin, Grégory; Jacques, Mario
Forming biofilms may be a survival strategy of Shiga toxin-producing Escherichia coli to enable it to persist in the environment and the food industry. Here, we evaluate and characterize the biofilm-forming ability of 39 isolates of Shiga toxin-producing Escherichia coli isolates recovered from human infection and belonging to seropathotypes A, B, or C. The presence and/or production of biofilm factors such as curli, cellulose, autotransporter, and fimbriae were investigated. The polymeric matrix of these biofilms was analyzed by confocal microscopy and by enzymatic digestion. Cell viability and matrix integrity were examined after sanitizer treatments. Isolates of the seropathotype A (O157:H7 and O157:NM), which have the highest relative incidence of human infection, had a greater ability to form biofilms than isolates of seropathotype B or C. Seropathotype A isolates were unique in their ability to produce cellulose and poly-N-acetylglucosamine. The integrity of the biofilms was dependent on proteins. Two autotransporter genes, ehaB and espP, and two fimbrial genes, z1538 and lpf2, were identified as potential genetic determinants for biofilm formation. Interestingly, the ability of several isolates from seropathotype A to form biofilms was associated with their ability to agglutinate yeast in a mannose-independent manner. We consider this an unidentified biofilm-associated factor produced by those isolates. Treatment with sanitizers reduced the viability of Shiga toxin-producing Escherichia coli but did not completely remove the biofilm matrix. Overall, our data indicate that biofilm formation could contribute to the persistence of Shiga toxin-producing Escherichia coli and specifically seropathotype A isolates in the environment. PMID:26712549
Martin, I E; Tyler, S D; Tyler, K D; Khakhria, R; Johnson, W M
A total of 121 representative Escherichia coli O157:H7 and O157:NM (nonmotile) isolates were characterized by ribotype, phage type, verotoxin genotype, and genomic fingerprints generated by pulsed-field gel electrophoresis. Ribotyping was not able to discriminate between O157:H7 isolates, and phage typing and pulsed-field gel electrophoresis were the most valuable and discriminatory techniques. PMID:8904445
Vogeleer, Philippe; Tremblay, Yannick D N; Jubelin, Grégory; Jacques, Mario; Harel, Josée
Forming biofilms may be a survival strategy of Shiga toxin-producing Escherichia coli to enable it to persist in the environment and the food industry. Here, we evaluate and characterize the biofilm-forming ability of 39 isolates of Shiga toxin-producing Escherichia coli isolates recovered from human infection and belonging to seropathotypes A, B, or C. The presence and/or production of biofilm factors such as curli, cellulose, autotransporter, and fimbriae were investigated. The polymeric matrix of these biofilms was analyzed by confocal microscopy and by enzymatic digestion. Cell viability and matrix integrity were examined after sanitizer treatments. Isolates of the seropathotype A (O157:H7 and O157:NM), which have the highest relative incidence of human infection, had a greater ability to form biofilms than isolates of seropathotype B or C. Seropathotype A isolates were unique in their ability to produce cellulose and poly-N-acetylglucosamine. The integrity of the biofilms was dependent on proteins. Two autotransporter genes, ehaB and espP, and two fimbrial genes, z1538 and lpf2, were identified as potential genetic determinants for biofilm formation. Interestingly, the ability of several isolates from seropathotype A to form biofilms was associated with their ability to agglutinate yeast in a mannose-independent manner. We consider this an unidentified biofilm-associated factor produced by those isolates. Treatment with sanitizers reduced the viability of Shiga toxin-producing Escherichia coli but did not completely remove the biofilm matrix. Overall, our data indicate that biofilm formation could contribute to the persistence of Shiga toxin-producing Escherichia coli and specifically seropathotype A isolates in the environment.
Martin, E C; Gentry, T J
There are numerous regulatory-approved Escherichia coli enumeration methods, but it is not known whether differences in media composition and incubation conditions impact the diversity of E. coli populations detected by these methods. A study was conducted to determine if three standard water quality assessments, Colilert(®) , USEPA Method 1603, (modified mTEC) and USEPA Method 1604 (MI), detect different populations of E. coli. Samples were collected from six watersheds and analysed using the three enumeration approaches followed by E. coli isolation and genotyping. Results indicated that the three methods generally produced similar enumeration data across the sites, although there were some differences on a site-by-site basis. The Colilert(®) method consistently generated the least diverse collection of E. coli genotypes as compared to modified mTEC and MI, with those two methods being roughly equal to each other. Although the three media assessed in this study were designed to enumerate E. coli, the differences in the media composition, incubation temperature, and growth platform appear to have a strong selective influence on the populations of E. coli isolated. This study suggests that standardized methods of enumeration and isolation may be warranted if researchers intend to obtain individual E. coli isolates for further characterization.
Runnels, P L; Moon, H W
Previous reports have demonstrated that heat-stable (A-type) capsule on piliated enterotoxigenic Escherichia coli enhances colonization of enterotoxigenic E. coli in the small intestine and enhances virulence of enterotoxigenic E. coli. In this report, four encapsulated enterotoxigenic E. coli strains and one encapsulated nonenterotoxigenic strain of E. coli and their nonencapsulated mutants were tested for adhesion to isolated intestinal epithelial cells or brush borders from neonatal pigs. The enterotoxigenic E. coli also expressed the K99 pilus antigen. The nonencapsulated mutants of the four enterotoxigenic E. coli adhered in higher numbers than did the encapsulated parental strains. Both the encapsulated and nonencapsulated forms of enterotoxigenic E. coli 431 grown at 18 degrees C (K99 production suppressed) adhered poorly to the isolated cells. The nonenterotoxigenic E. coli 1793 which does not express K99 antigen also adhered poorly in both encapsulated and nonencapsulated forms. Fab fragments of anticapsular immunoglobulin G failed to block the effect of capsule on adherence of strain 431. The results indicated that K99 was the principal mediator of in vitro adhesion of the enterotoxigenic E. coli strains and that capsule impedes the in vitro adhesion. They also suggested that the capsular enhancement of colonization by such strains in vivo probably is by some mechanism other than enhanced adhesion to epithelium. PMID:6147310
Runnels, P L; Moon, H W
Previous reports have demonstrated that heat-stable (A-type) capsule on piliated enterotoxigenic Escherichia coli enhances colonization of enterotoxigenic E. coli in the small intestine and enhances virulence of enterotoxigenic E. coli. In this report, four encapsulated enterotoxigenic E. coli strains and one encapsulated nonenterotoxigenic strain of E. coli and their nonencapsulated mutants were tested for adhesion to isolated intestinal epithelial cells or brush borders from neonatal pigs. The enterotoxigenic E. coli also expressed the K99 pilus antigen. The nonencapsulated mutants of the four enterotoxigenic E. coli adhered in higher numbers than did the encapsulated parental strains. Both the encapsulated and nonencapsulated forms of enterotoxigenic E. coli 431 grown at 18 degrees C (K99 production suppressed) adhered poorly to the isolated cells. The nonenterotoxigenic E. coli 1793 which does not express K99 antigen also adhered poorly in both encapsulated and nonencapsulated forms. Fab fragments of anticapsular immunoglobulin G failed to block the effect of capsule on adherence of strain 431. The results indicated that K99 was the principal mediator of in vitro adhesion of the enterotoxigenic E. coli strains and that capsule impedes the in vitro adhesion. They also suggested that the capsular enhancement of colonization by such strains in vivo probably is by some mechanism other than enhanced adhesion to epithelium.
Kim, Sara; Woo, Gun-Jo
To compare the characteristics and to identify the epidemiological relationships of Escherichia coli isolated from organic and conventional vegetables, the antimicrobial resistance and genetic properties of E. coli were investigated from 2010 to 2011. E. coli was isolated from 1 of 111 (0.9%) organic vegetables and from 20 of 225 (8.9%) conventional vegetables. The majority of strains were isolated from the surrounding farming environment (n=27/150 vs. 49/97 in organic vs. conventional samples). The majority of the vegetable strains were isolated from the surrounding farming environments. E. coli isolated from organic vegetables showed very low antimicrobial resistance rates except for cephalothin, ranging from 0% to 17.9%, while the resistance rates to cephalothin (71%) were extremely high in both groups. E. coli isolates expressed various resistance genes, which most commonly included blaTEM, tet(A), strA, strB, and qnrS. However, none of the isolates harbored tet(D), tet(E), tet(K), tet(L), tet(M), or qnrA. The transferability of tet gene, tet(A), and tet(B) was identified in tetracycline-resistant E. coli, and the genetic relationship was confirmed in a few cases from different sources. With regard to the lower antimicrobial resistance found in organic produce, this production mode seems able to considerably reduce the selection of antimicrobial-resistant bacteria on vegetables.
Holvoet, Kevin; Callens, Benedicte; Dewulf, Jeroen; Uyttendaele, Mieke
Fresh produce is known to carry nonpathogenic epiphytic microorganisms. During agricultural production and harvesting, leafy greens can become contaminated with antibiotic-resistant pathogens or commensals from animal and human sources. As lettuce does not undergo any inactivation or preservation treatment during processing, consumers may be exposed directly to all of the (resistant) bacteria present. In this study, we investigated whether lettuce or its production environment (irrigation water, soil) is able to act as a vector or reservoir of antimicrobial-resistant Escherichia coli. Over a 1-year period, eight lettuce farms were visited multiple times and 738 samples, including lettuce seedlings (leaves and soil), soil, irrigation water, and lettuce leaves were collected. From these samples, 473 isolates of Escherichia coli were obtained and tested for resistance to 14 antimicrobials. Fifty-four isolates (11.4%) were resistant to one or more antimicrobials. The highest resistance rate was observed for ampicillin (7%), followed by cephalothin, amoxicillin-clavulanic acid, tetracycline, trimethoprim, and streptomycin, with resistance rates between 4.4 and 3.6%. No resistance to amikacin, ciprofloxacin, gentamicin, or kanamycin was observed. One isolate was resistant to cefotaxime. Among the multiresistant isolates (n = 37), ampicillin and cephalothin showed the highest resistance rates, at 76 and 52%, respectively. E. coli isolates from lettuce showed higher resistance rates than E. coli isolates obtained from soil or irrigation water samples. When the presence of resistance in E. coli isolates from lettuce production sites and their resistance patterns were compared with the profiles of animal-derived E. coli strains, they were found to be the most comparable with what is found in the cattle reservoir. This may suggest that cattle are a potential reservoir of antimicrobial-resistant E. coli strains in plant primary production. PMID:23974140
Holvoet, Kevin; Sampers, Imca; Callens, Benedicte; Dewulf, Jeroen; Uyttendaele, Mieke
Fresh produce is known to carry nonpathogenic epiphytic microorganisms. During agricultural production and harvesting, leafy greens can become contaminated with antibiotic-resistant pathogens or commensals from animal and human sources. As lettuce does not undergo any inactivation or preservation treatment during processing, consumers may be exposed directly to all of the (resistant) bacteria present. In this study, we investigated whether lettuce or its production environment (irrigation water, soil) is able to act as a vector or reservoir of antimicrobial-resistant Escherichia coli. Over a 1-year period, eight lettuce farms were visited multiple times and 738 samples, including lettuce seedlings (leaves and soil), soil, irrigation water, and lettuce leaves were collected. From these samples, 473 isolates of Escherichia coli were obtained and tested for resistance to 14 antimicrobials. Fifty-four isolates (11.4%) were resistant to one or more antimicrobials. The highest resistance rate was observed for ampicillin (7%), followed by cephalothin, amoxicillin-clavulanic acid, tetracycline, trimethoprim, and streptomycin, with resistance rates between 4.4 and 3.6%. No resistance to amikacin, ciprofloxacin, gentamicin, or kanamycin was observed. One isolate was resistant to cefotaxime. Among the multiresistant isolates (n = 37), ampicillin and cephalothin showed the highest resistance rates, at 76 and 52%, respectively. E. coli isolates from lettuce showed higher resistance rates than E. coli isolates obtained from soil or irrigation water samples. When the presence of resistance in E. coli isolates from lettuce production sites and their resistance patterns were compared with the profiles of animal-derived E. coli strains, they were found to be the most comparable with what is found in the cattle reservoir. This may suggest that cattle are a potential reservoir of antimicrobial-resistant E. coli strains in plant primary production.
Mishra, Raghavendra Prasad; Jain, Udit; Singh, Rakesh Kumar
Aim: This study was planned to study the genotypes of verocytotoxigenic Escherichia coli (VTEC) in fecal samples of deer due to its public health significance. Materials and Methods: A total of 160 fecal samples of deer were taken from Mathura district and Kanpur Zoo and screened for VTEC genes by polymerase chain reaction (PCR). Results: All fecal samples were positive for E. coli. All the E. coli isolates were screened by PCR to detect virulence genes stx1, stx2, eaeA, and hlyA. Of these, 15 isolates were found positive for VTEC having one or more genes in different combinations. Conclusion: Genes such as stx1, stx2, eaeA, and hlyA were prevalent in VTEC isolates from feces of deer. The presence of VTEC isolates having virulent genes may pose a threat to public health. PMID:27651685
Byappanahalli, Muruleedhara N.; Yan, Tao; Hamilton, Matthew J.; Ishii, Satoshi; Fujioka, Roger S.; Whitman, Richard L.; Sadowsky, Michael J.
While genotypically-distinct naturalized Escherichia coli strains have been shown to occur in riparian soils of Lake Michigan and Lake Superior watersheds, comparative analyses of E. coli populations in diverse soils across a range of geographic and climatic conditions have not been investigated. The main objectives of this study were to: (a) examine the population structure and genetic relatedness of E. coli isolates collected from different soil types on a tropical island (Hawaii), and (b) determine if E. coli populations from Hawaii and temperate soils (Indiana, Minnesota) shared similar genotypes that may be reflective of biome-related soil conditions. DNA fingerprint and multivariate statistical analyses were used to examine the population structure and genotypic characteristics of the E. coli isolates. About 33% (98 of 293) of the E. coli from different soil types and locations on the island of Oahu, Hawaii, had unique DNA fingerprints, indicating that these bacteria were relatively diverse; the Shannon diversity index for the population was 4.03. Nearly 60% (171 of 293) of the E. coli isolates from Hawaii clustered into two major groups and the rest, with two or more isolates, fell into one of 22 smaller groups, or individual lineages. Multivariate analysis of variance of 89, 21, and 106 unique E. coli DNA fingerprints for Hawaii, Indiana, and Minnesota soils, respectively, showed that isolates formed tight cohesive groups, clustering mainly by location. However, there were several instances of clonal isolates being shared between geographically different locations. Thus, while nearly identical E. coli strains were shared between disparate climatologically- and geographically-distinct locations, a vast majority of the soil E. coli strains were genotypically diverse and were likely derived from separate lineages. This supports the hypothesis that these bacteria are not unique and multiple genotypes can readily adapt to become part of the soil autochthonous
Regua, A H; Bravo, V L; Leal, M C; Lobo Leite, M E
Escherichia coli was isolated in 382 (94 per cent) of 406 children from 0 to 3 years of age who had been hospitalized for diarrhoea at the Hospital Municipal Salles Neto, Rio de Janeiro. Enteropathogenic Escherichia coli strains were isolated in 67 samples (18 per cent), distributed among the serogroups that were tested as follows: 0111 (33 per cent); 0125 (19 per cent); 0126, 0127, and 0142 (9 per cent); 0128 and 0119 (8 per cent); 055 (5 per cent); 0114 (2 per cent). No strains of EPEC belonging to serogroups 086, 0126, and 0158 were found. Among the samples in which EPEC strains were isolated, 15.0 per cent were children living in dwellings which had piped systems of water supply and drain, whereas with regard to those living in houses without such facilities, this percentage raised to 24 per cent. Similar results were found when the availability of water supply of drainage were taken separately.
Picco, Natalia Y; Alustiza, Fabrisio E; Bellingeri, Romina V; Grosso, María C; Motta, Carlos E; Larriestra, Alejandro J; Vissio, Claudina; Tiranti, Karina I; Terzolo, Horacio R; Moreira, Ana R; Vivas, Adriana B
The aim of this study was to perform a current molecular characterization of bovine pathogenic Escherichia coli strains isolated from random samplings in Argentinean dairy farms. Rectal swabs were obtained from 395 (63.7%) healthy and 225 (36.3%) diarrheic calves, belonging to 45 dairy farms in Cordoba Province, Argentina. E. coli isolates were examined for virulence genes (f5, f41, f17, sta, stb, lt, eae, vt) using PCR and the prevalence of E. coli virulence profiles was spatially described in terms of spatial distribution. A total of 30.1% isolates were found to be positive for at least one of the virulence genes. Depending on the different gene combinations present, 11 virulence profiles were found. Most of the isolates analyzed had a single gene, and no combination of fimbrial and enterotoxin gene was predominant. There was no association between the frequency and distribution of E. coli virulence genes and calf health status. Most of the virulence profiles were compatible with ETEC strains and showed a homogeneous distribution over the sampled area. A clustering pattern for E. coli virulence profiles could not be recognized. This work provides updated information on the molecular characterization of pathogenic E. coli strains from dairy herds in Cordoba, Argentina. These findings would be important to formulate prevention programs and effective therapies for diarrhea in calves caused by E. coli.
Williams, A; Christley, R M; McKane, S A; Roberts, V L H; Clegg, P D; Williams, N J
The aim of this study was to determine whether hospitalisation of horses leads to increased antimicrobial resistance in equine faecal Escherichia coli isolates. E. coli were cultured from faecal samples of horses on admission and after 7 days of hospitalisation; antimicrobial susceptibility was determined for eight antimicrobial agents. Resistance profiles of E. coli isolates were grouped into clusters, which were analysed to determine resistance patterns. Resistance to 7/8 antimicrobial agents and multi-drug resistance (MDR; resistance to ≥3 antimicrobial classes) were significantly higher after 7 days of hospitalisation. Forty-eight resistance profiles were identified; 15/48 were present on day 0 only, 16/48 on day 7 only and 17/48 at both times of sampling. There was a significant association between day 7 profiles and resistance detected to an increased number of antimicrobial agents. Hospitalisation of horses for 7 days resulted in alterations in equine faecal E. coli antimicrobial resistance profiles.
Borges, C A; Beraldo, L G; Maluta, R P; Cardozo, M V; Barboza, K B; Guastalli, E A L; Kariyawasam, S; DebRoy, C; Ávila, F A
Wild birds are carriers of Escherichia coli. However, little is known about their role as reservoirs for extra-intestinal pathogenic E. coli (ExPEC). In this work we investigated E. coli strains carrying virulence genes related to human and animal ExPEC isolated from free-living wild birds treated in a veterinary hospital. Multidrug resistance was found in 47.4% of the strains, but none of them were extended-spectrum beta-lactamase producers. Not only the virulence genes, but also the serogroups (e.g. O1 and O2) detected in the isolates of E. coli have already been implicated in human and bird diseases. The sequence types detected were also found in wild, companion and food animals, environmental and human clinical isolates in different countries. Furthermore, from the 19 isolates, 17 (89.5%) showed a degree of pathogenicity on an in vivo infection model. The isolates showed high heterogeneity by pulsed-field gel electrophoresis indicating that E. coli from these birds are clonally diverse. Overall, the results showed that wild birds can be reservoirs and/or vectors of highly pathogenic and multidrug-resistant E. coli that have the potential to cause disease in humans and poultry.
Doyle, M P; Schoeni, J L
A total of 896 samples of retail fresh meats and poultry was assayed for Escherichia coli serogroup O157:H7 by a hydrophobic grid membrane filter-immunoblot procedure developed specifically to isolate the organism from foods. The procedure involves several steps, including selective enrichment, filtration of enrichment culture through hydrophobic grid membrane filters, incubation of each filter on nitrocellulose paper on selective agar, preparation of an immunoblot (by using antiserum to E. coli O157:H7 culture filtrate) of each nitrocellulose paper, selection from the filters of colonies which corresponded to immunopositive sites on blots, screening of isolates by a Biken test for precipitin lines from metabolites and antiserum to E. coli O157:H7 culture filtrate, and confirmation of isolates as Vero cell cytotoxic E. coli O157:H7 by biochemical, serological, and Vero cell cytotoxicity tests. E. coli O157:H7 was isolated from 6 (3.7%) of 164 beef, 4 (1.5%) of 264 pork, 4 (1.5%) of 263 poultry, and 4 (2.0%) of 205 lamb samples. One of 14 pork samples and 5 of 17 beef samples contaminated with the organism were from Calgary, Alberta, Canada, grocery stores, whereas all other contaminated samples were from Madison, Wis., retail outlets. This is the first report of the isolation of E. coli O157:H7 from food other than ground beef, and results indicate that the organism is not a rare contaminant of fresh meats and poultry. Images PMID:3322190
Shiga Toxin producing Escherichia coli (STEC) are a subtype of pathogenic E. coli and, in particular, STEC isolates of E. coli serotype O157:H7 are recognized as a major foodborne pathogen that can cause infections ranging from having simple intestinal discomfort to bloody diarrhea and life threaten...
Cimdins, Annika; Lüthje, Petra; Li, Fengyang; Ahmad, Irfan; Brauner, Annelie
ABSTRACT Strains of Escherichia coli exhibit diverse biofilm formation capabilities. E. coli K-12 expresses the red, dry, and rough (rdar) morphotype below 30°C, whereas clinical isolates frequently display the rdar morphotype semiconstitutively. We sequenced the genomes of eight E. coli strains to subsequently investigate the molecular basis of semiconstitutive rdar morphotype expression. PMID:28126929
Adib, N; Ghanbarpour, R; Solatzadeh, H; Alizade, H
Escherichia coli (E. coli) strains are the major cause of urinary tract infections (UTI) and belong to the large group of extra-intestinal pathogenic E. coli. The purposes of this study were to determine the antibiotic resistance profile, virulence genes and phylogenetic background of E. coli isolates from UTI cases. A total of 137 E. coli isolates were obtained from UTI samples. The antimicrobial susceptibility of confirmed isolates was determined by disk diffusion method against eight antibiotics. The isolates were examined to determine the presence and prevalence of selected virulence genes including iucD, sfa/focDE, papEF and hly. ECOR phylo-groups of isolates were determined by detection of yjaA and chuA genes and fragment TspE4.C2. The antibiogram results showed that 71% of the isolates were resistant to cefazolin, 60.42% to co-trimoxazole, 54.16% to nalidixic acid, 36.45% to gentamicin, 29.18% to ciprofloxacin, 14.58% to cefepime, 6.25% to nitrofurantoin and 0.00% to imipenem. Twenty-two antibiotic resistance patterns were observed among the isolates. Virulence genotyping of isolates revealed that 58.39% isolates had at least one of the four virulence genes. The iucD gene was the most prevalent gene (43.06%). The other genes including sfa/focDE, papEF and hly genes were detected in 35.76%, 18.97% and 2.18% isolates, respectively. Nine combination patterns of the virulence genes were detected in isolates. Phylotyping of 137 isolates revealed that the isolates fell into A (45.99%), B1 (13.14%), B2 (19.71%) and D (21.16%) groups. Phylotyping of multidrug resistant isolates indicated that these isolates are mostly in A (60.34%) and D (20.38%) groups. In conclusion, the isolates that possessed the iucD, sfa/focDE, papEF and hly virulence genes mostly belonged to A and B2 groups, whereas antibiotic resistant isolates were in groups A and D. Escherichia coli strains carrying virulence factors and antibiotic resistance are distributed in specific phylogenetic
Inoue, Ikuo; Shibata, Sanae; Fukata, Tsuneo
The aim of this study was to determine the antimicrobial resistance of Escherichia coli isolated from the uteri of bitches with pyometra, and 38 E. coli isolates were used. The antimicrobials used were ampicillin (ABPC), amoxicillin/clavulanic acid, gentamicin, minocycline, cefazolin, levofloxacin (LVFX), trimethoprim-sulfamethoxazole (ST) and fosfomycin (FOM). Resistance to ABPC occurred most frequently, followed by LVFX and ST. Multi-drug resistance, defined as resistance against 3 or more classes of antimicrobials, was found in 23.7% of all isolates. Nine out of 13 resistant strains were multi-drug resistant, but no strain was found to be resistant to FOM. This suggests that FOM should be administered for E. coli from pyometra.
Hartel, Peter G.; Frick, Elizabeth A.; Funk, Adrienne L.; Hill, Jennifer L.; Summer, Jacob D.; Gregory, M. Brian
Factors affecting bacterial source tracking are important to understand because they affect the amount of sampling needed to describe fecal sources in a watershed adequately. The study area was a 76-kilometer reach of the Chattahoochee River and its tributaries in Metropolitan Atlanta, Georgia. Escherichia coli was isolated from water samples collected during baseflow and stormflow conditions from four mainstem and eight tributary sites; 262 isolates were ribotyped and assessed for their similarity. The vast majority of the E. coli ribotype patterns were unshared, whether the comparisons were between baseflow and stormflow conditions at one location, or between one location and another. The data suggest that either baseflow and stormflow conditions affected sharing of ribotype patterns, or that the sample size was too small to characterize the sharing adequately. Regardless, the results suggest that a large sampling of E. coli isolates is needed during various flow conditions from watersheds with complex land-use patterns for adequate bacterial source tracking.
Mtambo, Madundo M. A.; Muhairwa, Amandus P.; Lupindu, Athumani M.; Olsen, John E.
While pathogenic types of Escherichia coli are well characterized, relatively little is known about the commensal E. coli flora. In the current study, antimicrobial resistance in commensal E. coli and distribution of ERIC-PCR genotypes among isolates of such bacteria from cattle and cattle attendants on cattle farms in Tanzania were investigated. Seventeen E. coli genomes representing different ERIC-PCR types of commensal E. coli were sequenced in order to determine their possible importance as a reservoir for both antimicrobial resistance genes and virulence factors. Both human and cattle isolates were highly resistant to tetracycline (40.8% and 33.1%), sulphamethazole-trimethoprim (49.0% and 8.8%) and ampicillin (44.9% and 21.3%). However, higher proportion of resistant E. coli and higher frequency of resistance to more than two antimicrobials was found in isolates from cattle attendants than isolates from cattle. Sixteen out of 66 ERIC-PCR genotypes were shared between the two hosts, and among these ones, seven types contained isolates from cattle and cattle attendants from the same farm, suggesting transfer of strains between hosts. Genome-wide analysis showed that the majority of the sequenced cattle isolates were assigned to phylogroups B1, while human isolates represented phylogroups A, C, D and E. In general, in silico resistome and virulence factor identification did not reveal differences between hosts or phylogroups, except for lpfA and iss found to be cattle and B1 phylogroup specific. The most frequent plasmids replicon genes found in strains from both hosts were of IncF type, which are commonly associated with carriage of antimicrobial and virulence genes. Commensal E. coli from cattle and attendants were found to share same genotypes and to carry antimicrobial resistance and virulence genes associated with both intra and extraintestinal E. coli pathotypes. PMID:27977751
Arthur, Terrance M; Ahmed, Rafiq; Chase-Topping, Margo; Kalchayanand, Norasak; Schmidt, John W; Bono, James L
Previous reports have indicated that a small proportion of cattle shedding high levels of Escherichia coli O157:H7 is the main source for transmission of this organism between animals. Cattle achieving a fecal shedding status of 10(4) CFU of E. coli O157:H7/gram or greater are now referred to as supershedders. The aim of this study was to investigate the contribution of E. coli O157:H7 strain type to supershedding and to determine if supershedding was restricted to a specific set of E. coli O157:H7 strains. Fecal swabs (n = 5,086) were collected from cattle at feedlots or during harvest. Supershedders constituted 2.0% of the bovine population tested. Supershedder isolates were characterized by pulsed-field gel electrophoresis (PFGE), phage typing, lineage-specific polymorphism assay (LSPA), Stx-associated bacteriophage insertion (SBI) site determination, and variant analysis of Shiga toxin, tir, and antiterminator Q genes. Isolates representing 52 unique PFGE patterns, 19 phage types, and 12 SBI clusters were obtained from supershedding cattle, indicating that there is no clustering to E. coli O157:H7 genotypes responsible for supershedding. While being isolated directly from cattle, this strain set tended to have higher frequencies of traits associated with human clinical isolates than previously collected bovine isolates with respect to lineage and tir allele, but not for SBI cluster and Q type. We conclude that no exclusive genotype was identified that was common to all supershedder isolates.
Kobayashi, Renata K T; Aquino, Ivani; Ferreira, Ana Lívia da S; Vidotto, Marilda C
Escherichia coli strains designated as avian pathogenic E. coli (APEC) are responsible for avian colibacillosis, an acute and largely systemic disease that promotes significant economic losses in poultry industry worldwide because of mortality increase, medication costs, and condemnation of carcasses. APEC is a subgroup of extraintestinal pathogenic E. coli pathotype, which includes uropathogenic E. coli, neonatal meningitis E. coli, and septicemic E. coli. We isolated E. coli from commercial chicken carcasses in a Brazilian community and compared by polymerase chain reaction-defined phylogenetic group (A, B1, B2, or D) with APEC strains isolated from sick chickens from different poultry farms. A substantial number of strains assigned to phylogenetic E. coli reference collection group B2, which is known to harbor potent extraintestinal human and animal E. coli pathogens, were identified as APEC (26.0%) in both commercial chicken carcasses and retail poultry meat (retail poultry E. coli [RPEC]) (21.25%). The majority of RPEC were classified as group A (35%), whereas the majority of APEC were groups B1 (30.8) and A (27.6%). APEC and RPEC presented the genes pentaplex, iutA, hly, iron, ompT, and iss, but with different virulence profiles. The similarity between APEC and RPEC indicates RPEC as potentially pathogenic strains and supports a possible zoonotic risk for humans.
Schierack, Peter; Römer, Antje; Jores, Jörg; Kaspar, Heike; Guenther, Sebastian; Filter, Matthias; Eichberg, Jürgen; Wieler, Lothar H.
Our understanding of the composition of Escherichia coli populations in wild boars is very limited. In order to obtain insight into the E. coli microflora of wild boars, we studied E. coli isolates from the jejunums, ileums, and colons of 21 wild boars hunted in five geographic locations in Germany. Ten isolates per section were subjected to clonal determination using pulsed-field gel electrophoresis. One representative isolate per clone was further investigated for virulence traits, phylogenetic affiliation, and antimicrobial susceptibility. Macrorestriction analysis of 620 isolates revealed a range of clone diversity among the sections and animals, with up to 9 and 16 different clones per section and animal, respectively. Most of the clones for a given animal were shared between two adjacent intestinal sections. The overall highest clonal diversity was observed within the colon. While the astA gene was present in a large number of clones, other virulence genes and hemolytic ability were detected only sporadically. Clones of all four ECOR groups dominated the intestinal sections. Phylogenetic analysis and the occurrence of virulence genes correlated with the isolation frequencies for clones. All E. coli clones from wild boars were susceptible to all antimicrobial agents tested. In conclusion, though several parameters (including an animal-specific and highly diverse E. coli clone composition, the simultaneous occurrence of single clones in two adjacent intestinal sections of a given animal, and a higher E. coli diversity in the large intestine than in the small intestine) of E. coli populations of wild boars were similar to those of previously described E. coli populations of conventionally reared domestic pigs, our data also indicate possible differences, as seen for the E. coli diversity in the large intestine, the occurrence of certain virulence genes and phylogenetic groups, and antimicrobial susceptibilities. PMID:19060173
Feng, Peter C H; Councell, Terry; Keys, Christine; Monday, Steven R
The 13 Shiga-toxigenic Escherichia coli (STEC) strains isolated from wholesale spinach and lettuce consisted mostly of serotypes that have not been implicated in illness. Among these strains, however, were two O113:H21 that carried virulence genes common to this pathogenic serotype (stx(2), ehxA, saa, and subAB), suggesting that their presence in ready-to-eat produce may be of health concern.
Runnels, P L; Moon, H W; Schneider, R A
When isolated intestinal epithelial cells from neonatal and older pigs, calves, and mice were tested for adhesion by K99+ enterotoxigenic Escherichia coli, cells from older animals were resistant to adhesion. PMID:6103878
Georges, M C; Wachsmuth, I K; Birkness, K A; Moseley, S L; Georges, A J
Escherichia coli strains were isolated from 778 children with diarrhea and 151 well children in the Central African Republic over a period of 1 year. These 929 strains were assayed for heat-labile and heat-stable enterotoxin production and were hybridized (probed) with structural genes for these enterotoxins. Twenty-four isolates from diarrheal patients and one isolate from a well child were found to be toxigenic by assay and probe. Minor discrepancies were encountered with both assays and probes during initial screening procedures, but the two methodologies were ultimately comparable. Images PMID:6350346
Ryu, Seung-Hee; Lee, Jip-Ho; Park, Sang-Hoon; Song, Mi-Ok; Park, Sun-Hee; Jung, Hyo-Won; Park, Geon-Yong; Choi, Sung-Min; Kim, Moo-Sang; Chae, Young-Zoo; Park, Seog-Gee; Lee, Young-Ki
A total of 4330 food samples of which microbiological standard for Escherichia coli is negative in Korea were determined for the frequency of E. coli. Ninety six samples (2.2%) were positive for E. coli. Detection rate of E. coli varied significantly by food type and ranged from 0.3% to 10.9%. Seasoned raw meat (yukhoe) and cold bean-soup had the highest prevalence for E. coli (10.9%) followed by gimbap (5.2%), meat broth for cold noodle (2.9%) and sprout (2.1%). E. coli isolates (n=96) were investigated for their phenotypic and genotypic antimicrobial resistance patterns. Seventeen E. coli isolates (17.7%) were resistant to one or more antimicrobial agents tested. High rates of resistance to the following drugs were observed: tetracycline (15.6%), streptomycin (12.5%), ampicillin (10.4%), nalidixic acid (9.4%) and ticarcillin (9.4%). All ampicillin resistant isolates were screened for extended-spectrum β-lactamase (ESBL) production by the combination disk test. None of the E. coli isolates produced ESBLs. Seventeen out of 96 E. coli isolates which were resistant to at least one antibiotic were investigated by PCR for the presence of 3 classes of antimicrobial resistance genes (tetracycline, aminoglycosides and beta-lactams). The tetracycline resistance genes tetA and tetB were found in 7 and 5 isolates, respectively. The aminoglycoside resistance genes, strA/B, aphA1, aadA and aac(3)-IV were found in 9, 5, 2 and 2 isolates, respectively. The beta-lactam resistance gene, bla(TEM) was found in 7 isolates. Results of this study show that 13 E. coli isolates were multidrug resistant (to three or more antibiotics) and 12 isolates carried at least one antimicrobial resistance gene. These isolates can act as the reservoir for antimicrobial resistance genes and facilitate the dissemination of these genes to other pathogenic and commensal bacteria. Adequate intervention to reduce microbial contamination of these foods is strongly recommended.
Soliman, Ahmed M; Khalifa, Hazim O; Ahmed, Ashraf M; Shimamoto, Toshi; Shimamoto, Tadashi
The first occurrence of New Delhi metallo-β-lactamase 5 (NDM-5), carried on an IncI1-Iγ-type plasmid of >93kb in a multidrug-resistant Escherichia coli strain in Kafr El-Sheikh, Egypt, is reported. The strain was isolated from a wound pus swab from a patient diagnosed with a fracture of the right femur. This E. coli strain was found to belong to sequence type (ST) 5018 and also to carry other resistance genes, including blaCTX-M-15, blaCMY-42, blaOXA-1, and aac(6')-Ib-cr.
Brennan, Fiona P.; Abram, Florence; Chinalia, Fabio A.; Richards, Karl G.; O'Flaherty, Vincent
Soils are typically considered to be suboptimal environments for enteric organisms, but there is increasing evidence that Escherichia coli populations can become resident in soil under favorable conditions. Previous work reported the growth of autochthonous E. coli in a maritime temperate Luvic Stagnosol soil, and this study aimed to characterize, by molecular and physiological means, the genetic diversity and physiology of environmentally persistent E. coli isolates leached from the soil. Molecular analysis (16S rRNA sequencing, enterobacterial repetitive intergenic consensus PCR, pulsed-field gel electrophoresis, and a multiplex PCR method) established the genetic diversity of the isolates (n = 7), while physiological methods determined the metabolic capability and environmental fitness of the isolates, relative to those of laboratory strains, under the conditions tested. Genotypic analysis indicated that the leached isolates do not form a single genetic grouping but that multiple genotypic groups are capable of surviving and proliferating in this environment. In physiological studies, environmental isolates grew well across a broad range of temperatures and media, in comparison with the growth of laboratory strains. These findings suggest that certain E. coli strains may have the ability to colonize and adapt to soil conditions. The resulting lack of fecal specificity has implications for the use of E. coli as an indicator of fecal pollution in the environment. PMID:20154122
Since the time of the first microbe hunters, primary culture and isolation of bacteria has been a foundation of microbiology. Like other microbial methods, bacterial culture and isolation methodologies continue to develop. Although fundamental concepts like selection and enrichment are as relevant t...
Saeidi, Saeide; Amini Boroujeni, Negar; Ahmadi, Hassan; Hassanshahian, Mehdi
Background: The extended-spectrum beta-lactamase (ESBL) -producing Escherichia coli isolates make many serious infections, especially urinary tract infections. Objectives: The purpose of this study was to determine the antibacterial activities of some natural plant extracts against ESBL-producing E. coli isolates, which harbor the TEM gene in urine samples of the patients who have urinary tract infections. Materials and Methods: Evaluation has to be exactly determined for both methods of disk diffusion test and polymerase chain reaction (PCR), separately. We evaluated 120 strains of E. coli isolates from the urine culture of the patients in Boo-Ali Hospital (Zahedan, south-eastern Iran) who were suffering from urinary tract infections. The ESBL-producing E. coli isolates were evaluated by disk diffusion test and PCR through TEM gene detection. The minimal inhibitory concentration (MIC) of commonly used antibiotics including ceftazidime, ceftriaxon, amikacin, gentamicin and ciprofloxacin along with the MIC of the alcoholic extract of different natural plants including Myrtus communis L (Myrtaceae), Amaranthus retraflexus (Amaranthaceae), Cyminum cuminum L (Apiaceae), Marrubium vulgare (Laminaceae) and Peganum. harmala (Zygrophyllaceae) against the ESBL-producing E. coli isolates, which harbor the TEM genes, were determined using the microdulition method. Results: Results of this study showed that in disk diffusion method, 80 samples of E. coli produced ESBLs. In PCR method, the TEM gene distribution in the isolated ESBL-producing organisms was 50 (41.6%). Amikacin was the most effective anti-bacterial agent and ciprofloxacin was the least effective against E. coli isolates. All the natural plant extracts mentioned above, especially P. harmala, were effective against the selected isolates of ESBL-producing E. coli. The most frequent ESBL rate producing E. coli isolates (32 out of 50) had MIC of 2.5 mg/mL in ethanol extract of P. harmala. Conclusions: The alcoholic
Pattnaik, Dipti; Neogi, Dhruba Kumar; Jena, Jagadananda; Mallick, Bandana
Introduction Resistance to common antibiotics is a matter of grave concern in treating infections in hospital settings especially in Intensive Care Units (ICUs). One of the most commonly used and effective group of antibiotics, cephalosporins, exhibit resistance due to production of Extended Spectrum Beta- Lactamases (ESBLs). The prevalence of ESBL producing Escherichia coli (E.coli) has increased throughout the world and is a major cause of treatment failure in ICUs. As per our knowledge studies were not available on the prevalence of ESBL producing E.coli in ICUs of this region. Aim To determine the prevalence of ESBLs among Escherichia coli isolates in ICUs of a tertiary care hospital. Materials and Methods A cross sectional study was conducted over a period of 4 years (Sept 2011 to Sept 2015) in the Department of Microbiology, Kalinga Institute of Medical Sciences (KIMS), Bhubaneswar. Consecutive non-duplicate isolates of E.coli recovered from 6800 clinical samples of patients admitted to different Intensive Care Units (ICUs) were subjected to ESBL screening test and then to CLSI recommended Phenotypic Confirmatory Disc Diffusion Tests (PCDDT) for ESBL production determination. Results Out of 6800 samples, 1038 were E.coli isolates and 452(44%) were resistant to third generation cephalosporins. ESBL producing Escherichia coli among them were 276 (61.1%). Paediatric ICU showed the highest prevalence of ESBL E.coli at 80.9%. The highest prevalence of ESBL E.coli was in urine samples (82.6%) followed by pus (9.8%). The most effective antibiotic for ESBL producers was imipenem (96.7% sensitive), followed by amikacin (88.4%) and piperacillin- tazobactum (87%). Conclusion This study has highlighted the high prevalence of ESBL producing E.coli in the ICUs of our hospital. An in depth analysis of their antibiogram will be helpful in formulating the antibiotic policy and prevent spread of ESBL strains. It is recommended that ESBL testing should be done routinely to
Li, Y F; Su, N; Chen, S Y; Hu, W X; Li, F F; Jiang, Z P; Yu, X Q
Escherichia coli is the most common cause of Gram-negative peritonitis resulting in peritoneal function deterioration as well as poor clinical outcome in continuous ambulatory peritoneal dialysis (PD) patients. In this study, we analyzed the phylogenetic background and genetic profile of the E. coli isolates and sought to determine the characteristics of specific bacteria associated with peritonitis. E. coli isolates from 56 episodes of peritonitis in 46 PD patient cases and rectal isolates from 57 matched PD control patient cases were compared for both phylogenetic groups and the presence of virulence factors (VFs). There were no significant differences in terms of demographic data between the peritonitis and control groups. Peritonitis isolates exhibited a significantly greater prevalence of 8 VFs. In multivariate logistic regression analysis, kpsMT II (group 2 capsule synthesis) was the strongest VF predictor of peritonitis (OR = 8.02; 95%CI = 3.18-20.25; P < 0.001), followed by traT (serum-resistance-associated outer membrane protein) (OR = 3.83; 95%CI = 1.33-11.03; P = 0.013). The pathogenic groups of E. coli contained a higher concentration of individual VFs compared to the commensal groups. The prevalence of pathogenic E. coli was much higher in peritoneal isolates than rectal isolates (64.3 vs 31.6%, P = 0.001). Our results indicate that the E. coli peritonitis and rectal isolates are different in PD patients. The specific VFs associated with peritonitis isolates may directly contribute to the pathogenesis of peritonitis.
Freeman, R.; Sisson, P. R.; Jenkins, D. R.; Ward, A. C.; Lightfoot, N. F.; O'Brien, S. J.
Thirty-six encoded isolates of Escherichia coli. 32 of which were of serotype O157, were examined by pyrolysis mass spectrometry (PyMS). Thirty-one of the serotype O157 isolates possessed the flagellar antigen H7 and produced Verocytotoxin (VT), the other isolate serotyped as H45 and was non-toxigenic. Eighteen of the VT-producing E. coli (VTEC) isolates were from sporadic disease in residents of the Northern Region. Standard principal component (PC) and canonical variate (CV) analysis of the data distinguished only the four non-O157 isolates from the remainder which were indistinguishable by this approach. A similarity matrix based on differences between individual CV means distinguished a further ten isolates. The matrix correctly clustered 2 pairs of isolates from siblings and 4 isolates from an affected family. A further 5 clusters of 3 or more isolates and 6 pairs of isolates were defined. These groupings proved to be homogenous for toxin phenotype but occasionally entrained isolates of dissimilar phage type. However, in general, PyMS-derived clustering of apparently sporadic isolates accorded with geographical locations as determined by postcode. PyMS, which is a quick and high volume capacity phenotypic technique, may be a useful addition to existing methods in the investigation of the epidemiology of sporadic VTEC disease. PMID:7781731
Yang, Hanchun; Chen, Sheng; White, David G.; Zhao, Shaohua; McDermott, Patrick; Walker, Robert; Meng, Jianghong
Escherichia coli isolates from diseased piglets (n = 89) and chickens (n = 71) in China were characterized for O serogroups, virulence genes, antimicrobial susceptibility, class 1 integrons, and mechanisms of fluoroquinolone resistance. O78 was the most common serogroup identified (63%) among the chicken E. coli isolates. Most isolates were PCR positive for the increased serum survival gene (iss; 97%) and the temperature-sensitive hemagglutinin gene (tsh; 93%). The O serogroups of swine E. coli were not those typically associated with pathogenic strains, nor did they posses common characteristic virulence factors. Twenty-three serogroups were identified among the swine isolates; however, 38% were O nontypeable. Overall, isolates displayed resistance to nalidixic acid (100%), tetracycline (98%), sulfamethoxazole (84%), ampicillin (79%), streptomycin (77%), and trimethoprim-sulfamethoxazole (76%). Among the fluoroquinolones, resistance ranged between 64% to levofloxacin, 79% to ciprofloxacin, and 95% to difloxacin. DNA sequencing of gyrA, gyrB, parC, and parE quinolone resistance-determining regions of 39 nalidixic acid-resistant E. coli isolates revealed that a single gyrA mutation was found in all of the isolates; mutations in parC together with double gyrA mutations conferred high-level resistance to fluoroquinolones (ciprofloxacin MIC, ≥8 μg/ml). Class 1 integrons were identified in 17 (19%) isolates from swine and 42 (47%) from chickens. The majority of integrons possessed genes conferring resistance to streptomycin and trimethoprim. These findings suggest that multiple-antimicrobial-resistant E. coli isolates, including fluoroquinolone-resistant variants, are commonly present among diseased swine and chickens in China, and they also suggest the need for the introduction of surveillance programs in China to monitor antimicrobial resistance in pathogenic bacteria that can be potentially transmitted to humans from food animals. PMID:15297487
Parul, Singh; Bist, Basanti; Sharma, Barkha; Jain, Udit; Yadav, Janardan K.
Aim: The present study was conducted to find the association among virulence determinants of verotoxic Escherichia coli (VTEC) isolated from cattle calf feces. Materials and Methods: A total of 216 cattle calf fecal samples were collected aseptically and processed under required conditions for the isolation of E. coli. The isolates were further subjected to multiplex polymerase chain reaction (mPCR) for the detection of virulent genes. All the VTEC isolates were serotyped at the Central Research Institute, Kasauli, Himachal Pradesh. The VTEC isolates were observed for the enterohemolysin production on washed sheep blood agar (wSBA). Results: A total of 177 presumptive E. coli were isolated from 216 calf fecal samples revealing an overall prevalence of E. coli to be 81.94%. A total of 32 (14.81%) isolates were detected as VTEC through mPCR. The prevalence of verotoxin genes vt1, vt2, and combination of vt1+vt2 in the VTEC isolates was found to be 12 (37.5%), 14 (43.75%), and 6 (18.75%), respectively. Other virulent genes eaeA and hlyA were found in 6 and 11 VTEC strains with prevalence values of 18.75% and 34.37%, respectively. A total of 13 different O serogroups were revealed in serotyping of 32 VTEC isolates. Out of 32 VTEC strains, only 26 (81.25%) were enterohemolytic on wSBA as they produced the characteristic small, turbid zone of hemolysis around the streaking line. Although enterohemolysin production has been attributed to the presence of hlyA gene, only 11 of 26 enterohemolysin producing VTEC were found to be harboring the hlyA gene (11/26) 42.03%. Conclusion: The present study concludes that there might be an association between the presence of verotoxin genes and enterohemolysin production in VTEC group of E. coli. PMID:27651684
Ma, Jun; Li, Liang; Fang, Liang-Xing; Zhang, Qijing; Liu, Ya-Hong; Liao, Xiao-Ping
Previous study indicated that the multi-resistance gene cfr was mainly found in gram-positive bacteria, such as Staphylococcus and Enterococcus, and was sporadically detected in Escherichia coli. Little is known about the prevalence and transmission mechanism of cfr in E. coli. In this study, the presence of cfr in E. coli isolates collected during 2010–2012 from food-producing animals in Guangdong Province of China was investigated, and the cfr-positive E. coli isolates were characterized by PFGE, plasmid profiling, and genetic environment analysis. Of the 839 E. coli isolates, 10 isolates from pig were cfr positive. All the cfr-positive isolates presented a multi-resistance phenotype and were genetically divergent as determined by PFGE. In 8 out of the 10 strains, the cfr gene was located on plasmids of ∼30 kb. Restriction digestion of the plasmids with EcoRI and sequence hybridization with a cfr-specific probe revealed that the cfr-harboring fragments ranged from 6 to 23 kb and a ∼18 kb cfr-carrying fragment was common for the plasmids that were ∼30 kb. Four different genetic environments of cfr were detected, in which cfr is flanked by two identical copies of IS26, which may loop out the intervening sequence through homologous recombination. Among the 8 plasmids of ∼30 kb, 7 plasmids shared the same genetic environment. These results demonstrate plasmid-carried cfr in E. coli and suggest that transposition and homologous recombination mediated by IS26 might have played a rule in the transfer of the cfr gene in E. coli. PMID:25036029
Johnson, Timothy J; Wannemuehler, Yvonne; Kariyawasam, Subhashinie; Johnson, James R; Logue, Catherine M; Nolan, Lisa K
Escherichia coli strains that cause disease outside the intestine are known as extraintestinal pathogenic E. coli (ExPEC) and include pathogens of humans and animals. Previously, the genome of avian-pathogenic E. coli (APEC) O1:K1:H7 strain O1, from ST95, was sequenced and compared to those of several other E. coli strains, identifying 43 genomic islands. Here, the genomic islands of APEC O1 were compared to those of other sequenced E. coli strains, and the distribution of 81 genes belonging to 12 APEC O1 genomic islands among 828 human and avian ExPEC and commensal E. coli isolates was determined. Multiple islands were highly prevalent among isolates belonging to the O1 and O18 serogroups within phylogenetic group B2, which are implicated in human neonatal meningitis. Because of the extensive genomic similarities between APEC O1 and other human ExPEC strains belonging to the ST95 phylogenetic lineage, its ability to cause disease in a rat model of sepsis and meningitis was assessed. Unlike other ST95 lineage strains, APEC O1 was unable to cause bacteremia or meningitis in the neonatal rat model and was significantly less virulent than uropathogenic E. coli (UPEC) CFT073 in a mouse sepsis model, despite carrying multiple neonatal meningitis E. coli (NMEC) virulence factors and belonging to the ST95 phylogenetic lineage. These results suggest that host adaptation or genome modifications have occurred either in APEC O1 or in highly virulent ExPEC isolates, resulting in differences in pathogenicity. Overall, the genomic islands examined provide targets for further discrimination of the different ExPEC subpathotypes, serogroups, phylogenetic types, and sequence types.
Gai, Wenyan; Wang, Junwei; Wang, Juan; Cui, Zhigang; Qu, Zhina; Cui, Jinghua; Du, Xiaoli; Huang, Xiumei; Zhao, Jianmei
We aimed to understand the distribution of Escherichia coli in poultry and to reveal the virulence factors, the drug resistance and molecular epidemic regularity and characteristics of isolate strains from 6 provinces in China and to complete the characteristics of E. coli for the risk assessment. A total of 87 E. coli isolates were analyzed with 7 virulence genes by PCR drug sensitivity test in 13 kinds of antimicrobial agents and analyzed with PFGE and MLST genotyping. The PFGE genotyping of 87 isolates yielded 75 PFGE type. MLST analysis of isolates identified the 39 STs, the 7 housekeeping genes had the different variation. The most prevalent virulence genes were iucD (74.7%), followed by iss (55.2%), Irp2 (43.7%), tsh (28.7%), cva (19.5%), papC (9.2%) and vat (8.1%). All isolates were resistant to two or three antimicrobial agents highly resistant to SXT, TE (85.06%), SF (83.91%), AM (66. 67%), to fluoroquinolones (ENR, 63.22%, NOR, 50.57%) and to GM (57.47%). E. coli strains resistant spectrum was wide gene was polymorphism the distribution had a certain timeliness and regional in part region of China. These were a solid foundation for the epidemiological investigation and traceability laid. PMID:25785064
Reznikoff, W S; Thornton, K P
A trp-lac fusion strain of Escherichia coli in which the lac structural genes are part of the tryptophan operon has been used to isolate trp regulatory mutants. This was accomplished by isolating lac(+) colonies on either lactose-minimal agar or lactose-MacConkey indicator agar. Seventy-seven of 78 lac(+) isolates contained mutations which mapped near the ara locus and most of these isolates were found to be 5-methyltryptophan-resistant after introduction of an F-trp episome. The lac(+) phenotypes of these 77 isolates were therefore probably the result of trpR(-) mutations. The one remaining isolate carried a mutation which was not part of the trp regulatory system.
Subramanian, Sreedhar; Roberts, Carol L; Hart, C Anthony; Martin, Helen M; Edwards, Steve W; Rhodes, Jonathan M; Campbell, Barry J
There is increasing evidence that Escherichia coli organisms are important in Crohn's disease (CD) pathogenesis. In CD tissue they are found within macrophages, and the adherent-invasive CD ileal E. coli isolate LF82 can replicate inside macrophage phagolysosomes. This study investigates replication and antibiotic susceptibility of CD colonic E. coli isolates inside macrophages. Replication of CD colonic E. coli within J774-A1 murine macrophages and human monocyte-derived macrophages (HMDM) was assessed by culture and lysis after gentamicin killing of noninternalized bacteria and verified by electron microscopy (EM). All seven CD colonic isolates tested replicated within J774-A1 macrophages by 3 h (6.36-fold +/- 0.7-fold increase; n = 7 isolates) to a similar extent to CD ileal E. coli LF82 (6.8-fold +/- 0.8-fold) but significantly more than control patient isolates (5.2-fold +/- 0.25-fold; n = 6; P = 0.006) and E. coli K-12 (1.0-fold +/- 0.1-fold; P < 0.0001). Replication of CD E. coli HM605 within HMDM (3.9-fold +/- 0.7-fold) exceeded that for K-12 (1.4-fold +/- 0.2-fold; P = 0.03). EM showed replicating E. coli within macrophage vacuoles. Killing of HM605 within J774-A1 macrophages following a 3-h incubation with antibiotics at published peak serum concentrations (C(max)) was as follows: for ciprofloxacin, 99.5% +/- 0.2%; rifampin, 85.1% +/- 6.6%; tetracycline, 62.8% +/- 6.1%; clarithromycin, 62.1% +/- 5.6% (all P < 0.0001); sulfamethoxazole, 61.3% +/- 7.0% (P = 0.0007); trimethoprim, 56.3% +/- 3.4% (P < 0.0001); and azithromycin, 41.0% +/- 10.5% (P = 0.03). Ampicillin was not effective against intracellular E. coli. Triple antibiotic combinations were assessed at 10% C(max), with ciprofloxacin, tetracycline, and trimethoprim causing 97% +/- 0.0% killing versus 86% +/- 2.0% for ciprofloxacin alone. Colonic mucosa-associated E. coli, particularly CD isolates, replicate within macrophages. Clinical trials are indicated to assess the efficacy of a combination
Carvalho, Vania M; Osugui, Lika; Setzer, Ariela P; Lopez, Rodrigo P G; Pestana de Castro, Antonio F; Irino, Kinue; Catão-Dias, José L
Diseases caused by extraintestinal pathogenic Escherichia coli (ExPEC) in wild felids are rarely reported. Although urinary tract infections are infrequently reported in domestic cats, such infections when present are commonly caused by ExPEC. The present work characterized ExPEC strains isolated from 2 adult felines, a snow leopard (Panthera uncia) and a black leopard (Panthera pardus melas), that died from secondary bacteremia associated with urinary tract infections. Isolates from both animals were classified into the B2 phylogenetic group and expressed virulence genotypes that allowed them to cause severe disease. In addition, strains from the black leopard showed multidrug resistance.
Talebiyan, Reza; Kheradmand, Mehdi; Khamesipour, Faham; Rabiee-Faradonbeh, Mohammad
Antimicrobial agents are used extremely in order to reduce the great losses caused by Escherichia coli infections in poultry industry. In this study, 318 pathogenic Escherichia coli (APEC) strains isolated from commercial broiler flocks with coli-septicemia were examined for antimicrobials of both veterinary and human significance by disc diffusion method. Multiple resistances to antimicrobial agents were observed in all the isolates. Resistance to the antibiotics was as follows: Tylosin (88.68%), Erythromycin (71.70%), Oxytetracycline (43.40%), Sulfadimethoxine-Trimethoprim (39.62%), Enrofloxacin (37.74%), Florfenicol (35.85%), Chlortetracycline (33.96%), Doxycycline (16.98%), Difloxacin (32.08%), Danofloxacin (28.30%), Chloramphenicol (20.75%), Ciprofloxacin (7.55%), and Gentamicin (5.66%). This study showed resistance against the antimicrobial agents that are commonly applied in poultry, although resistance against the antibiotics that are only applied in humans or less frequently used in poultry was significantly low. This study emphasizes on the occurrence of multiple drug resistant E. coli among diseased broiler chickens in Iran. The data revealed the relative risks of using antimicrobials in poultry industry. It also concluded that use of antibiotics must be limited in poultry farms in order to reduce the antibiotic resistances. PMID:25548716
Talebiyan, Reza; Kheradmand, Mehdi; Khamesipour, Faham; Rabiee-Faradonbeh, Mohammad
Antimicrobial agents are used extremely in order to reduce the great losses caused by Escherichia coli infections in poultry industry. In this study, 318 pathogenic Escherichia coli (APEC) strains isolated from commercial broiler flocks with coli-septicemia were examined for antimicrobials of both veterinary and human significance by disc diffusion method. Multiple resistances to antimicrobial agents were observed in all the isolates. Resistance to the antibiotics was as follows: Tylosin (88.68%), Erythromycin (71.70%), Oxytetracycline (43.40%), Sulfadimethoxine-Trimethoprim (39.62%), Enrofloxacin (37.74%), Florfenicol (35.85%), Chlortetracycline (33.96%), Doxycycline (16.98%), Difloxacin (32.08%), Danofloxacin (28.30%), Chloramphenicol (20.75%), Ciprofloxacin (7.55%), and Gentamicin (5.66%). This study showed resistance against the antimicrobial agents that are commonly applied in poultry, although resistance against the antibiotics that are only applied in humans or less frequently used in poultry was significantly low. This study emphasizes on the occurrence of multiple drug resistant E. coli among diseased broiler chickens in Iran. The data revealed the relative risks of using antimicrobials in poultry industry. It also concluded that use of antibiotics must be limited in poultry farms in order to reduce the antibiotic resistances.
Doregiraee, Fatemeh; Alebouyeh, Masoud; Nayeri Fasaei, Bahar; Charkhkar, Saeed; Tajedin, Elahe; Zali, Mohammad Reza
Aim: The purpose of this study was to investigate the prevalence of enteropathogenic Escherichia coli (EPEC) and shiga toxin producing E. coli (STEC) strains in healthy broilers in Iran. Background: STEC and EPEC strains as diarrheagenic E. coli are among the most prevalent causative agents in acute diarrhea. Domestic animals, mainly cattle and sheep, have been implicated as the principal reservoirs of these pathotypes; however their prevalence among the broilers is varied among different countries. Patients and methods: A total of 500 cloacal swab samples from broilers of five different poultry houses (A-E) were collected to investigate the presence of stx1, stx2, hly, eae, and bfp virulence genes among the E. coli isolates by polymerase chain reaction. The shiga toxin encoding strains were evaluated serologically to detect their interaction with a commercial antiserum against O157 antigen. Results: Out of the 500 collected samples, 444 E. coli strains were isolated. Three strains (0.67%) presented at least one of the studied virulence genes (stx2, hly and eae), two strains were identified as STEC (stx2+, O157:nonH7) and one as an atypical EPEC strains (eae+ bfp-). Conclusion: The study established the presence of STEC and atypical EPEC in healthy broilers in Iran. Poultry might serve as vectors for transmission of pathogenic E. coli to human populations. PMID:26744615
Sabaté, Montserrat; Prats, Guillem; Moreno, Eva; Ballesté, Elisenda; Blanch, Anicet R; Andreu, Antonia
To gain insight into whether Escherichia coli isolated from humans and resistant to some common antimicrobial agents are derived from animals, 85 E. coli strains were selected by ERIC-PCR from human and animal wastewater samples. Phylogroup, pathogenicity islands (PAIs), resistance to quinolones, fluoroquinolones and presence of extended-spectrum beta-lactamases (ESBLs) were analyzed. Among the total, 55% were resistant to nalidixic acid and 38% to ciprofloxacin; 12% produced ESBLs. Chicken-derived strains were associated with quinolone and fluoroquinolone resistance and presence of ESBLs, while human strains were associated with susceptibility. Group B2 E. coli strains were associated with human origin, susceptibility to fluoroquinolones and presence of PAIs, whereas groups A, B1 and D showed a low virulence profile and a high level of antimicrobial resistance. In both human and animal wastewater, E. coli A, B1 and D were prevalent, and strains from both origins showed a similar virulence profile in each phylogroup. These findings led us to hypothesize that abusive antibiotic use in food animal production may promote the development of resistance among these intestinal E. coli phylogroups, which could later be transmitted to humans through the food supply. The low prevalence of E. coli group B2 in the animal gut may explain, at least in part, the absence of emergence of resistant B2 isolates.
Shin, Hyun-Ho; Cho, Seung-Hak
Objectives: This study aimed to characterize the prevalence of antibiotic resistance in Escherichia coli isolates from the fecal samples of fishery workers who work in fish farms and often use antibiotics for the feeding fishes. Methods: Seventy-three E. coli strains isolated from the fecal samples of fishery workers and 180 isolates from a control group of restaurant workers were tested for antibiotic resistance by agar disk diffusion with 16 antimicrobial agents. Results: About 30% of isolates from each group showed antimicrobial resistance to ampicillin, and 60% of isolates from fishery workers and 41% from restaurant workers were resistant to tetracycline. The isolates showed higher resistance to cephalothin and cefoxitin than to other cephem antibiotics and to gentamicin than to other aminogycosides. Our data indicated that fecal E. coli isolates from fishery workers showed higher antibiotic resistance than those of non-fishery workers (restaurant workers), especially to cephalothin, tetracycline, and trimethoprim–sulfamethoxazole (p < 0.05). However, rates of multidrug resistance were similar among the fishery workers and restaurant workers. Conclusion: Frequent use of antibiotics may cause increased antibiotic resistance in the human microbiome. PMID:24159534
Saidi, Bamusi; Mafirakureva, Prettimore; Mbanga, Joshua
Colibacillosis, a disease caused by avian pathogenic Escherichia coli (APEC), can lead to great economic losses in the poultry industry. The aim of this study was to determine the prevalence of antibiotic resistance and antibiotic resistance patterns in APEC in Zimbabwe. From 503 chickens diagnosed with colibacillosis, 103 E. coli isolates were obtained. Isolation and identification of E. coli were carried out using microscopy and biochemical tests. The disc diffusion method was used to determine antibiotic susceptibility of the isolates to 8 commercial antibiotics. Many isolates exhibited resistance to more than one antibiotic. Antibiogram profiles indicated maximum resistance to tetracycline (100%), bacitracin (100%), and cloxacillin (100%) and a high prevalence of resistance to ampicillin (94.1%). However; there were high prevalences of sensitivity to ciprofloxacin (100%) and gentamycin (97.1%). The isolates showed moderate rates of sensitivity to chloramphenicol and neomycin. All isolates in this study showed multidrug resistance because they were all resistant to 3 or more antibiotics. Seven multidrug resistance patterns were observed. The most common pattern (resistance to ampicillin, bacitracin, cloxacillin, and tetracycline) was exhibited by 30 isolates. Our findings show that there is emerging drug resistance in APEC associated with colibacillosis in Zimbabwe. The observed high level of multidrug resistance could hamper the treatment of colibacillosis in Zimbabwe.
Zeighami, Habib; Haghi, Fakhri; Hajiahmadi, Fahimeh; Kashefiyeh, Mehdi; Memariani, Mojtaba
Multi-drug-resistant (MDR) diarrheagenic Escherichia coli (DEC) has rapidly spread worldwide and represents the most serious threat to the management of diarrhea in developing countries. During the period from March 2011 to January 2012, a total of 450 stool samples of diarrheal children aged 0-60 months were studied. In order to detect enterotoxigenic E. coli (ETEC) and enterohemorrhagic E. coli (EHEC) simultaneously, a mixture of four primer pairs specific for eltB, estA, vt1, and vt2 genes was used in a multiplex PCR. Antimicrobial susceptibility testing was performed as the Clinical and Laboratory Standards Institute (CLSI) guidelines. A total of 140 (31·1%) DEC were isolated from 450 stool samples. Diarrheagenic E. coli exhibited high-level resistance to aztreonam (80·7%), amoxicillin (74·4%), and tetracycline (69·3%). Also, 86·4% of E. coli isolates were resistant to at least three different classes of antimicrobial agents and considered as MDR. The frequency of ETEC and EHEC pathotypes was 46·4 and 12·1%, respectively and all of these isolates were MDR. In conclusion, MDR ETEC continues to be an important agent associated with diarrhea in children from Tabriz, Iran.
Milley, D G; Sekla, L H
A colony enzyme-linked immunosorbent assay using the hydrophobic grid membrane filter format was developed for the isolation of verotoxigenic Escherichia coli from human and food samples. The method utilizes monoclonal antibodies directed against the verotoxins and is sensitive to all verotoxin 1- and/or 2-producing serotypes. E. coli that produced a minimum of 2 x 10(2) and 2 x 10(3) 50% cytotoxic doses per ml of verotoxins 1 and 2, respectively, were detectable. In a method comparison using human stool specimens, this procedure isolated 29% more E. coli O157 than did the standard sorbitol-MacConkey agar procedure, with no false-positive reactions. When applied to meat, 11 of 20 samples positive for verotoxin by polymyxin extraction yielded verotoxigenic E. coli of a variety of serotypes including O157:H7. Four false positives were noted. This procedure provides a sensitive means for the isolation of verotoxigenic E. coli and should facilitate recovery of those serotypes that are otherwise indistinguishable from nonpathogenic strains. PMID:8285713
Rasmussen, Mette Marie; Opintan, Japheth A.; Frimodt-Møller, Niels; Styrishave, Bjarne
The use of antibiotics in food animals is of public health concern, because resistant zoonotic pathogens can be transmitted to humans. Furthermore, global trade with food may rapidly spread multi-resistant pathogens between countries and even continents. The purpose of the study was to investigate whether imported chicken meat and meat from locally reared chicken are potential sources for human exposure to multi resistant Escherichia coli isolates. 188 samples from imported and locally produced chicken meat were sampled and analyzed. 153 bacteria isolates were successfully cultured and identified as E. coli using MALDI-ToF. Of these 109 isolates were from meat whereas the remaining 44 were isolated from the cloaca of locally reared live chickens. Antimicrobial susceptibility test was done on the identified E. coli isolates. Additionally, beta-lactamases production (ESBL and/or AmpC) were phenotypically confirmed on all isolates showing resistance to cefpodoxime. Beta-lactamase producing (BLP) E. coli meat isolates were further genotyped. Antimicrobial resistance to four antibiotic markers with highest resistance was detected more frequently in isolates from local chickens compared to imported chickens (tetracycline 88.9% vs. 57.5%, sulphonamide 75.0% vs. 46.6%, ampicillin 69.4% vs. 61.6% and trimethoprim 66.7% vs. 38.4%). Beta-lactamase production was found in 29 E. coli meat isolates, with 56.9% of them being multiple drug resistant (≥ 3). The predominant phylogroup identified was B1 followed by A and D, with similar distribution among the isolates from meat of locally reared chickens and imported chickens. Beta-lactamase producing genotype blaCTX-M-15 (50%; 10/20) was the most frequently drug resistant gene detected. More BLP E. coli isolates were found in imported chicken meat compared to locally reared chickens, demonstrating that these isolates may be spreading through food trade. In conclusion, both imported and locally produced chicken meats are potential
Rasmussen, Mette Marie; Opintan, Japheth A; Frimodt-Møller, Niels; Styrishave, Bjarne
The use of antibiotics in food animals is of public health concern, because resistant zoonotic pathogens can be transmitted to humans. Furthermore, global trade with food may rapidly spread multi-resistant pathogens between countries and even continents. The purpose of the study was to investigate whether imported chicken meat and meat from locally reared chicken are potential sources for human exposure to multi resistant Escherichia coli isolates. 188 samples from imported and locally produced chicken meat were sampled and analyzed. 153 bacteria isolates were successfully cultured and identified as E. coli using MALDI-ToF. Of these 109 isolates were from meat whereas the remaining 44 were isolated from the cloaca of locally reared live chickens. Antimicrobial susceptibility test was done on the identified E. coli isolates. Additionally, beta-lactamases production (ESBL and/or AmpC) were phenotypically confirmed on all isolates showing resistance to cefpodoxime. Beta-lactamase producing (BLP) E. coli meat isolates were further genotyped. Antimicrobial resistance to four antibiotic markers with highest resistance was detected more frequently in isolates from local chickens compared to imported chickens (tetracycline 88.9% vs. 57.5%, sulphonamide 75.0% vs. 46.6%, ampicillin 69.4% vs. 61.6% and trimethoprim 66.7% vs. 38.4%). Beta-lactamase production was found in 29 E. coli meat isolates, with 56.9% of them being multiple drug resistant (≥ 3). The predominant phylogroup identified was B1 followed by A and D, with similar distribution among the isolates from meat of locally reared chickens and imported chickens. Beta-lactamase producing genotype blaCTX-M-15 (50%; 10/20) was the most frequently drug resistant gene detected. More BLP E. coli isolates were found in imported chicken meat compared to locally reared chickens, demonstrating that these isolates may be spreading through food trade. In conclusion, both imported and locally produced chicken meats are potential
Edquist, Petra; Sandegren, Linus; Adler, Marlen; Tängdén, Thomas; Drobni, Mirva; Olsen, Björn; Melhus, Åsa
Members of the Enterobacteriaceae with extended-spectrum beta-lactamases (ESBLs) of the CTX-M type have disseminated rapidly in recent years and have become a threat to public health. In parallel with the CTX-M type expansion, the consumption and widespread use of silver-containing products has increased. To determine the carriage rates of silver resistance genes in different Escherichia coli populations, the presence of three silver resistance genes (silE, silP, and silS) and genes encoding CTX-M-, TEM-, and SHV-type enzymes were explored in E. coli isolates of human (n = 105) and avian (n = 111) origin. The antibiotic profiles were also determined. Isolates harboring CTX-M genes were further characterized, and phenotypic silver resistance was examined. The silE gene was present in 13 of the isolates. All of them were of human origin. Eleven of these isolates harbored ESBLs of the CTX-M type (P = 0.007), and eight of them were typed as CTX-M-15 and three as CTX-M-14. None of the silE-positive isolates was related to the O25b-ST131 clone, but 10 out of 13 belonged to the ST10 or ST58 complexes. Phenotypic silver resistance (silver nitrate MIC > 512 mg/liter) was observed after silver exposure in 12 of them, and a concomitant reduced susceptibility to piperacillin-tazobactam developed in three. In conclusion, 12% of the human E. coli isolates but none of the avian isolates harbored silver resistance genes. This indicates another route for or level of silver exposure for humans than that caused by common environmental contamination. Since silE-positive isolates were significantly more often found in CTX-M-positive isolates, it is possible that silver may exert a selective pressure on CTX-M-producing E. coli isolates. PMID:25128339
Sütterlin, Susanne; Edquist, Petra; Sandegren, Linus; Adler, Marlen; Tängdén, Thomas; Drobni, Mirva; Olsen, Björn; Melhus, Asa
Members of the Enterobacteriaceae with extended-spectrum beta-lactamases (ESBLs) of the CTX-M type have disseminated rapidly in recent years and have become a threat to public health. In parallel with the CTX-M type expansion, the consumption and widespread use of silver-containing products has increased. To determine the carriage rates of silver resistance genes in different Escherichia coli populations, the presence of three silver resistance genes (silE, silP, and silS) and genes encoding CTX-M-, TEM-, and SHV-type enzymes were explored in E. coli isolates of human (n = 105) and avian (n = 111) origin. The antibiotic profiles were also determined. Isolates harboring CTX-M genes were further characterized, and phenotypic silver resistance was examined. The silE gene was present in 13 of the isolates. All of them were of human origin. Eleven of these isolates harbored ESBLs of the CTX-M type (P = 0.007), and eight of them were typed as CTX-M-15 and three as CTX-M-14. None of the silE-positive isolates was related to the O25b-ST131 clone, but 10 out of 13 belonged to the ST10 or ST58 complexes. Phenotypic silver resistance (silver nitrate MIC > 512 mg/liter) was observed after silver exposure in 12 of them, and a concomitant reduced susceptibility to piperacillin-tazobactam developed in three. In conclusion, 12% of the human E. coli isolates but none of the avian isolates harbored silver resistance genes. This indicates another route for or level of silver exposure for humans than that caused by common environmental contamination. Since silE-positive isolates were significantly more often found in CTX-M-positive isolates, it is possible that silver may exert a selective pressure on CTX-M-producing E. coli isolates.
Ho, Pak-Leung; Chu, Yuki Pui-Shan; Lo, Wai-U; Chow, Kin-Hung; Law, Pierra Y; Tse, Cindy Wing-Sze; Ng, Tak-Keung; Cheng, Vincent Chi-Chung; Que, Tak-Lun
Previous work on the subclones within Escherichia coli ST131 predominantly involved isolates from Western countries. This study assessed the prevalence and antimicrobial resistance attributed to this clonal group. A total of 340 consecutive, non-duplicated urinary E. coli isolates originating from four clinical laboratories in Hong Kong in 2013 were tested. ST131 prevalence among the total isolates was 18.5 % (63/340) and was higher among inpatient isolates (23.0 %) than outpatient isolates (11.8 %, P<0.001), and higher among isolates from patients aged ≥65 years than from patients aged 18-50 years and 51-64 years (25.4 vs 3.4 and 4.0 %, respectively, P<0.001). Of the 63 ST131 isolates, 43 (68.3 %) isolates belonged to the H30 subclone, whereas the remaining isolates belonged to H41 (n = 17), H54 (n = 2) and H22 (n = 1). All H30 isolates were ciprofloxacin-resistant, of which 18.6 % (8/43) belonged to the H30-Rx subclone. Twenty-six (41.3 %) ST131 isolates were ESBL-producers, of which 19 had blaCTX-M-14 (12 non-H30-Rx, two H30-Rx and five H41), six had blaCTX-M-15 (five non-H30-Rx and one H30-Rx) and one was blaCTX-M-negative (H30). In conclusion, ST131 accounts for a large share of the antimicrobial-resistant E. coli isolates from geriatric patients. Unlike previous reports, ESBL-producing ST131 strains mainly belonged to non-H30-Rx rather than the H30-Rx subclone, with blaCTX-M-14 as the dominant enzyme type.
Sullivan, M A; Bock, R M
Nonsense mutations in lacI have been shown to be useful as indicators of the efficiency of nonsense suppression. From strains containing supE and a lacI nonsense mutation, selection for LacI- mutants has resulted in the isolation of four antisuppressor mutations. Tn10 insertions linked to these mutations were isolated and used to group the four mutations into three loci. The asuA1 and asuA2 mutations are linked to trp, reduce suppression by supE approximately twofold, and affect a variety of suppressors. The asuB3 mutation was mapped by P1 cotransduction to rpsL but does not confer resistance to streptomycin. The asuC4 mutation reduced suppression by supE by 95% and was shown biochemically to result in the loss of two pseudouridine modifications from the 3' side of the anticodon stem and loop of tRNA2Gln. This mutation is linked to purF, suggesting that it is a new allele of hisT. Images PMID:3918006
Kudva, I T; Hatfield, P G; Hovde, C J
The isolation and characterization of Escherichia coli O157:H7 and non-O157 Shiga toxin-producing E. coli (STEC) strains from sheep are described. One flock was investigated for E. coli O157:H7 over a 16-month period that spanned two summer and two autumn seasons. Variation in the occurrence of E. coli O157:H7-positive sheep was observed, with animals being culture positive only in the summer months but not in the spring, autumn, or winter. E. coli O157:H7 isolates were distinguished by pulsed-field gel electrophoresis (PFGE) of chromosomal DNA and toxin gene restriction fragment length polymorphism (RFLP) analysis. Ten PFGE patterns and five RFLP patterns, identified among the isolates, showed that multiple E. coli O157:H7 strains were isolated from one flock, that a single animal simultaneously shed multiple E. coli O157:H7 strains, and that the strains shed by individuals changed over time. E. coli O157:H7 was isolated only by selective enrichment culture off 10 g of ovine feces. In contrast, strains of eight STEC serotypes other than O157:H7 were cultured from feces of sheep from a separate flock without enrichment. The predominant non-O157 STEC serotype found was O91:NM (NM indicates nonmotile), and others included O128:NM, O88:NM, O6:H49, and O5:NM. Irrespective of serotype, 98% of the ovine STEC isolates possessed various combinations of the virulence-associated genes for Shiga toxin(s) and the attaching-and-effacing lesion (stx1, stx2, and eae), suggesting their potential for human pathogenicity. The most common toxin-eae genotype was positive for stx1, stx2, and eae. A Vero cell cytotoxicity assay demonstrated that 90% of the representative STEC isolates tested expressed the toxin gene. The report demonstrates that sheep transiently shed a variety of STEC strains, including E. coli O157:H7, that have potential as human pathogens. PMID:9157149
Janezic, Kristopher J; Ferry, Blake; Hendricks, Eric W; Janiga, Brian A; Johnson, Tiffany; Murphy, Samantha; Roberts, Morgan E; Scott, Sarah M; Theisen, Alexandra N; Hung, Kai F; Daniel, Steven L
A common member of the intestinal microbiota in humans and animals is Escherichia coli. Based on the presence of virulence factors, E. coli can be potentially pathogenic. The focus of this study was to isolate E. coli from untreated surface waters (37 sites) in Illinois and Missouri and determine phenotypic and genotypic diversity among isolates. Water samples positive for fecal coliforms based on the Colisure® test were streaked directly onto Eosin Methylene Blue (EMB) agar (37°C) or transferred to EC broth (44.5°C). EC broth cultures producing gas were then streaked onto EMB agar. Forty-five isolates were identified as E. coli using API 20E and Enterotube II identification systems, and some phenotypic variation was observed in metabolism and fermentation. Antibiotic susceptibility of each isolate was also determined using the Kirby-Bauer Method. Differential responses to 10 antimicrobial agents were seen with 7, 16, 2, and 9 of the isolates resistant to ampicillin, cephalothin, tetracycline, and triple sulfonamide, respectively. All of the isolates were susceptible or intermediate to amoxicillin, ciprofloxacin, polymyxin B, gentamicin, imipenem, and nalidixic acid. Genotypic variation was assessed through multiplex Polymerase Chain Reaction for four virulence genes (stx1 and stx2 [shiga toxin], eaeA [intimin]; and hlyA [enterohemolysin]) and one housekeeping gene (uidA [β-D-glucuronidase]). Genotypic variation was observed with two of the isolates possessing the virulence gene (eaeA) for intimin. These findings increase our understanding of the diversity of E. coli in the environment which will ultimately help in the assessment of this organism and its role in public health. PMID:23539437
Johnson, J R; Weissman, S J; Stell, A L; Trintchina, E; Dykhuizen, D E; Sokurenko, E V
Escherichia coli NU14, a cystitis isolate used to study the pathogenesis of cystitis and to develop a FimH (type 1 fimbrial adhesin) vaccine, was assessed for extended virulence genotype, phylogenetic background, and FimH sequence and binding phenotype(s). NU14 exhibited the same virulence genotype and was derived from the same (meningitis- and cystitis-associated) subclone of E. coli O18:K1:H7 as the archetypal neonatal bacterial meningitis (NBM) isolate RS218. NU14 also displayed the same Ser62Ala FimH polymorphism as did NBM isolates RS218 and IHE3034-conferring both collagen binding and a distinct monomannose binding capability (which characterizes uropathogenic but not commensal E. coli and dramatically increases adherence to uroepithelial cells). These findings establish that strain NU14 exhibits numerous urovirulence-associated traits and derives from the single most prevalent clonal group in acute cystitis. They provide further evidence of clonal and pathotypic similarities between cystitis and NBM isolates of E. coli O18:K1:H7.
Mateus, Luisa; Henriques, Sofia; Merino, Carolina; Pomba, Constança; Lopes da Costa, Luís; Silva, Elisabete
Pyometra is the most common diestrual uterine disease of bitches. Escherichia coli is the most frequent bacterium isolated from the uterine content of pyometra uteri and it is associated with the most severe clinical signs, leading to endotoxemia and sepsis. In this study, canine E. coli isolates from pyometra (n=31), cystitis (n=23) and fecal (n=26) origin were compared regarding the prevalence of 23 potential virulence traits (15 virulence factor (VF) genes and 8 pathogenicity associated islands-PAIs), detected by PCR assays. Overall, there was a considerable overlap between pyometra, cystitis and fecal isolates regarding the phylogenetic grouping and virulence traits. Virulence traits more prevalent in pyometra than in cystitis and fecal isolates included two PAIs (PAI IV536 and PAI ICFT073) and three VF genes (sfa/focDE, fyuA and chuA). Regardless the isolates' origin, the average number of virulence traits per strain was higher in B2 than in the other phylogenetic groups (A, B1 and D). The prevalence of phylogenetic group B2 was significantly higher in pyometra (94%) than in cystitis (48%) and fecal (39%) isolates. In conclusion, pyometra isolates have a high potential of virulence and a broad virulence genotype, although being similar to a subset of cystitis and fecal isolates. This leads to the suggestion that cystitis and fecal isolates may be able to induce pyometra in receptive hosts.
Puño-Sarmiento, Juan; Medeiros, Leonardo; Chiconi, Carolina; Martins, Fernando; Pelayo, Jacinta; Rocha, Sérgio; Blanco, Jorge; Blanco, Miguel; Zanutto, Marcelo; Kobayashi, Renata; Nakazato, Gerson
Escherichia coli are gut microbiota bacteria that can cause disease in some humans and other animals, including dogs and cats that humans often keep as pets. Diarrheagenic E. coli (DEC) strains are classified into six categories: enteropathogenic (EPEC), enterotoxigenic (ETEC), Shiga toxin-producing (STEC), enteroinvasive (EIEC), enteroaggregative (EAEC), and diffuse-adhering E. coli (DAEC). In this study 144 and 163 E. coli colonies were isolated from the fecal samples of 50 dogs and 50 cats, respectively, with and without diarrhea from a Veterinary Hospital (clinical isolates). The virulence factors were determined using multiplex Polymerase Chain Reaction. Adherence assays, antibacterial susceptibility and serotyping (somatic or flagellar antigens) were performed on DEC isolates. We found 25 (17.4%) and 4 (2.5%) DEC strains isolated from dogs and cats, respectively. Only the EPEC and EAEC pathotypes were found in both animals. Meanwhile, genes from other pathotypes (STEC, EIEC, and ETEC) were not found in these clinical isolates. All of the DEC strains showed mannose-resistant adherence to HEp-2 and HeLa cells, and aggregative adherence was predominant in these isolates. Multiresistant strains to antimicrobials were found in most DEC strains including usual and unusual antimicrobials in veterinary practices. The serotypes of these DEC isolates were variable. The ONT serotype was predominant in these isolates. Some serotypes found in our study were described to human DEC. Here, we demonstrate that pets carry virulent DEC genes, which are mainly strains of EPECs and EAECs. The presence of these virulence factors in isolates from animals without diarrhea suggests that pets can act as a reservoir for human infection.
Meyer, R. C.; Rhoades, H. E.; Simon, J.
Newborn, germfree piglets were susceptible to Escherichia coli associated with human, nonenteric infections and should provide a useful model in the study of generalized E. coli infections. PMID:4557565
Mercado, E. C.; Rodríguez, S. M.; Elizondo, A. M.; Marcoppido, G.; Parreño, V.
Shiga toxin-producing Escherichia coli belonging to serotype O26:H11 was isolated from a 2-month-old guanaco with severe watery diarrhea. E. coli colonies carried the stx1 and eae genes, showed localized adherence to HEp-2 cells, and produced enterohemolysin. A serological response to lipopolysaccharide O26 was observed at the onset of diarrhea. PMID:15472347
Ortega-Paredes, D; Barba, P; Zurita, J
Colistin resistance mediated by the mcr-1 gene has been reported worldwide, but to date not from the Andean region, South America. We report the first clinical isolate of Escherichia coli harbouring the mcr-1 gene in Ecuador. The strain was isolated from peritoneal fluid from a 14-year-old male with acute appendicitis, and subjected to molecular analysis. The minimum inhibitory concentration of colistin for the strain was 8 mg/ml and it was susceptible to carbapenems but resistant to tigecycline. The strain harboured mcr-1 and bla CTX-M-55 genes and was of sequence type 609. The recognition of an apparently commensal strain of E. coli harbouring mcr-1 serves as an alert to the presence in the region of this recently described resistance mechanism to one of the last line of drugs available for the treatment of multi-resistant Gram-negative infections.
Orden, José Antonio; Ruiz-Santa-Quiteria, José Antonio; García, Silvia; Cid, Dolores; de la Fuente, Ricardo
The in vitro activities of several cephalosporins and quinolones against 195 strains of Escherichia coli isolated from dairy calves affected by neonatal diarrhea were determined. One hundred thirty-seven of these strains produced one or more potential virulence factors (F5, F41, F17, cytotoxic necrotizing factor, verotoxin, and the eae gene), but the remaining 58 strains did not produce any of these factors. From 11 to 18% of the E. coli strains were resistant to cephalothin, nalidixic acid, enoxacin, and enrofloxacin. However, cefuroxime, cefotaxime, and cefquinome were highly effective against the E. coli isolates tested. Some significant differences (P < 0.05) in resistance to quinolones between the strains producing potential virulence factors and nonfimbriated, nontoxigenic, eae-negative strains were found. Thus, eae-positive, necrotoxigenic, and verotoxigenic (except for nalidixic acid) E. coli strains were significantly more sensitive to nalidixic acid, enoxacin, and enrofloxacin than nonfimbriated, nontoxigenic, eae-negative strains. Moreover, eae-positive strains were significantly more sensitive to enoxacin and enrofloxacin than F5-positive strains. Thus, the results of this study suggest that the bovine E. coli strains that produce some potential virulence factors are more sensitive to quinolones than those that do not express these factors. PMID:10049259
Sacristán, C; Esperón, F; Herrera-León, S; Iglesias, I; Neves, E; Nogal, V; Muñoz, M J; de la Torre, A
The aim of this study was to determine the presence of virulence genes and antibiotic resistance profiles in 164 Escherichia coli strains isolated from birds (feral pigeons, hybrid ducks, house sparrows and spotless starlings) inhabiting urban and rural environments. A total of eight atypical enteropathogenic E. coli strains were identified: one in a house sparrow, four in feral pigeons and three in spotless starlings. Antibiotic resistance was present in 32.9% (54) of E. coli strains. The dominant type of resistance was to tetracycline (21.3%), ampicillin (19.5%) and sulfamethoxazole (18.9%). Five isolates had class 1 integrons containing gene cassettes encoding for dihydrofolate reductase A (dfrA) and aminoglycoside adenyltransferase A (aadA), one in a feral pigeon and four in spotless starlings. To our knowledge, the present study constitutes the first detection of virulence genes from E. coli in spotless starlings and house sparrows, and is also the first identification worldwide of integrons containing antibiotic resistance gene cassettes in E. coli strains from spotless starlings and pigeons.
Marinho, Catarina; Silva, Nuno; Pombo, Sofia; Santos, Tiago; Monteiro, Ricardo; Gonçalves, Alexandre; Micael, Joana; Rodrigues, Pedro; Costa, Ana Cristina; Igrejas, Gilberto; Poeta, Patrícia
The prevalence of antibiotic resistance and the implicated mechanisms of resistance were evaluated in Enterococcus spp. and Escherichia coli, isolated from a total of 250 faecal samples of echinoderms collected from Azorean waters (Portugal). A total of 144 enterococci (120 Enterococcus faecium, 14 E. hirae, 8 E. faecalis, 2 E. gallinarum) and 10 E. coli were recovered. High percentages of resistance in enterococci were found for erythromycin, ampicillin, tetracyclin and ciprofloxacin. The erm(A) or erm(B), tet(M) and/or tet(L), vat(D), aac(6')-aph(2″) and aph(3')-IIIa genes were found in isolates resistant to erythromycin, tetracycline, quinupristin/dalfopristin, high-level gentamicin and high-level kanamycin, respectively. Resistance in E. coli isolates was detected for streptomycin, amikacin, tetracycline and tobramycin. The aadA gene was found in streptomycin-resistant isolates and tet(A)+tet(B) genes in tetracycline-resistant isolates. The data recovered are essential to improve knowledge about the dissemination of resistant strains through marine ecosystems and the possible implications involved in transferring these resistances either to other animals or to humans.
Monroy, Maria A R; Knöbl, Terezinha; Bottino, José A; Ferreira, Claudete S Astolfi; Ferreira, Antonio J Piantino
Thirty isolates of Escherichia coli from broiler breeders with salpingitis were studied. Using the slide agglutination test, the isolates were found to belong to serogroups O1, O2, O5, O36, O45, O53 and O78. Pathogenicity for day-old chicks was determined by air sac inoculation and isolates were categorized as having high, intermediate or low virulence. Growth on iron starvation medium was observed together with aerobactin production. Based on the results of in vitro adherence tests, attachment to oviduct epithelium from old birds was found to be superior to that observed using corresponding material from young birds. DNA hybridization testing for type 1, P, and S fimbriae revealed predominant expression of type 1, correlating with mannose-sensitive hemagglutination using guinea-pig erythrocytes. In this study, P and S fimbriae were not considered to be important adherence factors. Study findings would suggest that, as far as salpingitis is concerned, type 1 fimbriae can play an important role in E. coli infection in breeders. An interesting result to emerge from the study was the observation that E. coli isolates were completely resistant to serum from young breeders, whereas they were completely sensitive using serum from older breeders. Based on serogroups involved, pathogenicity for day-old chicks and virulence indicators, the salpingitis isolates were similar to those from cases of chronic respiratory disease.
Rocha, Rafael dos Santos; Leite, Lana Oliveira; de Sousa, Oscarina Viana; Vieira, Regine Helena Silva dos Fernandes
The contamination of seafood by bacteria of fecal origin, especially Escherichia coli, is a widely documented sanitary problem. The objective of the present study was to isolate E. coli strains from the gills, muscle, and body surface of farmed Nile tilapias (Oreochromis niloticus) fresh-marketed in supermarkets in Fortaleza (Ceará, Brazil), to determine their susceptibility to antibiotics of different families (amikacin, gentamicin, imipenem, cephalothin, cefotaxime, ciprofloxacin, aztreonam, ampicillin, nalidixic acid, tetracycline, and sulfametoxazol-trimetoprim), and to determine the nature of resistance by plasmid curing. Forty-four strains (body surface = 25, gills = 15, muscle = 4) were isolated, all of which were susceptible to amikacin, aztreonam, cefotaxime, ciprofloxacin, gentamicin, and imipenem. Gill and body surface samples yielded 11 isolates resistant to ampicillin, tetracycline, and sulfametoxazol-trimetoprim, 4 of which of plasmidial nature. The multiple antibiotic resistance index was higher for strains isolated from body surface than from gills. The overall high antibiotic susceptibility of E. coli strains isolated from fresh-marketed tilapia was satisfactory, although the occasional finding of plasmidial resistance points to the need for close microbiological surveillance of the farming, handling, and marketing conditions of aquaculture products. PMID:24808957
Timonin, M E; Poissant, J; McLoughlin, P D; Hedlin, C E; Rubin, J E
The feral horses of Sable Island are a geographically isolated population located ∼160 km off the east coast of Nova Scotia, Canada. Because these horses have no contact with domestic animals, have minimal contact with people, and have never received antimicrobials, they offer a unique opportunity to study the dissemination of antimicrobial resistance in unmanaged populations. As part of an ongoing multidisciplinary and individual-based monitoring program, we collected feces from 508 geolocalized horses (92% of the total population) between July and September 2014. We selectively cultured Escherichia coli on MacConkey and CHROMagar ESBL media. Antimicrobial susceptibilities were determined, and organisms resistant to β-lactam antimicrobials were screened for β-lactamase genes by PCR. Escherichia coli was recovered from 146 (28.7%) individuals, and the majority of isolates (97%) were susceptible to all drugs tested. Resistance to tetracycline was most common, including organisms isolated from 4 (2.7%) of the colonized horses. A single isolate resistant to ampicillin, ceftriaxone, and ceftiofur was identified, which possessed the CTX-M-1 gene. Our findings demonstrate that although antimicrobial resistance is not common in this remote population, clinically relevant resistance genes are present.
Vollmerhausen, T L; Katouli, M
Urinary tract infection (UTI) is common amongst children and recurs in 10-30 % of cases. The differences between Escherichia coli strains causing UTI among hospitalised children and adults remains to be fully elucidated. Here, we examined the genetic relatedness and virulence gene (VG) profiles of a collection of E. coli causing UTI among hospitalised children and adults. Genetic relatedness among the strains was investigated using random amplified polymorphic DNA (RAPD) analysis and the strains were characterised using a combination of phylogenetic grouping, the ability to form biofilm and the presence of antigen 43 (Ag43) and its five known alleles, as well 20 VGs associated with uropathogenic E. coli (UPEC). RAPD analysis resolved six major clusters, with two clusters (A and B) consisting almost exclusively of E. coli isolated from children. Isolates from children had a higher prevalence of alpha-haemolysin (hlyA, p < 0.05) and group II capsular polysaccharide synthesis genes (kpsMT II, p < 0.01) than adults. In contrast, E. coli strains from adults had a higher prevalence of invasive ibeA (p < 0.05) and Ag43 (agn43) (p < 0.05) genes, and produced significantly (p < 0.001) more biofilm than E. coli from children. Adult isolates also carried significantly (p < 0.05) more agn43 allele RS218 compared to isolates from children, which carried significantly (p < 0.05) more of the agn43 allele bCFT073. Our results suggest that bacterial virulence factors play an important role in UTI among hospitalised children; however, further research will determine whether these findings apply to a larger cohort and other clinical settings for UTI in children and adults.
Sun, J D; Huang, S F; Yang, S S; Pu, S L; Zhang, C M; Zhang, L P
Although heteroresistance is common in a wide range of microorganisms, carbapenem heteroresistance among invasive Escherichia coli infections has not been reported. The objective of this study was to evaluate the clinical significance of carbapenem heteroresistance and to identify risk factors for its acquisition. A case-control study was conducted at a 3200-bed teaching hospital in Chongqing, southwestern China. Successive and non-duplicate nosocomial E. coli isolates (n = 332) were obtained from July 2011 to June 2013. Bloodstream isolates made up 50.6% of the strains collected. The rates of heteroresistance were 25.0% to imipenem, 17.2% to ertapenem, and 3.9% to meropenem. The population analysis profile revealed the presence of subpopulations with higher carbapenem resistance, showing MICs ranging from 2.0-128.0mg/L. Male gender, invasive intervention, antibiotic use and bacterial extended-spectrum β-lactamase (ESBL) production contributed to invasive infections by carbapenem-heteroresistant E. coli (CHEC). The production of ESBL was identified as the common independent risk factor for heteroresistance to both ertapenem and imipenem. Pulsed-ﬁeld gel electrophoresis revealed clonal diversity among the CHEC isolates. Most importantly, characterization of two successive E. coli strains isolated from the same patient indicated that carbapenem resistance evolved from heteroresistance. In conclusion, the high prevalence of heteroresistance to carbapenem among invasive E. coli merits great attention. Routine detection of ESBLs and the prudent use of imipenem and ertapenem are advocated. The early targeted intervention should be formulated to reduce CHEC infection and carbapenem resistance of E. coli.
Jiang, Yun; Yin, Shuang; Dudley, Edward G; Cutter, Catherine N
Shiga toxin-producing Escherichia coli (STEC) strains, including those of O157:H7 and the "big six" serogroups (i.e., O26, O45, O103, O111, O121, and O145) are food-borne pathogens that pose a serious health threat to humans. Ruminants, especially cattle, are a major reservoir for O157 and non-O157 STEC. In the present study, 115 E. coli strains isolated from small and very small beef processing plants were screened for virulence genes (stx1, stx2, eae) using a multiplex polymerase chain reaction (PCR). Thirteen (11.3%) of the 115 isolates tested positive for stx1, stx2, or eae genes, but only 4 (3.5%) tested positive for either stx1 or stx2. A multiplex PCR reaction targeting eight O-serogroups (O26, O45, O103, O111, O113, O121, O145, O157) identified 12 isolates as O26, O103, O111, or O145, with E. coli O26 being the most predominant serogroup (61.5%). The thirteen isolates were further analyzed using Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) subtyping. Consistent with previous studies, CRISPR alleles from strains of the same serogroup were similar in their spacer content and order, regardless of the isolation source. A completely different CRISPR allele was observed in one isolate ("7-J") which exhibited a different O-serogroup (O78). Our results confirmed previous findings that CRISPR loci are conserved among phylogenetically-related strains. In addition, 8 E. coli O26 isolates and a collection of 42 E. coli O26 isolates were screened for 12 enterohemorrhagic E. coli-specific genes. Seven genes (ECs848-Hypothetical Protein, ECs2226-Hypothetical Protein, ECs3857-nleB, ECs3858-Hypothetical Protein, ECs4552-escF, ECs4553-Hypothetical Protein, and ECs4557-sepL) were found in all 50 isolates. An additional 5 genes (ECs1322-ureA urease subunit γ, ECs1323-ureB urease subunit β, ECs1326-ureF, ECs1561-Hypothetical Protein, and ECs1568-Hypothetical Protein) were found to be highly prevalent in isolates from human sources, while lower in
Ikuta, Nilo; De Oliveira Solla Sobral, Fabiana; Lehmann, Fernanda Kieling Moreira; da Silveira, Proença Vinicius; de Carli, Silvia; Casanova, Yara Silva; Celmer, Álvaro José; Fonseca, André Salvador Kazantzi; Lunge, Vagner Ricardo
Avian pathogenic Escherichia coli (APEC) isolates are currently differentiated from nonpathogenic strains by classical PCR of virulence genes. This study improves the detection of the five main virulence genes used for APEC detection with the development of duplex and single Taqman real-time PCR to these targets. Primers and probes targeted to ompT, hlyF, iroN, iutA, and iss genes were designed and used in the implementation of single (iss) and duplex (hlyF/ompT and iroN/iutA) Taqman PCR assays. All five virulence genes of E coli strains were successfully detected by classical and Taqman real-time (single and duplex) PCR. A panel of 111 E coli isolates, obtained from avian samples collected in different Brazilian regions between 2010 and 2011, were further tested by both assays. Complete agreement was observed in the detection of four genes, ompT, hlyF, iron, iutA, but not for iss. This issue was addressed by combining the forward primer of the classical PCR to the new iss reverse primer and probe, resulting in complete agreement for all five genes. In total, 61 (55%) Brazilian E. coli isolates were detected as APEC, and the remaining 50 (45%) as avian fecal E. coli (AFEC). In conclusion, classical and Taqman real-time PCR presented exactly the same analytical performance for the differentiation of APEC and AFEC isolates. The developed real-time Taqman PCR assays could be used for the detection and differentiation of APEC isolates.
Nakhjavani, Farrokh Akbari; Emaneini, Mohammad; Hosseini, Hossein; Iman-Eini, Hossein; Aligholi, Marzieh; Jabalameli, Fereshteh; Haghi-Ashtiani, Mohammad Taghi; Taherikalani, Morovat; Mirsalehian, Akbar
Diarrhoea continues to be one of the most common causes of morbidity and mortality among infants and children in developing countries. To investigate the incidence, antimicrobial resistance and genetic relationships of enteropathogenic Escherichia coli (EPEC) in children with diarrhoea, a total of 612 stool specimens were collected in Tehran, Iran, and cultured to isolate strains of EPEC. The disc diffusion method was used to determine the susceptibility of the isolates according to the Clinical and Laboratory Standards Institute (CLSI) guidelines. The presence of eae, stx and bfp-A genes was determined by PCR. The genetic relationships between EPEC isolates were determined by pulsed-field gel electrophoresis (PFGE). Out of the 412 strains of E. coli obtained from 612 diarrhoeal stool specimens, 23 (5.6 %) were identified as EPEC, of which seven (30.4 %) were classified as typical strains of EPEC and 16 (69.6 %) were classified as atypical. Out of the 23 EPEC isolates, 69.5 % were resistant to ampicillin, 39.1 % were resistant to tetracycline and cotrimoxazole, 30.4 % were resistant to cefpodoxime, ceftazidime, ceftriaxone and aztreonam, and 26.1 % were resistant to imipenem. The isolates were classified into 21 pulsotypes by PFGE profiles. The present study shows that typical and atypical EPEC isolates displayed considerable heterogeneity in PFGE profiles and EPEC infections were only sporadic in Tehran. Overall 69 % of isolates were resistant to at least one of the antibiotics tested.
Grillo-Puertas, M; Martínez-Zamora, MG; Rintoul, MR; Soto, SM; Rapisarda, VA
K-12 Escherichia coli cells grown in static media containing a critical phosphate (Pi) concentration ≥25 mM maintained a high polyphosphate (polyP) level in stationary phase, impairing biofilm formation, a phenomenon that is triggered by polyP degradation. Pi concentration in human urine fluctuates according to health state. Here, the influence of environmental Pi concentration on the occurrence of virulence traits in uropathogenic E. coli (UPEC) isolated from acute prostatitis patients was evaluated. After a first screening, 3 isolates were selected according to differential biofilm formation profiles depending on media Pi concentration. For each isolate, biofilm positive and negative conditions were established. Regardless of the isolate, biofilm formation capacity was accompanied with curli and cellulose production and expression of some key virulence factors associated with adhesion. When the selected isolates were grown in their non-biofilm-forming condition, low concentrations of nalidixic acid and ciprofloxacin induced biofilm formation. Interestingly, similar to laboratory strains, polyP degradation induced biofilm formation in the selected isolates. Data demonstrated the complexity of UPEC responses to environmental Pi and the importance of polyP metabolism in the virulence of clinical isolates. PMID:26083279
Novais, Ângela; Pires, João; Ferreira, Helena; Costa, Luísa; Montenegro, Carolina; Vuotto, Claudia; Donelli, Gianfranco; Coque, Teresa M.
The characterization of a broad representative sample of ST131 Escherichia coli isolates from different origins and settings (1991 to 2010) revealed that this clonal group has likely diversified recently and that the expansion of particular variants has probably been favored by the capture of diverse, multidrug-resistant IncFII plasmids (pC15-1a, pEK499, pKF3-140-like). The low ability to adhere and to grow as biofilm that was detected in this study suggests unknown mechanisms for the persistence of this clonal group which need to be further explored. PMID:22491693
Meier, C; Oelschlaeger, T A; Merkert, H; Korhonen, T K; Hacker, J
Escherichia coli isolates that cause meningitis in newborns are able to invade the circulation and subsequently cross the blood-brain barrier. One mechanism for traversing the blood-brain barrier might involve transcytosis through the endothelial cells. The ability of the meningitis isolate E. coli IHE3034, of serotype 018:K1:H7, to invade epithelial (T24) and endothelial (EA-hy926) cells was investigated by the standard gentamicin survival assay and by electron microscopy. Human bladder epithelial and endothelial cells were efficiently invaded by strain IHE3034, whereas epithelial human colon Caco-2 cells, canine kidney MDCK cells, and the opossum [correction of opposum] epithelial kidney cell line OK were not invaded. The ability to invade human epithelial cells of the bladder could also be demonstrated for several other newborn meningitis E. coli strains and one septicemic E. coli strain. Studies utilizing inhibitors which act on eukaryotic cells revealed a dependence on microfilaments as well as on microtubules in the process of E. coli IHE3034 entry into T24 and EA-hy926 cells. These results indicated that cell cytoskeletal rearrangements are involved in bacterial uptake and suggest that there are either two pathways (microtubule dependent and microfilament dependent) or one complex pathway involving both microtubules and microfilaments. The intracellular IHE3034 organisms were contained in a host-membrane-confined compartment mainly as single microorganisms. Intracellular replication of 1HE3034 was not detected, nor did the number of intracellular bacteria decrease significantly during a 48-h period. The ability of E. coli O18:K1 to invade and survive within certain eukaryotic cells may be another virulence factor of meningitis-associated E. coli. PMID:8698457
Khalil, Uzma; Younus, Mahwish; Asghar, Naeem; Siddiqui, Fariha; Gómez-Duarte, Oscar G; Wren, Brendan W; Bokhari, Habib
Enteroaggregative Escherichia coli (EAEC) are a leading cause of diarrhea among children. The objective of this study was to define the frequency of EAEC among diarrheal children from flood-affected areas as well as sporadic cases, determine multidrug resistance, and evaluation of virulence using an in vivo model of pathogenesis. Stool samples were collected from 225 diarrheal children from 2010 to 2011 from flood-affected areas as well as from sporadic cases in Pakistan. Identified EAEC isolates were characterized by phylogrouping, antibiotic resistance patterns including the extended-spectrum beta lactamase spectrum, single nucleotide polymorphism detection in gyrA and parC, and virulence potential using wax worm, G. mellonella. A total of 35 (12.5%) confirmed EAEC isolates were identified among 225 E. coli isolates. EAEC isolates displayed high resistance to tetracycline, ampicillin, and cefaclor. A total of 34.28% were ESBL positive. Single nucleotide polymorphism detection revealed 37.14% and 68.57% isolates were positive for SNPs in gyrA (A660 -T660 ) and parC (C330 -T330 ), respectively. Phylogrouping revealed that B2 phylogroup was more prevalent among all EAEC isolates tested followed by D, A, B1, and non-typeable (NT). Infection of G. mellonella with EAEC showed that killing infective dose was 100% higher than E. coli DH5 alpha control. EAEC are prevalent among Pakistani children with diarrhea, they are highly resistant to antibiotics, and predominantly fall into B2 phylogroup. Epidemiologic surveillance of EAEC and other E. coli pathotypes is critical to assess not only the role of these pathogens in diarrheal disease but also to determine the extent of multidrug resistance among the population.
Blanco, J E; Blanco, M; Blanco, J; Mora, A; Balaguer, L; Mouriño, M; Juarez, A; Jansen, W H
A total of 305 Escherichia coli strains isolated from diarrheic and healthy rabbits in 10 industrial fattening farms from different areas of Spain were serotyped, biotyped, and tested for the presence of the eae gene and toxin production. The characteristics found in strains isolated from healthy rabbits were generally different from those observed in E. coli strains associated with disease. Thus, strains with the eae gene (74% versus 22%); strains belonging to serogroups O26, O49, O92, O103, and O128 (64% versus 12%); rhamnose-negative strains (51% versus 5%); and rhamnose-negative O103 strains with eae genes present (41% versus 1%) were significantly (P < 0.001 in all cases) more frequently detected in isolates from diarrheic animals than in those from healthy rabbits. Whereas a total of 35 serogroups and 17 biotypes were distinguished, the majority of the strains obtained from diarrheic rabbits belonged to only four serobiotypes, which in order of frequency were O103:B14 (72 strains), O103:B6 (16 strains), O26:B13 (12 strains), and O128:B30 (12 strains). These four serobiotypes accounted for 48% (112 of 231) and 5% (4 of 74) of the E. coli strains isolated from diarrheic and healthy rabbits, respectively. Only six strains were toxigenic (three CNF1+, two CNF2+, and one VT1+). We conclude that enteropathogenic E. coli strains that possess the eae gene are a common cause of diarrhea in Spanish rabbit farms and that the rhamnose-negative highly pathogenic strains of serotype O103:K-:H2 and biotype B14 are especially predominant. Detection of the eae gene is a useful method for the identification of enteropathogenic E. coli strains from rabbits. However, a combination of serogrouping and biotyping may be sufficient to accurately identify the highly pathogenic strains for rabbits. PMID:8940455
Stephenson, Stacy A M; Brown, Paul D
Quinolone resistance is generally caused by chromosomal mutations, but has been more recently found associated with the plasmid-mediated qnr genes. The objective of this study was to screen and analyse polymorphisms of integrons in clinical isolates of Escherichia coli in Jamaica. Previous studies in Jamaica identified fluoroquinolone resistance in predominantly uropathogenic E. coli clinical isolates: 45% harbouring qnrA, qnrB and/or qnrS, and 17% were (Extended-spectrum beta-lactamase) ESBL-producers. These isolates were analysed for the presence and variation of class 1 and 2 integrase genes, 5'- and 3'- conserved segments and the Orf513 recombinase gene by primer-specific polymerase chain reaction (PCR) and restriction fragment-length polymorphism (RFLP). Results indicated integron-encoded integrases in 93% of isolates primarily harbouring class 1 integrase genes; four of 58 isolates carried both classes. The Orf513 and 5'- and 3'-conserved segment (CS) regions were identified in 83% and 55% of the isolates respectively. RFLP evaluation of the 5'- and 3'-CS regions in int1-positive strains yielded two main types. The reduced diversity, but wide dispersion of class 1 integrons harbouring qnr genes may give rise to the conservation of the mobile genetic elements in which they are carried.
Yamamoto, Shiori; Nakano, Motoki; Kitagawa, Wataru; Tanaka, Michiko; Sone, Teruo; Hirai, Katsuya; Asano, Kozo
The emergence of multiple-antibiotic-resistance bacteria is increasing, which is a particular concern on livestock farms. We previously isolated 1,347 antimicrobial-resistant (AMR) Escherichia coli strains from the feces of beef cattle on 14 Japanese farms. In the present study, the genetic backgrounds and phylogenetic relationships of 45 AMR isolates were characterized by the chromosome phylotype, AMR phenotype, AMR genotype, and plasmid type. These isolates were classified into five chromosome phylotypes, which were closely linked to the farms from which they were isolated, suggesting that each farm had its own E. coli phylotype. AMR phenotype and plasmid type analyses yielded 8 and 14 types, all of which were associated with the chromosomal phylotype and, thus, to the original farms. AMR genotype analysis revealed more variety, with 16 types, indicating both inter- and intra-farm diversity. Different phylotype isolates from the same farm shared highly similar plasmid types, which indicated that plasmids with AMR genes could be transferred between phylotypes, thereby generating multi-antibiotic-resistant microorganisms. This ecological study demonstrated that the chromosome phylotype was strongly correlated with the farm from which they were isolated, while the AMR phenotype, genotype, and plasmid type were generally correlated with the chromosome phylotype and farm source.
Giammanco, A; Maggio, M; Giammanco, G; Morelli, R; Minelli, F; Scheutz, F; Caprioli, A
Fifty-five Escherichia coli strains belonging to enteropathogenic E. coli (EPEC) serogroups were examined for phenotypic and genetic factors associated with virulence. The strains were isolated in Italy from children with diarrhea and identified as EPEC by clinical laboratories using commercially available antisera. O:H serotyping showed that 35 strains (27 of O26, O111, and O128 serogroups) belonged to 11 serotypes considered to be classical EPEC O:H serotypes. The other 20 isolates were classified as 15 nonclassical EPEC O:H serotypes. All the potential EPEC virulence factors associated with bacterial adhesion (localized adherence, fluorescentactin staining test positivity, presence of the attaching and effacing [eaeA] gene), the production of verotoxin, and the positivity with the enterohemorrhagic E. coli probe were significantly more frequent among isolates belonging to classical than nonclassical serotypes. Strains displaying an aggregative adhesion and hybridizing with the enteroaggregative DNA probe were found in serogroups O86, O111, and O126. Verotoxin-producing isolates belonged to serogroups O26, O111, and O128. Only one of the isolates hybridized with the EPEC adherence factor (EAF) probe, but 33 strains gave positive results with the eae probe, confirming that the former is more suitable in epidemiological studies in European countries. These results indicate that up to 75% of strains identified as EPEC by commercial antisera may possess potential virulence properties and/or belong to classical EPEC O:H serotypes and suggest that O grouping is still a useful diagnostic tool for presumptive identification of diarrheagenic E. coli in clinical laboratories. PMID:8904439
Vranic, Sabina Mahmutovic; Uzunovic, Aida
Objectives: The aim of this study was to examine antimicrobial resistance of Escherichia coli strains isolated from urine in outpatient population. Material and methods: We performed a retrospective study for tree months period, between January 1st and March 31st, 2015, at the Department of Microbiology and Parasitology, Faculty of Medicine, University of Sarajevo. We determined the E. coli antimicrobial resistance in 556 first urine samples from outpatient population of Hrasno community in Sarajevo, Bosnia and Herzegovina. E. coli is the most frequent agent causing urinary tract infections in outpatients as well. The standard methods of descriptive statistics were performed for data analysis. Results: We observed the highest antimicrobial resistance of E. coli for ampicillin (82,79%), followed by trimethoprim-sulfamethoxazole (40,86%), nalidixic acid (19,35%), cephazolin (7,52%), nitrofurantoin (5,37%), gentamicin (2,15%) and ciprofloxacin (4,30%). Conclusions: The results of study showed that E. coli has the highest resistance to ampicillin and trimethoprim-sulfamethoxazole in outpatient population of Hrasno community. PMID:27147918
Delong, E F; Steinhauer, D; Israel, A; Nealson, K H
Genes necessary for luminescence (lux genes) in the marine bacterium Photobacterium leiognathi, strain PL721, were isolated and expressed in Escherichia coli. A 15-kb fragment obtained from a partial digestion of PL721 DNA with HindIII was cloned into the plasmid pACYC184, resulting in the hybrid plasmid pSD721. When pSD721 was transformed into E. coli ED8654, the resulting transformants were luminous with no additions to the cells, indicating that it contained the structural genes coding for the alpha and beta subunits of luciferase (luxA and luxB), and for components involved in aldehyde biosynthesis. Hybridization analysis with luxA and luxB 32P probes confirmed the location of these two genes on the 15-kb insert. When pSD721 was transformed into four different strains of E. coli, luminescence expression varied widely in amount and in pattern. In some strains, luminescence developed like an autoinducible system, and at maximum induction was very bright, even with no addition of aldehyde, while in others, luminescence was 100-fold less, and no induction was seen. In no case was luminescence affected by shifts in temperature, osmolarity, or iron concentration. These results indicate that, while the complete lux regulon is apparently contained on the 15-kb cloned fragment, the regulation of the lux regulon in pSD721 is subject to host controls by E. coli, controls which vary widely among different E. coli strains.
Randall, L P; Mueller-Doblies, D; Lemma, F L; Horton, R A; Teale, C J; Davies, R H
1. A field study was performed to investigate the presence and characteristics of ciprofloxacin-resistant, extended spectrum β-lactamase (ESBL) and AmpC Escherichia coli from turkeys in Great Britain. E. coli were isolated from ~9000 boot swab samples from 27 different farms owned by four different companies. Between 1 and 14 visits were made to each farm (mean 3) at between 0 and 15 m intervals (mean ~5 m). 2. CHROMagar ECC with and without ciprofloxacin or cephalosporin antibiotics was used as selective isolation media. Representative isolates with different phenotypes were tested for mutations in gyrA and for: qnrA, B, S, qepA and aac(6')-Ib genes, for ESBL phenotype, the presence of bla genes and plasmid type, and for ampC genes Representative ciprofloxacin-resistant and CTX-M isolates were further tested for serotype and PFGE type. On ciprofloxacin selective media 55% of samples yielded ciprofloxacin resistant E. coli and of those further analysed, most had ciprofloxacin MICs >4 mg/l and mutations in gyrA. 3. For the different companies, the mean number of samples per farm with cefoxitin- or cefotaxime-resistant isolates ranged from 1·0% to 61·9% and 4·7% to 31·7% respectively. Cefotaxime-resistance was most commonly associated with an ESBL phenotype, a CTX-M-1 or CTX-M-14 sequence type and an I1-γ or K plasmid inc type. The mechanism of cefoxitin resistance was not determined for most isolates, but where determined it was bla . 4. PFGE and serotyping showed clonally-related isolates persisting over multiple visits suggesting both more prudent use of antibiotics and improved farm hygiene are needed to address the issue of antimicrobial resistance in isolates from turkeys.
Suhaimi, Sheril Norliana; Phang, Lai-Yee; Maeda, Toshinari; Abd-Aziz, Suraini; Wakisaka, Minato; Shirai, Yoshihito; Hassan, Mohd Ali
Bioconverting glycerol into various valuable products is one of glycerol's promising applications due to its high availability at low cost and the existence of many glycerol-utilizing microorganisms. Bioethanol and biohydrogen, which are types of renewable fuels, are two examples of bioconverted products. The objectives of this study were to evaluate ethanol production from different media by local microorganism isolates and compare the ethanol fermentation profile of the selected strains to use of glucose or glycerol as sole carbon sources. The ethanol fermentations by six isolates were evaluated after a preliminary screening process. Strain named SS1 produced the highest ethanol yield of 1.0 mol: 1.0 mol glycerol and was identified as Escherichia coli SS1 Also, this isolated strain showed a higher affinity to glycerol than glucose for bioethanol production. PMID:24031858
Carlos, Camila; Alexandrino, Fabiana; Vieira, Monica A M; Stoppe, Nancy C; Sato, Maria Inês Z; Gomes, Tânia A T; Ottoboni, Laura M M
The aim of this work was to verify the presence of seven virulence factors (ST, LT, eae, stx(1), stx(2), INV and EAEC) among Escherichia coli strains isolated from healthy humans, bovines, chickens, sheep, pigs and goats, from two sewage treatment plants and from the Tietê River. We have found a high prevalence of eae, stx(1) and stx(2) in ruminants. The EAEC gene was only found in humans and sewage. No strains presented ST, LT or INV. BOX-PCR fingerprints revealed a high diversity among the strains analysed and a non-clonal origin of strains that presented the same virulence factors. Therefore, we concluded that ruminants may constitute an important reservoir of most diarrheagenic E. coli in Brazil, except for EAEC strains. These results emphasize the importance of the identification of the animal source of fecal contamination for the correct water risk assessment.
Ferdous, Mithila; Zhou, Kai; de Boer, Richard F.; Friedrich, Alexander W.; Kooistra-Smid, Anna M. D.; Rossen, John W. A.
In 2011, a Shiga toxin-producing Enteroaggregative Escherichia coli (EAEC Stx2a+) O104:H4 strain caused a serious outbreak of acute gastroenteritis and hemolytic-uremic syndrome (HUS) in Germany. In 2013, E. coli O104:H4 isolates were obtained from a patient with HUS and her friend showing only gastrointestinal complaints. The antimicrobial resistance and virulence profiles of these isolates together with three EAEC Stx2a+ O104:H4 isolates from 2011 were determined and compared. Whole-genome sequencing (WGS) was performed for detailed characterization and to determine genetic relationship of the isolates. Four additional genomes of EAEC Stx2a+ O104:H4 isolates of 2009 and 2011 available on NCBI were included in the virulence and phylogenetic analysis. All E. coli O104:H4 isolates tested were positive for stx2a, aatA, and terD but were negative for escV. All, except one 2011 isolate, were positive for aggR and were therefore considered EAEC. The EAEC Stx2a+ O104:H4 isolates of 2013 belonged to sequence type (ST) ST678 as the 2011 isolates and showed slightly different resistance and virulence patterns compared to the 2011 isolates. Core-genome phylogenetic analysis showed that the isolates of 2013 formed a separate cluster from the isolates of 2011 and 2009 by 27 and 20 different alleles, respectively. In addition, only a one-allele difference was found between the isolate of the HUS-patient and that of her friend. Our study shows that EAEC Stx2a+ O104:H4 strains highly similar to the 2011 outbreak clone in their core genome are still circulating necessitating proper surveillance to prevent further outbreaks with these potentially pathogenic strains. In addition, WGS not only provided a detailed characterization of the isolates but its high discriminatory power also enabled us to discriminate the 2013 isolates from the isolates of 2009 and 2011 expediting the use of WGS in public health services to rapidly apply proper infection control strategies. PMID:26696970
Ferdous, Mithila; Zhou, Kai; de Boer, Richard F; Friedrich, Alexander W; Kooistra-Smid, Anna M D; Rossen, John W A
In 2011, a Shiga toxin-producing Enteroaggregative Escherichia coli (EAEC Stx2a+) O104:H4 strain caused a serious outbreak of acute gastroenteritis and hemolytic-uremic syndrome (HUS) in Germany. In 2013, E. coli O104:H4 isolates were obtained from a patient with HUS and her friend showing only gastrointestinal complaints. The antimicrobial resistance and virulence profiles of these isolates together with three EAEC Stx2a+ O104:H4 isolates from 2011 were determined and compared. Whole-genome sequencing (WGS) was performed for detailed characterization and to determine genetic relationship of the isolates. Four additional genomes of EAEC Stx2a+ O104:H4 isolates of 2009 and 2011 available on NCBI were included in the virulence and phylogenetic analysis. All E. coli O104:H4 isolates tested were positive for stx2a, aatA, and terD but were negative for escV. All, except one 2011 isolate, were positive for aggR and were therefore considered EAEC. The EAEC Stx2a+ O104:H4 isolates of 2013 belonged to sequence type (ST) ST678 as the 2011 isolates and showed slightly different resistance and virulence patterns compared to the 2011 isolates. Core-genome phylogenetic analysis showed that the isolates of 2013 formed a separate cluster from the isolates of 2011 and 2009 by 27 and 20 different alleles, respectively. In addition, only a one-allele difference was found between the isolate of the HUS-patient and that of her friend. Our study shows that EAEC Stx2a+ O104:H4 strains highly similar to the 2011 outbreak clone in their core genome are still circulating necessitating proper surveillance to prevent further outbreaks with these potentially pathogenic strains. In addition, WGS not only provided a detailed characterization of the isolates but its high discriminatory power also enabled us to discriminate the 2013 isolates from the isolates of 2009 and 2011 expediting the use of WGS in public health services to rapidly apply proper infection control strategies.
Han, Jennifer H.; Nachamkin, Irving; Tolomeo, Pam; Mao, Xiangqun; Bilker, Warren B.; Lautenbach, Ebbing
The objective of this study was to characterize the temporal variability of fluoroquinolone resistance mechanisms among Escherichia coli colonizing the gastrointestinal tract of hospitalized patients. Patients with new fluoroquinolone-resistant E. coli (FQREC) colonization were followed with serial fecal sampling until discharge or death. Genetic mechanism(s) of resistance for all FQREC isolates were characterized, including mutations in gyrA and parC and efflux pump overexpression. Of 451 subjects, 73 (16.2%) became newly colonized with FQREC. There was significant variability in regard to temporal changes in resistance mechanisms and levofloxacin MICs among isolates from individual patients. Compared to patients with transient colonization, patients with persistent colonization were more likely to have a urinary catheter (P=0.04), diarrhea (P=0.04), and a longer duration of hospitalization (22 and 9.0 mean days, respectively; P=0.01) prior to sampling. Our data demonstrate the significant variability of resistance mechanisms in colonizing E. coli isolates among hospitalized patients. PMID:23719087
Stephan, R.; Untermann, F.
Fourteen verotoxin-producing Escherichia coli strains isolated from stool samples of 14 different asymptomatic human carriers were further characterized. A variety of serotypes was found, but none of the strains belonged to serogroup O157. Only one isolate carried most of the virulence genes that are associated with increased pathogenicity. PMID:10203524
Background Escherichia coli (E. coli) is commonly used as a surrogate for pathogens in research to identify sources of agricultural contamination and to characterize how pathogens persist on plant surfaces. However, E. coli strains are highly diverse, exhibiting differences in physical, chemical and...
Lalak, Anna; Wasyl, Dariusz; Zając, Magdalena; Skarżyńska, Magdalena; Hoszowski, Andrzej; Samcik, Ilona; Woźniakowski, Grzegorz; Szulowski, Krzysztof
Resistance to β-lactams is considered one of the major global problems and recently it became the most frequently studied topic in the area of antimicrobial resistance. The study was focused on phenotypic and genetic characterisation of commensal Escherichia coli (E. coli), including those producing cephalosporinases, isolated from gut flora of healthy slaughter animals. E. coli were cultured simultaneously on MacConkey agar (MCA) and cefotaxime supplemented MCA. The isolates were confirmed with ONPG and indol tube tests as well as PCR targeting uspA gene. Microbroth dilution method was applied for determination of Minimal Inhibitory Concentrations and interpreted according to EUCAST epidemiological cut-off values. Cephalosporin resistance phenotypes were defined by E-tests (BioMerieux) and relevant gene amplicons from selected strains were sequenced. A total of 298 E. coli isolates with cephalosporin resistance (ESC) found in 99 ones, were obtained from 318 cloacal or rectal swabs deriving from broilers, layers, turkeys, pigs and cattle. Both extended spectrum β-lactamase (ESBL) and ampC-cephalosporinase resistance phenotypes were noted in all tested animal species but cattle. At least one of the analysed genes was identified in 90 out of 99 cephalosporin-resistant isolates: blaTEM (n=44), blaCMY (n=38), blaCTX-M (n=33) and blaSHV (n=12). None of the phenotypes was identified in nine isolates. Sequencing of PCR products showed occurrence of ESBL-genes: blaCTX-M-1/-61, blaSHV-12, blaTEM-1,-52/-92,-135 and ampC-gene blaCMY-2. They were located on numerous and diverse plasmids and resistance transferability was proved by electroporation of blaSHV-12 and blaCTX-M-1/-61 located on X1 plasmids. Detection of cephalosporin resistant E. coli confirms the existence of resistance genes reservoir in farm animals and their possible spread (i.e. via IncX1 plasmids) to other bacteria including human and animal pathogens. The identified genetic background indicates on
Karczmarczyk, Maria; Martins, Marta; Quinn, Teresa; Leonard, Nola; Fanning, Séamus
Eleven multidrug-resistant Escherichia coli isolates (comprising 6 porcine and 5 bovine field isolates) displaying fluoroquinolone (FQ) resistance were selected from a collection obtained from the University Veterinary Hospital (Dublin, Ireland). MICs of nalidixic acid and ciprofloxacin were determined by Etest. All showed MICs of nalidixic acid of >256 μg/ml and MICs of ciprofloxacin ranging from 4 to >32 μg/ml. DNA sequencing was used to identify mutations within the quinolone resistance-determining regions of target genes, and quantitative real-time PCR (qRT-PCR) was used to evaluate the expression of the major porin, OmpF, and component genes of the AcrAB-TolC efflux pump and its associated regulatory loci. Decreased MIC values to nalidixic acid and/or ciprofloxacin were observed in the presence of the efflux pump inhibitor phenylalanine-arginine-β-naphthylamide (PAβN) in some but not all isolates. Several mutations were identified in genes coding for quinolone target enzymes (3 to 5 mutations per strain). All isolates harbored GyrA amino acid substitutions at positions 83 and 87. Novel GyrA (Asp87 → Ala), ParC (Ser80 → Trp), and ParE (Glu460 → Val) substitutions were observed. The efflux activity of these isolates was evaluated using a semiautomated ethidium bromide (EB) uptake assay. Compared to wild-type E. coli K-12 AG100, isolates accumulated less EB, and in the presence of PAβN the accumulation of EB increased. Upregulation of the acrB gene, encoding the pump component of the AcrAB-TolC efflux pump, was observed in 5 of 11 isolates, while 10 isolates showed decreased expression of OmpF. This study identified multiple mechanisms that likely contribute to resistance to quinolone-based drugs in the field isolates studied.
Nontongana, Nolonwabo; Sibanda, Timothy; Ngwenya, Elvis; Okoh, Anthony I
Escherichia coli is a widespread bacterium encompassing a variety of strains, ranging from highly pathogenic strains, causing worldwide outbreaks of severe diseases to avirulent, well characterized safe laboratory strains. This study evaluated the prevalence and antibiogram profiles of E. coli pathotypes isolated from the Kat River and Fort Beaufort abstraction water. A total of 171 out of 278 confirmed E. coli isolates were positive for at least one pathogenic determinant and these included enteropathogenic E. coli (6%), enterotoxigenic E. coli (47%), uropathogenic E. coli (2%), neonatal meningitis E. coli (5%), diffusely adherent E. coli (1%) and enterohaemorrhagic E. coli (1%). Interestingly, enteroinvasive and enteroaggregative E. coli were not detected. The phenotypic antibiogram profiles of the isolates revealed that all were resistant to penicillin G, while 98% and 38% of the pathotypes were resistant to ampicillin and trimethoprim-sulphamethoxazole, respectively. About 8% of the isolates were resistant to streptomycin. More than half of the isolates exhibited multiple antibiotic resistance with 44% being resistant to three antibiotics and 8% resistant to four antibiotics. We conclude that the Kat River is a reservoir of potentially virulent antibiotic resistant E. coli strains that can cause serious health risks to humans who drink raw water from this river, or in the case that consumption of treated drinking water coincides with failed drinking water processes.
Nontongana, Nolonwabo; Sibanda, Timothy; Ngwenya, Elvis; Okoh, Anthony I.
Escherichia coli is a widespread bacterium encompassing a variety of strains, ranging from highly pathogenic strains, causing worldwide outbreaks of severe diseases to avirulent, well characterized safe laboratory strains. This study evaluated the prevalence and antibiogram profiles of E. coli pathotypes isolated from the Kat River and Fort Beaufort abstraction water. A total of 171 out of 278 confirmed E. coli isolates were positive for at least one pathogenic determinant and these included enteropathogenic E. coli (6%), enterotoxigenic E. coli (47%), uropathogenic E. coli (2%), neonatal meningitis E. coli (5%), diffusely adherent E. coli (1%) and enterohaemorrhagic E. coli (1%). Interestingly, enteroinvasive and enteroaggregative E. coli were not detected. The phenotypic antibiogram profiles of the isolates revealed that all were resistant to penicillin G, while 98% and 38% of the pathotypes were resistant to ampicillin and trimethoprim-sulphamethoxazole, respectively. About 8% of the isolates were resistant to streptomycin. More than half of the isolates exhibited multiple antibiotic resistance with 44% being resistant to three antibiotics and 8% resistant to four antibiotics. We conclude that the Kat River is a reservoir of potentially virulent antibiotic resistant E. coli strains that can cause serious health risks to humans who drink raw water from this river, or in the case that consumption of treated drinking water coincides with failed drinking water processes. PMID:25119699
Zanotto, Carlo; Bissa, Massimiliano; Illiano, Elena; Mezzanotte, Valeria; Marazzi, Francesca; Turolla, Andrea; Antonelli, Manuela; De Giuli Morghen, Carlo; Radaelli, Antonia
The emergence and diffusion of antibiotic-resistant bacteria has been a major public health problem for many years now. In this study, antibiotic-resistance of coliforms and Escherichia coli were investigated after their isolation from samples collected in a municipal wastewater treatment plant in the Milan area (Italy) along different points of the treatment sequence: inflow to biological treatment; outflow from biological treatment following rapid sand filtration; and outflow from peracetic acid disinfection. The presence of E. coli that showed resistance to ampicillin (AMP) and chloramphenicol (CAF), used as representative antibiotics for the efficacy against Gram-positive and Gram-negative bacteria, was evaluated. After determining E. coli survival using increasing AMP and CAF concentrations, specific single-resistant (AMP(R) or CAF(R)) and double-resistant (AMP(R)/CAF(R)) strains were identified among E. coli colonies, through amplification of the β-lactamase Tem-1 (bla) and acetyl-transferase catA1 (cat) gene sequences. While a limited number of CAF(R) bacteria was observed, most AMP(R) colonies showed the specific resistance genes to both antibiotics, which was mainly due to the presence of the bla gene sequence. The peracetic acid, used as disinfection agent, showed to be very effective in reducing bacteria at the negligible levels of less than 10 CFU/100 mL, compatible with those admitted for the irrigation use of treated waters.
Shamsi, Hamid; Mardani, Karim; Ownagh, Abdolghaffar
Escherichia coli isolates from chickens with colibacillosis were assigned to phylogenetic groups based on multiplex polymerase chain reaction (PCR) and antibacterial resistance of E. coli belonging to these groups was examined. Furthermore, the gyrA gene of isolates was sequenced and a phylogenetic tree was generated. A total of 84 E. coli isolates were grouped using multiplex PCR of TSPE4.C2, chuA, yjaA, and gadA molecular markers. Four phylogenetic groups were identified with strains divided as follows: 16 in group A (19.05%), 17 in group B1 (20.24%), 23 in group B2 (27.38%), and 28 in group D (33.33%). Escherichia coli isolates belonging to phylogenetic groups B2 and D were resistant to Soltrim and Flumequine unlike the majority of E. coli isolates that belonged to groups A and B1, and which were susceptible to these antibiotics. The phylogenetic results based on gyrA gene sequences from multiplex PCR revealed that E. coli phylogenetic grouping was in accordance with the clusters obtained in the phylogenetic tree. In conclusion, the comparative sequence analysis of gyrA sequences provides a firm framework for an accurate classification of E. coli and related taxa and may constitute a pertinent phylogenetic marker for E. coli.
Rodríguez-Pérez, Celia; Quirantes-Piné, Rosa; Uberos, José; Jiménez-Sánchez, Cecilia; Peña, Alejandro; Segura-Carretero, Antonio
Phenolic compounds from a cranberry extract were isolated in order to assess their contribution to the antibacterial activity against uropathogenic strains of Escherichia coli (UPEC). With this purpose, a total of 25 fractions from a cranberry extract were isolated using semipreparative high performance liquid chromatography (HPLC) and characterized based on the results obtained by reversed-phase HPLC coupled to mass spectrometry detection. Then, the effects on UPEC surface hydrophobicity and biofilm formation of the cranberry extract as well as the purest fractions (a total of 13) were tested. As expected, the whole extract presented a powerful antibacterial activity against UPEC while the selected fractions presented a different behavior. Myricetin and quercitrin significantly decreased (p < 0.05) E. coli biofilm formation compared with the control, while dihydroferulic acid glucuronide, procyanidin A dimer, quercetin glucoside, myricetin and prodelphinidin B led to a significant decrease of the surface hydrophobicity compared with the control. The results suggest that apart from proanthocyanidins, other compounds, mainly flavonoids, can act against E. coli biofilm formation and also modify UPEC surface hydrophobicity in vitro, one of the first steps of adhesion.
Jones, Suzanna E; Burgos, Jonathan M; Lutnesky, Marvin M F; Sena, Johnny A; Kumar, Sanath; Jones, Lindsay M; Varela, Manuel F
Antimicrobial agent usage is common in animal agriculture for therapeutic and prophylactic purposes. Selective pressure exerted by these antimicrobials on soil bacteria could result in the selection of strains that are resistant due to chromosomal- or plasmid-derived genetic components. Multiple antimicrobial resistances in Escherichia coli and the direct relationship between antimicrobial agent use over time has been extensively studied, yet the relationship between the age of an animal agriculture environment such as a dairy farm and antibiotic resistance remains unclear. Therefore, we tested the hypothesis that antimicrobial-resistance profiles of E. coli isolated from dairy farm topsoil correlate with dairy farm age. E. coli isolated from eleven dairy farms of varying ages within Roosevelt County, NM were used for MIC determinations to chloramphenicol, nalidixic acid, penicillin, tetracycline, ampicillin, amoxicillin/clavulanic acid, gentamicin, trimethoprim/sulfamethoxazole, cefotaxime, and ciprofloxacin. The minimum inhibitory concentration values of four antibiotics ranged 0.75 to >256 μg/ml, 1 to >256 μg/ml, 12 to >256 μg/ml, and 0.75 to >256 μg/ml for chloramphenicol, nalidixic acid, penicillin, and tetracycline, respectively. The study did not show a direct relationship between antibiotic resistance and the age of dairy farms.
Sáez-López, Emma; Cossa, Anélsio; Benmessaoud, Rachid; Madrid, Lola; Moraleda, Cinta; Villanueva, Sonia; Tligui, Houssain; Moiane, Benilde; Alami, Hassan; Massora, Sérgio; Bezad, Rachid; Mandomando, Inacio; Bosch, Jordi; Vila, Jordi; Bassat, Quique; Soto, Sara M.
The relevance of vaginal colonization of pregnant women by Escherichia coli is poorly understood, despite these strains sharing a similar virulence profile with other extraintestinal pathogenic E. coli producing severe obstetric and neonatal infections. We characterized the epidemiology, antimicrobial susceptibility and virulence profiles of 84 vaginal E. coli isolates from pregnant women from Rabat (Morocco) and Manhiça (Mozambique), two very distinct epidemiological settings. Low levels of antimicrobial resistance were observed to all drugs tested, except for trimethoprim-sulfamethoxazole in Manhiça, where this drug is extensively used as prophylaxis for opportunistic HIV infections. The most prevalent virulence factors were related to iron acquisition systems. Phylogroup A was the most common in Rabat, while phylogroups E and non-typeable were the most frequent in Manhiça. Regardless of the apparently “low virulence” of these isolates, the frequency of infections is higher and the outcomes more devastating in constrained-resources conditions, especially among pregnant women and newborns. PMID:27387665
Teshager, T; Herrero, I A; Porrero, M C; Garde, J; Moreno, M A; Domínguez, L
Antimicrobial resistance can make the efficient treatment of bacterial infections in humans and animals more difficult. Antimicrobial use in food animals may be one of the factors contributing to resistance. The Spanish surveillance network VAV has established a baseline of antimicrobial resistance in Escherichia coli strains from healthy pigs. Minimum inhibitory concentration and patterns of resistance to antimicrobials used in animals and humans were determined for 205 faecal strains isolated in a sampling frame of four slaughterhouses in Spain from 220 pigs in 1998. Higher levels of resistance were seen against antimicrobial agents authorised for use in food animals especially tetracycline, sulphonamides, trimethoprim and amoxycillin. All isolates were susceptible to antimicrobials employed mainly in humans such as ceftazidime, cefotaxime, imipenem, aztreonam and amikacin.
Jardine, Claire M; Janecko, Nicol; Allan, Mike; Boerlin, Patrick; Chalmers, Gabhan; Kozak, Gosia; McEwen, Scott A; Reid-Smith, Richard J
We conducted a cross-sectional study to determine the prevalence of antimicrobial resistance (AMR) in fecal Escherichia coli isolates from raccoons (Procyon lotor) living in Ontario, Canada. From June to October 2007, we trapped raccoons in three areas: one primarily urban site around Niagara, one primarily rural site north of Guelph, and one at the Toronto Zoo. In addition, we conducted a longitudinal study at the Toronto Zoo site to investigate the temporal dynamics of fecal E. coli and AMR in raccoons. Reduced susceptibility to ≥1 antimicrobial agent was detected in E. coli isolates from 19% of 16 raccoons at the urban site, 17% of 29 raccoons from the rural site, and 42% of 130 samples collected from 59 raccoons at the zoo site. Raccoons from the zoo site were significantly more likely to shed E. coli with reduced susceptibility to ≥1 antimicrobial agent than animals from the rural site (odds ratio [OR], 3.41; 95% confidence interval [CI], 1.17 to 12.09; P = 0.02). Resistance to expanded-spectrum cephalosporins (and the associated bla(CMY-2) gene) was detected in two animals from the zoo site and one animal from the rural site. Serotyping and pulsed-field gel electrophoresis analysis show that raccoons on the zoo grounds harbor a diverse assemblage of E. coli, with rapid bacterial turnover within individuals over time. Our study indicates that raccoons may shed resistant bacteria of public health significance and that raccoons have the potential to disseminate these bacteria throughout their environment.
Guerra, Julio A; Romero-Herazo, Yesenia C; Arzuza, Octavio; Gómez-Duarte, Oscar G
Enterotoxigenic Escherichia coli (ETEC) are major causes of childhood diarrhea in low and middle income countries including Colombia, South America. To understand the diversity of ETEC strains in the region, clinical isolates obtained from northern Colombia children were evaluated for multiple locus sequencing typing, serotyping, classical and nonclassical virulence genes, and antibiotic susceptibility. Among 40 ETEC clinical isolates evaluated, 21 (52.5%) were positive for LT gene, 13 (32.5%) for ST gene, and 6 (15%) for both ST and LT. The most prevalent colonization surface antigens (CS) were CS21 and CFA/I identified in 21 (50%) and 13 (32.5%) isolates, respectively. The eatA, irp2, and fyuA were the most common nonclassical virulence genes present in more than 60% of the isolates. Ampicillin resistance (80% of the strains) was the most frequent phenotype among ETEC strains followed by trimethoprim-sulfamethoxazole resistance (52.5%). Based on multiple locus sequencing typing (MLST), we recognize that 6 clonal groups of ETEC clinical isolates circulate in Colombia. ETEC clinical isolates from children in northern Colombia are highly diverse, yet some isolates circulating in the community belong to well-defined clonal groups that share a unique set of virulence factors, serotypes, and MLST sequence types.
Guerra, Julio A.; Romero-Herazo, Yesenia C.; Arzuza, Octavio; Gómez-Duarte, Oscar G.
Enterotoxigenic Escherichia coli (ETEC) are major causes of childhood diarrhea in low and middle income countries including Colombia, South America. To understand the diversity of ETEC strains in the region, clinical isolates obtained from northern Colombia children were evaluated for multiple locus sequencing typing, serotyping, classical and nonclassical virulence genes, and antibiotic susceptibility. Among 40 ETEC clinical isolates evaluated, 21 (52.5%) were positive for LT gene, 13 (32.5%) for ST gene, and 6 (15%) for both ST and LT. The most prevalent colonization surface antigens (CS) were CS21 and CFA/I identified in 21 (50%) and 13 (32.5%) isolates, respectively. The eatA, irp2, and fyuA were the most common nonclassical virulence genes present in more than 60% of the isolates. Ampicillin resistance (80% of the strains) was the most frequent phenotype among ETEC strains followed by trimethoprim-sulfamethoxazole resistance (52.5%). Based on multiple locus sequencing typing (MLST), we recognize that 6 clonal groups of ETEC clinical isolates circulate in Colombia. ETEC clinical isolates from children in northern Colombia are highly diverse, yet some isolates circulating in the community belong to well-defined clonal groups that share a unique set of virulence factors, serotypes, and MLST sequence types. PMID:24877071
Budič, Maruška; Rijavec, Matija; Petkovšek, Ziva; Zgur-Bertok, Darja
Bacteriocins are antimicrobial peptides generally active against bacteria closely related to the producer. Escherichia coli produces two types of bacteriocins, colicins and microcins. The in vitro efficacy of isolated colicins E1, E6, E7, K and M, was assessed against Escherichia coli strains from patients with bacteraemia of urinary tract origin. Colicin E7 was most effective, as only 13% of the tested strains were resistant. On the other hand, 32%, 33%, 43% and 53% of the tested strains exhibited resistance to colicins E6, K, M and E1. Moreover, the inhibitory activity of individual colicins E1, E6, E7, K and M and combinations of colicins K, M, E7 and E1, E6, E7, K, M were followed in liquid broth for 24 hours. Resistance against individual colicins developed after 9 hours of treatment. On the contrary, resistance development against the combined action of 5 colicins was not observed. One hundred and five E. coli strains from patients with bacteraemia were screened by PCR for the presence of 5 colicins and 7 microcins. Sixty-six percent of the strains encoded at least one bacteriocin, 43% one or more colicins, and 54% one or more microcins. Microcins were found to co-occur with toxins, siderophores, adhesins and with the Toll/Interleukin-1 receptor domain-containing protein involved in suppression of innate immunity, and were significantly more prevalent among strains from non-immunocompromised patients. In addition, microcins were highly prevalent among non-multidrug-resistant strains compared to multidrug-resistant strains. Our results indicate that microcins contribute to virulence of E. coli instigating bacteraemia of urinary tract origin.
Balière, C.; Rincé, A.; Delannoy, S.; Fach, P.
ABSTRACT Shiga toxin-producing Escherichia coli (STEC) and enteropathogenic E. coli (EPEC) strains may be responsible for food-borne infections in humans. Twenty-eight STEC and 75 EPEC strains previously isolated from French shellfish-harvesting areas and their watersheds and belonging to 68 distinguishable serotypes were characterized in this study. High-throughput real-time PCR was used to search for the presence of 75 E. coli virulence-associated gene targets, and genes encoding Shiga toxin (stx) and intimin (eae) were subtyped using PCR tests and DNA sequencing, respectively. The results showed a high level of diversity between strains, with 17 unique virulence gene profiles for STEC and 56 for EPEC. Seven STEC and 15 EPEC strains were found to display a large number or a particular combination of genetic markers of virulence and the presence of stx and/or eae variants, suggesting their potential pathogenicity for humans. Among these, an O26:H11 stx1a eae-β1 strain was associated with a large number of virulence-associated genes (n = 47), including genes carried on the locus of enterocyte effacement (LEE) or other pathogenicity islands, such as OI-122, OI-71, OI-43/48, OI-50, OI-57, and the high-pathogenicity island (HPI). One O91:H21 STEC strain containing 4 stx variants (stx1a, stx2a, stx2c, and stx2d) was found to possess genes associated with pathogenicity islands OI-122, OI-43/48, and OI-15. Among EPEC strains harboring a large number of virulence genes (n, 34 to 50), eight belonged to serotype O26:H11, O103:H2, O103:H25, O145:H28, O157:H7, or O153:H2. IMPORTANCE The species E. coli includes a wide variety of strains, some of which may be responsible for severe infections. This study, a molecular risk assessment study of E. coli strains isolated from the coastal environment, was conducted to evaluate the potential risk for shellfish consumers. This report describes the characterization of virulence gene profiles and stx/eae polymorphisms of E. coli
Nawaz, Mohamed; Sung, Kidon; Kweon, Ohgew; Khan, Saeed; Nawaz, Samia; Steele, Roger
Fifty-five nalidixic acid-resistant Escherichia coli strains were isolated from imported shrimp. Purified PCR amplicons of gyrA, gyrB, parC and parE from the template DNA of all isolates were sequenced and analysed for point mutations that confer resistance to nalidixic acid and ciprofloxacin. Point mutations in the quinolone resistance-determining regions (QRDRs) of GyrA at positions 68, 83 and 87 and in ParC at positions 80 and 84 as well as in the non-QRDR of GyrA at positions 112, 127, 128 and 154 along with point mutations in parE at position 476 conferred resistance to these antibiotics. Computational modelling and analysis of the different point mutations and their role in the enhanced resistance to these antibiotics indicated that only mutation at codons 83 (Ser→Ile) and 87 (Asp→Asn) played a vital role in increasing the minimum inhibitory concentration (MIC) to these drugs compared with other mutations. Ethidium bromide experiments indicated higher efflux pump activities in quinolone-resistant E. coli strains compared with their quinolone-sensitive counterparts. Class 1 integrons measuring 0.7-2.3kb were amplified and sequenced from the template DNA of the isolates. Sequence analysis of the 2.0kb and 1.7kb integrons indicated the presence of resistance determinants for trimethoprim (dfrA12 and dfrA17) and aminoglycosides (aadA2 and aadA5). These results indicate that use of nalidixic acid, ciprofloxacin and other antibiotics in shrimp aquaculture ponds may select E. coli resistant to these antibiotics and that imported shrimp is a reservoir of multiple antibiotic-resistant E. coli.
Chang, Jiyoung; Yu, Jinkyung; Lee, Hyeyoung; Ryu, Hyejin; Park, Kanggyun; Park, Yeon-Joon
Recently, serotype O75 was found to be prominent among the non-ST131 ciprofloxacin-resistant Escherichia coli, and they were all lactose non-fermenters. In this study, we investigated the prevalence and characteristics of lactose non-fermenters in urinary isolates of E. coli. A total of 167 E. coli isolates was collected. Antimicrobial susceptibility tests were determined by VITEK 2 (bioMerieux, France). The lactose non-fermenters underwent PCR-based O typing, multilocus sequence typing (MLST) analysis, phylogenetic grouping. For ciprofloxacin-resistant isolates, the resistance mechanisms were investigated. Thirty-three (19.7%) isolates were lactose non-fermenters and the ciprofloxacin resistance rate was significantly higher than in lactose fermenters (66.7% vs. 31.6%, P = 0.0002). According to the serotype, O75 was the most common (42.4%, 14/33) and was followed by O16 (5/33), O2 (4/33), O25b (3/33), O15 (1/33), O6 (1/33), O1 (1/33). All the O75 isolates were ciprofloxacin-resistant and belonged to ST1193. By MLST, they were resolved into 11 STs. ST1193 was the most common (14/33) and was followed by ST131 (8/33). Interestingly, 8 isolates of ST131 were divided into three O types [O16 (4 isolates), O25b (3), and non-typeable (1)]. The ciprofloxacin resistance rate was high in isolates of O75-ST1193 and O25b-ST131 but low in O16-ST131 and O2-ST95. All the ciprofloxacin-resistant isolates showed identical triple mutations in gyrA and parC but the serotype O25b isolates had an additional mutation in parC (E84V). Only one isolate harbored aac(6')-Ib-cr variant and no qnr gene was detected. Continuous monitoring of the prevalence and clonal composition of the lactose non-fermenters is needed.
Vieira, Regine H S Dos F; Carvalho, Edirsana M R; Carvalho, Fatima C T; Silva, Camila M; Sousa, Oscarina V; Rodrigues, Dalia P
This study aimed to test the susceptibility of Escherichia coli strains isolated from the water, bottom sediments and individuals cultivated in shrimp farm ponds, to antibiotics belonging to different families, namely B-Lactams: Imipenem (IPM; 10 micro g), Ampicillin (AMP; 10 micro g), Cephalothin (CEP; 30 micro g), Cefoxitin (FOX; 30 micro g), Ceftriaxone (CRO; 30 micro g); Tetracycline: Tetracycline (TCY; 30 micro g); Aminoglycosides: Gentamicin (GEN; 10 micro g), Amikacin (AMK; 30 micro g); Chloramphenicol: Chloramphenicol (CHO; 30 micro g); Fluoroquinolones: Ciprofloxacin (CIP; 5 micro g); Nitrofurans: Nitrofurantoin (NIT; 300 micro g); Sulfonamides: Trimethoprim-Sulfamethoxazole (SXT; 30 micro g); Quilononas: Nalidixic Acid (NAL; 30 micro g). In the laboratory, the method of dissemination (Test Kirby-Bauer) was performed in order to fulfill the antibiogram tests. The results showed high indices of resistance to Imipenem, Cephalothin and Ampicillin. Chloramphenicol, Nitrofurantoin, Cefoxitin, Ceftiaxone and Ciprofloxacin have displayed the highest index of sensitive strains. The antibiotic resistance index (ARI) and the multiple resistance index (MAR) varied within the ranges of 0.068-0.077 and 0.15-0.39, respectively. More than 90.5% of strains of Escherichia coli showed a variety of resistance profiles to the tested antibiotics. The high indices of resistance may be a consequence of indiscriminate use of antibiotics, but also the transfer of resistance through mobile genetic elements found in shrimp farms.
Maluta, Renato Pariz; Stella, Ariel Eurides; Riccardi, Kátia; Rigobelo, Everlon Cid; Marin, José Moacir; Carvalho, Marileda Bonafim; de Ávila, Fernando Antonio
Pathogenic strains of Escherichia coli are the most common bacteria associated with urinary tract infections in both humans and companion animals. Standard biochemical tests may be useful in demonstrating detailed phenotypical characteristics of these strains. Thirteen strains of E. coli isolated from dogs with UTIs were submitted to biochemical tests, serotyping for O and H antigens and antimicrobial resistance testing. Furthermore, the presence of papC, sfa, and afa genes was evaluated by PCR, and genetic relationships were established using enterobacterial repetitive intergenic consensus PCR (ERIC-PCR). The antimicrobial that showed the highest resistance rate among the isolates was nalidixic acid (76.9%), followed by cephalotin (69.2%), sulfamethoxazole + trimethoprim (61.5%), tetracycline (61.5%), streptomycin (53.8%), ciprofloxacin (53.8%), ampicillin (46.2%), gentamicin (30.8%) and chloramphenicol (23.1%). No isolate was resistant either to meropenem or nitrofurantoin. Among the five clusters that were identified using ERIC-PCR, one cluster (A) had only one strain, which belonged to a serotype with zoonotic potential (O6:H31) and showed the genes papC+, sfa+, afa-. Strains with the genes papC-, sfa+, afa- were found in two other clusters (C and D), whereas all strains in clusters B and E possessed papC-, sfa-, afa- genes. Sucrose and raffinose phenotypic tests showed some ability in discriminating clusters A, B and C from clusters D and E. PMID:24031842
Ohkoshi, Yasuo; Sato, Toyotaka; Suzuki, Yuuki; Yamamoto, Soh; Shiraishi, Tsukasa; Ogasawara, Noriko; Yokota, Shin-Ichi
In recent years, multidrug resistance of Escherichia coli has become a serious problem. However, resistance to fosfomycin (FOM) has been low. We screened E. coli clinical isolates with reduced susceptibility to FOM and characterized molecular mechanisms of resistance and reduced susceptibility of these strains. Ten strains showing reduced FOM susceptibility (MIC ≥ 8 μg/mL) in 211 clinical isolates were found and examined. Acquisition of genes encoding FOM-modifying enzyme genes (fos genes) and mutations in murA that underlie high resistance to FOM were not observed. We examined ability of FOM incorporation via glucose-6-phosphate (G6P) transporter and sn-glycerol-3-phosphate transporter. In ten strains, nine showed lack of growth on M9 minimum salt agar supplemented with G6P. Eight of the ten strains showed fluctuated induction by G6P of uhpT that encodes G6P transporter expression. Nucleotide sequences of the uhpT, uhpA, glpT, ptsI, and cyaA shared several deletions and amino acid mutations in the nine strains with lack of growth on G6P-supplemented M9 agar. In conclusion, reduction of uhpT function is largely responsible for the reduced sensitivity to FOM in clinical isolates that have not acquired FOM-modifying genes or mutations in murA. However, there are a few strains whose mechanisms of reduced susceptibility to FOM are still unclear.
Ohkoshi, Yasuo; Sato, Toyotaka; Suzuki, Yuuki; Yamamoto, Soh; Shiraishi, Tsukasa; Ogasawara, Noriko
In recent years, multidrug resistance of Escherichia coli has become a serious problem. However, resistance to fosfomycin (FOM) has been low. We screened E. coli clinical isolates with reduced susceptibility to FOM and characterized molecular mechanisms of resistance and reduced susceptibility of these strains. Ten strains showing reduced FOM susceptibility (MIC ≥ 8 μg/mL) in 211 clinical isolates were found and examined. Acquisition of genes encoding FOM-modifying enzyme genes (fos genes) and mutations in murA that underlie high resistance to FOM were not observed. We examined ability of FOM incorporation via glucose-6-phosphate (G6P) transporter and sn-glycerol-3-phosphate transporter. In ten strains, nine showed lack of growth on M9 minimum salt agar supplemented with G6P. Eight of the ten strains showed fluctuated induction by G6P of uhpT that encodes G6P transporter expression. Nucleotide sequences of the uhpT, uhpA, glpT, ptsI, and cyaA shared several deletions and amino acid mutations in the nine strains with lack of growth on G6P-supplemented M9 agar. In conclusion, reduction of uhpT function is largely responsible for the reduced sensitivity to FOM in clinical isolates that have not acquired FOM-modifying genes or mutations in murA. However, there are a few strains whose mechanisms of reduced susceptibility to FOM are still unclear. PMID:28197413
Day, Michael W; Jackson, Lydgia A; Akins, Darrin R; Dyer, David W; Chavez-Bueno, Susana
Neonatal bacteremia Escherichia coli strains commonly belong to the K1 capsular type. Their ability to cause invasive neonatal disease appears to be determined by other virulence factors that have yet to be identified. We report here the genome sequences of four E. coli neonatal bacteremia isolates, including that of the archetypal strain RS218.
Berman, Hillary; Barberino, Maria Goreth; Moreira, Edson Duarte; Riley, Lee; Reis, Joice N
The clinical management of meningitis caused by Escherichia coli is greatly complicated when the organism becomes resistant to broad-spectrum antibiotics. We sought to characterize the antimicrobial susceptibilities, sequence types (ST), and presence of known drug resistance genes of E. coli isolates that caused meningitis between 1996 and 2011 in Salvador, Brazil. We then compared these findings to those for E. coli isolates from community-acquired urinary tract infections (UTI) that occurred during the same time period and in the same city. We found that 19% of E. coli isolates from cases of meningitis and less than 1% of isolates from UTI were resistant to third-generation cephalosporins. The sequence types of E. coli isolates from cases of meningitis included ST131, ST69, ST405, and ST62, which were also found among isolates from UTI. Additionally, among the E. coli isolates that were resistant to third-generation cephalosporins, we found genes that encode the extended-spectrum beta-lactamases CTX-M-2, CTX-M-14, and CTX-M-15. These observations demonstrate that compared to E. coli strains isolated from cases of community-acquired UTI, those isolated from cases of meningitis are more resistant to third-generation cephalosporins, even though the same sequence types are shared between the two forms of extraintestinal infections.
Maciel, Jonas Fernandes; Matter, Letícia Beatriz; Trindade, Michele Martins; Camillo, Giovana; Lovato, Maristela; de Ávila Botton, Sônia; Castagna de Vargas, Agueda
In this study an avian colisepticemia outbreak was investigated. Two isolates from a chicken with colisepticemia were characterized for antimicrobial susceptibility and virulence factors profile. For this purpose 7 antimicrobial and 29 genes (fimH, hrlA/hek, iha, papC, sfa/focCD, tsh, mat, tia, gimB, ibeA, chuA, fyuA, ireA, iroN, irp2, iucD, sitD. chr., sitD. ep., iss, neuC, ompA, traT, astA, hlyA, sat, vat, pic, malX, cvi/cva) were tested. The outbreak happened in a hick chicken breeding located in the northwestern region of Rio Grande do Sul state in South of Brazil and caused 28.3% (102 deads of a total of 360 chickens) of mortality rate. Escherichia coli isolates obtained from the avian spleen and liver belong to the same phylogenetic group A and present resistance to all antimicrobials tested (ampicillin, tetracycline, gentamicin, neomycin, sulfa + trimethoprim, enrofloxacin, and norfloxacin). Both isolates harbor virulence factors related to adhesion (fimH, papC, mat), invasion (tia), iron acquisition system (iroN) and serum resistance (iss, ompA, traT), showing that these groups are important for Avian Pathogenic E. coli (APEC). However, they present different virulence profiles for some genes, whereas liver-isolate carries more hrlA/hek (adhesin), gimB (invasin), sitD ep. (iron acquisition system), sat (toxin) and hylA (toxin) genes, the spleen-isolate harbors fyuA (iron acquisition system) gene. Here, we highlight a coinfection by different strains of APEC in the same animal with colisepticemia, the great antimicrobial resistance of these bacterial isolates and the genetic traits that modulate the virulence for high mortality rate of chickens for human consumption.
Palmer, Christine E; Bratcher, Christy L; Singh, Manpreet; Wang, Luxin
In addition to Escherichia coli O157:H7, shiga toxin-producing E. coli (STEC) O26 was added to the zero-tolerance adulterant list together with other 5 non-O157 STEC serogroups in 2012. Four farm O26 isolates were used in this study; they were obtained from a on-farm survey study conducted in Alabama. The presence of 3 major pathogenic genes (stx1, stx2, and eaeA) was determined through multiplex polymerase chain reaction (PCR). Two major pathogenic gene profiles were observed: 3 of the farm isolates contain only the eaeA gene whereas 1 farm isolate has both the eaeA and the stx1 genes. No significant difference was seen among the 4 farm isolates in the antibiotic resistance tests. To test their survival in ground beef and environmental samples, 2 inoculums were prepared and inoculated at various concentrations into samples of ground beef, bovine feces, bedding materials, and trough water. One inoculum was made of 3 farm isolates containing only the eaeA gene and another inoculum contained the isolate with both the eaeA and stx1 genes. Inoculated beef samples were stored at 4 °C for 10 d and the inoculated environmental samples were stored at ambient temperature for 30 d. Results showed that virulence gene profiles do not have an impact on O26's ability to survive in ground beef and in environment (P > 0.05). The inoculation levels, sample types as well as the storage times are the major factors that impact O26 survival (P < 0.05).
Karczmarczyk, Maria; Walsh, Ciara; Slowey, Rosemarie; Leonard, Nola; Fanning, Séamus
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%), blaTEM (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, blaOXA-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 IV536 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. PMID:21856840
Hamamoto, Kouta; Ueda, Shuhei; Yamamoto, Yoshimasa; Hirai, Itaru
Genotyping and characterization of bacterial isolates are essential steps in the identification and control of antibiotic-resistant bacterial infections. Recently, one novel genotyping method using three genomic guided Escherichia coli markers (GIG-EM), dinG, tonB, and dipeptide permease (DPP), was reported. Because GIG-EM has not been fully evaluated using clinical isolates, we assessed this typing method with 72 E. coli collection of reference (ECOR) environmental E. coli reference strains and 63 E. coli isolates of various genetic backgrounds. In this study, we designated 768 bp of dinG, 745 bp of tonB, and 655 bp of DPP target sequences for use in the typing method. Concatenations of the processed marker sequences were used to draw GIG-EM phylogenetic trees. E. coli isolates with identical sequence types as identified by the conventional multilocus sequence typing (MLST) method were localized to the same branch of the GIG-EM phylogenetic tree. Sixteen clinical E. coli isolates were utilized as test isolates without prior characterization by conventional MLST and phylogenetic grouping before GIG-EM typing. Of these, 14 clinical isolates were assigned to a branch including only isolates of a pandemic clone, E. coli B2-ST131-O25b, and these results were confirmed by conventional typing methods. Our results suggested that the GIG-EM typing method and its application to phylogenetic trees might be useful tools for the molecular characterization and determination of the genetic relationships among E. coli isolates.
Andrade, Vanessa da Costa; Zampieri, Bruna Del Busso; Ballesteros, Eliete Rodrigues; Pinto, Aline Bartelochi; de Oliveira, Ana Julia Fernandes Cardoso
Bacterial resistance is a rising problem all over the world. Many studies have showed that beach sands can contain higher concentration of microorganisms and represent a risk to public health. This paper aims to evaluate the densities and resistance to antimicrobials of Escherichia coli strains, isolated from seawater and samples. The hypothesis is that microorganisms show higher densities in contaminated beach sands and more antimicrobial resistance than the water column. Density, distribution, and antimicrobial resistance of bacteria E. coli were evaluate in seawater and sands from two recreational beaches with different levels of pollution. At the beach with higher degree of pollution (Gonzaguinha), water samples presented the highest densities of E. coli; however, higher frequency of resistant strains was observe in wet sand (71.9 %). Resistance to a larger number of antimicrobial groups was observe in water (betalactamics, aminoglycosides, macrolides, rifampicins, and tetracyclines) and sand (betagalactamics and aminoglycosids). In water samples, highest frequencies of resistance were obtain against ampicilin (22.5 %), streptomycin (15.0 %), and rifampicin (15.0 %), while in sand, the highest frequencies were observe in relation to ampicilin (36.25 %) and streptomycin (23.52 %). At the less polluted beach, Ilha Porchat, highest densities of E. coli and higher frequency of resistance were obtain in wet and dry sand (53.7 and 53.8 %, respectively) compared to water (50 %). Antimicrobial resistance in strains isolated from water and sand only occurred against betalactamics (ampicilin and amoxicilin plus clavulanic acid). The frequency and variability of bacterial resistance to antimicrobials in marine recreational waters and sands were related to the degree of fecal contamination in this environment. These results show that water and sands from beaches with a high index of fecal contamination of human origin may be potential sources of contamination by pathogens
Sargeant, J M; Shi, X; Sanderson, M W; Renter, D G; Nagaraja, T G
This study investigated the prevalence and distribution of Escherichia coli O157 genetic types within and among feedlots using pulsed-field gel electrophoresis to separate XbaI-digested DNA. The study population consisted of 300 pens of cattle in 30 feedlots in Kansas that were sampled (feces, water, and water sediment) within a month of being shipped for slaughter. The prevalence of E. coli O157 was 8.5% in feces, 3.1% in water, and 4.5% in water sediment samples. A total of 424 E. coli O157 isolates were characterized by pulsed-field gel electrophoresis, and 139 subtypes (100% Dice similarity with no band differences) were identified. The majority of subtypes (70/139) was identified only once, but nine were identified 10 or more times. Identical subtypes were recovered from both feces and water tanks in 10 feedlots. The majority of subtypes were identified in only one feedlot, and the number of subtypes ranged from one to 23 within a feedlot and from one to seven within a pen. There were 10 feedlots with at least 15 positive samples. In these 10 feedlots, the most common subtype accounted for 16.9-78.6% of the isolates. Common subtypes differed among feedlots. In eight of the 10 feedlots, the most common subtype was identified in multiple pens. The results support a complex ecology for E. coli O157 in feedlot operations, with factors associated with exposure and transmission likely acting at a common level for multiple feedlots, within feedlots, and within pens of cattle.
Nicoletti, M; Superti, F; Conti, C; Calconi, A; Zagaglia, C
Lactose-negative Escherichia coli strains were isolated at high frequency from children with diarrhea in Somalia during a 2-year study on diarrheal diseases. Sixty-four of these strains, considered to be a representative sample, were characterized for virulence factors, plasmid profiles, and antibiotic resistance. Of these strains, 5 were recognized as enteroinvasive E. coli (they were serotyped as O135:K-:H-), 6 belonged to classical enteropathogenic E. coli serotypes, 9 were able to adhere to tissue culture cells (of these, 4 showed a pattern of localized adherence and 1 was an enteropathogenic strain), 18 were both adherent and hemolytic, and 8 were simply hemolytic. None hybridized with 32P-labeled heat-labile or heat-stable (a and b) enterotoxin gene probes or produced moderate or high-level cytotoxic effects on HeLa cells. Of the 64 strains examined, 24 produced mannose-resistant hemagglutination with human, chicken, and monkey erythrocytes. One of these was serotyped as O4:K-:H8, and a rabbit O antiserum raised against this strain allowed us to establish that 23 strains had the same O antigen. The 23 O4 strains were hemolytic and were not enterotoxic for rabbit ileal loops, and intact bacteria were able to destroy tissue culture cell monolayers very rapidly. The uniformity of the antibiotic resistance pattern and of the plasmid DNA content, together with the fact that they were isolated in different years and in different children, suggests that the O4 strains must be epidemiologically relevant in Somalia. A possible diarrheagenic role for the adherent-hemolytic E. coli strains is also discussed. Images PMID:3281977
Paixão, A C; Ferreira, A C; Fontes, M; Themudo, P; Albuquerque, T; Soares, M C; Fevereiro, M; Martins, L; Corrêa de Sá, M I
Poultry colibacillosis due to Avian Pathogenic Escherichia coli (APEC) is responsible for several extra-intestinal pathological conditions, leading to serious economic damage in poultry production. The most commonly associated pathologies are airsacculitis, colisepticemia, and cellulitis in broiler chickens, and salpingitis and peritonitis in broiler breeders. In this work a total of 66 strains isolated from dead broiler breeders affected with colibacillosis and 61 strains from healthy broilers were studied. Strains from broiler breeders were typified with serogroups O2, O18, and O78, which are mainly associated with disease. The serogroup O78 was the most prevalent (58%). All the strains were checked for the presence of 11 virulence genes: 1) arginine succinyltransferase A (astA); ii) E.coli hemeutilization protein A (chuA); iii) colicin V A/B (cvaA/B); iv) fimbriae mannose-binding type 1 (fimC); v) ferric yersiniabactin uptake A (fyuA); vi) iron-repressible high-molecular-weight proteins 2 (irp2); vii) increased serum survival (iss); viii) iron-uptake systems of E.coli D (iucD); ix) pielonefritis associated to pili C (papC); x) temperature sensitive haemaglutinin (tsh), and xi) vacuolating autotransporter toxin (vat), by Multiplex-PCR. The results showed that all genes are present in both commensal and pathogenic E. coli strains. The iron uptake-related genes and the serum survival gene were more prevalent among APEC. The adhesin genes, except tsh, and the toxin genes, except astA, were also more prevalent among APEC isolates. Except for astA and tsh, APEC strains harbored the majority of the virulence-associated genes studied and fimC was the most prevalent gene, detected in 96.97 and 88.52% of APEC and AFEC strains, respectively. Possession of more than one iron transport system seems to play an important role on APEC survival.
Shridhar, Pragathi B; Noll, Lance W; Shi, Xiaorong; Cernicchiaro, Natalia; Renter, David G; Bai, J; Nagaraja, T G
Escherichia coli O104:H4, an hybrid pathotype of Shiga toxigenic and enteroaggregative E. coli, involved in a major foodborne outbreak in Germany in 2011, has not been detected in cattle feces. Serogroup O104 with H type other than H4 has been reported to cause human illnesses, but their prevalence and characteristics in cattle have not been reported. Our objectives were to determine the prevalence of E. coli O104 in feces of feedlot cattle, by culture and PCR detection methods, and characterize the isolated strains. Rectal fecal samples from a total of 757 cattle originating from 29 feedlots were collected at a Midwest commercial slaughter plant. Fecal samples, enriched in E. coli broth, were subjected to culture and PCR methods of detection. The culture method involved immunomagnetic separation with O104-specific beads and plating on a selective chromogenic medium, followed by serogroup confirmation of pooled colonies by PCR. If pooled colonies were positive for the wzxO104 gene, then colonies were tested individually to identify wzxO104-positive serogroup and associated genes of the hybrid strains. Extracted DNA from feces were also tested by a multiplex PCR to detect wzxO104-positive serogroup and associated major genes of the O104 hybrid pathotype. Because wzxO104 has been shown to be present in E. coli O8/O9/O9a, wzxO104-positive isolates and extracted DNA from fecal samples were also tested by a PCR targeting wbdDO8/O9/O9a, a gene specific for E. coli O8/O9/O9a serogroups. Model-adjusted prevalence estimates of E. coli O104 (positive for wzxO104 and negative for wbdDO8/O9/O9a) at the feedlot level were 5.7% and 21.2%, and at the sample level were 0.5% and 25.9% by culture and PCR, respectively. The McNemar's test indicated that there was a significant difference (P < 0.01) between the proportions of samples that tested positive for wzxO104 and samples that were positive for wzxO104, but negative for wbdDO8/O9/O9a by PCR and culture methods. A total of 143
Shridhar, Pragathi B.; Noll, Lance W.; Shi, Xiaorong; Cernicchiaro, Natalia; Renter, David G.; Bai, J.; Nagaraja, T. G.
Escherichia coli O104:H4, an hybrid pathotype of Shiga toxigenic and enteroaggregative E. coli, involved in a major foodborne outbreak in Germany in 2011, has not been detected in cattle feces. Serogroup O104 with H type other than H4 has been reported to cause human illnesses, but their prevalence and characteristics in cattle have not been reported. Our objectives were to determine the prevalence of E. coli O104 in feces of feedlot cattle, by culture and PCR detection methods, and characterize the isolated strains. Rectal fecal samples from a total of 757 cattle originating from 29 feedlots were collected at a Midwest commercial slaughter plant. Fecal samples, enriched in E. coli broth, were subjected to culture and PCR methods of detection. The culture method involved immunomagnetic separation with O104-specific beads and plating on a selective chromogenic medium, followed by serogroup confirmation of pooled colonies by PCR. If pooled colonies were positive for the wzxO104 gene, then colonies were tested individually to identify wzxO104-positive serogroup and associated genes of the hybrid strains. Extracted DNA from feces were also tested by a multiplex PCR to detect wzxO104-positive serogroup and associated major genes of the O104 hybrid pathotype. Because wzxO104 has been shown to be present in E. coli O8/O9/O9a, wzxO104-positive isolates and extracted DNA from fecal samples were also tested by a PCR targeting wbdDO8/O9/O9a, a gene specific for E. coli O8/O9/O9a serogroups. Model-adjusted prevalence estimates of E. coli O104 (positive for wzxO104 and negative for wbdDO8/O9/O9a) at the feedlot level were 5.7% and 21.2%, and at the sample level were 0.5% and 25.9% by culture and PCR, respectively. The McNemar’s test indicated that there was a significant difference (P < 0.01) between the proportions of samples that tested positive for wzxO104 and samples that were positive for wzxO104, but negative for wbdDO8/O9/O9a by PCR and culture methods. A total of 143
Carbonari, Claudia C; Deza, Natalia; Flores, Mario; Gasparini, Alejandra; Manfredi, Eduardo; Rivas, Marta
We describe the first isolation of an enteroaggregative Escherichia coli (EAEC) O104:H4 strain associated with an acute diarrhea case in Argentina. Two multiplex PCRs (mPCR) were performed as screening of genes mPCR1 (eae, lt, and st) and mPCR2 (IpaH, aggR, stx1 and stx2). A mPCR to detect the rfbO104, fliCH4 and terD genes, and PCR assays for the detection of pCVD432 plasmid, aaiC and lpfO113 genes were included. Biochemical and antimicrobial susceptibility assays as well as serotyping were performed. The identified E. coli strain was susceptible to all antimicrobials tested and harbored the aggR, aaiC, pCVD432 plasmid, lpfO113, rfbO104, fliCH4 and terD genes. Although serotype EAEC O104:H4 rarely spreads and sporadic cases have been reported, global concern increased after the large-scale outbreak in Europe in 2011. The finding of EAEC O104:H4 reinforces the need for improved methodologies for the detection of all E. coli pathotypes.
Martins, R; Queiroz, J A; Sousa, F
6S RNA is an abundant non-coding RNA in Escherichia coli (E. coli), but its function has not been discovered until recently. The first advance on 6S RNA function was the demonstration of its ability to bind the σ(70)-holoenzyme form of RNA polymerase, inhibiting its activity and consequently the transcription process. The growing interest in the investigation of non-coding small RNAs (sRNA) calls for the development of new methods for isolation and purification of RNA. This work presents an optimized RNA extraction procedure and describes a new affinity chromatography method using a histidine support to specifically purify 6S RNA from other E. coli sRNA species. The RNA extraction procedure was optimized, and a high yield was obtained in the separation of sRNA and ribosomal RNA (rRNA) from total RNA (RNAt). This improved method takes advantage of its simplicity and significant cost reduction, since some complex operations have been eliminated. A purification strategy was also developed to separate 6S RNA from an sRNA mixture. Pure RNA can be advantageously obtained using the histidine-affinity chromatography method, aiming at its application to structural or functional studies.
Dar, S. H.; Qureshi, S.; Palanivelu, M.; Muthu, S.; Mehrotra, S.; Jan, M. H.; Chaudhary, G. R.; Kumar, H.; Saravanan, R.; Narayanan, K.
Ascending infection of the uterus with Gram-negative bacteria is responsible for postpartum endometritis in cattle and buffalo and can adversely affect fertility. Development of a laboratory animal model for bovine endometritis would facilitate the understanding of the pathogenesis as it is difficult to conduct controlled experimentation in the native host. In the present study, 30 virgin Swiss Albino mice (5-8 weeks old) were used to evaluate the pathogenic potential of Escherichia coli, isolated from the normally calved postpartum buffalo to induce endometritis. Mice in the diestrus phase of the estrous cycle were randomly allotted to one of the following four intravaginal inoculation (100 μL) treatments: EG (experimental group)-1: sterile normal saline; EG-2, -3 and -4: E. coli@ 1.5 × 104, 105 and 106 CFU/ml, respectively. The animals were then scarified 36 h post-inoculation to study gross and microscopical lesions. Gross changes were confined to EG-4. Acute endometritis was recorded in 50% of the EG-3 and 66.7% of the EG-4. The rate of acute endometritis development was significantly higher in EG-4 (P<0.05) as compared to the other groups. The present study demonstrated that the animal model for bubaline endometritis can be developed in mice by intravaginal inoculation of E. coli@ 1.5 × 106 CFU/ml at diestrus. Ease of intravaginal inoculation, apparent absence of systemic involvement and high infective rate are the advantages of the model over other studies. PMID:27822246
Touchon, Marie; Charpentier, Sophie; Pognard, Dominique; Picard, Bertrand; Arlet, Guillaume; Rocha, Eduardo P C; Denamur, Erick; Branger, Catherine
Clustered, regularly interspaced, short palindromic repeats (CRISPRs) are implicated in defence against foreign DNA in various archaeal and bacterial species. They have also been associated with a slower spread of antibiotic resistance. However, experimental and evolutionary studies raise doubts about the role of CRISPRs as a sort of immune system in Escherichia coli. We studied a collection of 263 natural E. coli isolates from human and animal hosts, representative of the phylogenetic and lifestyle diversity of the species and exhibiting various levels of plasmid-encoded antibiotic resistance. We characterized the strains in terms of CRISPRs, performed replicon typing of the plasmids and tested for class 1 integrons to explore the possible association between CRISPRs and the absence of plasmids and mobile antibiotic resistance determinants. We found no meaningful association between the presence/absence of the cas genes, reflecting the activity of the CRISPRs, and the presence of plasmids, integrons or antibiotic resistance. No CRISPR in the collection contained a spacer that matched an antibiotic resistance gene or element involved in antibiotic resistance gene mobilization, and 79.8 % (210/263) of the strains lacked spacers matching sequences in the 2282 plasmid genomes available. Hence, E. coli CRISPRs do not seem to be efficient barriers to the spread of plasmids and antibiotic resistance, consistent with what has been reported for phages, and contrary to reports concerning other species.
Touchon, Marie; Charpentier, Sophie; Pognard, Dominique; Picard, Bertrand; Arlet, Guillaume; Rocha, Eduardo P C; Denamur, Erick; Branger, Catherine
Clustered, regularly interspaced, short palindromic repeats (CRISPRs) are implicated in defence against foreign DNA in various archaeal and bacterial species. They have also been associated with a slower spread of antibiotic resistance. However, experimental and evolutionary studies raise doubts about the role of CRISPRs as a sort of immune system in Escherichia coli. We studied a collection of 263 natural E. coli isolates from human and animal hosts, representative of the phylogenetic and lifestyle diversity of the species and exhibiting various levels of plasmid-encoded antibiotic resistance. We characterized the strains in terms of CRISPRs, performed replicon typing of the plasmids and tested for class 1 integrons to explore the possible association between CRISPRs and the absence of plasmids and mobile antibiotic resistance determinants. We found no meaningful association between the presence/absence of the cas genes, reflecting the activity of the CRISPRs, and the presence of plasmids, integrons or antibiotic resistance. No CRISPR in the collection contained a spacer that matched an antibiotic resistance gene or element involved in antibiotic resistance gene mobilization, and 79.8% (210/263) of the strains lacked spacers matching sequences in the 2282 plasmid genomes available. Hence, E. coli CRISPRs do not seem to be efficient barriers to the spread of plasmids and antibiotic resistance, consistent with what has been reported for phages, and contrary to reports concerning other species.
Wagner, Bruce A; Straw, Barbara E; Fedorka-Cray, Paula J; Dargatz, David A
A body of evidence exists that suggests that antimicrobial use in food animals leads to resistance in both pathogenic and commensal bacteria. This study focused on the impact of three different antimicrobial regimes (low-level continuous, pulse, and no antimicrobial) for two antimicrobials (chlortetracycline and tylosin) on the presence of Salmonella spp. and on the prevalence of antimicrobial resistance of both Salmonella spp. and nonspecific Escherichia coli in fecal samples from feeder swine. The prevalence of fecal samples positive for Salmonella spp. significantly decreased between the samples taken at feeder placement compared to samples taken when the animals were close to market weight. Differences in resistance of Salmonella spp. did not appear to be influenced by dosing treatment including the control. Analysis of antimicrobial resistance examining both susceptibility and resistance, as well as MIC outcomes, demonstrated that only resistance to cephalothin increased in E. coli under the pulse chlortetracycline treatment. These results suggest that the dosing regimes examined in this study did not lead to an increase in either the prevalence of Salmonella spp. or the prevalence of antimicrobial resistance in isolates of Salmonella spp. or E. coli.
Prim, Núria; Rivera, Alba; Rodríguez-Navarro, Judith; Español, Montserrat; Turbau, Miquel; Coll, Pere; Mirelis, Beatriz
Colistin resistance was detected in 53 of 10,011 Escherichia coli (0.5%) by prospective phenotypic testing of consecutive clinical isolates in a single hospital in Barcelona, Spain (2012-15). The mcr-1 gene was retrospectively identified by PCR and sequencing in 15 of 50 available isolates. Each isolate had a unique PFGE pattern except for two. This clonal diversity supports the hypothesis of horizontal dissemination of the mcr-1 gene in the local study population.
Background The aim of our study was to investigate the possible etiology of avian colibacillosis by examining Escherichia coli isolates from fecal samples of healthy broilers. Findings Seventy-eight E. coli isolates from fecal samples of healthy broilers in Japan were subjected to analysis of phylogenetic background, virulence-associated gene profiling, multi-locus sequence typing (MLST), and antimicrobial resistance profiling. Phylogenetic analysis demonstrated that 35 of the 78 isolates belonged to group A, 28 to group B1, one to group B2, and 14 to group D. Virulence-associated genes iutA, iss, cvaC, tsh, iroN, ompT, and hlyF were found in 23 isolates (29.5%), 16 isolates (20.5%), nine isolates (11.5%), five isolates (6.4%), 19 isolates (24.4%), 23 isolates (29.5%), and 22 isolates (28.2%) respectively. Although the genetic diversity of group D isolates was revealed by MLST, the group D isolates harbored iutA (10 isolates, 71.4%), iss (6 isolates, 42.9%), cvaC (5 isolates, 35.7%), tsh (3 isolates, 21.4%), hlyF (9 isolates, 64.3%), iroN (7 isolates, 50.0%), and ompT (9 isolates, 64.3%). Conclusions Our results indicated that E. coli isolates inhabiting the intestines of healthy broilers pose a potential risk of causing avian colibacillosis. PMID:25061511
Ladely, Scott R.; Bono, James L.; Plumblee, Jodie R.; Hall, M. Carolina; Genzlinger, Linda L.; Cook, Kimberly L.
Transmissible colistin resistance conferred by the mcr-1 gene-bearing IncI2 plasmid has been recently reported in Escherichia coli in the United States. We report here the completed genome sequence of a second E. coli strain isolated from swine in the United States that carried the mcr-1 gene on an IncI2-type plasmid. PMID:27834721
Davoodabadi, Abolfazl; Soltan Dallal, Mohammad Mehdi; Lashani, Elahe; Tajabadi Ebrahimi, Maryam
Background: Among the enteric pathogens, diarrheagenic Escherichia coli are important causes of diarrhea in children in both developing and industrialized countries. Some Lactobacillus species are commonly used as probiotics, with effects especially against acute diarrhea in childhood. Objectives: The aim of this study was to explore antimicrobial activity of Lactobacillus strains isolated from fecal flora of healthy breast-fed infants against five diarrheagenic E. coli pathotypes such as enteroaggregative E. coli (EAEC), enterohaemorrhagic E. coli (EHEC) enteroinvasive E. coli (EIEC), enteropathogenic E. coli (EPEC) and enterotoxigenic E. coli (ETEC). Materials and Methods: Fecal samples were collected from seven healthy breast-fed infants between 1 to 18 months of age in Tehran city, Iran. Identification of Lactobacillus isolates was performed by biochemical and 16S rRNA gene sequencing methods. An agar well diffusion assay was used for detection of antimicrobial activity of Lactobacillus isolates against five diarrheagenic E. coli pathotypes. Results: A total of 20 Lactobacillus isolates were identified from stool samples. Lactobacillus fermentum was the most frequently isolated strain, followed by L. plantarum and L. rhamnosus. Seven Lactobacillus strains including L. fermentum (four isolates), L. paracasei (one isolate), L. plantarum (one isolate) and L. rhamnosus (one isolate) had a mild inhibitory activity against diarrheagenic E. coli. The mechanism of inhibitory activity of Lactobacillus strains appeared to be due to the production of organic acids or hydrogen peroxide. Conclusions: Our findings show that Lactobacillus strains with human origin had a mild inhibitory activity against the diarrheagenic E. coli, and these strains may be useful as probiotic candidates in prevention of intestinal infections caused by diarrheagenic E. coli. PMID:26865944
Baranzoni, Gian Marco; Fratamico, Pina M; Rubio, Fernando; Glaze, Thomas; Bagi, Lori K; Albonetti, Sabrina
Shiga toxin-producing Escherichia coli (STEC) strains belonging to serogroup O104 have been associated with sporadic cases of illness and have caused outbreaks associated with milk and sprouts. An outbreak that occurred in Europe in 2011 linked to fenugreek sprouts was caused by E. coli O104:H4 that had characteristics of an enteroaggregative E. coli (EAEC) but carried the gene that encoded for Shiga toxin 2. In this study, methods were developed for detection of this enteroaggregative STEC O104, as well as STEC O104 in sprouts. Multiplex PCR assays for enteroaggregative STEC O104:H4 targeted the stx2, aggR, and wzx104 genes, and for STEC O104 targeted the stx1-2, ehxA, and wzx104 genes. After incubating artificially contaminated sprouts at 4 °C for 48 h and overnight enrichment in modified buffered peptone water with pyruvate supplemented with three antibiotics (mBPWp), the pathogens were detected in all samples inoculated at a level of ca. 100CFU/25 g. Several samples inoculated at lower concentrations of ca. 10CFU/25 g were negative by the PCR assays, and this could have been due to cells not surviving or not being able to recover after the stress treatment at 4 °C for 48 h. For isolation of the pathogens, immunomagnetic separation (IMS) using magnetic beads coated with antibodies against O104 were employed, and this was followed by plating the beads onto mRBA and CHROMagar STEC O104 for isolation of E. coli O104:H4 and mRBA and CHROMagar STEC for isolation of E. coli O104:H7. Presumptive colonies were confirmed by agglutination using latex particles attached to antibodies against serogroup O104 and by the multiplex PCR assays. The methodologies described in this study for detection of enteroaggregative STEC O104:H4 and STEC O104 include the use of IMS and latex reagents for serogroup O104, and they enhance the ability to detect and isolate these pathogens from sprouts and potentially other foods, as well.
Gupta, Neerja; Ali, Arif
Ten Escherichia coli strains isolated from five different aquatic environments representing three distinct geographical regions of India showed significantly high levels of tolerance to the inorganic form of mercury, i.e., mercuric chloride (HgCl(2)). MRD14 isolated from the Dal Lake (Kashmir) could tolerate the highest concentration of HgCl(2), i.e., 55 microg/mL, and MRF1 from the flood water of the Yamuna River (Delhi) tolerated the lowest concentration, i.e., 25 microg/mL. All ten strains revealed the presence of a plasmid of approximately 24 kb, and transformation of the isolated plasmids into the mercury-sensitive competent cells of E. coli DH5alpha rendered the transformants resistant to the same concentration of mercury as the wild-type strains. Mating experiments were performed to assess the self-transmissible nature of these promiscuous plasmids. The transfer of mercury resistance from these wild-type strains to the mercury-sensitive, naladixic acid-resistant E. coli K12 (F(-) lac(+)) strain used as a recipient was observed in six of the nine strains tested. Transconjugants revealed the presence of a plasmid of approximately 24 kb. An evaluation of the mechanism of mercury resistance in the three most efficient strains (MRG12, MRD11, and MRD14) encountered in our study was determined by cold vapor atomic absorption spectroscopy (CV-AAS), and it was noted that resistance to HgCl(2) was conferred by conversion of the toxic ionic form of mercury (Hg(++)) to the nontoxic elemental form (Hg(0)) in all three strains. MRD14 volatilized mercury most efficiently.
Aslam, Mueen; Toufeer, Mehdi; Narvaez Bravo, Claudia; Lai, Vita; Rempel, Heidi; Manges, Amee; Diarra, Moussa Sory
Extraintestinal Pathogenic Escherichia coli (ExPEC) have the potential to spread through fecal waste resulting in the contamination of both farm workers and retail poultry meat in the processing plants or environment. The objective of this study was to characterize ExPEC from retail poultry meats purchased from Alberta, Canada and to compare them with 12 human ExPEC representatives from major ExPEC lineages. Fifty-four virulence genes were screened by a set of multiplex PCRs in 700 E. coli from retail poultry meat samples. ExPEC was defined as the detection of at least two of the following virulence genes: papA/papC, sfa, kpsMT II and iutA. Genetic relationships between isolates were determined using pulsed field gel electrophoresis (PFGE). Fifty-nine (8.4%) of the 700 poultry meat isolates were identified as ExPEC and were equally distributed among the phylogenetic groups A, B1, B2 and D. Isolates of phylogenetic group A possessed up to 12 virulence genes compared to 24 and 18 genes in phylogenetic groups B2 and D, respectively. E. coli identified as ExPEC and recovered from poultry harbored as many virulence genes as those of human isolates. In addition to the iutA gene, siderophore-related iroN and fyuA were detected in combination with other virulence genes including those genes encoding for adhesion, protectin and toxin while the fimH, ompT, traT, uidA and vat were commonly detected in poultry ExPEC. The hemF, iss and cvaC genes were found in 40% of poultry ExPEC. All human ExPEC isolates harbored concnf (cytotoxic necrotizing factor 1 altering cytoskeleton and causing necrosis) and hlyD (hemolysin transport) genes which were not found in poultry ExPEC. PFGE analysis showed that a few poultry ExPEC isolates clustered with human ExPEC isolates at 55-70% similarity level. Comparing ExPEC isolated from retail poultry meats provides insight into their virulence potential and suggests that poultry associated ExPEC may be important for retail meat safety
Escherichia coli is a commonly used indicator organism for detecting the presence of fecal-borne pathogenic microorganisms in water supplies. The importance of E. coli as an indicator organism has led to numerous studies looking at surface characteristics and transport behavior of this important mi...
Previous reports have indicated that a small proportion of cattle shedding high levels of Escherichia coli O157:H7 is the main source for transmission of this organism between animals. Cattle achieving a fecal shedding status of 10**4 CFU of E. coli O157: H7/gram or greater are now referred to as su...
Shiga toxin-producing Escherichia coli (STEC) O104 strains have been associated with sporadic cases of illness and have caused outbreaks associated with milk and sprouts. E. coli O104:H21 caused an outbreak associated with milk in the U.S. in 1994. In this study, next generation sequencing techno...
Kogoma, T; Farr, S B; Joyce, K M; Natvig, D O
Mu dX phage was used to isolate three gene fusions to the lacZ gene (soi::lacZ; soi for superoxide radical inducible) that were induced by treatment with superoxide radical anion generators such as paraquat and plumbagin. The induction of beta-galactosidase in these fusion strains with the superoxide radical generating agents required aerobic metabolism. Hyperoxygenation (i.e., bubbling of cultures with oxygen gas) also induced the fusions. On the other hand, hydrogen peroxide did not induce the fusions at concentrations that are known to invoke an adaptive response. Introduction of oxyR, htpR, or recA mutations did not affect the induction. Two of the fusion strains exhibited increased sensitivity to paraquat but not to hydrogen peroxide. The third fusion strain showed no increased sensitivity to either agent. All three fusions were located in the 45- to 61-min region of the Escherichia coli chromosome. PMID:2838846
A novel phage, (phi)241, specific for Escherichia coli O157:H7 was isolated from an industrial cucumber fermentation where both acidity (pH less than or equal to 3.7) and salinity (greater than or equal to 5% NaCl) were high. The phage belongs to the Myoviridae family. Its latent period was 15 min a...
Amarillas, Luis; Chaidez, Cristobal; Lugo, Yadira
The bacteriophage phiJLA23 was isolated from an animal feces sample and lytic activity was demonstrated against the Escherichia coli O157:H7 strain. We report the complete nucleotide sequence of bacteriophage phiJLA23, information which may be useful for determining whether this phage is a candidate for biocontrol or another biotechnological application. PMID:23469347
Ikeda, Tetsuya; Yamamoto, Shiori; Kabeya, Hidenori; Sugiyama, Hiromu; Takai, Shinji
ABSTRACT Shiga toxin-producing Escherichia coli (STEC) is one of the major foodborne pathogens. Having observed the wide distribution of this pathogen in wild deer, we report here the draft genome sequence of five STEC strains isolated from wild deer (Cervus nippon yesoensis) in Hokkaido, Japan. PMID:28254967
Guiral, Elisabet; Sáez-López, Emma; Bosch, Jordi; Goncé, Anna; López, Marta; Sanz, Sergi; Vila, Jordi
The virulence markers and the antimicrobial resistance profiles of 78 Escherichia coli isolates causing obstetric infections accompanied by sepsis or not were studied. Adhesion-related virulence factors were the most prevalent markers. Low rates of resistance to the antimicrobial agents used as first-line therapy suggest their correct implementation in stewardship guidelines. PMID:25740771
Specific Airs 12 METHODS AND MATERIALS 14 Bacterial Strains 14 Culture Media and Diluents 15 Ultraviolet Light Bacteriophage Induction 15 Plaque...coli 026 strain H-19 were carried on a bacteriophage . Escherichia coli 026 strain H-19 does not produce heat-labile or heat-stable enterotoxin and is...strain that had become toxinogenic after coculti- vation with Escherichia coli 026 strain H-19. Attempts to isolate the bacteriophage from Escherichia
Cuba, Gabriel Trova; Pignatari, Antonio Carlos Campos; Patekoski, Katya Silva; Luchesi, Lucimila Jorge; Kiffer, Carlos Roberto Veiga
Since antimicrobial resistance among uropathogens against current first line agents has affected the management of severe urinary tract infection, we determined the likelihood that antibiotic regimens achieve bactericidal pharmacodynamic exposures using Monte Carlo simulation for five antimicrobials (ciprofloxacin, ceftriaxone, piperacillin/tazobactam, ertapenem, and meropenem) commonly prescribed as initial empirical treatment of inpatients with severe community acquired urinary tract infections. Minimum inhibitory concentration determination by Etest was performed for 205 Brazilian community urinary tract infection Escherichia coli strains from 2008 to 2012 and 74 E. coli bloodstream strains recovered from a surveillance study. Pharmacodynamic exposure was modeled via a 5000 subject Monte Carlo simulation. All isolates were susceptible to ertapenem and meropenem. Piperacillin/tazobactam, ceftriaxone and ciprofloxacin showed 100%, 97.5% and 83.3% susceptibility among outpatient isolates and 98.6%, 75.7% and 64.3% among inpatient isolates, respectively. Against outpatient isolates, all drugs except ciprofloxacin (82.7% in aggressive and 77.6% in conservative scenarios) achieved high cumulative fraction of response: carbapenems and piperacillin/tazobactam cumulative fraction of responses were close to 100%, and ceftriaxone cumulative fraction of response was 97.5%. Similar results were observed against inpatients isolates for carbapenems (100%) and piperacillin/tazobactam (98.4%), whereas ceftriaxone achieved only 76.9% bactericidal cumulative fraction of response and ciprofloxacin 61.9% (aggressive scenario) and 56.7% (conservative scenario) respectively. Based on this model, standard doses of beta-lactams were predicted to deliver sufficient pharmacodynamic exposure for outpatients. However, ceftriaxone should be avoided for inpatients and ciprofloxacin empirical prescription should be avoided in both inpatients and outpatients with complicated urinary tract
Ahmed, Ashraf M; Shimamoto, Toshi; Shimamoto, Tadashi
Avian pathogenic Escherichia coli (APEC) causes extensive mortality in poultry flocks, leading to extensive economic losses. To date, little information is available on the molecular basis of antimicrobial resistance in APEC in Africa. Therefore, the objective of this study was to characterize the virulence and antimicrobial resistance of multidrug-resistant APEC isolated from septicemic broilers in Egypt at the molecular level. Among 91 non-repetitive E. coli isolates, 73 (80.2%) carried three or more of the APEC virulence genes iroN, ompT, iss, iutA, and hlyF. All 73 APEC isolates showed multidrug resistance phenotypes, particularly against ampicillin, tetracycline, spectinomycin, streptomycin, kanamycin, and trimethoprim/sulfamethoxazole. PCR and DNA sequencing identified class 1 and class 2 integrons in 34 (46.6%) and seven (9.6%) isolates, respectively. The β-lactamase-encoding genes, bla(TEM-1), bla(TEM-104), bla(CMY-2), bla(OXA-30), bla(CTX-M-15), and bla(SHV-2); tetracycline resistance genes, tet(A), tet(B), tet(C), tet(D), and tet(E); the plasmid-mediated quinolone resistance genes, qnrA1, qnrB2, qnrS1, and aac(6')-Ib-cr, and florfenicol resistance gene, floR, were also identified in 69 (94.5%), 67 (91.8%), 47 (64.4%), and 13 (17.8%) isolates, respectively. To the best of our knowledge, this is the first report of molecular characterization of antimicrobial resistance in APEC strains from Africa.
Xu, Guofeng; Jiang, Yue; An, Wei; Wang, Hongdong; Zhang, Xiuying
Three KPC-2-producing Escherichia coli (E1, E2, and E3) were recovered from water samples of an urban river in the city of Harbin, China. Antimicrobial susceptibility was determined by broth microdilution. Molecular characterization and genetic relatedness of the isolates were determined by polymerase chain reaction (PCR), pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST) and PCR-directed phylotyping. Plasmids were analyzed by conjugation, S1-PFGE, Southern blotting and PCR-based replicon typing (PBRT). The genetic environment of the bla KPC-2 gene was determined using PCR and sequencing. PCR analyses revealed that the E1 isolate carried the bla KPC-2, bla CMY-2, bla TEM-1, bla CTX-M-14, and qnrB2 genes and belonged to sequence type ST410, phylogenetic type A; the E2 isolate was assigned to ST131-B2 and carried the bla KPC-2, bla TEM-1, bla CTX-M-3, bla DHA-1, aac(6')-Ib-cr, and qnrS1 genes; while the E3 isolate was of ST648-D and possessed bla KPC-2, bla TEM-1, bla OXA-1, bla CTX-M-15, armA, and aac(6')-Ib-cr genes. PFGE demonstrated that each of the three KPC-2-producing E. coli isolates exhibited an individual XbaI patterns. The bla KPC-2 gene was located on plasmids of 60-140 kb with IncA/C, IncN, or non-typeable replicon types. The genetic environment of bla KPC-2 of the three strains was consistent with the genetic structure of bla KPC-2 on the plasmid pKP048.
Patterson, Edward I; Swarbrick, Crystall M D; Roman, Noelia; Forwood, Jade K; Raidal, Shane R
Expression of recombinant beak and feather disease virus (BFDV) capsid-associated protein (Cap) has relied on inefficient techniques that typically produce low yields or use specialized expression systems, which greatly increase the cost and expertise required for mass production. An Escherichia coli system was used to express recombinant BFDV Cap derived from two isolates of BFDV, from a Long-billed Corella (Cacatua tenuirostris) and an Orange-bellied parrot (OBP; Neophema chrysogaster). Purification by affinity and size exclusion chromatography was optimized through an iterative process involving screening and modification of buffer constituents and pH. A buffer containing glycerol, β-mercaptoethanol, Triton X-100, and a high concentration of NaCl at pH 8 was used to increase solubility of the protein. The final concentration of the corella-isolated BFDV protein was fifteen- to twenty-fold greater than that produced in previous publications using E. coli expression systems. Immunoassays were used to confirm the specific antigenicity of recombinant Cap, verifying its validity for use in continued experimentation as a potential vaccine, a reagent in diagnostic assays, and as a concentrated sample for biological discoveries.
Lopes, Lucia M.; Fabbricotti, Sandra H.; Ferreira, Antonio J. P.; Kato, Maria A. M. F.; Michalski, Jane; Scaletsky, Isabel C. A.
One hundred twelve diffusely adherent Escherichia coli strains isolated from children in a case control study were evaluated for virulence-associated characteristics, serotyping, antibiotic resistance, and plasmid profiles. Half of the strains hybridized with the probes for icuA (aerobactin) and fimH (type 1 pili); daaE (F1845 fimbriae), afa (afimbrial Dr adhesin), agg-3A (aggregative adhesion fimbria type III fimbriae), pap (P fimbriae), astA (EAST1 toxin), and shET1 (Shigella enterotoxin 1) sequences were present in <20% of the strains. The shET1 gene was noted most frequently in strains isolated from patients. A minority (7%) of the strains produced hemolysin or colicin or showed cytotoxic effects on Vero cells. Forty-five different serotypes were found. The majority (70%) of the strains presented multiple antibiotic resistance. Antibiotic resistance and diffuse adherence were located on the same conjugative plasmids. These results suggest that the transfer of these potential virulence markers could be important in the epidemiology of diffusely adherent E. coli. PMID:15815034
Tostes, Renata; Goji, Noriko; Amoako, Kingsley; Chui, Linda; Kastelic, John; DeVinney, Rebekah; Stanford, Kim; Reuter, Tim
Clinical outcomes of Shiga toxin (stx)-producing Escherichia coli infection are largely determined by virulence gene subtypes. This study used a polymerase chain reaction (PCR)-pyrosequencing assay to analyze single-nucleotide polymorphisms for subtyping three major virulence genes (stx1, stx2, eae) of pathogenic E. coli (O157, O26, O111, and O103) isolated from cattle over a 2-year interval (n = 465) and human clinical cases (n = 42) in western Canada. Most bovine isolates were PCR positive for at least one target virulence gene (367/465), whereas 100% of human isolates harbored eae in combination with at least one stx gene. Four Shiga toxin (1a, 2a, 2c, and 2e) and four eae (λ/γ1-eae, ɛ-eae, θ/γ2-eae, and β-eae) subtypes were identified in over 25 distinct virulence genotypes. Among cattle isolates, every serogroup, but O103, presented a dominant genotype (O157: stx1a+stx2a+λ/γ1-eae, O26: β-eae alone, and O111: stx1a+θ/γ2-eae). Similar patterns were found in human isolates, although it was not possible to establish a clear genotypic association between the two sources. Many O157 and non-O157 cattle isolates lacked stx genes; the absence was greater in non-O157 (75/258) and O157:non-H7 (19/40) than in O157:H7 strains (1/164). In addition, there was a greater diversity of virulence genotypes of E. coli isolated from cattle than those of human diseases, which could be due to sample characteristics (e.g., source and clinical condition). However, the majority of cattle strains had virulence profiles identical to those of clinical cases. Consequently, determining the presence of certain stx (stx1a and stx2a) and eae (λ/γ1-eae) subtypes known to cause human disease would be a valuable tool for risk assessment and prediction of disease outcome along the farm-to-fork continuum.
Obaid, Jamil M A S; Mansour, Samira R; Elshahedy, Mohammed S; Rabie, Tarik E; Azab, Adel M H
Uropathogenic Escherichia coli are the major causative agent of urinary tract infection--they may simultaneously express a number of virulence factors to cause disease. The aim of this study was to investigate the relation between virulence factors content of fifteen UPEC isolates and their pathogenic potential. The isolates belonged to the five serotypes O78:K80, O114:K90, O142:K86, O164 and O157. Nine of the virulence factors have been explored, ibeA, pap, sfa/foc, cnfl, hly, fyuA, pil, ompT and traT. Virulence factors profiling of the isolates revealed a different content ranging from 22% to 100% of the virulence genes explored. The pathogenic capacity of all fifteen isolates when tested on Vero cells showed that the cytotoxicity for all tested strains on Vero cells was approximately equal and enhanced after growth in syncase broth, leading mainly to cell lysis. The toxic effects reduced slightly after heat treatment of the toxin, and greatly after formalin detoxification, but not all the deleterious effect was abolished. Endotoxin also has cytotoxic effects on Vero cells, but longer time is needed for cytolysis which is greatly diminished with formalin treatment. In conclusion, our study revealed that pathogenic strains of UPEC can exert their pathogenic effect on live cells or system with limited virulence factors gene content.
Karam, Mohammad Reza Asadi; Rezaei, Ali Akbar; Siadat, Seyed Davar; Habibi, Mehri; Bouzari, Saeid
Background: Diarrhea, caused by enteroaggregative Escherichia coli (EAEC), is an important infection leading toillness and death. Numerous virulent factors have been described in EAEC. However, their prevalence was highly variable among EAECs of distinct geographic locations. Studies have shown that dispersin (antiaggregation protein, aap) is one of the important and abundant virulent factors in EAEC. In this study, we aimed to determine the presence, conservation, and immunogenicity of aap gene in EAEC isolated from Iranian patients. Methods: PCR amplification of aap gene in the EAEC isolates was performed, and the aap gene was cloned in pBAD-gIIIA vector. The sequence of aap gene was analyzed using the ExPASy and BLAST tools. The expression of aap gene was performed in E. coli Top10, and expression confirmation was carried out by SDS-PAGE and Western-blot techniques. Rabbits were immunized with purified dispersin protein emulsified with Freund’s adjuvant. Sera were collected and examined for antibody response. Finally, in vitro efficacy of dispersin and anti-dispersin was evaluated. Results: The results of PCR showed the presence of aap gene in all of the EAEC isolates with significant homology. Finally, the significant difference between the levels of IgG response in dispersin-injected rabbits and control group was observed. Conclusion: Our results were in accordance with other studies that reported the presence of dispersin in the EAEC isolates with high conservation and immunogenicity. Hence, dispersin could be a promising candidate for any probable prevention against EAEC infections. PMID:27155019
Haenni, Marisa; Saras, Estelle; Auvray, Frédéric; Forest, Karine; Oswald, Eric; Madec, Jean-Yves
We report the discovery of a CTX-M-15-producing Escherichia coli (STEC) of serogroup O111:H8, a major serotype responsible for human enterohemorrhagic Escherichia coli (EHEC) infections. In line with the recent CTX-M-15/O104:H4 E. coli outbreak, these data may reflect an accelerating spread of resistance to expanded-spectrum cephalosporins within the E. coli population, including STEC isolates. PMID:22156432
Mantilla-Calderon, David; Jumat, Muhammad Raihan; Wang, Tiannyu; Ganesan, Pugalenthi; Al-Jassim, Nada
The emergence of resistance to last-resort antibiotics is a public health concern of global scale. Besides direct person-to-person propagation, environmental pathways might contribute to the dissemination of antibiotic-resistant bacteria and antibiotic resistance genes (ARGs). Here, we describe the incidence of blaNDM-1, a gene conferring resistance to carbapenems, in the wastewater of the city of Jeddah, Saudi Arabia, over a 1-year period. blaNDM-1 was detected at concentrations ranging from 104 to 105 copies/m3 of untreated wastewater during the entire monitoring period. These results indicate the ubiquity and high incidence of blaNDM-1 in the local wastewater. To track the bacteria carrying blaNDM-1, we isolated Escherichia coli PI7, a strain of sequence type 101 (ST101), from wastewater around the Hajj event in October 2013. Genome sequencing of this strain revealed an extensive repertoire of ARGs as well as virulence and invasive traits. These traits were further confirmed by antibiotic resistance profiling and in vitro cell internalization in HeLa cell cultures. Given that this strain remains viable even after a certain duration in the sewerage, and that Jeddah lacks a robust sanitary infrastructure to fully capture all generated sewage, the presence of this bacterium in the untreated wastewater represents a potential hazard to the local public health. To the best of our knowledge, this is the first report of a blaNDM-1-positive E. coli strain isolated from a nonnosocomial environment in Saudi Arabia and may set a priority concern for the need to establish improved surveillance for carbapenem-resistant E. coli in the country and nearby regions. PMID:27324770
Álvarez-Suárez, María-Elena; Otero, Andrés; García-López, María-Luisa; Dahbi, Ghizlane; Blanco, Miguel; Mora, Azucena; Blanco, Jorge; Santos, Jesús A
The aim of this study was to characterize a collection of 44 Shiga toxin-producing (STEC) and enteropathogenic Escherichia coli (EPEC) isolated from goat milk and goat farm environment. Of the 19 STEC isolates, five (26.3%) carried the stx1 gene, four (21.1%) the stx2 gene and 10 (52.6%) presented both stx genes. Six (31.6%) STEC strains were eae-positive and belonged to serotypes related to severe human disease (O157:H7 and O5:HNM). Another seven STEC strains were of serotype O146:H21 and three of serotype O166:H28, also linked to human disease. The STEC strains isolated from goat milk were of serotypes potentially pathogenic for humans. All the 25 EPEC isolates were considered atypical (aEPEC) and one aEPEC strain was of serotype O26:H11, a serotype frequently isolated in children with diarrhea. Multilocus sequence typing (MLST) was carried out with seven housekeeping genes and 23 sequence types (ST) were detected, 14 of them newly described. Twelve STs grouped STEC isolates and 11 STs grouped EPEC isolates. Genetic typing by pulsed field gel electrophoresis (PFGE) resulted in 38 patterns which grouped in 10 clusters. Well-defined groups were also observed for strains of pathogenic serotypes. In conclusion, strains of STEC and aEPEC belonging to serotypes related to severe human disease have been detected in goat milk and the goat farm environment. Ruminants are an important reservoir of STEC strains and the role of these animals as carriers of other pathogenic types of E. coli seems to be an emerging concern.
Varga, Csaba; Rajíc, Andrijana; McFall, Margaret E.; Avery, Brent P.; Reid-Smith, Richard J.; Deckert, Anne; Checkley, Sylvia L.; McEwen, Scott A.
The objective of this study was to determine the prevalence of antimicrobial resistance in generic Escherichia coli isolates obtained from 90 Alberta finisher swine farms. Up to 5 isolates were obtained from each of 269 pooled fecal samples and were classified as susceptible or resistant according to Clinical and Laboratory Standards Institute guidelines. Of the 1322 isolates, 166 (12.6%) were susceptible to all 15 antimicrobials. No resistance to amikacin, ceftiofur, ceftriaxone, or ciprofloxacin, antimicrobials of importance in human medicine, was observed. Relatively low frequencies of resistance were observed to gentamicin (1.1%), amoxicillin/clavulanic acid (0.7%), and cefoxitin (0.7%). Higher frequencies of resistance were observed for tetracycline (78.9%), sulfisoxazole (49.9%), streptomycin (49.6%), ampicillin (30.6%), chloramphenicol (17.6%), kanamycin (10%), and trimethoprim/sulfamethoxazole (6.4%). Among the isolates resistant to ≥ 2 antimicrobial classes, 20.8%, 20.6%, 18.2%, 7.0%, 1.8%, 0.2%, and 0.2% were resistant to 2, 3, 4, 5, 6, 7, and 8 antimicrobials, respectively. The most common multidrug-resistance patterns (resistance to ≥ 2 antimicrobial classes) were streptomycin-tetracycline (9.4%), streptomycin-sulfisoxazole-tetracycline (6.2%), and ampicillin-streptomycin-sulfisoxazole-tetracycline (6.1%). More clustering (higher intra-class correlation coefficients) in antimicrobial resistance was observed for isolates at the same visit than for isolates from different visits in the same farm, indicating that sampling more farms, testing fewer isolates per visits, and taking longer periods between visits may be appropriate and more efficient for a better understanding of potential shifts in resistance over time. PMID:18505207
Baron, S; Delannoy, S; Bougeard, S; Larvor, E; Jouy, E; Balan, O; Fach, P; Kempf, I
This study investigated antimicrobial resistance, screened for the presence of virulence genes involved in intestinal infections, and determined phylogenetic groups of Escherichia coli isolates from untreated poultry and poultry treated with ceftiofur, an expanded-spectrum cephalosporin. Results show that none of the 76 isolates appeared to be Shiga toxin-producing E. coli or enteropathogenic E. coli. All isolates were negative for the major virulence factors/toxins tested (ehxA, cdt, heat-stable enterotoxin [ST], and heat-labile enterotoxin [LT]). The few virulence genes harbored in isolates generally did not correlate with isolate antimicrobial resistance or treatment status. However, some of the virulence genes were significantly associated with certain phylogenetic groups.
Moredo, Fabiana A; Cappuccio, Javier A; Insarralde, Lucas; Perfumo, Carlos J; Quiroga, María A; Leotta, Gerardo A
The purpose of this work was to characterize 47 Escherichia coli strains isolated from 32 pigs diagnosed with postweaning diarrhea and three pigs with edema disease by PCR. Forty two (95.5 %) of the strains isolated from diarrheic pigs were characterized as enterotoxigenic E. coli (ETEC) and 2 (4.5 %) as Shiga toxin-producing E. coli (STEC). Fourteen (33.3 %) ETEC strains were positive for est/estII/fedA genes. The most complex genotype was eltA/estI/faeG/aidA. Strains isolated from pigs with ED were classified as porcine STEC and were stx2e/aidA carriers. Eleven (25 %) strains carried the gene encoding adhesin protein AIDA-I. However, genes coding for F5, F6, F41, intimin and Paa were not detected. The development of vaccines generating antibodies against prevalent E. coli adhesins in Argentina could be useful for the prevention of PWD and ED.
Nataro, James P.; Kaper, James B.
Escherichia coli is the predominant nonpathogenic facultative flora of the human intestine. Some E. coli strains, however, have developed the ability to cause disease of the gastrointestinal, urinary, or central nervous system in even the most robust human hosts. Diarrheagenic strains of E. coli can be divided into at least six different categories with corresponding distinct pathogenic schemes. Taken together, these organisms probably represent the most common cause of pediatric diarrhea worldwide. Several distinct clinical syndromes accompany infection with diarrheagenic E. coli categories, including traveler’s diarrhea (enterotoxigenic E. coli), hemorrhagic colitis and hemolytic-uremic syndrome (enterohemorrhagic E. coli), persistent diarrhea (enteroaggregative E. coli), and watery diarrhea of infants (enteropathogenic E. coli). This review discusses the current level of understanding of the pathogenesis of the diarrheagenic E. coli strains and describes how their pathogenic schemes underlie the clinical manifestations, diagnostic approach, and epidemiologic investigation of these important pathogens. PMID:9457432
Alikhani, Mohammad Yousef; Sedighi, Iraj; Zamani, Alireza; Aslani, Mohammad Mehdi; Sadrosadat, Taravat
Diarrhoeagenic Escherichia coli (DEC) represents a main group of enteric pathogens that cause human diarrhoea. Because it is not simply distinguished from normal flora by simple laboratory methods, modern molecular diagnostic assays are necessary. Although it is neither necessary nor applicable to perform PCR for all patients, it is of many advantages to verify the prevalence of DEC in different areas by this method. Knowing the prevalence of DEC in an area, we can focus on few pathogens and narrow our antimicrobial treatment. The aim of this study is to evaluate the contribution of the different DEC categories in children diarrhoea in the west of Iran.The stool specimens of 251 children with diarrhoea were collected from June to September 2007. Polymerase chain reaction (PCR) was performed to determine the presence of enteropathogenic (EPEC), enterotoxigenic (ETEC), entero-invasive (EIEC), Shiga toxin-producing (STEC) and entero-aggregative (EAEC) strains. ETEC strains were isolated from 13 and EAEC strains from 16 children. STEC was detected in 7 children, and no EIEC was isolated. Finally, EPEC strains were isolated in 41 cases. EAEC and EPEC are the most frequent DECs in children less than 10 years of age in West of Iran.
Gahlot, Sunayna; Joshi, Amita; Singh, Pratap; Tuteja, Renu; Dua, Meenakshi; Jogawat, Abhimanyu; Kumar, Manoj; Raj, Sumit; Dayaman, Vikram; Johri, Atul Kumar; Tuteja, Narendra
Piriformospora indica, a root endophytic fungus identified in the Indian Thar desert, colonizes the roots of plants and provides resistance towards biotic stress as well as tolerance to abiotic stress in the plants. Despite its positive impact on the host, little is known about the P. indica genes that are involved in salt stress tolerance. Therefore this study was conducted to identify and isolate high salinity-tolerance genes from P. indica. Thirty-six salinity-tolerance genes were obtained by functional screening, based on random over expression of a P. indica cDNA library in Escherichia coli grown on medium supplemented with 0.6 M NaCl. The salinity tolerance conferred by these 36 genes in bacteria was further confirmed by using another strain of E. coli (DH5α) transformants. However when the expression of these 36 genes was analysed in P. indica using quantitative RT-PCR, we found only six genes were up-regulated by salt stress. These six genes are involved in different cellular processes, such as metabolism, energy and biosynthetic processes, DNA repair, regulation of protein turnover, transport and salt stress tolerance. This work presents the basis for further molecular analyses of the mechanisms of salt tolerance in P. indica and for the use of this endophyte to confer salt tolerance to plants.
Background Diffusely adherent Escherichia coli (DAEC) have been considered a diarrheagenic category of E. coli for which several potential virulence factors have been described in the last few years. Despite this, epidemiological studies involving DAEC have shown inconsistent results. In this work, two different collections of DAEC possessing Afa/Dr genes, from children and adults, were studied regarding characteristics potentially associated to virulence. Results DAEC strains were recovered in similar frequencies from diarrheic and asymptomatic children, and more frequently from adults with diarrhea (P < 0.01) than from asymptomatic adults. Association with diarrhea (P < 0.05) was found for SAT-positive strains recovered from children and for curli-positive strains recovered from adults. Mixed biofilms involving DAEC and a Citrobacter freundii strain have shown an improved ability to form biofilms in relation to the monocultures. Control strains have shown a greater diversity of Afa/Dr adhesins and higher frequencies of cellulose, TTSS, biofilm formation and induction of IL-8 secretion than strains from cases of diarrhea in children. Conclusions DAEC strains possessing Afa/Dr genes isolated from children and adults represent two different bacterial populations. DAEC strains carrying genes associated to virulence can be found as part of the normal microbiota present in asymptomatic children. PMID:23374248
Wang, Gui-Qin; Wu, Cong-Ming; Du, Xiang-Dang; Shen, Zhang-Qi; Song, Li-Hua; Chen, Xia; Shen, Jian-Zhong
To assess the prevalence of antimicrobial resistance and class I integrons in Escherichia coli strains (n=58) isolated from bovine mastitis in Inner Mongolia, antimicrobial susceptibility and the presence of various types of integrons were characterized. Most isolates were susceptible to amikacin, colistin, ceftazidime, gentamicin and kanamycin, while those also exhibited high resistant incidence rates to ampicillin, amoxicillin, sulfadiazine and sulfamethoxydiazine. The integrase gene of integrons was amplified by PCR using degenerate primers. The integrons were confirmed by restriction fragment length polymorphism (RFLP) analysis of positive PCR products. Neither class II nor class III integron was detected, while 56.90% (n=33) of the isolates were positive for the presence of intI1 gene. Sequencing analysis of gene cassettes revealed that seven gene cassettes were found, which encoded resistance to trimethoprim (dfrA1 and dfrA17), aminoglycosides (aacA4, aadA1 and aadA5) and chloramphenicol (catB3), respectively. Of them, the gene cassette array dfrA17-aadA5 was found most prevalent (62.96%). The percentage of positive-integron among the isolates whose resistant profile was relatively broad (n> or =7) is 100.00%, while the one in narrow-profile isolates (n=2-6) is 30.56%. The correlation analysis revealed the incidence of integrons among the isolates were highly related to the resistant profile, indicating integrons play an important role in the dissemination and spread of the antimicrobial resistant strains.
Escherichia coli is a bacterial species which inhabits the gastrointestinal tract of man and warm-blooded animals. Because of the ubiquity of this bacterium in the intestinal flora, it serves as an important indicator organism of fecal contamination. E. coli, aside from serving a...
Escherichia coli, a member of the Enterobacteriaceae family, is a part of the normal flora of the intestinal tract of humans and a variety of animals. E. coli strains are classified on the basis of antigenic differences in two surface components (serotyping), the somatic antigen (O) of the lipopoly...
Evans, D J; Evans, D G; DuPont, H L; Orskov, F; Orskov, I
Enterotoxigenic Escherichia coli isolates obtained in Mexico from adult subjects with diarrhea and from healthy controls were examined for the production of heat-stable enterotoxin (ST) and heat-labile enterotoxin (LT) after serial passage in the laboratory. Isolates were found to be either stable for the production of ST and LT or unstable with respect to ST, LT, or both. Unilateral loss of either ST or LT production allowed classification of E. coli isolates into four groups according to stability/instability of enterotoxin production. Fewer serotypes, with more representative isolates, were in group I (stable) than in group IV (completely unstable). Isolates from Dacca, Bangladesh, could be similarly classified into stability groups. There is an apparent relationship between serotype, stability of enterotoxin production, particularly LT, and isolation from diarrhea cases as opposed to isolation from healthy controls. PMID:328392
Lobos, Olga; Padilla, Carlos
Vaginal infections such as vulvovaginal candiadiasis, trichomoniasis and bacterial vaginosis are common worldwide. Accurate diagnosis and prescription of appropriate treatments are important since these infections are linked to adverse outcomes for women during pregnancy and for newborns. Several aetiological agents are responsible for these infectious diseases; however, the presence of Escherichia coli in these infections is controversial. Thus, it is important to identify some phenotypic and genotypic properties of E. coli strains isolated from vaginal infections. Forty-six E. coli strains isolated from vaginal fluid as the sole micro-organism, and 20 other E. coli strains isolated from other samples (urinary tract infections, otitis and septicaemia) were analysed by several phenotypic tests. In addition, genotypic features were studied by RAPD-PCR techniques. Biochemical tests showed that the E. coli strains isolated from vaginal fluid could be grouped into a single cluster which is subdivided into two phenogroups. Analysis of the dendrogram based on fragment length polymorphisms of genomic DNA indicated that E. coli isolates from vaginal infections form a single cluster with two subdivisions. Further studies are needed to analyse the molecular structure and virulence characteristics of these E. coli strains in order to determine their potential role in vaginal infections.
Pereira, Andrea S; Andrade, Soraya S; Monteiro, Jussimara; Sader, Helio S; Pignatari, Antonio C C; Gales, Ana C
Increasing quinolone resistance has been reported worldwide, mainly among clinical isolates of Escherichia coli. The objectives of this study were to determine the susceptibility profile, the genetic relatedness, and the prevalence of the qnr gene among ciprofloxacin-resistant Escherichia coli isolated from distinct Brazilian hospitals. A total of 144 ciprofloxacin-resistant Escherichia coli were isolated from 17 Brazilian hospitals between January/2002 and June/2003. The antimicrobial susceptibility testing was performed by microdilution according to NCCLS. The presence of the qnr gene was initially screened by colony blotting, and then confirmed by PCR followed by DNA sequencing. Ninety-five urinary ciprofloxacin-resistant Escherichia coli were further selected for molecular typing by pulsed-field gel electrophoresis (PFGE). Imipenem and meropenem showed the highest susceptibility rates (100.0% for both compounds) followed by amikacin (91.0%) and piperacillin/tazobactan (84.8%). A single ciprofloxacin-resistant Escherichia coli isolate was positive for qnr among the 144 ciprofloxacin-resistant Escherichia coli. Forty-six PFGE patterns were observed among the 95 ciprofloxacin-resistant Escherichia coli type. This study shows that therapeutic options are limited for treatment of ciprofloxacin-resistant Escherichia coli due to the presence of additional mechanisms of antimicrobial resistance, such as ESBL production. The qnr gene was uncommon among ciprofloxacin-resistant Escherichia coli clinical isolates, but its identification might indicate the emergence of this mechanism of quinolone resistance in Brazil. The great genomic variability found among the ciprofloxacin-resistant Escherichia coli highlights the importance of the appropriate use of quinolone to restrict the selection of resistant isolates.
Salit, I E; Vavougios, J; Hofmann, T
Bacteria which attach to different mucous membranes should have differing specificities of adherence in vitro. Human Escherichia coli isolates from blood and urine (pathogens) and from stool and throat (commensals) were characterized as to the patterns of hemagglutination (HA), as well as the structure and function of their pili. Bacterial HA was done in microtiter plates and on slides after bacterial growth in broth or agar. Human erythrocytes were agglutinated by 95% of the pathogens and 65 to 70% of the commensals grown in broth or agar. Mannose-resistant HA was characteristically caused by pathogens, and commensals characteristically caused mannose-sensitive HA of guinea pig cells. Strains often had both mannose-resistant and mannose-sensitive reactions, or even a mannose-paradoxical reaction. Pathogens more often caused HA, but titers were lower than those for commensals. Slide HA was less sensitive than the microtiter method. All isolates were piliated. Commensals also had more pili than pathogens when grown in broth (117.8 versus 38.3 pili per bacterium), but pathogens had more pili after growth on agar (32.1 versus 8.1 pili per bacterium). Isolates causing high-titer HA had large numbers of pili (greater than 85 pili per bacterium), but some well-piliated strains were non-hemagglutinating. Pili were purified from seven E. coli strains from different sites of isolation and with different erythrocyte-binding specificity. Pili usually migrated as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. However, more than one type of pilus could be copurified from some strains since there were two or more bands after separation in octyl-glucoside and two different amino terminal sequences. Protein sequencing was done on five different pili: four resembled type 1 pili and one was a P fimbria. The type 1-like pili (strains 2239 and 9353) had an initial variable sequence of 1 to 5 residues, followed by a common region of 21 residues. The P
Del Rio-Avila, C; Rosario, C; Arroyo-Escalante, S; Carrillo-Casas, E M; Díaz-Aparicio, E; Suarez-Güemes, F; Silva-Sanchez, J; Xicohtencatl-Cortes, J; Maravilla, P; Hernández-Castro, R
Forty-two enrofloxacin-resistant Escherichia coli strains isolated from eggs and first-week mortality associated with yolk sac infection of two vertically integrated poultry companies of Central Mexico in 1997 and 2005 were characterised. E. coli resistance to 19 antibiotics was determined, as well as the minimum inhibitory concentrations (broth dilution) for ciprofloxacin. The presence of gyrA,B, parC,E chromosomal point mutations, qnrA,B,S plasmid genes and the aminoglycoside acetyltransferase aac(6')-Ib-cr were determined by PCR and sequencing. Resistance to ampicillin (95%), piperacillin (95%), gatifloxacin (95%), levofloxacin (95%), ampicillin/sulbactam (90%), cefazolin (85%), trimethoprim/sulfamethoxazole (80%), amoxicillin/clavulanic acid (80%), aztreonam (80%), cefepime (80%), cefotaxime (80%), ceftazidime (80%), ceftriaxone (80%) and cefoxitin (75%) was high in the 2005 strains and 19 (95%) strains were resistant to 7 or more antimicrobials. The strains from 1997 expressed high rates of resistance only to the fluoroquinolones and 4 strains (18%) expressed resistance to 7 or more antimicrobials. All strains had a gyrA mutation (Ser83Leu) and a parC mutation (Ser80Ile or Ser80Arg) and 41 (97.6%) strains had a second gyrA mutation (Asp87Asn, Asp87Tyr or Asp87Gly). Only two (4.7%) strains had a parE mutation (Ser458Ala). A total of 10 strains were positive for the aac(6')-Ib wild-type gene, 6 strains for the aac(6')-Ib-cr variant and 6 strains possessed both the wild type and the variant. No gyrB mutations or qnrA,B,S genes were detected. This is the first report in Latin America of chromosomal and plasmid quinolone resistance genes in E. coli strains recovered from poultry.
Calhau, Vera; Mendes, Catarina; Pena, Angelina; Mendonça, Nuno; Da Silva, Gabriela Jorge
Escherichia coli is simultaneously an indicator of water contamination and a human pathogen. This study aimed to characterize the virulence and resistance of E. coli from municipal and hospital wastewater treatment plants (WWTPs) in central Portugal. From a total of 193 isolates showing reduced susceptibility to cefotaxime and/or nalidixic acid, 20 E. coli with genetically distinct fingerprint profiles were selected and characterized. Resistance to antimicrobials was determined using the disc diffusion method. Extended spectrum β-lactamase and plasmid-mediated quinolone resistance genes, phylogroups, pathogenicity islands (PAIs) and virulence genes were screened by polymerase chain reaction (PCR). CTX-M producers were typed by multilocus sequence typing. Resistance to beta-lactams was associated with the presence of bla(TEM), bla(SHV), bla(CTX-M-15) and bla(CTX-M-32). Plasmid-mediated quinolone resistance was associated with qnrA, qnrS and aac(6')-Ib-cr. Aminoglycoside resistance and multidrug-resistant phenotypes were also detected. PAI IV(536), PAI II(CFT073), PAI II(536) and PAI I(CFT073), and uropathogenic genes iutA, papAH and sfa/foc were detected. With regard to the clinical ST131 clone, it carried bla(CTX-M-15), blaTEM-type, qnrS and aac(6')-lb-cr; IncF and IncP plasmids, and virulence factors PAI IV(536), PAI I(CFT073), PAI II(CFT073), iutA, sfa/foc and papAH were identified in the effluent of a hospital plant. WWTPs contribute to the dissemination of virulent and resistant bacteria in water ecosystems, constituting an environmental and public health risk.
McConnel, Craig S; Stenkamp-Strahm, Chloe M; Rao, Sangeeta; Linke, Lyndsey M; Magnuson, Roberta J; Hyatt, Doreene R
Escherichia coli O157 (EcO157) infections can lead to serious disease and death in humans. Although the ecology of EcO157 is complex, ruminant animals serve as an important reservoir for human infection. Dairy cattle are unique because they may be a source of contamination for milk, meat, and manure-fertilized crops. Foodborne dairy pathogens such as EcO157 are of primary importance to public health. Antimicrobial resistance (AMR) is a complex phenomenon that complicates the treatment of serious bacterial infections and is of increasing concern. In the face of recommended use restrictions for antimicrobial agents in livestock operations, current AMR patterns in known foodborne pathogens should be documented. The objective of this study was to document AMR patterns in EcO157 isolates from dairies in northern Colorado using antimicrobial agents commonly found on dairies and representative of medically important antimicrobial drug classes. Seventy-five EcO157 isolates were recovered from three dairies. Six isolates were resistant to at least 1 of the 10 tested antimicrobial agents: four were resistant to streptomycin, sulfisoxazole, and tetracycline; one was resistant to streptomycin and tetracycline; and one was resistant to only tetracycline. All resistant isolates were from a single dairy. Overall, a low prevalence (8%) of AMR was observed among the 75 EcO157 isolates. No significant effects on AMR profiles due to virulence genes, parity, or previous antimicrobial treatments within the current lactation period were detected. The results of this study provide background information for future comparative studies investigating AMR trends. Future studies should include more participating farms and more samples and should control for potential confounding factors of AMR that may underlie individual farm variation.
Soto, S. M.; Bosch, J.; Jimenez de Anta, M. T.; Vila, J.
Neonatal meningitis and septicemia caused by Escherichia coli are still major health problems in industrialized countries. Forty-seven E. coli strains causing neonatal sepsis were analyzed. Twenty-two and 25 strains caused early (detected from 0 to 3 days after birth) and late (detected from 4 to 28 days after birth) infections, respectively. Only the ibeA gene was significantly more prevalent in the strains causing early infections. PMID:18160454
Yao, Yufeng; Xie, Yi; Kim, Kwang Sik
Escherichia coli is a major cause of enteric/diarrheal diseases, urinary tract infections, and sepsis. E. coli K1 is the leading gram-negative organism causing neonatal meningitis, but the microbial basis of E. coli K1 meningitis is incompletely understood. Here we employed comparative genomic hybridization to investigate 11 strains of E. coli K1 isolated from the cerebrospinal fluid (CSF) of patients with meningitis. These 11 strains cover the majority of common O serotypes in E. coli K1 isolates from CSF. Our data demonstrated that these 11 strains of E. coli K1 can be categorized into two groups based on their profile for putative virulence factors, lipoproteins, proteases, and outer membrane proteins. Of interest, we showed that some open reading frames (ORFs) encoding the type III secretion system apparatus were found in group 2 strains but not in group 1 strains, while ORFs encoding the general secretory pathway are predominant in group 1 strains. These findings suggest that E. coli K1 strains isolated from CSF can be divided into two groups and these two groups of E. coli K1 may utilize different mechanisms to induce meningitis. PMID:16552050
Carregaro, Adriano Bonfim; Santurio, Deise Flores; de Sá, Mariangela Facco; Santurio, Janio Moraes; Alves, Sydney Hartz
This study evaluated the in vitro antibacterial activity of essential oils from Lippia graveolens (Mexican oregano), Origanum vulgaris (oregano), Thymus vulgaris (thyme), Rosmarinus officinalis (rosemary), Cymbopogon nardus (citronella), Cymbopogon citratus (lemongrass), and Eucalyptus citriodora (eucalyptus) against Escherichia coli (n = 22) strains isolated from Alouatta spp. feces. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined for each isolate using the broth microdilution technique. Essential oils of Mexican oregano (MIC mean = 1818 μg mL−1; MBC mean = 2618 μg mL−1), thyme (MIC mean = 2618 μg mL−1; MBC mean = 2909 μg mL−1), and oregano (MIC mean = 3418 μg mL−1; MBC mean = 4800 μg mL−1) showed the best antibacterial activity, while essential oils of eucalyptus, rosemary, citronella, and lemongrass displayed no antibacterial activity at concentrations greater than or equal to 6400 μg mL−1. Our results confirm the antimicrobial potential of some essential oils, which deserve further research. PMID:27313638
Lara, Valéria Maria; Carregaro, Adriano Bonfim; Santurio, Deise Flores; de Sá, Mariangela Facco; Santurio, Janio Moraes; Alves, Sydney Hartz
This study evaluated the in vitro antibacterial activity of essential oils from Lippia graveolens (Mexican oregano), Origanum vulgaris (oregano), Thymus vulgaris (thyme), Rosmarinus officinalis (rosemary), Cymbopogon nardus (citronella), Cymbopogon citratus (lemongrass), and Eucalyptus citriodora (eucalyptus) against Escherichia coli (n = 22) strains isolated from Alouatta spp. feces. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined for each isolate using the broth microdilution technique. Essential oils of Mexican oregano (MIC mean = 1818 μg mL(-1); MBC mean = 2618 μg mL(-1)), thyme (MIC mean = 2618 μg mL(-1); MBC mean = 2909 μg mL(-1)), and oregano (MIC mean = 3418 μg mL(-1); MBC mean = 4800 μg mL(-1)) showed the best antibacterial activity, while essential oils of eucalyptus, rosemary, citronella, and lemongrass displayed no antibacterial activity at concentrations greater than or equal to 6400 μg mL(-1). Our results confirm the antimicrobial potential of some essential oils, which deserve further research.
Mercer, A A; Morelli, G; Heuzenroeder, M; Kamke, M; Achtman, M
Escherichia coli K1 isolates of various O types were previously assigned to different clonal groups. Members of the two clones defined by membrane pattern 9 (MP9) and serotypes O18:K1 and O1:K1 had been found to be very similar to each other. The plasmid contents of these bacteria confirmed this conclusion. Both groups carried a self-transmissible plasmid of the FI incompatibility group that coded for colicin production and a major outer membrane protein called the plasmid-coded protein (PCP). The size of this plasmid varied from 76 to 96 megadaltons, but restriction endonuclease digestion and DNA heteroduplex analysis revealed that these plasmids were highly related. O18:K1 bacteria of MP6 had previously been determined to represent a subclone, related to but different from O18:K1 MP9 bacteria. These MP6 bacteria carried a different, smaller IncFI plasmid which did not code for colicin production or the PCP protein. This smaller plasmid was primarily related to the larger plasmid within the regions of DNA encoding incompatibility, replication, and conjugation. O1:K1 bacteria of MP5 contained other unrelated plasmids in agreement with the previous conclusion that they are unrelated to O1:K1 bacteria of MP9. The bacteria examined had been isolated from two continents over a time span of 38 years, and the results attest to conservative inheritance of plasmids within bacteria of common descent.
Wang, Yang; Yi, Li; Wang, Yuxin; Wang, Yuanguo; Cai, Ying; Zhao, Wenpeng; Ding, Chan
The isolation and identification, genetic typing, antibiotic sensitivity, and biofilm formation of avian Escherichia coli in central China was studied. A total of 256 isolates of E. coli were obtained, and classified into groups: A (50.78%, 130/256), B1 (11.72%, 30/256), B2 (17.58%, 45/256), and D (19.92%, 51/256). Drug susceptibility testing revealed that the strains showed a high drug resistance rate against penicillin, aztreonam, rifampicin, kanamycin, clindamycin, and gentamicin, with 92.19% of strains exhibiting multi-drug resistance. A biofilm assay revealed that 81.64% of isolates could form biofilms. Of the total isolates, 25.39% of isolates showed strong biofilm-formation ability, 31.25% showed moderate biofilm-formation ability, 28.90% showed weak biofilm-formation ability, and 18.36% were unable to form biofilms. Most adhesion-associated genes were distributed among 5 or 8 genes in strong biofilm-forming ability isolates. However, adhesion-associated genes distributed among 1 or 4 genes were found in weak biofilm-forming ability isolates and non-ability isolates. The results showed a high drug resistance rate and biofilm formation ability in E.coli strains isolated from poultry. The isolates which have strong biofilm-forming ability were mostly belong to pathogenic E. coli (B2, D). Furthermore, it was the first report to demonstrate a positive correlation between adhesion-encoding genes and biofilms phenotype.
Grande Burgos, María José; Fernández Márquez, Maria Luisa; Pérez Pulido, Rubén; Gálvez, Antonio; Lucas López, Rosario
Eggs may contain extraintestinal pathogenic (ExPEC) and diarrheogenic (DEC) Escherichia coli which in addition may carry antibiotic resistance. The wide use of biocides and disinfectants in the food industry may induce biocide tolerance in bacteria. The aim of the present study was to evaluate biocide tolerance and antibiotic resistance in E. coli from hen egg shells. A total of 27 isolates obtained from a screening of 180 eggs were studied. Seven isolates carried both eae and bfpA genes of typical enteropathogenic E. coli (EPEC) strains, while 14 isolates only carried eae associated with atypical EPEC strains. Shiga toxin genes stx and stx2 were detected in four isolates. Heat-stable and heat-labile enterotoxin genes as well as aggR were also detected. Several isolates had minimum inhibitory concentrations (MICs) that were higher than the wild-type for the biocide hexadecylpyridinium chloride (HDP, 18.52%) or the commercial disinfectant P3 oxonia (OX, 14.81%). Antibiotic resistance was detected for ampicillin (37.03%), streptomycin (37.03%), tetracycline (37.03%), chloramphenicol (11.11%), nalidixic acid (18.51%) and trimethoprim-sulfamethoxazole (14.81%). Eight isolates (29.63%) were biocide tolerant and antibiotic resistant. Efflux pump genes detected included acrB (96.29%), mdfA (85.18%) and oxqA (37.03%), in addition to quaternary ammonium compound (QAC) resistance genes qacA/B (11.11%) and qacE (7.40%). Antibiotic resistance genes detected included blaCTX-M-2 (22.22%), blaTEM (3.70%), blaPSE (3.70%), tet(A) (29.63%), tet(B) (29.63%), tet(C) (7.40%), tet(E) (11.11%), aac(6')-Ib (3.70%), sul1 (14.81%), dfrA12 (3.70%) and dfrA15 (3.70%). Most isolates (96.30%) carried more than one genetic determinant of resistance. The most frequent combinations were efflux pump components acrB and mdfA with tetracycline resistance genes (33.33% of isolates). Isolates carrying QAC resistance genes also carried between 4 and 8 of the additional antimicrobial resistance genes
Gu, Zhenyu; Weidenhaupt, Marianne; Ivanova, Natalia; Pavlov, Michail; Xu, Bingze; Su, Zhi-Guo; Janson, Jan-Christer
New methods for the chromatographic isolation of inclusion bodies directly from crude Escherichia coli homogenates and for the refolding of denatured protein are presented. The traditional method of differential centrifugation for the isolation of purified inclusion bodies is replaced by a single gel-filtration step. The principle is that the exclusion limit of the gel particles is chosen such that only the inclusion bodies are excluded, i.e., all other components of the crude homogenate penetrate the gel under the conditions selected. In the novel column refolding process, a decreasing gradient of denaturant (urea or Gu-HCl), combined with an increasing pH gradient, is introduced into a gel-filtration column packed with a gel medium that has an exclusion limit lower than the molecular mass of the protein to be refolded. A limited sample volume of the protein, dissolved in the highest denaturant concentration at the lowest pH of the selected gradient combination, is applied to the column. During the course of elution, the zone of denatured protein moves down the column at a speed approximately threefold higher than that of the denaturant. This means that the protein sample will gradually pass through areas of increasingly lower denaturant concentrations and higher pH, which promotes refolding into the native conformation. The shape and slope of the gradients, as well as the flow rate, will influence the refolding rate and can be adjusted for different protein samples. The principle is illustrated using a denatured recombinant scFv fusion protein obtained from E. coli inclusion bodies.
Manjarrez-Hernandez, H. A.; Gavilanes-Parra, S.; Chavez-Berrocal, E.; Navarro-Ocaña, A.; Cravioto, A.
Enteropathogenic Escherichia coli (EPEC) produces a characteristic attaching and effacing (A/E) lesion in the small intestines of infected children. The immune response to EPEC infection remains poorly characterized. The molecular targets that elicit protective immunity against EPEC disease are unknown. In this study protein antigens from EPEC were identified using secretory immunoglobulin A (sIgA) antibodies isolated from milk from Mexican women by Western blot analysis. Purified sIgA antibodies, which inhibit the adherence of EPEC to cells, reacted to many EPEC proteins, the most prominent of which were intimin (a 94-kDa outer membrane protein) and two unknown proteins with apparent molecular masses of 80 and 70 kDa. A culture supernatant protein of 110 kDa also reacted strongly with the sIgA antibodies. The molecular size of this protein and its reactivity with specific anti-EspC antiserum suggest that it is EPEC-secreted protein C (EspC). These EPEC surface protein antigens were consistently recognized by all the different sIgA samples obtained from 15 women. Screening of clinical isolates of various O serogroups from cases of severe infantile diarrhea revealed that all EPEC strains able to produce the A/E lesion showed expression of intimin and the 80- and 70-kDa proteins. Such proteins reacted strongly with the purified sIgA pool. Moreover, nonvirulent E. coli strains were unable to generate a sIgA response. The immunogenic capacities of the 80- and 70-kDa proteins as virulence antigens have not been previously reported. The strong sIgA response to intimin and the 80- and 70-kDa proteins obtained in this study indicates that such antigens stimulate intestinal immune responses and may elicit protective immunity against EPEC disease. PMID:10948121
Miranda, J M; Vázquez, B I; Fente, C A; Calo-Mata, P; Cepeda, A; Franco, C M
The presence of Escherichia coli, Staphylococcus aureus, and Listeria monocytogenes was determined in 55 samples of organic poultry meat and in 61 samples of conventional poultry meat. A total of 220 E. coli, 192 S. aureus, and 71 L. monocytogenes strains were analyzed by an agar disk diffusion assay for their resistance to ampicillin, cephalothin, chloramphenicol, ciprofloxacin, doxycycline, fosfomycin, gentamicin, nitrofurantoin, streptomycin, and sulfisoxazole (E. coli); chloramphenicol, ciprofloxacin, clindamycin, doxycycline, erythromycin, gentamicin, nitrofurantoin, oxacillin, and sulfisoxazole (S. aureus); and chloramphenicol, doxycycline, erythromycin, gentamicin, sulfisoxazole, and vancomycin (L. monocytogenes). The results indicated a significantly higher (P < 0.0001) prevalence of E. coli but not of S. aureus and L. monocytogenes in organic poultry meat as compared with conventional poultry meat. E. coli isolated from organic poultry meat exhibited lower levels of antimicrobial resistance against 7 of the 10 antimicrobials tested as compared with isolates recovered from conventional meat. In the case of S. aureus and L. monocytogenes isolated from conventional poultry, antimicrobial resistance was significantly higher only for doxycycline as compared with strains isolated from organic poultry. In the case of E. coli, the presence of multiresistant strains was significantly higher (P < 0.0001) in conventional poultry meat as compared with organic poultry meat. Organically farmed poultry samples showed significantly lower development of antimicrobial resistance in intestinal bacteria such as E. coli.
Rocha-Gracia, R C; Cortés-Cortés, G; Lozano-Zarain, P; Bello, F; Martínez-Laguna, Y; Torres, C
The presence of extended spectrum β-lactamase (ESBL) and plasmid-mediated AmpC β-lactamase (pAmpC) producing Escherichia coli, along with the mechanisms of antimicrobial resistance and the molecular types of isolates, was investigated in faecal samples from 53 healthy dogs in Mexico. Samples were inoculated on Levine agar plates with 2 µg/mL cefotaxime for recovery of cefotaxime-resistant (CTX(R)) E. coli. CTX(R)E. coli isolates were recovered from 9/53 (17%) samples; one isolate was characterised from each positive sample. ESBL producing E. coli isolates were detected in 3/53 (6%) samples; these isolates carried the blaCTX-M-15 gene and one isolate also carried blaSHV-2. These three ESBL-positive E. coli isolates belonged to phylogroup A and sequence types ST617, ST410 or ST3944. The remaining 6/53 (11%) samples contained pAmpC positive isolates; these isolates carried the blaCMY-2 gene, which encodes CMY-2 β-lactamase. These six isolates belonged to phylogroups A (n = 2), B1 (n = 1) and D (n = 3), and sequences types ST1431, ST57, ST93 and ST4565. One CMY-2 β-lactamase positive E. coli isolate of lineage ST93 had the -32 mutation in the chromosomal ampC promoter/attenuator region. Five ESBL/pAmpC positive E. coli isolates carried class 1 integrons (dfrA17-aadA5, aadA and aadA/aadB arrays were detected in three isolates) and one isolate carried a class 2 integron (dfrA12-sat2-aadA1). The aac(6')Ib-cr, aac(3)-II, qnrB19, tet(A), tet(B), cmlA, and sul3 genes were also detected. All studied isolates showed unrelated PFGE-patterns. To our knowledge, this is the first description of ESBL-producing E. coli and the second of pAmpC-producing E. coli from healthy dogs in America. Our results suggest the potential zoonotic role of dogs in the transmission to humans of ESBL and pAmpC E. coli in the household environment.
Rzewuska, Magdalena; Czopowicz, Michał; Kizerwetter-Świda, Magdalena; Chrobak, Dorota; Błaszczak, Borys; Binek, Marian
The antimicrobial susceptibility of Escherichia coli isolates associated with various types of infections in dogs and cats was determined. The studied isolates were most frequently susceptible to fluoroquinolones and the extended-spectrum cephalosporins (ESCs), antimicrobials commonly used in treatment of infections in companion animals. However, an increase in the percentage of strains resistant to β-lactam antibiotics including ESCs was noted between January 2007 and December 2013. The frequency of multidrug-resistant (MDR) E. coli isolation (66.8% of isolates) is alarming. Moreover, the statistically significant increase of the percentage of MDR isolates was observed during the study period. No difference in the prevalence of multidrug resistance was found between bacteria causing intestinal and extraintestinal infections and between canine and feline isolates. Nonhemolytic E. coli isolates were MDR more often than hemolytic ones. Our study showed the companion animals in Poland as an important reservoir of MDR bacteria. These results indicate that continuous monitoring of canine and feline E. coli antimicrobial susceptibility is required. Furthermore, introduction and application of recommendations for appropriate use of antimicrobials in small animal practice should be essential to minimize the emergence of multidrug resistance among E. coli in companion animals. PMID:25667937
Rebello, Raquel Costa de Luca; Regua-Mangia, Adriana Hamond
Escherichia coli contamination in aquatic ecosystems has emerged as a relevant concern of public health impact, especially in developing areas. In this study, E. coli isolates were recovered from residential, industrial, agricultural, hospital wastewaters and recreational waters and, further characterized according to diarrheagenic potential, phylotyping and antimicrobial resistance phenotype. Among the total 178 E. coli isolates, antimicrobial resistance was detected in 37% to at least one of the 11 antimicrobials tested. The highest percentage of resistant E. coli was recovered from agricultural wastewaters (57.7%) followed by recreational waters (56.4%), hospital (34.5%), residential (22.7%) and industrial wastewaters (22.2%). Twenty-three resistance profiles (I-XXIII) were detected and 17 isolates exhibited the MDR phenotype. 11.2% of the total E. coli isolates carried diarrheagenic markers: astA (7.3%, 13/178), stx1 (2.8%, 05/178), escV (2.2%, 04/178) and estIa (0.6%, 01/178). All isolates harbored the uidA gene. E. coli isolates were mostly found in phylogenetic groups A (91.6%, 163/178) followed by groups D (5%, 09/178) and B2 (3.4%, 06/178). Specific gene combinations characterized E. coli pathotypes as ETEC (01/20), ATEC (04/20) and STEC (05/20) which belonged to A (75%, 15/20), D (15%, 03/20) and B2 (10%, 02/20) phylogroups. Our results revealed the widespread distribution of E. coli in aquatic systems in Rio de Janeiro. The circulation of pathogenic E. coli and antimicrobial resistance within bacterial population represents high risk to ecosystem and human health and highlights epidemiological surveillance and sanitary improvement.
Ibekwe, A M; Murinda, Shelton E; DebRoy, Chitrita; Reddy, Gudigopura B
Escherichia coli populations originating from swine houses through constructed wetlands were analyzed for potential pathogens, antimicrobial susceptibility patterns, and genotypic diversity. Escherichia coli isolates (n = 493) were screened for the presence of the following virulence genes: stx1, stx2 and eae (Shiga toxin-producing E. coli [STEC]), heat-labile enterotoxin (LT) genes and heat stable toxin STa and STb (enterotoxigenic E. coli (ETEC), cytotoxin necrotizing factors 1 and 2 (cnf1 and cnf2 [necrotoxigenic E. coli- NTEC]), as well as O and H antigens, and the presence of the antibiotic resistance genes blaTEM, blaSHV, blaCMY-2, tet A, tet B, tet C, mph(A), aadA, StrA/B, sul1, sul2 and sul3. The commensal strains were further screened for 16 antimicrobials and characterized by BOX AIR-1 PCR for unique genotypes. The highest antibiotic resistance prevalence was for tetracycline, followed by erythromycin, ampicillin, streptomycin, sulfisoxazole and kanamycin. Our data showed that most of the isolates had high distribution of single or multidrug-resistant (MDR) genotypes. Therefore, the occurrence of MDR E. coli in the wetland is a matter of great concern due to possible transfer of resistance genes from nonpathogenic to pathogenic strains or vice versa in the environment.
Cook, Angela; Reid-Smith, Richard J; Irwin, Rebecca J; McEwen, Scott A; Young, Virginia; Ribble, Carl
This study estimated the prevalence of Salmonella, Campylobacter, and Escherichia coli isolates in fresh retail grain-fed veal obtained in Ontario, Canada. The prevalence and antimicrobial resistance patterns were examined for points of public health significance. Veal samples (n = 528) were collected from February 2003 through May 2004. Twenty-one Salmonella isolates were recovered from 18 (4%) of 438 samples and underwent antimicrobial susceptibility testing. Resistance to one or more antimicrobials was found in 6 (29%) of 21 Salmonella isolates; 5 (24%) of 21 isolates were resistant to five or more antimicrobials. No resistance to antimicrobials of very high human health importance was observed. Ampicillin-chloramphenicolstreptomycin-sulfamethoxazole-tetracycline resistance was found in 5 (3%) of 21 Salmonella isolates. Campylobacter isolates were recovered from 5 (1%) of 438 samples; 6 isolates underwent antimicrobial susceptibility testing. Resistance to one or more antimicrobials was documented in 3 (50%) of 6 Campylobacter isolates. No Campylobacter isolates were resistant to five or more antimicrobials or category I antimicrobials. E. coli isolates were recovered from 387 (88%) of 438 samples; 1,258 isolates underwent antimicrobial susceptibility testing. Resistance to one or more antimicrobials was found in 678 (54%) of 1,258 E. coli isolates; 128 (10%) of 1,258 were resistant to five or more antimicrobials. Five (0.4%) and 7 (0.6%) of 1,258 E. coli isolates were resistant to ceftiofur and ceftriaxone, respectively, while 34 (3%) of 1,258 were resistant to nalidixic acid. Ciprofloxacin resistance was not detected. There were 101 different resistance patterns observed among E. coli isolates; resistance to tetracycline alone (12.7%, 161 of 1,258) was most frequently observed. This study provides baseline prevalence and antimicrobial resistance data and highlights potential public health concerns.
Escherichia coli O157:H7 has been linked to waterborne outbreaks in the United States and abroad. Methods employed to detect this pathogen typically are cultural based and take advantage of phenotypic traits that are specific for this serotype, including slow sorbitol fermentati...
Escherichia coli O157 H:7 has been linked to waterborne outbreaks in the United States and abroad. Methods employed to detect this pathogen typically are cultural based and take advantage of phenotypic traits that are specific for this serotype. These phenotypic characteristics...
Beloin, Christophe; Roux, Agnès; Ghigo, Jean-Marc
Escherichia coli is a predominant species among facultative anaerobic bacteria of the gastrointestinal tract. Both its frequent community lifestyle and the availability of a wide array of genetic tools contributed to establish E. coli as a relevant model organism for the study of surface colonization. Several key factors, including different extracellular appendages, are implicated in E. coli surface colonization and their expression and activity are finely regulated, both in space and time, to ensure productive events leading to mature biofilm formation. This chapter will present known molecular mechanisms underlying biofilm development in both commensal and pathogenic E. coli. PMID:18453280
Ghanbarpour, Reza; Askari, Nasrin; Ghorbanpour, Masoud; Tahamtan, Yahya; Mashayekhi, Khoobyar; Afsharipour, Narjes; Darijani, Nasim
The aim of the present study was to determine the analysis of virulence genes and antimicrobial profile of diarrheagenic Escherichia coli isolated from diseased lambs. Two hundred ninety E. coli isolates were recovered from 300 rectal swabs of diarrheic lambs and were confirmed by biochemical tests. The pathotype determination was done according to the presence of genes including f5, f41, LTI, STI, bfp, ipaH, stx 1 , stx 2 , eae, ehlyA, cnf 1 , cnf 2 , cdIII, cdIV, and f17 by PCR method. Sixty-six isolates (23.72%) possessed the STI gene and categorized into entrotoxigenic E. coli (ETEC). Nine isolates (3.1%) and five isolates (1.72%) were positive for the cnf1 and cnf2 genes which categorized into necrotoxic E. coli (NTEC). Hundred and seventeen isolates (40.34%) harbored stx 1 and/or stx 2 and classified as Shiga toxin-producing E. coli (STEC). Thirteen isolates (4.48%) were assigned to atypical entropathogenic E. coli (aEPEC) and possessed eae gene. Two isolates (0.68%) were positive for ipaH gene and were assigned to entroinvasive E. coli (EIEC). Statistical analysis showed a specific association between eae gene and STEC pathotype (P < 0.0001). The most prevalent resistance was observed against lincomycin (96.5%) and the lowest resistance was against kanamycine (56.02%), respectively. The high prevalence of STEC and ETEC indicates that diarrheic lambs represent an important reservoir for humans. ETEC may play an important role for frequent occurrence of diarrhea in lambs observed in this region. Due to high antibiotic resistance, appropriate control should be implemented in veterinary medicine to curb the development of novel resistant isolates.
Ahmed, Ashraf M; Shimabukuro, Hirofumi; Shimamoto, Tadashi
Sixty-nine Escherichia coli and 10 Salmonella isolates, recovered from retail chicken meat in Hiroshima prefecture, Japan, were assayed for antimicrobial susceptibility, the presence of integrons and antimicrobial resistance genes. Twenty-eight out of 69 (40.6%) of E. coli and all 10 Salmonella isolates were exhibited multidrug resistance phenotypes. The most commonly reported resistance phenotypes were against ampicillin, streptomycin, spectinomycin, kanamycin, tetracycline, and trimethoprim/sulfamethoxazole. PCR screening for integrons showed that 8 (11.6%) of the E. coli isolates were positive for the class 1 integrons and 1 isolate (1.4%) was positive for the class 2 integrons. Among the 10 Salmonella isolates, 9 were positive for class 1 integrons and none was positive for class 2 integrons. The identified antibiotic resistance gene cassettes within the class 1 integrons were dfrA1, dfrA7, aadA1, aadB, and catB3, while dfrA1, sat2, and aadA1 were identified within class 2 integron. The beta-lactamase resistance gene bla(TEM-1) was identified in 12 (17.3%) of E. coli isolates and in only one of the Salmonella isolates. The bla(CMY-2) gene, encoding AmpC beta-lactamase, was detected in 16 (23.2%) of the E. coli isolates only. Conjugation experiments demonstrated that there was plasmid-mediated transfer of bla(CMY-2) and bla(TEM-1). These results highlighted the role of retail chicken meat as a potential source for multidrug-resistant strains of E. coli and Salmonella. To the best of our knowledge, this is the 1st report of isolation and molecular characterization of multidrug-resistant strains of E. coli from retail chicken meat in Japan.
Micenková, Lenka; Beňová, Alžbeta; Frankovičová, Lucia; Bosák, Juraj; Vrba, Martin; Ševčíková, Alena; Kmeťová, Marta; Šmajs, David
Escherichia coli is the most common cause of bloodstream infections and community-acquired sepsis. The main aim of this study was to determine virulence characteristics of E. coli isolates from hemocultures of patients with a primary disease of urogenital tract, digestive system, a neoplastic blood disease, or other conditions. Results from a set of 314 E. coli isolates from hemocultures were compared to data from a previously published analysis of 1283 fecal commensal E. coli isolates. Genetic profiling of the 314 E. coli isolates involved determination of phylogenetic group (A, B1, B2, D, C, E, and F), identification of 21 virulence factors, as well as 30 bacteriocin-encoding determinants. Pulsed-field gel electrophoresis was used to analyze clonal character of the hemoculture-derived isolates. The E. coli isolates from hemocultures belonged mainly to phylogenetic groups B2 (59.9%) and D (21.0%), and less frequently to phylogroups A (10.2%) and B1 (5.7%). Commonly detected virulence factors included adhesins (fimA 92.0%, pap 47.1%, and sfa 26.8%), and iron-uptake encoding genes (fyuA 87.9%, fepC 79.6%, aer 70.7%, iucC 68.2%, and ireA 13.7%), followed by colibactin (pks island 31.5%), and cytotoxic necrotizing factor (cnf1 11.1%). A higher frequency of microcin producers (and microcin M determinant) and a lower frequency of colicin Ib and microcin B17 was found in hemoculture-derived isolates compared to commensal fecal isolates. E. coli isolates from hemocultures harbored more virulence genes compared to fecal E. coli isolates. In addition, hemoculture E. coli isolates from patients with primary diagnosis related to urogenital tract were clearly different and more virulence genes were detected in these isolates compared to both fecal isolates and hemoculture-derived isolates from patients with blood and gastrointestinal diseases.
Krizman, Manja; Avgustin, Jerneja Ambrozic; Zdovc, Irena; Golob, Majda; Trkov, Marija; Ciglenecki, Urska Jamnikar; Biasizzo, Majda; Kirbis, Andrej
Antibiotics have always appeared miraculous, saving innumerable lives. However, the unwise use of antimicrobial drugs has led to the appearance of resistant bacteria. The purpose of this study was to evaluate antimicrobial resistance in Escherichia coli (n =160) isolated from food of animal origin. The focus was on E. coli -producing extended-spectrum β-lactamases. E. coli was chosen because it is a part of the normal microbiota in mammals and can enter the food chain during slaughtering and food manipulation. Subsequently, its resistance genes can be transferred to pathogenic bacteria and human microbiota. Phenotypic and genotypic analyses of selected antimicrobial resistances were carried out together with a molecular analysis of virulence genes. E. coli isolates from food of animal origin were compared with clinical E. coli strains isolated from the human intestinal tract. Extended-spectrum β-lactamase-producing E. coli isolates were found in 9.4% of food isolates and in 1.8% of intestinal isolates. Phylogenetically, the majority of food (86.3%) and intestinal E. coli (58.1%) isolates were found to belong to the commensal phylogenetic groups A and B1. The distribution of 4 of 14 analyzed virulence factors was similar in the food and intestinal isolates. Strains isolated from food in Slovenia harbored resistance genes and virulence factors, which can constitute a problem for food safety if not handled properly.
Bogyiová, E; Kmetová, M; Biros, E; Siegfried, L
P-fimbriae, S-fimbriae and AFA-adhesins are virulence factors responsible for adherence of Escherichia coli strains to extraintestinal host-cell surface. Detection of pap-, sfa- and afa-specific sequences performed by PCR revealed 74% pap+, 65% sfa+, and 8.3% afa+ strains in a group of 84 extraintestinal E. coli isolates. Detection in a group of fecal strains showed 29% pap+, 21% sfa+ and 4% afa+ strains. pap together with sfa were found as the most frequent combination (56%) among extraintestinal isolates probably due to localization of pap- and sfa-operons on a common pathogenicity island. The occurrence of afa-specific sequence among 56 urine strains was 11%, although no afa+ strain was detected among 28 gynecological isolates. No strains with detected adhesin operons were found among twenty (24%) extraintestinal E. coli strains.
Jurado, Sonia; Medina, Alberto; de la Fuente, Ricardo; Ruiz-Santa-Quiteria, José A; Orden, José A
The aim of the present study was evaluate how oral administration of enrofloxacin affected the frequency of resistance to different antimicrobials in commensal Escherichia coli isolates from healthy chickens. A further objective of this study was to characterize the mechanisms of resistance in these isolates. A trend towards increased resistance to enrofloxacin, doxycycline and amoxicillin of E. coli isolates from chickens after enrofloxacin administration was observed. The increase in the resistance to doxycycline and amoxicillin was probably due to a co-selection of tetracycline and β-lactam resistance genes by the administration of enrofloxacin. The detection of tetM was much higher than expected (50%), which indicates that this gene may play an important role in tetracycline resistance in E. coli from chickens.
Maal-Bared, Rasha; Bartlett, Karen H; Bowie, William R; Hall, Eric R
This study examined the distribution of antibiotic resistant Escherichia coli and E. coli O157 isolated from water, sediment and biofilms in an intensive agricultural watershed (Elk Creek, British Columbia) between 2005 and 2007. It also examined physical and chemical water parameters associated with antibiotic resistance. Broth microdilution techniques were used to determine minimum inhibitory concentrations (MIC) for E. coli (n=214) and E. coli O157 (n=27) recovered isolates for ampicillin, cefotaxime, ciprofloxacin, nalidixic acid, streptomycin and tetracycline. Both E. coli and E. coli O157 isolates showed highest frequency of resistance to tetracycline, ampicillin, streptomycin and nalidixic acid; respectively. For E. coli, the highest frequency of resistance was observed at the most agriculturally-impacted site, while the lowest frequency of resistance was found at the headwaters. Sediment and river rock biofilms were the most likely to be associated with resistant E. coli, while water was the least likely. While seasonality (wet versus dry) had no relationship with resistance frequency, length of biofilm colonization of the substratum in the aquatic environment only affected resistance frequency to nalidixic acid and tetracycline. Multivariate logistic regressions showed that water depth, nutrient concentrations, temperature, dissolved oxygen and salinity had statistically significant associations with frequency of E. coli resistance to nalidixic acid, streptomycin, ampicillin and tetracycline. The results indicate that antibiotic resistant E. coli and E. coli O157 were prevalent in an agricultural stream. Since E. coli is adept at horizontal gene transfer and prevalent in biofilms and sediment, where ample opportunities for genetic exchange with potential environmental pathogens present themselves, resistant isolates may present a risk to ecosystem, wildlife and public health.
Süzük, Serap; Kaşkatepe, Banu; Avcıküçük, Havva; Aksaray, Sebahat; Başustaoğlu, Ahmet
Determination of treatment protocols for infections according to antimicrobial susceptibility test (AST) results is are important for controlling the problem of antibiotic resistance. Two standards are widely used in the world. One of them is Clinical Laboratory Standards Institute (CLSI) standards used in Turkey for many years and the other is the European Committee on Antimicrobial Susceptibility Testing (EUCAST) standards which is used in European Union member countries and came into use in 2015 in Turkey. Since the EUCAST standards had higher clinical sensitivity limits particularly for gram-negative bacilli compared to CLSI (2009) standards, there will be some changes in antibiotic resistance profiles of Turkey with the use of EUCAST. CLSI has changed zone diameters after 2009 versions and the differences between the two standards were brought to a minimum level. Knowledge of local epidemiological data is important to determine empirical therapy which will be used in urinary tract infections (UTI). The aim of this study was to determine the differences of antibiotic susceptibility zone diameters based on our local epidemiological data among uropathogenic Escherichia coli isolates according to EUCAST 2014 and CLSI 2014 standards. A total of 298 E.coli strains isolated from urine samples as the cause of uncomplicated acute UTI agents, were included in the study. Isolates were identified by conventional methods and with BBL Crystal E/NF ID System (Becton Dickinson, USA). AST was performed with Kirby Bauer disk diffusion method and results were evaluated and interpreted according to the CLSI 2014 and EUCAST 2014 standards. According to the results, susceptibility rates of isolates against amikacin (100%) and trimethoprim-sulfamethoxazole (63.09%) were identical in both standards. However, statistically significant differences were observed between CLSI and EUCAST standards in terms of susceptibilities against gentamicin (91.95% and 84.56%, respectively; p= 0
Zeighami, Habib; Haghi, Fakhri; Hajiahmadi, Fahimeh
Integrons are considered to play a significant role in the evolution and spread of antibiotic resistance genes. A total of 349 clinical isolates of Escherichia coli and Klebsiella pneumoniae were investigated for molecular characterization of integrons and betalactamases. Antimicrobial susceptibility testing was also performed as the Clinical and Laboratory Standards Institute (CLSI) guidelines. The frequency of extended spectrum betalactamases (ESBL) or metallo-betalactamases (MBL)-producing isolates, patient demographics, and the susceptibility to various antimicrobial agents were described. BlaCTX-M was the most frequently detected betalactamase in all isolates. Moreover, MBL producing K. pneumoniae carried blaIMP and blaVIM at 100 and 41·6%, respectively but no MBL-positive E. coli was detected. Class 1 integrons were more frequent among E. coli and K. pneumoniae isolates in comparison with class 2 integrons and the frequency of intI2 in K. pneumoniae was significantly higher than E. coli isolates. Five different resistance gene arrays were identified among class 1 integrons. Dihydrofolate reductase (dfrA) and aminoglycoside adenyltransferase (aad) gene cassettes were found to be predominant in the class 1 integrons. These results indicate that class 1 integrons are widespread among ESBL-producing isolates of K. pneumoniae and E. coli and appropriate surveillance and control measures are essential to prevent further dissemination of these elements among Enterobacteriaceae in our country.
Mosquito, Susan; Ruiz, Joaquim; Pons, María J; Durand, David; Barletta, Francesca; Ochoa, Theresa J
Diarrhoeagenic Escherichia coli (DEC) are an important cause of diarrhoea in children and are associated with high antibiotic resistance. However, there are few studies on the molecular mechanisms of resistance in this group of bacteria. The aim of this study was to determine the mechanisms associated with antibiotic resistance in the most common phenotypes of DEC. A total of 369 E. coli strains [commensal strains and DEC from children with ('DEC-diarrhoea') or without ('DEC-control') diarrhoea] isolated from children aged <1 year in periurban districts of Lima, Peru, were analysed. In total, 154 ampicillin-resistant strains (36 commensals, 33 DEC-control and 85 DEC-diarrhoea) were studied by PCR for the most prevalent resistance mechanisms to ampicillin, trimethoprim/sulfamethoxazole (SXT), tetracycline and chloramphenicol as well as for integrase types 1 and 2. In addition, restriction fragment length polymorphism was performed for SXT-resistant strains. Commensal strains were more frequently resistant to nalidixic acid and ciprofloxacin (68% and 28%, respectively) than DEC strains (23% and 2%, respectively) (P<0.05). DEC-diarrhoea strains were more frequently SXT-resistant (78%) compared with DEC-control strains (65%) and commensal strains (60%) (P<0.05). The most frequent mechanisms of antibiotic resistance in DEC strains were: for β-lactams, bla(TEM) (31%; 37/118); for SXT, sul2 (48%; 49/103); for tetracycline, tetA (27%; 23/84); and for chloramphenicol, cat (80%; 28/35). The genes sul1 and dfrA1, related to SXT resistance, were more frequent in the DEC-diarrhoea group (41% and 28%, respectively) than in the other two groups (P<0.05). There was a high diversity of resistance genes in DEC, including symptomatic strains.
Ghorbani-Dalini, Sadegh; Kargar, Mohammad; Doosti, Abbas; Abbasi, Pejman; Sarshar, Meysam
Resistance to oxyimino cephalosporins antibiotics in Enterobacteriaceae is primarily done by the extended spectrum β-lactamases (ESBLs). Clear identification of risk factors for ESBLs-producing infections is necessary. Therefore, efficient strategies can be developed to decrease outbreak of these infections. The aim of this study was to determine the antibacterial susceptibility and ESBLs pattern of diarrhogenic Escherichia coli (E. coli) strains isolated from adult patients. In the present study, diarrheogenic E. coli strains were isolated from 54 patients from the University of Medical Sciences hospitals in Shiraz. Antimicrobial susceptibility testing was done by disk diffusion method by CLSI criteria. The presence of blaTEM, blaSHV and blaCTX-M genes was investigated by PCR using designated primers. The prevalence of ESBLs-producer E. coli strains was 12.96%. Antimicrobial resistance testing showed a high resistance to cefexime, trimethoprim-sulfamethoxazole, ampicillin and penicillin. Overall, β-lactamase genes were identified in 52 (96.30%) isolates which were identified as 45 (83.33%) blaTEM, 17 (31.48%) blaSHV and 11 (20.37%) blaCTX-M. ESBLs-producer E. coli is very prevalent in Diarrheogenic strains isolated from adult patients. Also, this study clearly showed that the blaTEM gene for ESBLs-producer E. coli was widespread in Iran. PMID:26664394
Burlingame, R P; Wyman, L; Chapman, P J
Mutants of Escherichia coli defective in catabolism of 3-phenylpropionate, 3-(3-hydroxyphenyl)propionate, or both were isolated after mutagenesis with ethylmethane sulfonate. Nine phenotypically distinct classes of mutants were identified, including strains lacking each of the first five enzyme activities for the degradation of these compounds and mutants pleiotropically negative for some of these activities. Characterization of these mutants was greatly facilitated by the use of indicator media in which accumulation of 3-(2,3-dihydroxyphenyl)propionate or 2-hydroxy-6-ketononadienedioic acid led to the formation of dark red or bright yellow colors, respectively, in the medium. Assays with wild-type and mutant strains indicated that 3-phenylpropionate (or its dihydrodiol), but none of the hydroxylated derivatives tested, induced the synthesis of enzymes for its conversion to 3-(2,3-dihydroxyphenyl)propionate. The remaining enzymes were induced by the 2- or 3-hydroxy or 2,3-dihydroxy derivatives of 3-phenylpropionate, with the 2-hydroxy compound acting as an apparent gratuitous inducer. Metabolism to nonaromatic intermediates appeared to be unnecessary for full induction of any pathway enzyme. One unusual class of mutants, in which 2-keto-4-pentenoate hydratase appeared to be uninducible, indicated a level of control not previously shown in meta-fission catabolic pathways. PMID:3531186
Cunha, Marcos Paulo Vieira; de Oliveira, Maria Gabriela Xavier; de Oliveira, Mirela Caroline Vilela; da Silva, Ketrin Cristina; Gomes, Cleise Ribeiro; Moreno, Andrea Micke; Knöbl, Terezinha
Avian Pathogenic Escherichia coli (APEC) has been studied for decades because of its economic impact on the poultry industry. Recently, the zoonotic potential of APEC and multidrug-resistant strains have emerged. The aim of this study was to characterize 225 APEC isolated from turkeys presenting airsacculitis. The results showed that 92% of strains presented a multidrug-resistance (MDR), and the highest levels of resistance were to sulfamethazine (94%) and tetracycline (83%). Half of these strains were classified in phylogenetic group B2, followed by B1 (28.6%), A (17.1%), and D (4.8%). The prevalence of virulence genes was as follows: salmochelin (iroN, 95%), increased serum survival (iss, 93%), colicin V (cvi/cva, 67%), aerobactin (iucD, 67%), temperature-sensitive haemagglutinin (tsh, 56%), iron-repressible protein (irp2, 51%), invasion brain endothelium (ibeA, 31%), vacuolating autotransporter toxin (vat, 24%), K1 antigen (neuS, 19%), enteroaggregative heat-stable cytotoxin (astA, 17%), and pilus associated with pyelonephritis (papC, 15%). These results demonstrate that the majority of the investigated strains belonged to group B2 and were MDR. These data suggest that turkeys may serve as a reservoir of pathogenic and multidrug-resistance strains, reinforcing the idea that poultry plays a role in the epidemiological chain of ExPEC. PMID:25105155
Sassi, Asma; Loucif, Lotfi; Gupta, Sushim Kumar; Dekhil, Mazouz; Chettibi, Houria
Here, we report the first autochthonous cases of infections caused by blaNDM-5 New Delhi metallo-β-lactamase-producing Escherichia coli strains recovered from urine and blood specimens of three patients from Algeria between January 2012 and February 2013. The three isolates belong to sequence type 2659 and they coexpress blaCTX-M-15 with the blaTEM-1 and blaaadA2 genes. PMID:24982080
The aadA genes, encoding resistance to streptomycin and spectinomycin, have been found as gene cassettes in different gram-negative and gram-positive bacterial species. The present study has revealed the sequence of a new gene, aadA5, integrated as a gene cassette together with the trimethoprim resistance gene dfr7 in a class 1 integron. The integron was located on a plasmid and was identified in a pathogenic porcine Escherichia coli isolate. PMID:10582907
Yang, Tong; Zeng, Zhenling; Rao, Lili; Chen, Xiaojie; He, Dandan; Lv, Luchao; Wang, Jing; Zeng, Li; Feng, Minsha; Liu, Jian-Hua
This study was performed to determine the prevalence of plasmid-mediated quinolone resistance (PMQR) determinants and characterize the ciprofloxacin resistance in Escherichia coli isolated from different sources in China. PMQR determinants were detected by PCR amplification and sequencing in 2297 E. coli isolates randomly collected from animals, food and humans during 2004 to 2011. MICs of ciprofloxacin were determined by agar dilution method. Of the 2297 E. coli isolates, 43.6% harbored at least one PMQR gene. The most common PMQR gene was oqxAB (29.3%), followed by qnr (13.6%), aac(6')-Ib-cr (11.6%), and qepA (3.3%). 12.0% isolates carried two or more PMQR genes. The prevalence of PMQR genes in food animal isolates increased over time, from 38.7% in 2004 to 69.8% in 2011. The prevalence of PMQR/ciprofloxacin resistance among isolates from pig, chicken, duck, companion animals, animal food and human volunteers were 65.2%/69.6%, 42.4%/60.0%, 59.4%/65.0%, 28.6%/57.5%, 29.3%/25.6%, and 14.0/8.7%, respectively. Most isolates carrying qnr along showed susceptible to ciprofloxacin, and only 21.6% the isolates exhibited resistance to ciprofloxacin, which was significantly lower than those carrying other PMQR genes (65.2-89.9%) and those that do not (43.1%) (p<0.01). In conclusion, high frequency of ciprofloxacin resistance and PMQR genes was observed among E. coli isolates of different origins in China, with oqxAB being the most frequent. qnr-positive E. coli isolates have relatively low ciprofloxacin resistance rate compared with other PMQR determinants-carrying isolates and PMQR-negative isolates.
Wang, Lian-Hui; Liu, Pan-Pan; Wei, Dan-Dan; Liu, Yang; Wan, La-Gen; Xiang, Tian-Xin; Zhang, Yu-Juan
Here we report five cases of NDM-7-producing Escherichia coli from patients with bacteriuria in a teaching hospital in mainland China. Two isolates belonged to sequence type 131 (ST131), simultaneously carrying blaCTX-M-15, blaSHV-11, blaTEM-1 and qnrS1. The blaNDM-7 gene was located on a conjugative IncX3-type plasmid bearing blaTEM-1 and qnrS1. These findings indicate the spread of NDM-7 metallo-β-lactamase in a highly resistant and virulent E. coli sequence type in China.
Dobiasova, Hana; Dolejska, Monika; Jamborova, Ivana; Brhelova, Eva; Blazkova, Lucie; Papousek, Ivo; Kozlova, Marketa; Klimes, Jiri; Cizek, Alois; Literak, Ivan
Commensal Escherichia coli isolates from healthy zoo animals kept in Ostrava Zoological Garden, Czech Republic, were investigated to evaluate the dissemination of extended-spectrum beta-lactamase (ESBL) and plasmid-mediated quinolone resistance (PMQR) genes. A total of 160 faecal samples of various animal species were inoculated onto MacConkey agar with cefotaxime (2 mg L(-1)) or ciprofloxacin (0.05 mg L(-1)) to obtain ESBL- or PMQR-positive E. coli isolates. Clonality of E. coli isolates was investigated by multilocus sequence typing and pulsed-field gel electrophoresis. Plasmids carrying ESBL or PMQR genes were typed by PCR-based replicon typing, plasmid multilocus sequence typing and restriction fragment length polymorphism. Forty-nine (71%, n = 69) cefotaxime-resistant and 15 (16%, n = 94) ciprofloxacin-resistant E. coli isolates harboured ESBL or PMQR genes. Isolates were assigned to 18 sequence types (ST) and 20 clusters according to their macrorestriction patterns by pulsed-field gel electrophoresis. The genes blaCTX -M-1 and qnrS1 were detected on highly related IncI1 plasmids assigned to clonal complex 3 (ST3, ST38) and on non-related IncN plasmids of ST1 and ST3, respectively. The gene qnrS1 was located on related IncX1 plasmids. Dissemination of antibiotic resistance is associated with spreading of particular E. coli clones and plasmids of specific incompatibility groups among various animal species.
Jure, María A; Condorí, Marina S; Pérez Terrazzino, Gabriela; Catalán, Mariana G; López Campo, Alejandro; Zolezzi, Gisella; Chinen, Isabel; Rivas, Marta; Castillo, Marta
Escherichia coli O157 is an emergent pathogen associated with diarrhea, hemorrhagic colitis and hemolytic uremic syndrome. Meat products constitute an important transmission source of this microorganism. The aims of this study were to characterize E. coli O157 isolated from cattle and meat products collected from abattoirs and retail stores, to establish the clonal relatedness among regional isolates and to compare them with those in the national database. Between 2004 and 2013, 169 minced meat, 35 sausage and 216 carcass samples were analyzed. Thirteen E. coli O157 isolates were identified; 6 of which were O157:H7 and characterized as stx2c(vh-a)/eae/ehxA (n = 5) and stx2/eae/ehxA (n = 1). The 7 remaining isolates were non-toxigenic E. coli strains, and serotyped as O157:NT (n = 4), O157:NM (n = 1), O157:ND (n = 1) and O157:H16 (n = 1). The strains yielded different XbaI-PFGE patterns. Compared to the E. coli O157 isolates in the National Database, none of these patterns have been previously detected in strains of different origin in Argentina.
Luciani, Mirella; Di Febo, Tiziana; Zilli, Katiuscia; Di Giannatale, Elisabetta; Armillotta, Gisella; Manna, Laura; Minelli, Fabio; Tittarelli, Manuela; Caprioli, Alfredo
Monoclonal antibodies (MAbs) specific for the lipopolysaccharide (LPS) of Escherichia coli O104:H4 were produced by fusion of Sp2/O-Ag-14 mouse myeloma cells with spleen cells of Balb/c mice, immunized with heat-inactivated and sonicated E. coli O104:H4 bacterial cells. Four MAbs specific for the E. coli O104:H4 LPS (1E6G6, 1F4C9, 3G6G7, and 4G10D2) were characterized and evaluated for the use in a method for the detection of E. coli O104:H4 in milk samples that involves antibody conjugation to magnetic microbeads to reduce time and increase the efficiency of isolation. MAb 1E6G6 was selected and coupled to microbeads, then used for immuno-magnetic separation (IMS); the efficiency of the IMS method for E. coli O104:H4 isolation from milk was evaluated and compared to that of the EU RL VTEC conventional culture-based isolation procedure. Milk suspensions also containing other pathogenic bacteria that could potentially be found in milk (Campylobacter jejuni, Listeria monocytogenes, and Staphylococcus aureus) were also tested to evaluate the specificity of MAb-coated beads. Beads coated with MAb 1E6G6 showed a good ability to capture the E. coli O104:H4, even in milk samples contaminated with other bacteria, with a higher number of E. coli O104:H4 CFU reisolated in comparison with the official method (121 and 41 CFU, respectively, at 10(3) E. coli O104:H4 initial load; 19 and 6 CFU, respectively, at 10(2) E. coli O104:H4 initial load; 1 and 0 CFU, respectively, at 10(1) E. coli O104:H4 initial load). The specificity was 100%.
Silva, Nuno; Igrejas, Gilberto; Figueiredo, Nicholas; Gonçalves, Alexandre; Radhouani, Hajer; Rodrigues, Jorge; Poeta, Patrícia
A total of 44 Escherichia coli and 64 enterococci recovered from 77 intestinal samples of wild European rabbits in Portugal were analyzed for resistance to antimicrobial agents. Resistance in E. coli isolates was observed for ampicillin, tetracycline, sulfamethoxazole/trimethoprim, streptomycin, gentamicin, tobramycin, nalidixic acid, ciprofloxacin and chloramphenicol. None of the E. coli isolates produced extended-spectrum beta-lactamases (ESBLs). The bla(TEM), aadA, aac(3)-II, tet(A) and/or tet(B), and the catA genes were demonstrated in all ampicillin, streptomycin, gentamicin, tetracycline, and chloramphenicol-resistant isolates respectively, and the sul1 and/or sul2 and/or sul3 genes in 4 of 5 sulfamethoxazole/trimethoprim resistant isolates. Of the enterococcal isolates, Enterococcus faecalis was the most prevalent detected species (39 isolates), followed by E. faecium (21 isolates) and E. hirae (4 isolates). More than one-fourth (29.7%) of the isolates were resistant to tetracycline; 20.3% were resistant to erythromycin, 14.1% were resistant to ciprofloxacin and 10.9% were resistant to high-level-kanamycin. Lower level of resistance (<10%) was detected for ampicillin, quinupristin/dalfopristin and high-level-gentamicin, -streptomycin. No vancomycin-resistance was detected in the enterococci isolates. Resistance genes detected included aac(6')-aph(2''), ant(6)-Ia, tet(M) and/or tet(L) in all gentamicin, streptomycin and tetracycline-resistant isolates respectively. The aph(3')-IIIa gene was detected in 6 of 7 kanamycin-resistant isolates, the erm(B) gene in 11 of 13 erythromycin-resistant isolates and the vat(D) gene in the quinupristin/dalfopristin-resistant E. faecium isolate. This survey showed that faecal bacteria such as E. coli and enterococci of wild rabbits could be a reservoir of antimicrobial resistance genes.
da Silva, Ketrin Cristina; Cunha, Marcos Paulo Vieira; Cerdeira, Louise; de Oliveira, Maria Gabriela Xavier; de Oliveira, Mirela Caroline Vilela; Gomes, Cleise Ribeiro; Lincopan, Nilton; Knöbl, Terezinha; Moreno, Andrea Micke
This study reports the high-virulence phylogenetic backgrounds of CMY-2- and CTX-M-2-producing avian pathogenic Escherichia coli strains isolated from turkeys sent to slaughter and condemned by airsacculitis in Brazil. Among 300 air sac samples, seven E. coli strains produced plasmid-mediated CMY-2-type AmpC, of which three carried also the blaCTX-M-2 Extended Spectrum Beta-Lactamase encoding gene. Interestingly, the transfer of the blaCMY-2 gene was positive for three E. coli strains, being associated with the presence of IncI1 plasmids. The complete sequence of the representative pJB10 plasmid revealed that the blaCMY-2 gene was within a transposon-like element in the classical genetic environment consisting of tnpA-blaCMY-2-blc-sugE structure. This plasmid with 94-kb belonged to the sequence type (ST) 12 among IncI1 plasmids, which has been associated with the worldwide spread of blaCMY-2 among Salmonella enterica and E. coli. Furthermore, to the best of our knowledge, this is the first complete sequence of a CMY-2-encoding plasmid derived from an Escherichia coli isolated from food-producing animals in Latin America.
Carter, John M.; Lin, Andrew; Clotilde, Laurie; Lesho, Matthew
Molecular methods have emerged as the most reliable techniques to detect and characterize pathogenic Escherichia coli. These molecular techniques include conventional single analyte and multiplex PCR, PCR followed by microarray detection, pulsed-field gel electrophoresis (PFGE), and whole genome sequencing. The choice of methods used depends upon the specific needs of the particular study. One versatile method involves detecting serogroup-specific markers by hybridization or binding to encoded microbeads in a suspension array. This molecular serotyping method has been developed and adopted for investigating E. coli outbreaks. The major advantages of this technique are the ability to simultaneously serotype E. coli and detect the presence of virulence and pathogenicity markers. Here, we describe the development of a family of multiplex molecular serotyping methods for Shiga toxin-producing E. coli, compare their performance to traditional serotyping methods, and discuss the cost-benefit balance of these methods in the context of various food safety objectives. PMID:27242670
Pianciola, L; D'Astek, B A; Mazzeo, M; Chinen, I; Masana, M; Rivas, M
Shiga toxin-producing Escherichia coli (STEC) are important food-borne pathogens associated with human diseases. In Argentina, O157:H7 is the dominant serotype in hemolytic uremic syndrome (HUS) cases. Previously, we have described the almost exclusive circulation of human E. coli O157 strains belonging to the hypervirulent clade 8 in Neuquén Province. The aim of the present study was to investigate, by a broad molecular characterization, if this particular distribution of E. coli O157 clades in Neuquén is similar to the situation in other regions of the country and if it may be originated in a similar profile in cattle, its main reservoir. Two-hundred and eighty O157 strains (54 bovine and 226 human) isolated between 2006 and 2008 in different regions of Argentina were studied. All strains harbored rfbO157, fliCH7, eae, and ehxA genes. The predominant genotype was stx2a/stx2c in human (76.1%) and bovine (55.5%) strains. All human isolates tested by Lineage-Specific Polymorphism Assay (LSPA-6), were lineage I/II; among bovine strains, 94.1% belonged to lineage I/II and 5.9% to lineage I. No LSPA-6 lineage II isolates were detected. Single nucleotide polymorphism (SNP) analysis has revealed the existence of nine clade phylogenetic groups. In our clinical strains collection, 87.6% belonged to the hypervirulent clade 8, and 12.4% were classified as clade 4/5. In bovine isolates, 59.3% strains were clade 8, 33.3% clade 4/5 and 7.4% clade 3. More than 80% of human strains showed the presence of 6 of the 7 virulence determinants described in the TW14359 O157 strain associated with the raw spinach outbreak in the U.S. in 2006. More than 80% of bovine strains showed the presence of 3 of these factors. The q933 allele, which has been related to high toxin production, was present in 98.2% of clinical strains and 75.9% of the bovine isolates. The molecular characterization of human STEC O157 strains allows us to conclude that the particular situation previously described
Carvalho, Vania M.; Gyles, Carlton L.; Ziebell, Kim; Ribeiro, Marcela A.; Catão-Dias, José L.; Sinhorini, Idércio L.; Otman, Jamile; Keller, Rogéria; Trabulsi, Luiz R.; Pestana de Castro, Antônio F.
Enteropathogenic Escherichia coli (EPEC) has been associated with infantile diarrhea and mortality in humans in developing countries. While diarrhea is also a major problem among primates kept in captivity, the role of E. coli is unclear. This study was designed to characterize diarrheagenic E. coli recovered from the feces of 56 New World nonhuman primates, primarily marmosets (Callithrix spp.). Seventeen of the 56 primates had signs of diarrhea and/or enteritis. E. coli recovered from feces from these animals was tested by PCR for genes encoding virulence factors of diarrheagenic E. coli and for patterns of adherence to HeLa cells. In addition, isolates were characterized by the fluorescence actin staining test and by their ability to induce attaching and effacing lesions. PCR for the eae gene was positive in 10 of the 39 (27%) apparently healthy animals and in 8 of the 17 (47%) animals with diarrhea and/or enteritis. Colonies of eae+ E. coli were serotyped and examined by PCR for genes encoding EPEC virulence markers. The eae+ E. coli isolates recovered from both healthy and sick nonhuman primates demonstrated virulence-associated attributes similar to those of EPEC strains implicated in human disease and are designated monkey EPEC. The results presented here indicate that EPEC may be a significant pathogen for nonhuman primates, deserving further investigation. The similarities between the affected animals investigated in this study and human EPEC infections suggest that marmosets may represent an important model for EPEC in humans. PMID:12624055
Jahanbakhsh, Seyedehameneh; Smith, Matthew G; Kohan-Ghadr, Hamid-Reza; Letellier, Ann; Abraham, Sam; Trott, Darren J; Fairbrother, John Morris
The aim of this study was to investigate the evolution with time of ceftiofur-resistant Escherichia coli clinical isolates from pigs in Québec, Canada, between 1997 and 2012 with respect to pathotypes, clones and antimicrobial resistance. Eighty-five ceftiofur-resistant E. coli isolates were obtained from the OIE (World Organisation for Animal Health) Reference Laboratory for Escherichia coli. The most prevalent pathovirotypes were enterotoxigenic E. coli (ETEC):F4 (40%), extraintestinal pathogenic E. coli (ExPEC) (16.5%) and Shiga toxin-producing E. coli (STEC):F18 (8.2%). Susceptibility testing to 15 antimicrobial agents revealed a high prevalence of resistance to 13 antimicrobials, with all isolates being multidrug-resistant. blaCMY-2 (96.5%) was the most frequently detected β-lactamase gene, followed by blaTEM (49.4%) and blaCTX-M (3.5%). Pulsed-field gel electrophoresis (PFGE) applied to 45 representative E. coli isolates revealed that resistance to ceftiofur is spread both horizontally and clonally. In addition, the emergence of extended-spectrum β-lactamase-producing E. coli isolates carrying blaCTX-M was observed in 2011 and 2012 in distinct clones. The most predominant plasmid incompatibility (Inc) groups were IncFIB, IncI1, IncA/C and IncFIC. Resistance to gentamicin, kanamycin and chloramphenicol as well as the frequency of blaTEM and IncA/C significantly decreased over the study period, whereas the frequency of IncI1 and multidrug resistance to seven antimicrobial categories significantly increased. These findings reveal that extended-spectrum cephalosporin-resistant porcine E. coli isolates in Québec belong to several different clones with diverse antimicrobial resistance patterns and plasmids. Furthermore, blaCMY-2 was the major β-lactamase gene in these isolates. From 2011, we report the emergence of blaCTX-M in distinct clones.
Johnson, James R; Porter, Stephen; Johnston, Brian; Kuskowski, Michael A; Spurbeck, Rachel R; Mobley, Harry L T; Williamson, Deborah A
Background. Extraintestinal Escherichia coli infections are common, costly, and potentially serious. A better understanding of their pathogenesis is needed. Methods. Sixty-seven E coli bloodstream isolates from adults with urosepsis (Seattle, WA; 1980s) underwent extensive molecular characterization and virulence assessment in 2 infection models (murine subcutaneous sepsis and moth larval lethality). Statistical comparisons were made among host characteristics, bacterial traits, and experimental virulence. Results. The 67 source patients were diverse for age, sex, and underlying medical and urological conditions. The corresponding E coli isolates exhibited diverse phylogenetic backgrounds and virulence profiles. Despite the E coli isolates' common bloodstream origin, they exhibited a broad range of experimental virulence in mice and moth larvae, in patterns that (for the murine model only) corresponded significantly with host characteristics and bacterial traits. The most highly mouse-lethal strains were enriched with classic "urovirulence" traits and typically were from younger women with anatomically and functionally normal urinary tracts. The 2 animal models corresponded poorly with one another. Conclusions. Host compromise, including older age and urinary tract abnormalities, allows comparatively low-virulence E coli strains to cause urosepsis. Multiple E coli traits predict both experimental and epidemiological virulence. The larval lethality model cannot be a substitute for the murine sepsis model.
Delport, Tiffany C; Harcourt, Robert G; Beaumont, Linda J; Webster, Koa N; Power, Michelle L
Greater interaction between humans and wildlife populations poses significant risks of anthropogenic impact to natural ecosystems, especially in the marine environment. Understanding the spread of microorganisms at the marine interface is therefore important if we are to mitigate adverse effects on marine wildlife. We investigated the establishment of Escherichia coli in the endangered Australian sea lion (Neophoca cinerea) by comparing fecal isolation from wild and captive sea lion populations. Fecal samples were collected from wild colonies March 2009-September 2010 and from captive individuals March 2011-May 2013. Using molecular screening, we assigned a phylotype to E. coli isolates and determined the presence of integrons, mobile genetic elements that capture gene cassettes conferring resistance to antimicrobial agents common in fecal coliforms. Group B2 was the most abundant phylotype in all E. coli isolates (n = 37), with groups A, B1, and D also identified. Integrons were not observed in E. coli (n = 21) isolated from wild sea lions, but were identified in E. coli from captive animals (n = 16), from which class I integrases were detected in eight isolates. Sequencing of gene cassette arrays identified genes conferring resistance to streptomycin-spectinomycin (aadA1) and trimethoprim (dfrA17, dfrB4). Class II integrases were not detected in the E. coli isolates. The frequent detection in captive sea lions of E. coli with resistance genes commonly identified in human clinical cases suggests that conditions experienced in captivity may contribute to establishment. Identification of antibiotic resistance in the microbiota of Australian sea lions provides crucial information for disease management. Our data will inform conservation management strategies and provide a mechanism to monitor microorganism dissemination to sensitive pinniped populations.
Thorsteinsdottir, T R; Haraldsson, G; Fridriksdottir, V; Kristinsson, K G; Gunnarsson, E
The prevalence of resistant bacteria in food products in Iceland is unknown, and little is known of the prevalence in production animals. The aim of this study was to investigate the prevalence and genetic relatedness of antimicrobial-resistant Escherichia coli from healthy pigs and broiler chicken, pork, broiler meat, slaughterhouse personnel and outpatients in Iceland. A total of 419 E. coli isolates were tested for antimicrobial susceptibility using a microbroth dilution method (VetMIC), and resistant strains were compared using pulsed-field gel electrophoresis (PFGE). All samples were screened for enrofloxacin-resistant strains with selective agar plates. The resistance rates among E. coli isolates were moderate to high from caecal and meat samples of pigs (54.1% and 28%), broilers (33.6% and 52%) and slaughterhouse personnel (39.1%), whereas isolates from outpatients showed moderate resistance rates (23.1%). Of notice was resistance to quinolones (minimum inhibitory concentrations: nalidixic acid > or = 32, ciprofloxacin > or = 0.12 and enrofloxacin > or = 0.5), particularly among broiler and broiler meat isolates (18.2% and 36%), as there is no known antimicrobial selection pressure in the broiler production in Iceland. The majority (78.6%) of the resistant E. coli isolates was genotypically different, based on PFGE fingerprint analyses and clustering was limited. However, the same resistance pattern and pulsotype were found among isolates from broiler meat and a slaughterhouse worker, indicating spread of antimicrobial-resistant E. coli from animals to humans. Diverse resistance patterns and pulsotypes suggest the presence of a large population of resistant E. coli in production animals in Iceland. This study gives baseline information on the prevalence of antimicrobial-resistant E. coli from production animals, and their food products in Iceland and the moderate to high resistance rates emphasize the need for continuing surveillance. Further studies on the
Amaya, E; Reyes, D; Paniagua, M; Calderón, S; Rashid, M-U; Colque, P; Kühn, I; Möllby, R; Weintraub, A; Nord, C E
Antibiotic-resistant bacteria have emerged due to the selective pressure of antimicrobial use in humans and animals. Water plays an important role in dissemination of these organisms among humans, animals and the environment. We studied the antibiotic resistance patterns among 493 Escherichia coli isolates from different aquatic environmental sources collected from October 2008 to May 2009 in León, Nicaragua. High levels of antibiotic resistance were found in E. coli isolates in hospital sewage water and in eight of 87 well-water samples. Among the resistant isolates from the hospital sewage, ampicillin, chloramphenicol, ciprofloxacin, nalidixic acid, trimethoprim-sulphamethoxazole was the most common multi-resistance profile. Among the resistant isolates from the wells, 19% were resistant to ampicillin, ceftazidime, ceftriaxone, cefotaxime, chloramphenicol, ciprofloxacin, gentamicin, nalidixic acid and trimethoprim-sulphamethoxazole. E. coli producing ESBL and harbouring bla(CTX-M) genes were detected in one of the hospital sewage samples and in 26% of the resistant isolates from the well-water samples. The bla(CTX-M-9) group was more prevalent in E. coli isolates from the hospital sewage samples and the bla(CTX-M-1) group was more prevalent in the well-water samples.
Gomes, Tânia A T; Elias, Waldir P; Scaletsky, Isabel C A; Guth, Beatriz E C; Rodrigues, Juliana F; Piazza, Roxane M F; Ferreira, Luís C S; Martinez, Marina B
Most Escherichia coli strains live harmlessly in the intestines and rarely cause disease in healthy individuals. Nonetheless, a number of pathogenic strains can cause diarrhea or extraintestinal diseases both in healthy and immunocompromised individuals. Diarrheal illnesses are a severe public health problem and a major cause of morbidity and mortality in infants and young children, especially in developing countries. E. coli strains that cause diarrhea have evolved by acquiring, through horizontal gene transfer, a particular set of characteristics that have successfully persisted in the host. According to the group of virulence determinants acquired, specific combinations were formed determining the currently known E. coli pathotypes, which are collectively known as diarrheagenic E. coli. In this review, we have gathered information on current definitions, serotypes, lineages, virulence mechanisms, epidemiology, and diagnosis of the major diarrheagenic E. coli pathotypes.
Li, Ming-Cheng; Wang, Fang
Abstract Ten (2.7%) Shiga toxin–producing Escherichia coli (STEC) were isolated from 370 samples of raw minced beef, mutton, pork, and chicken from the Jilin region of China; and additional 10 E. coli O157:H7 isolates were previously isolated from different Jilin regions. Seventeen of the isolates were multiresistant, exhibiting resistance to ampicillin, ciprofloxacin, tetracycline, sulfamethoxazole-trimethoprim, gentamycin, and streptomycin. Class 1 integrons were detected in nine (45.0%) of the STEC isolates and consisted of serogroups O157, O62, O113, O149, and O70. Integrons containing amplicons of a 0.5–1.5 or 1.0 kb gene cassette were found in seven (77.8%) of the integron-containing isolates. Sequencing analysis revealed that these gene cassettes encode genes conferring resistance to trimethoprim (dfrA1) and streptomycin (aadA1). The 0.5 kb cassette described here was found to encode a putative transporter peptide in the STEC. Seventeen isolates contained plasmids with different bands, and transfer by conjugation between strains of E. coli demonstrated that class 1 integrons located on mobile plasmids could contribute to the emergence and dissemination of antimicrobial resistance to ampicillin, gentamycin, streptomycin, and sulfamethoxazole-trimethoprim amongst STEC. These data revealed the high prevalence of antimicrobial-resistant STEC isolates in Jilin's surrounding regions, providing important and useful surveillance information reflecting antimicrobial selection pressure. PMID:21453117
Thani, Thani Suleiman; Symekher, Samwel Morris Lifumo; Boga, Hamadi; Oundo, Joseph
Introduction Safe water for human consumption is important, but there is a limited supply. Mombasa County has water shortages making residences rely on other sources of water including boreholes and wells. Microbiological evaluation of drinking water is important to reduce exposure to water borne enteric diseases. This cross sectional study aimed at determining the frequency and characterization of Escherichia coli (E. coli) pathotypes from water samples collected from wells and boreholes in Mombasa County. Methods One hundred and fifty seven (157) water samples were collected from four divisions of the county and a questionnaire administered. The samples were inoculated to double strength MacConkey broth and incubated at 370C for up to 48 hours. Positive results were compared to the 3 tube McCrady MPN table. The E. coli were confirmed by Eijkman's test and antibiotic susceptibility carried out. Using polymerase chain reaction (PCR), the E. coli were characterized to establish pathotypes. Results One hundred and thirty one (n = 131; 83.4%) samples had coliform bacteria with only 79 (60.3%) samples having E. coli. Significant values (<0.05) were noted when coliforms were compared to variables with E. Coli showing no significance when compared to similar variables. E. coli (n = 77; 100%) tested were sensitive to Gentamicin, while all (n = 77; 100%) isolates were resistant to Ampicillin. PCR typed isolates as enteroinvasive E. Coli (EIEC). Conclusion Findings suggest that coliforms and E. coli are major contaminants of wells and boreholes in Mombasa County. The isolates have a variety of resistant and sensitivity patterns to commonly used antibiotics. PMID:27200121
potential coverage of children in Peru by investigational vaccines . Enterotoxigenic Escherichia coli (ETEC) is one of the main causes of diarrhea in...important target for vaccine development (11). Diarrhea due to ETEC develops between 8 and 72 h after initial infection, usually due to the ingestion of...products in clinical specimens. Currently, derivatives of LT and the CFs are targets for the development of vaccines against ETEC. However, the great
Ahmed, W; Hodgers, L; Masters, N; Sidhu, J P S; Katouli, M; Toze, S
In this study, 200 Escherichia coli isolates from 22 rainwater tank samples in Southeast Queensland, Australia, were tested for the presence of 20 virulence genes (VGs) associated with intestinal and extraintestinal pathotypes. In addition, E. coli isolates were also classified into phylogenetic groups based on the detection of the chuA, yjaA, and TSPE4.C2 genes. Of the 22 rainwater tanks, 8 (36%) and 5 (23%) were positive for the eaeA (belonging to enteropathogenic E. coli [EPEC] and Shiga-toxigenic E. coli [STEC]) and ST1 (belonging to enterotoxigenic E. coli [ETEC]) genes, respectively. VGs (cdtB, cvaC, ibeA, kpsMT allele III, PAI, papAH, and traT) belonging to extraintestinal pathogenic E. coli (ExPEC) were detected in 15 (68%) of the 22 rainwater tanks. Of the 22 samples, 17 (77%) and 11 (50%) contained E. coli belonging to phylogenetic groups A and B1, respectively. Similarly, 10 (45%) and 16 (72%) contained E. coli belonging to phylogenetic groups B2 and D, respectively. Of the 96 of the 200 strains from 22 tanks that were VG positive, 40 (42%) were carrying a single VG, 36 (37.5%) were carrying two VGs, 17 (18%) were carrying three VGs, and 3 (3%) had four or more VGs. This study reports the presence of multiple VGs in E. coli strains belonging to the STEC, EPEC, ETEC, and ExPEC pathotypes in rainwater tanks. The public health risks associated with potentially clinically significant E. coli in rainwater tanks should be assessed, as the water is used for drinking and other, nonpotable purposes. It is recommended that rainwater be disinfected using effective treatment procedures such as filtration, UV disinfection, or simply boiling prior to drinking.
Uysal, Ahmet; Durak, Yusuf
Bacterial contamination in foods and antimicrobial resistance levels of common pathogenic strains causing food-borne disease are important in human health. Thus, typing technologies are important tools to determine primary sources of bacterial contamination. In this study, 40 Escherichia coli strains isolated from 85 food samples were evaluated in terms of genetic diversity, susceptibility to certain antibiotics, and plasmid profiles. Pulsed-field gel electrophoresis was used to identify the genetic relations of E. coli isolates. It was determined that the 40 E. coli strains revealed 32 different pulsotypes represented by 6 subtypes. Antibiotic susceptibility tests conducted by using a disc diffusion method against 15 antibiotics showed that although the isolates revealed 14 different types of resistance profiles, the strains showed the greatest resistance to ampicillin (77.5%), followed by ticarcillin-clavulanic acid (30%), tetracycline (22.5%), and cephalothin (14.5%). Plasmid isolations studies of the strains conducted by the method of alkaline lysis revealed that 18 (45%) of 40 E. coli strains contain 31 different plasmid bands ranging between 64.4 and 1 kb. The results showed that PFGE was a powerful method in tracking sources of food contamination and that the antibiotic resistance levels of food isolates were high and should be monitored.
Koczura, Ryszard; Mokracka, Joanna; Jabłońska, Lucyna; Gozdecka, Edyta; Kubek, Martyna; Kaznowski, Adam
The presence and persistence of antibiotic resistant bacteria in the environment is thought to be a growing threat to public health. The route of the spread of multiresistant bacteria from human communities to aquatic environment may lead through wastewater treatment plants that release treated wastewater to a water reservoir. In this study we used multiplex PCR assay to determine the frequency of integron presence in Escherichia coli isolates cultured from wastewater treatment plant (WWTP) (integrons were detected in 11% of E. coli isolates), river water upstream (6%) and downstream (14%) the discharge of WWTP, and clinical specimens (56%). Antimicrobial resistance of the integron-positive isolates, determined by disk diffusion method, varied between E. coli of different origin. Isolates from the downstream river, compared to those cultured from upstream river, were more frequently resistant to kanamycin, cephalotin, co-trimoxazole, trimethoprim, and fluoroquinolones. Moreover, they displayed broader resistance ranges, expressed as the number of classes of antimicrobials to which they were resistant. The results may suggest that WWTP effluent contributes to increased frequency of integron-positive E. coli isolates in the river downstream the WWTP and to their elevated resistance level.
De Carli, Silvia; Ikuta, Nilo; Lehmann, Fernanda Kieling Moreira; da Silveira, Vinicius Proença; de Melo Predebon, Gabriela; Fonseca, André Salvador Kazantzi; Lunge, Vagner Ricardo
Escherichia coli is a commensal bacterium of the bird's intestinal tract, but it can invade different tissues resulting in systemic symptoms (colibacillosis). This disease occurs only when the E. coli infecting strain presents virulence factors (encoded by specific genes) that enable the adhesion and proliferation in the host organism. Thus, it is important to differentiate pathogenic (APEC, avian pathogenic E. coli) and non-pathogenic or fecal (AFEC, avian fecal E. coli) isolates. Previous studies analyzed the occurrence of virulence factors in E. coli strains isolated from birds with colibacillosis, demonstrating a high frequency of the bacterial genes cvaC, iroN, iss, iutA, sitA, tsh, fyuA, irp-2, ompT and hlyF in pathogenic strains. The aim of the present study was to evaluate the occurrence and frequency of these virulence genes in E. coli isolated from poultry flocks in Brazil. A total of 138 isolates of E. coli was obtained from samples of different tissues and/or organs (spleen, liver, kidney, trachea, lungs, skin, ovary, oviduct, intestine, cloaca) and environmental swabs collected from chicken and turkey flocks suspected to have colibacillosis in farms from the main Brazilian producing regions. Total DNA was extracted and the 10 virulence genes were detected by traditional and/or real-time PCR. At least 11 samples of each gene were sequenced and compared to reference strains. All 10 virulence factors were detected in Brazilian E. coli isolates, with frequencies ranging from 39.9% (irp-2) to 68.8% (hlyF and sitA). Moreover, a high nucleotide similarity (over 99%) was observed between gene sequences of Brazilian isolates and reference strains. Seventy-nine isolates were defined as pathogenic (APEC) and 59 as fecal (AFEC) based on previously described criteria. In conclusion, the main virulence genes of the reference E. coli strains are also present in isolates associated with colibacillosis in Brazil. The analysis of this set of virulence factors can be
Galli, Lucía; Brusa, Victoria; Singh, Pallavi; Cataldi, Angel Adrián; Manning, Shannon; Peral-García, Pilar; Leotta, Gerardo Aníbal
Escherichia coli O157:H7 is an enteric pathogen associated with food safety threats and with significant morbidity and mortality worldwide. In Argentina, post-enteric hemolytic uremic syndrome (HUS) is endemic, with >70% of cases associated with E. coli O157 infection. To date the biological basis behind the severity among E. coli O157 infections is unknown. However, single nucleotide polymorphism (SNP) typing has helped to define nine E. coli O157:H7 clades, of which clade 8 strains are associated with severe disease cases. The aim of this study was to characterize a collection of 20 STEC O157:H7 strains isolated between 2011 and 2013 from ground beef and different environmental samples from butcher shops of Argentina. All strains harbored the eae, ehxA, fliCH7, efa, iha, and toxB genes, with stx2a/stx2c as the predominant genotype (75%). The XbaI-PFGE analysis showed that the E. coli O157 strains had high genetic diversity. Nine strains were grouped in four XbaI-PFGE clusters, whereas 11 strains showed unique XbaI-PFGE patterns. In contrast, the SNP analysis allowed us to separate the strains in two distinct lineages representing clade 8 (70%) and clade 6 (30%). Our results show the molecular characterization of E. coli O157 strains isolated from ground beef and environmental samples from Argentinean butcher shops.
Ibrahim, Delveen R; Dodd, Christine E R; Stekel, Dov J; Ramsden, Stephen J; Hobman, Jon L
Escherichia coli strains were isolated from a single dairy farm as a sentinel organism for the persistence of antibiotic resistance genes in the farm environment. Selective microbiological media were used to obtain 126 E. coli isolates from slurry and faeces samples from different farm areas. Antibiotic resistance profiling for 17 antibiotics (seven antibiotic classes) showed 57.9% of the isolates were resistant to between 3 and 15 antibiotics. The highest frequency of resistance was to ampicillin (56.3%), and the lowest to imipenem (1.6%), which appeared to be an unstable phenotype and was subsequently lost. Extended spectrum β-lactamase (ESBL) resistance was detected in 53 isolates and blaCTX-M, blaTEM and blaOXA genes were detected by PCR in 12, 4 and 2 strains, respectively. Phenotypically most isolates showing resistance to cephalosporins were AmpC rather than ESBL, a number of isolates having both activities. Phenotypic resistance patterns suggested co-acquisition of some resistance genes within subsets of the isolates. Genotyping using ERIC-PCR demonstrated these were not clonal, and therefore co-resistance may be associated with mobile genetic elements. These data show a snapshot of diverse resistance genes present in the E. coli population reservoir, including resistance to historically used antibiotics as well as cephalosporins in contemporary use.
Singh, Santosh Kumar; Seema, Kumari; Gupta, Minakshi
Escherichia coli mediated urinary tract infection has been reported to be most prevalent among patients of different class, gender and ages. Currently, multidrug resistant E. coli harboring several virulence factors are most perilous threats for patients especially for elders. The aim of this study was to determine the antibiotic resistance pattern, co-resistance and phenotypic virulence factors present in uropathogenic E. coli isolated from aged patients. Thirty-nine E. coli isolates were collected during May-June 2014 from patients between 50 to 80 years of age. Experiments have been carried out to determine the antibiotic resistance, co-resistances and phenotypic adherent factors present in each isolate. Clonal relatedness was also determined in the AmpC positive uropathogenic E. coli (UPEC). 97.43% isolates were found to be multidrug resistant and 41.02% of them were AmpC producer. AmpC producer group showed higher multiple antibiotic resistance index than AmpC non-producer (p value < 0.01) group. Interestingly, adherence factor Type 1 fimbriae were found among 84.61% of total isolates which were more prevalent in elderly female patients than males. Biofilm production studies revealed that 84.61% of total isolates are more common in elderly males. This study adds value for the proper empiric selection of antibiotic therapy as well as calls for continuous monitoring of the incidence of drug resistance virulent uropathogenic E. coli mediated urinary tract infection in elderly patients.
HOANG, Phuong Hoai; AWASTHI, Sharda Prasad; DO NGUYEN, Phuc; NGUYEN, Ngan Ly Hoang; NGUYEN, Dao Thi Anh; LE, Ninh Hoang; VAN DANG, Chinh; HINENOYA, Atsushi; YAMASAKI, Shinji
In this study, we attempted to isolate Escherichia coli from healthy adults in Ho Chi Minh City, Vietnam, and characterized its antimicrobial resistance profile, extended-spectrum β-lactamase (ESBL) genotype, phylogenetic grouping and virulence gene profile. A total of 103 E. coli isolates were obtained, and most of them were antimicrobial resistant such to streptomycin (80.6%), tetracycline (67.0%), ampicillin (65.0%), sulfamethoxsazole/trimethoprim (48.5%), nalidixic acid (43.7%), chloramphenicol (34.0%), cefotaxime (15.5%), ciprofloxacin (15.5%), kanamycin (12.6%), ceftazidime (10.7%), fosfomycin (4.9%) and gentamicin (2.9%). However, all these E. coli strains were susceptible to imipenem. Surprisingly, of 103 strains, 74 (71.8%) and 43 (41.7%) strains showed resistance to more than 3 and 5 classes of antimicrobials, respectively. Furthermore, 10 E. coli strains were ESBL-producers and positive for blaCTX-M genes (7 for blaCTX-M-9 and 3 for blaCTX-M-1), while five were additionally positive for blaTEM genes. S1-nuclease pulsed-field gel electrophoresis analysis revealed that 7 and 3 strains of E. coli carry blaCTX-M genes on their large plasmid and chromosome, respectively. Phylogenetic analysis exhibited that majority of the E. coli strains was grouped into A (44.7%), followed by B1 (23.3%), B2 (18.4%) and D (13.6%). Virulence genes associated with diarrheagenic E. coli, such as astA, EAF, eaeA, elt and eagg were also detected in ESBL-producing E. coli as well as antimicrobial resistant strains. These data suggest that commensal E. coli of healthy human could be a reservoir for antimicrobial resistance determinants and some of them might be harmful to human. PMID:28123141
Hoang, Phuong Hoai; Awasthi, Sharda Prasad; DO Nguyen, Phuc; Nguyen, Ngan Ly Hoang; Nguyen, Dao Thi Anh; LE, Ninh Hoang; VAN Dang, Chinh; Hinenoya, Atsushi; Yamasaki, Shinji
In this study, we attempted to isolate Escherichia coli from healthy adults in Ho Chi Minh City, Vietnam, and characterized its antimicrobial resistance profile, extended-spectrum β-lactamase (ESBL) genotype, phylogenetic grouping and virulence gene profile. A total of 103 E. coli isolates were obtained, and most of them were antimicrobial resistant such to streptomycin (80.6%), tetracycline (67.0%), ampicillin (65.0%), sulfamethoxsazole/trimethoprim (48.5%), nalidixic acid (43.7%), chloramphenicol (34.0%), cefotaxime (15.5%), ciprofloxacin (15.5%), kanamycin (12.6%), ceftazidime (10.7%), fosfomycin (4.9%) and gentamicin (2.9%). However, all these E. coli strains were susceptible to imipenem. Surprisingly, of 103 strains, 74 (71.8%) and 43 (41.7%) strains showed resistance to more than 3 and 5 classes of antimicrobials, respectively. Furthermore, 10 E. coli strains were ESBL-producers and positive for blaCTX-M genes (7 for blaCTX-M-9 and 3 for blaCTX-M-1), while five were additionally positive for blaTEM genes. S1-nuclease pulsed-field gel electrophoresis analysis revealed that 7 and 3 strains of E. coli carry blaCTX-M genes on their large plasmid and chromosome, respectively. Phylogenetic analysis exhibited that majority of the E. coli strains was grouped into A (44.7%), followed by B1 (23.3%), B2 (18.4%) and D (13.6%). Virulence genes associated with diarrheagenic E. coli, such as astA, EAF, eaeA, elt and eagg were also detected in ESBL-producing E. coli as well as antimicrobial resistant strains. These data suggest that commensal E. coli of healthy human could be a reservoir for antimicrobial resistance determinants and some of them might be harmful to human.
Manges, Amee R; Mende, Katrin; Murray, Clinton K; Johnston, Brian D; Sokurenko, Evgeni V; Tchesnokova, Veronika; Johnson, James R
Antimicrobial-resistant Escherichia coli are a concern for military health services. We studied 100 extended-spectrum beta-lactamase (ESBL)-producing and non-producing E. coli clinical and surveillance isolates from military personnel and civilians at Brooke Army Medical Center (2007-2011). Major E. coli lineages, most prominently ST10 (24%), ST131 (16%), and ST648 (8%), were distributed much as reported for other North American populations. ST131, represented mainly by its resistance-associated ST131-H30 clonal subset, was uniquely associated with a clinical origin, regardless of ESBL status. Thus, clonal background predicted resistance phenotype and clinical versus surveillance origin, and these findings could assist military clinicians and epidemiologists.
Munns, Krysty D.; Zaheer, Rahat; Xu, Yong; Stanford, Kim; Laing, Chad R.; Gannon, Victor P. J.; Selinger, L. Brent; McAllister, Tim A.
Cattle are the primary reservoir of the foodborne pathogen Escherichia coli O157:H7, with the concentration and frequency of E. coli O157:H7 shedding varying substantially among individual hosts. The term ‘‘super-shedder” has been applied to cattle that shed ≥104 cfu E. coli O157:H7/g of feces. Super-shedders have been reported to be responsible for the majority of E. coli O157:H7 shed into the environment. The objective of this study was to determine if there are phenotypic and/or genotypic differences between E. coli O157:H7 isolates obtained from super-shedder compared to low-shedder cattle. From a total of 784 isolates, four were selected from low-shedder steers and six isolates from super-shedder steers (4.01–8.45 log cfu/g feces) for whole genome sequencing. Isolates were phage and clade typed, screened for substrate utilization, pH sensitivity, virulence gene profiles and Stx bacteriophage insertion (SBI) sites. A range of 89–2473 total single nucleotide polymorphisms (SNPs) were identified when sequenced strains were compared to E. coli O157:H7 strain Sakai. More non-synonymous SNP mutations were observed in low-shedder isolates. Pan-genomic and SNPs comparisons did not identify genetic segregation between super-shedder or low-shedder isolates. All super-shedder isolates and 3 of 4 of low-shedder isolates were typed as phage type 14a, SBI cluster 3 and SNP clade 2. Super-shedder isolates displayed increased utilization of galactitol, thymidine and 3-O-β-D-galactopyranosyl-D-arabinose when compared to low-shedder isolates, but no differences in SNPs were observed in genes encoding for proteins involved in the metabolism of these substrates. While genetic traits specific to super-shedder isolates were not identified in this study, differences in the level of gene expression or genes of unknown function may still contribute to some strains of E. coli O157:H7 reaching high densities within bovine feces. PMID:27018858
Harada, Kazuki; Shimizu, Takae; Kataoka, Yasushi; Takahashi, Toshio
Orbifloxacin is a fluoroquinolone drug used widely in companion animal medicine. In this study, we firstly determined post-antibiotic effects (PAEs) and post-antibiotic sub-minimum inhibitory concentrations (MIC) effects (PA-SMEs) of orbifloxacin for two strains each of Escherichia coli and Pseudomonas aeruginosa from dogs, and these parameters were compared with those of enrofloxacin. At twice the MIC, the PAEs of orbifloxacin ranged from -0.28-0.93 h (mean, 0.29 h) for E. coli and -0.18-1.18 h (mean, 0.37 h) for P. aeruginosa. These parameters were not significantly different for E. coli and shorter for P. aeruginosa, compared to enrofloxacin (P < 0.05). Continued exposure to 0.1, 0.2, and 0.3 the MIC of orbifloxacin resulted in average PA-SMEs of 0.55, 1.11, and 2.03 h, respectively, for E. coli, and 1.04, 1.40, and 2.47 h, respectively, for P. aeruginosa. These PA-SMEs, which had no significant differences with those of enrofloxacin, were significantly longer than the corresponding PAEs (P < 0.05). These results suggest that the PA-SME of orbifloxacin for E. coli and P. aeruginosa can be meaningfully prolonged by increase of sub-MICs.
Agabou, A; Lezzar, N; Ouchenane, Z; Khemissi, S; Satta, D; Sotto, A; Lavigne, J-P; Pantel, A
The objectives of this study were to determine rates, patterns, and mechanisms of antibiotic resistance, and to assess connections between chicken commensal, human commensal, and pathogenic ciprofloxacin-resistant Escherichia coli isolates. All E. coli isolates collected from chickens, their farmers, and patients in the Constantine region (North-east Algeria) were analyzed for bla and plasmid-mediated quinolone resistance (PMQR) gene contents, phylogroups, Rep-PCR profiles, and multilocus sequence types. A high prevalence of resistance to fluoroquinolones (51.4 % to ciprofloxacin) was recorded in avian isolates. Of these, 22.2 % carried the aac(6')-Ib-cr gene, whereas lower resistance levels to these antibiotics were recorded in chicken farmers' isolates. None of the commensal isolates harbored the qnr, qepA, or oqxAB genes. One human pathogenic isolate was ertapenem-resistant and harbored the bla OXA-48 gene, 84 showed an extended-spectrum β-lactamase phenotype, with bla CTX-M-15 gene prevalent in 87.2 % of them. Seventy isolates were resistant to fluoroquinolones, with aac(6')-Ib-cr present in 72.8 %, qnrB in 5.7 %, and qnrS in 10 %. Three Rep-PCR profiles were common to chicken commensal and human pathogenic isolates (phylogroups D and B1; ST21, ST48, and ST471 respectively); one was found in both chicken and chicken-farmer commensal strains (D; ST108), while another profile was identified in a chicken-farmer commensal strain and a human pathogenic one (B1; ST19). These findings suggest clonal and epidemiologic links between chicken and human ciprofloxacin-resistant E. coli isolates and the important role that poultry may play in the epidemiology of human E. coli infections in the Constantine region.
Chigor, Vincent N; Umoh, Veronica J; Smith, Stella I; Igbinosa, Etinosa O; Okoh, Anthony I
We have assessed the prevalence of Escherichia coli O157 in diarrhoeal patients and surface waters from some selected sources in Zaria (Nigeria), evaluating the antibiotic susceptibility and plasmid profiles of 184 E. coli isolates, obtained from 228 water samples and 112 diarrhoeal stool specimens (collected from children aged <15 years), using standard methods. The detection rate of E. coli O157 in surface waters was 2.2% and its prevalence in children with diarrhoea was 5.4%. The most active antibiotics were gentamicin, chloramphenicol and fluoroquinolones. Seventy-nine (42.9%) of 184 E. coli isolates were resistant to four or more antibiotics. Multidrug resistance (MDR) was higher amongst aquatic isolates than the clinical isolates. Out of 35 MDR isolates (20 of which were O157 strains), 22 (62.9%) harboured plasmids all of which were no less than 2.1 kb in size. Amongst the 20 E. coli O157 strains, only seven (35.0%) contained multiple plasmids. An aquatic O157 isolate containing two plasmids was resistant to seven drugs, including ampicillin, cefuroxime, ciprofloxacin, cotrimoxazole, nalidixic acid, nitrofurantoin and tetracycline. Loss of plasmid correlated with loss of resistance to antibiotics in cured (mutant) strains selected in tetracycline (50 μg/mL)-nutrient agar plates. Our findings revealed that plasmids were prevalent in both the aquatic and clinical isolates, and suggest that the observed MDR is plasmid-mediated. The occurrence of plasmid-mediated multidrug resistant E. coli O157 in surface waters used as sources for drinking, recreation and fresh produce irrigation heightens public health concern.
Begum, Yasmin A.; Talukder, K. A.; Azmi, Ishrat J.; Shahnaij, Mohammad; Sheikh, A.; Sharmin, Salma; Svennerholm, A.-M.; Qadri, Firdausi
Background Enterotoxigenic Escherichia coli (ETEC) is a common cause of bacterial infection leading to acute watery diarrhea in infants and young children as well as in travellers to ETEC endemic countries. Ciprofloxacin is a broad-spectrum antimicrobial agent nowadays used for the treatment of diarrhea. This study aimed to characterize ciprofloxacin resistant ETEC strains isolated from diarrheal patients in Bangladesh. Methods A total of 8580 stool specimens from diarrheal patients attending the icddr,b Dhaka hospital was screened for ETEC between 2005 and 2009. PCR and Ganglioside GM1- Enzyme Linked Immuno sorbent Assay (ELISA) was used for detection of Heat labile (LT) and Heat stable (ST) toxins of ETEC. Antimicrobial susceptibilities for commonly used antibiotics and the minimum inhibitory concentration (MIC) of nalidixic acid, ciprofloxacin and azithromycin were examined. DNA sequencing of representative ciprofloxacin resistant strains was performed to analyze mutations of the quinolone resistance-determining region of gyrA, gyrB, parC and parE. PCR was used for the detection of qnr, a plasmid mediated ciprofloxacin resistance gene. Clonal variations among ciprofloxacin resistant (CipR) and ciprofloxacin susceptible (CipS) strains were determined by Pulsed-field gel electrophoresis (PFGE). Results Among 1067 (12%) ETEC isolates identified, 42% produced LT/ST, 28% ST and 30% LT alone. Forty nine percent (n = 523) of the ETEC strains expressed one or more of the 13 tested colonization factors (CFs) as determined by dot blot immunoassay. Antibiotic resistance of the ETEC strains was observed as follows: ampicillin 66%, azithromycin 27%, ciprofloxacin 27%, ceftriazone 13%, cotrimaxazole 46%, doxycycline 44%, erythromycin 96%, nalidixic acid 83%, norfloxacin 27%, streptomycin 48% and tetracycline 42%. Resistance to ciprofloxacin increased from 13% in 2005 to 34% in 2009. None of the strains was resistant to mecillinam. The MIC of the nalidixic acid and
Damavandi, Mohammad-Sadegh; Latif Pour, Mohammad
Introduction Extended-spectrum β-lactamases (ESBLs) are a set of plasmid-borne, various and quickly evolving enzymes that are a main therapeutic issue now-a-days for inpatient and outpatient treatment. Aim The aim of this study was to determine multi-drug resistance (MDR) and ESBLs producing E. coli strains, prevalence of class D Carbapenemases among ESBLs producing Escherichia coli isolates from educational hospitals in Shahrekord, Iran. Materials and Methods Uropathogenic Escherichia coli strains were isolated from patients with Urinary Tract Infections (UTIs). The agar disc diffusion test was used to characterize the antimicrobial sensitivity of the E. coli isolates. The ESBL positive strains were identified by phenotypic double-disk synergy test, by third-generation cephalosporin in combination with or without clavulanic acid. Multiplex PCR was carried out for detection of the three families of OXA-type carbapenamases including OXA-23, OXA-24, and OXA-48 in E. coli strains. Results All bacterial isolates were susceptible to meropenem. Ninety isolates produced ESBL, 55 E. coli isolates from inpatients, and 35 isolates from outpatients, with a significant association (p< 0.05). The prevalence of OXA-23, OXA-24, and OXA-48 in the ESBLs producing isolates was respectively 21%, 18%, and 11% for inpatients, and 10%, 8%, and 6% for outpatients. Conclusion ESBL-producing E. coli isolates are also a major threat in the clinical setting. The findings of this study indicated the high occurrence of ESBLs and multiple antibiotic resistance in E. coli isolates. PMID:27462579
Röderova, Magdalena; Halova, Dana; Papousek, Ivo; Dolejska, Monika; Masarikova, Martina; Hanulik, Vojtech; Pudova, Vendula; Broz, Petr; Htoutou-Sedlakova, Miroslava; Sauer, Pavel; Bardon, Jan; Cizek, Alois; Kolar, Milan; Literak, Ivan
Escherichia coli is a common commensal bacterial species of humans and animals that may become a troublesome pathogen causing serious diseases. The aim of this study was to characterize the quinolone resistance phenotypes and genotypes in E. coli isolates of different origin from one area of the Czech Republic. E. coli isolates were obtained from hospitalized patients and outpatients, chicken farms, retailed turkeys, rooks wintering in the area, and wastewaters. Susceptibility of the isolates grown on the MacConkey agar with ciprofloxacin (0.05 mg/L) to 23 antimicrobial agents was determined. The presence of plasmid-mediated quinolone resistance (PMQR) and ESBL genes was tested by PCR and sequencing. Specific mutations in gyrA, gyrB, parC, and parE were also examined. Multilocus sequence typing and pulsed-field gel electrophoresis were performed to assess the clonal relationship. In total, 1050 E. coli isolates were obtained, including 303 isolates from humans, 156 from chickens, 105 from turkeys, 114 from the rooks, and 372 from wastewater samples. PMQR genes were detected in 262 (25%) isolates. The highest occurrence was observed in isolates from retailed turkey (49% of the isolates were positive) and inpatients (32%). The qnrS1 gene was the most common PMQR determinant identified in 146 (56%) followed by aac(6′)-Ib-cr in 77 (29%), qnrB19 in 41 (16%), and qnrB1 in 9 (3%) isolates. All isolates with high level of ciprofloxacin resistance (>32 mg/L) carried double or triple mutations in gyrA combined with single or double mutations in parC. The most frequently identified substitutions were Ser(83)Leu; Asp(87)Asn in GyrA, together with Ser(80)Ile, or Glu(84)Val in ParC. Majority of these isolates showed resistance to beta-lactams and multiresistance phenotype was found in 95% isolates. Forty-eight different sequence types among 144 isolates analyzed were found, including five major clones ST131 (26), ST355 (19), ST48 (13), ST95 (10), and ST10 (5). No isolates
Vijay, Deepthi; Dhaka, Pankaj; Vergis, Jess; Negi, Mamta; Mohan, Vysakh; Kumar, Manesh; Doijad, Swapnil; Poharkar, Krupali; Kumar, Ashok; Malik, Satyaveer Singh; Barbuddhe, Sukhadeo Baliram; Rawool, Deepak B
Enteroaggregative Escherichia coli (EAEC) is an important pathotype that causes infection in humans and animals. EAEC isolates (n=86) recovered from diarrhoeal cases in human infants (37) and young animals (49) were characterized as 'typical' and/or 'atypical' EAEC strains employing PCR for virulence associated genes (cvd432, aaiA, astA, pilS, irp2, ecp, pic, aggR, aafA, aggA, and agg3A). Besides, biofilm formation ability of human and animal EAEC isolates was assessed using microtiter plate assay. In addition, the transcriptional profile of biofilm associated genes (fis and ecp) was also evaluated and correlated with biofilm formation assay for few selected EAEC isolates of human and animal origins. Overall, a diverse virulence gene profile was observed for the EAEC isolates of human and animal origins as none of the EAEC isolates revealed the presence of all the genes that were targeted. Nine 'typical' EAEC isolates were identified (6 from humans and 3 from animals) while, the majority of the isolates were 'atypical' EAEC strains. Isolation and identification of three 'typical' EAEC isolates from animals (canines) appears to be the first report globally. Further, based on the observations of the biofilm formation assay, the study suggested that human EAEC isolates in particular were comparatively more biofilm producers than that of the animal EAEC isolates. The fis gene was highly expressed in majority of 'typical' EAEC isolates and the ecp gene in 'atypical' EAEC isolates.
Jeon, Soo Jin; Cunha, Federico; Ginn, Amber; Jeong, KwangCheol Casey
ABSTRACT Escherichia coli is involved in the pathogenicity of metritis in cows. We report here the genome sequences of E. coli strains isolated at calving from the uterus, vagina, vulva, and rectoanal junction of a dairy cow that later developed metritis. The genomic similarities will give an insight into phylogenetic relationships among strains. PMID:28302783
Ringertz, S.; Bellete, B.; Karlsson, I.; Ohman, G.; Gedebou, M.; Kronvall, G.
A high level of antimicrobial resistance of bacteria has been detected at the Tikur Anbessa Hospital (TAH), Addis Ababa, for many years. In contrast, at the Karolinska Hospital (KH), Stockholm, the level of resistance is low. Reported are the results of an investigation of the correlation between antibiotic usage and the antimicrobial resistance rates of Escherichia coli isolates from patients with urinary tract infections in these hospitals. At TAH the strains of E. coli isolated were considerably more resistant to all seven antibiotics tested. The level of multiresistance was 63% at TAH and 7% at KH. There were no significant differences in the total amount of antibiotics used in the two hospitals, except for antituberculosis agents. The strain biotypes and antibiograms, together with the length of patients' hospitalization before a positive urine culture was obtained, suggest that the majority of the strains from TAH were of nosocomial origin. PMID:2189587
Ombarak, Rabee A; Hinenoya, Atsushi; Awasthi, Sharda Prasad; Iguchi, Atsushi; Shima, Ayaka; Elbagory, Abdel-Rahman M; Yamasaki, Shinji
The objectives of this study were to investigate prevalence and pathogenic potential of Escherichia coli contaminating raw milk and its products in Egypt. Out of 187 dairy products including 72 raw milk samples, 55 Karish cheese and 60 Ras cheese, 222 E. coli isolates including 111, 89 and 22 were obtained from 55 raw milk samples (76.4%), 41 Karish cheese (74.5%), and 13 Ras cheese (21.7%), respectively. Isolated E. coli strains were examined for 24 representative virulence genes present in diarrheagenic E. coli (DEC) and extraintestinal pathogenic E. coli (ExPEC). Among DEC and ExPEC virulence factors, genes for enteropathogenic E. coli (eaeA, bfpA, EAF), enterohemorrhagic E. coli (stx1, stx2, eaeA), enterotoxigenic E. coli (elt, est), enteroinvasive E. coli (invE), enteroaggregative E. coli (Eagg, astA), diffusely adherent E. coli (daaD), ExPEC (cdt-I to cdt-V, cnf1, cnf2, hlyA) and putative adhesins (efa1, iha, ehaA, saa, and lpfAO113) were screened by colony hybridization assay. Out of 222 E. coli strains, 104 (46.8%) isolated from 69 (36.9%) samples carried one or more virulence genes. The most prevalent gene detected was lpfAO113 (40.5%), followed by ehaA (32.4%,), astA (3.15%,), iha (1.80%), hlyA (1.35%), stx1 (0.90%), stx2 (0.90%), eaeA (0.45%), cdt-III (0.45%) and cnf2 (0.45%). Two strains isolated from Karish cheese harbored 5 virulence genes (stx1, stx2, iha, ehaA, lpfAO113). Stx subtype was determined to be stx1 (not stx1c or stx1d) and stx2d. Indeed, expression of hemolysin A, CDT-III, CNF-II, Stx1 and Stx2d was confirmed by blood agar plate, cytotoxicity assay and Western blotting, respectively. Among the 222 E. coli strains, 54 (48.6%), 38 (42.6%) and 12 (54.7%) isolated from raw milk, Karish cheese and Ras cheese were potentially virulent, respectively. O-genotyping indicated that most of the potentially virulent E. coli isolates did not belong to clinically important O serogroups except O75, O91 and O166, which have been associated with human
Pons, Maria J.; Mosquito, S.; Gomes, C.; del Valle, L.J.; Ochoa, T.J.; Ruiz, J.
Background Antibiotic resistance is an increasing problem, particularly in countries where antibiotic use is frequently not controlled. The aim of this study was to analyse the prevalence of the molecular mechanisms of quinolone-resistance in E. coli isolated from faeces of healthy Peruvian children or those presenting diarrhoea. Methods The presence of target mutations, transferable quinolone-resistance mechanisms and the role of Phe-Arg-β-Naphtylamyde inhibitible efflux pumps were studied in 96 Escherichia coli (46 diarrheogenic and 50 non-diarrheogenic) isolates exhibiting resistance or diminished susceptibility to quinolones. Results The most resistant phenotype, NalR and CipR, was most frequently present in isolates of healthy children. The distribution of quinolone resistance mechanisms between diarrheogenic (DEC) and commensal (non DEC) isolates was equitable, although the aac(6′)Ib-cr gene was mainly detected in DEC isolates: 17 (34%) vs non DEC isolates nine (20%). QnrB was present in five (10%) DEC vs three (6%) non DEC isolates. Conclusions Point mutations in gyrA and parC genes play a relevant role in quinolone resistance acquisition and highlight the role of efflux pumps also. This study provides knowledge about the molecular mechanisms involved in quinolone resistance in isolates in a non exposed population under high community antibiotic pressure. PMID:24306130
Adhikari, Achyut; Bary, Andy; Cogger, Craig; James, Caleb; Ünlü, Gülhan; Killinger, Karen
Pathogens exposed to agricultural production environments are subject to multiple stresses that may alter their survival under subsequent stress conditions. The objective of this study was to examine heat and starvation stress response of Escherichia coli O157:H7 strains isolated from agricultural matrices. Seven E. coli O157:H7 isolates from different agricultural matrices-soil, compost, irrigation water, and sheep manure-were selected, and two ATCC strains were used as controls. The E. coli O157:H7 isolates were exposed to heat stress (56°C in 0.1% peptone water for up to 1 h) and starvation (in phosphate-buffered saline at 37°C for 15 days), and their survival was examined. GInaFiT freeware tool was used to perform regression analyses of the surviving populations. The Weibull model was identified as the most appropriate model for response of the isolates to heat stress, whereas the biphasic survival curves during starvation were fitted using the double Weibull model, indicating the adaptation to starvation or a resistant subpopulation. The inactivation time during heating to achieve the first decimal reduction time (δ) calculated with the Weibull parameters was the highest (45 min) for a compost isolate (Comp60A) and the lowest (28 min) for ATCC strain 43895. Two of the nine isolates (ATCC 43895 and a manure isolate) had β < 1, indicating that surviving populations adapted to heat stress, and six strains demonstrated downward concavity (β > 1), indicating decreasing heat resistance over time. The ATCC strains displayed the longest δ2 (>1,250 h) in response to starvation stress, compared with from 328 to 812 h for the environmental strains. The considerable variation in inactivation kinetics of E. coli O157:H7 highlights the importance of evaluating response to stress conditions among individual strains of a specific pathogen. Environmental isolates did not exhibit more robust response to stress conditions in this study compared with ATCC strains.
Girardeau, Jean Pierre; Lalioui, Lila; Said, A Mohamed Ou; De Champs, Christophe; Le Bouguénec, Chantal
The afimbrial AfaE-VIII adhesin is common among Escherichia coli isolates from calves with intestinal and/or extraintestinal infections and from humans with sepsis or pyelonephritis. The virulence genotypes of 77 Escherichia coli afa-8 isolates from farm animals and humans were compared to determine whether any trait of commonality exists between isolates of the different host species. Over half of the extraintestinal afa-8 isolates were associated with pap and f17Ac adhesin genes and contained virulence genes (pap, hly, and cnf1) which are characteristic of human extraintestinal pathogenic E. coli (ExPEC). PapG, which occurs as three known variants (variants I to III), is encoded by the corresponding three alleles of papG. Among the pap-positive strains, new papG variants (papGrs) that differed from the isolates with genes for the three adhesin classes predominated over isolates with papG allele III, which in turn were more prevalent than those with allele II. The data showed the substantial prevalence of the enteroaggregative E. coli heat-stable enterotoxin gene (east1) among afa-8 isolates. Most of the afa-8 isolates harbored the high-pathogenicity island (HPI) present in pathogenic Yersinia; however, two-thirds of the HPI-positive strains shared a truncated HPI integrase gene. The presence of ExPEC-associated virulence factors (VFs) in extraintestinal isolates that carry genes typical of enteric strains and that express O antigens associated with intestinal E. coli is consistent with transfer of VFs and O-antigen determinants between ExPEC and enteric strains. The similarities between animal and human ExPEC strains support the hypothesis of overlapping populations, with members of certain clones or clonal groups including animal and human strains. The presence of multiple-antibiotic-resistant bovine afa-8 strains among such clones may represent a potential public health risk.
Beheshti Maal, Keivan; Soleimani Delfan, Abbas; Salmanizadeh, Sharareh
Background: Phage therapy or use of lytic bacteriophages for eliminating bacterial populations has been developed for several aspects of human affairs such as medicine, agriculture and food industries. Objectives: The high load of coliforms of treated wastewater effluents that are discharged into the rivers or agricultural lands is a serious concern of the Iran Department of Environment and the reduction of coliforms using phages to overcome this problem is an asset. This research aimed to isolate and identify specific lytic coliphages and investigate their effects on native and standard Escherichia coli strains as well as coliform populations in municipal wastewater. Materials and Methods: The wastewater sample was cultured on selective culture media to isolate a native coliform strain and characterized using molecular methods. River water was centrifuged and passed through a 0.45 μm filter and its lytic coliphages were enriched and purified against a native E. coli as well as a standard E. coli strain. Municipal wastewater was treated with isolated lytic coliphages and most probable number (MPN) reduction was examined. Results: E. coli SBSWF27, which is a native strain of E. coli from Isfahan municipal wastewater treatment plant, was isolated and characterized. Also two novel bacteriophages related to Myoviridae and Podoviridae families of bacteriophages from Zayandehrood River (Isfahan, Iran) were isolated. These coliphages had lytic effects on E. coli PTCC1399 and E. coli SBSWF27 as coliform's index. The myovirus had a hexagonal head measuring 27.28 nm and a noncontractile tail measuring 204.5 × 13.63 nm. The podovirus had an oval head measuring 98 × 35 nm and a tail, 14 nm in diameter. The treatment of municipal sewage with the coliphage mixture resulted in a 22-fold decrease of the coliform's MPN from 2400 to 110 after two hours of incubation. Conclusions: This is the first report on isolation and identification of two novel lytic myovirus and podovirus
Szunerits, Sabine; Zagorodko, Oleksandr; Cogez, Virginie; Dumych, Tetiana; Chalopin, Thibaut; Alvarez Dorta, Dimitri; Sivignon, Adeline; Barnich, Nicolas; Harduin-Lepers, Anne; Larroulet, Iban; Yanguas Serrano, Aritz; Siriwardena, Aloysius; Pesquera, Amaia; Zurutuza, Amaia; Gouin, Sébastien G.; Boukherroub, Rabah; Bouckaert, Julie
Shear force exerted on uropathogenic Escherichia coli adhering to surfaces makes type-1 fimbriae stretch out like springs to catch on to mannosidic receptors. This mechanism is initiated by a disruption of the quaternary interactions between the lectin and the pilin of the two-domain FimH adhesin and transduces allosterically to the mannose-binding pocket of FimH to increase its affinity. Mannose-specific adhesion of 14 E. coli pathovars was measured under flow, using surface plasmon resonance detection on functionalized graphene-coated gold interfaces. Increasing the shear had important differential consequences on bacterial adhesion. Adherent-invasive E. coli, isolated from the feces and biopsies of Crohn’s disease patients, consistently changed their adhesion behavior less under shear and displayed lower SPR signals, compared to E. coli opportunistically infecting the urinary tract, intestines or loci of knee and hip prostheses. We exemplified this further with the extreme behaviors of the reference strains UTI89 and LF82. Whereas their FimA major pilins have identical sequences, FimH of LF82 E. coli is marked by the Thr158Pro mutation. Positioned in the inter-domain region known to carry hot spots of mutations in E. coli pathotypes, residue 158 is indicated to play a structural role in the allosteric regulation of type-1 fimbriae-mediated bacterial adhesion. PMID:27043645
Tamang, Migma Dorji; Nam, Hyang-Mi; Chae, Myung Hwa; Kim, Su-Ran; Gurung, Mamata; Jang, Geum-Chan; Jung, Suk-Chan; Lim, Suk-Kyung
The aim of this study was to investigate the prevalence of plasmid-mediated quinolone resistance (PMQR) determinants in Escherichia coli isolated from food-producing animals and to characterize the PMQR-positive isolates. A total of 365 E. coli isolates which were either nalidixic acid resistant and ciprofloxacin susceptible (NAL(R)-CIP(S); n=185), or nalidixic acid and ciprofloxacin resistant (NAL(R)-CIP(R); n=180) were assessed for the presence of PMQR determinants by polymerase chain reaction. PMQR-positive isolates were further characterized by mutation analysis within the quinolone resistance-determining region (QRDR) of gyrA, gyrB, parC, and parE, phylogenetic group analysis, and pulsed-field gel electrophoresis (PFGE). Fourteen NAL(R)-CIP(S) (n=8) and NAL(R)-CIP(R) (n=6) E. coli isolates were positive for PMQR genes. Among them, qnrB4, qnrS1, and aac(6')-Ib-cr genes were detected in two (0.5%), eight (2.2%), and four (1.1%) isolates, respectively. None of the isolates harbored qnrA, qnrC, qnrD, and qepA genes. All but one PMQR-positive isolates harbored one or more point mutations in the QRDR of gyrA, and five of these isolates had additional mutations in the parC gene. Furthermore, one isolate each had additional substitutions in gyrB and parE genes, respectively. The most prevalent mutation was Ser83-Leu within the QRDR of gyrA. Phylogenetic analysis identified three major phylogenetic lineages, with phylogroups A (n=7) and D (n=4) being the most common phylogroups. None of the isolates belonged to virulent phylogroup B2. PFGE demonstrated that a combination of clonal and horizontal gene transmission is disseminating PMQR genes among the veterinary E. coli isolates in Korea. To our knowledge, this is the first report of occurrence of qnrB, qnrS, and aac(6')-Ib-cr genes in E. coli isolated from food-producing animals in Korea. Isolation of PMQR genes from food animals is a matter of concern since they could be transmitted to humans via food animals.
Introduction: The importance of Salmonella, Campylobacter, E.coli, and Enterococcus as carriers of antimicrobial resistance is well known, but limited work has been done to examine the relationship between this phenotypic characteristic and genotypic attributes among strains isolated in similar set...
Brusa, Victoria; Piñeyro, Pablo E; Galli, Lucía; Linares, Luciano H; Ortega, Emanuel E; Padola, Nora L; Leotta, Gerardo A
Shiga toxin-producing Escherichia coli (STEC) are foodborne pathogens, and beef cattle are recognized as the principal reservoir. The aims of this study were (1) to identify the most sensitive combination of selective enrichment broths and agars for STEC isolation in artificially inoculated ground beef samples, and (2) to evaluate the most efficient combination(s) of methods for naturally contaminated ground beef samples. A total of 192 ground beef samples were artificially inoculated with STEC and non-stx bacterial strains. A combination of four enrichment broths and three agars were evaluated for sensitivity, specificity, and positive predictive value for STEC isolation from experimentally inoculated samples. Enrichments with either modified tryptic soy broth (mTSB) containing 8 mg/L novobiocin (mTSB-8) or modified Escherichia coli (mEC) broth followed by isolation in MacConkey agar were the most sensitive combinations for STEC isolation of artificially inoculated samples. Independently, both enrichments media followed by isolation in MacConkey were used to evaluate ground beef samples from 43 retail stores, yielding 65.1% and 58.1% stx-positive samples by RT-PCR, respectively. No difference was observed in the isolate proportions between these two methods (8/25 [32%] and 8/28 [28.6%]). Identical serotypes and stx genotypes were observed in STEC strains isolated from the same samples by either method. In this study, no single enrichment protocol was sufficient to detect all STEC in artificially inoculated samples and had considerable variation in detection ability with naturally contaminated samples. Moreover, none of the single or combinations of multiple isolation agars used were capable of identifying all STEC serogroups in either artificially inoculated or naturally occurring STEC-contaminated ground beef. Therefore, it may be prudent to conclude that there is no single method or combination of isolation methods capable of identifying all STEC serogroups.
Zoolkifli, Nurliyana Wan; Mutalib, Sahilah Abd
Genomic DNA of Escherichia coli were characterized by enterobacterial repetitive intergenic consensus-Polymerase chain reaction (ERIC-PCR) and the presence of Shiga toxin gene-I (Stx1) and Shiga toxin gene-2 (Stx2). These isolates were originated from imported raw beef which are come from two countries namely Australia and India. The isolation of E. coli was conducted by using Eosin Methylene Blue Agar (EMBA). A total of 94 strains had been isolated from 30 samples of imported raw beefand 42 strains had been detected positively E. coli by doing biochemical tests. All strains had been tested and the results of biochemical tests showed that 3 strains were from Australia samples while the other 39 strains were from India samples. The biochemical tests used are Indole test, Methyl Red test, Voges-Proskauer test and Citrate test. All the 42 strains were examined for Shiga toxin (stx1 and stx2) gene detection by two pair primers which are stx2F (5'-TTCTTCGGTATCCTATTCCC-3'), stx2R (5'-ATGCATCTCTGGTCATTGTA-3'), stx1F (5'-CAGTTAATGTGGTGGCGAAG-3'), and stx1R (5'-CTGTCACAGTAACAACCGT-3'). The results showed that none of the strains are positive for Shiga toxin gene. Application of ERIC-PCR method towards E. coli had successfully shown the high diversity polymorphism in 21 different genome types of DNA with primers ERIC1R (5'- CACTTAGGGGTCCTCGAATGTA- 3') and ERIC2R (5'- AAGTAAGTGACTGGGGTGACGC- 3').
Triandala Sibero, Mada; Sabdaningsih, Aninditia; Cristianawati, Olvi; Nuryadi, Handung; Karna Radjasa, Ocky; Sabdono, Agus; Trianto, Agus
Irrational used of antibiotic in several decades ago causing resistant in bacteria and decreasing the cure rate of infectious diseases. Multidrug-resistant (MDR) Escherichia coli is known to cause various of infectious diseases such as urinary tract infection, nosocomial bloodstream infection, meningitis, bacteraemia, and gastrointestinal disease. Marine sponge-associated fungi have potential as source of new compound to combat MDR E. coli. The aims of this research were to isolate marine sponge-assosiated fungi, to screen potential fungi against MDR E. coli, to identify the potential fungi and its host sponge. There were 29 marine sponge-associated fungi successfully isolated from 9 sponges. Among 29 sponge-associated fungi screened, there were 7 isolates showed antibacterial activity against MDR E. coli. The best inhibition zone produced by MPS 14.1/MT 02 and MPS 14.3/MT 04 from sponge PP.SP.16.14. According to fungi identification result fungus MPS 14.1/MT 02 was identified as Trichoderma asperellum while MPS 14.3/MT 04 was identified as Trichoderma reesei. Sponge identification leaded the PP.SP.16.14 as Cinachyrella sp.
Piazza, Roxane M F; Delannoy, Sabine; Fach, Patrick; Saridakis, Halha O; Pedroso, Margareth Z; Rocha, Letícia B; Gomes, Tânia A T; Vieira, Mônica A M; Beutin, Lothar; Guth, Beatriz E C
Escherichia coli strains of serogroup O26 comprise two distinct groups of pathogens, characterized as enteropathogenic E. coli (EPEC) and enterohemorrhagic E. coli (EHEC). Among the several genes related to type III secretion system-secreted effector proteins, espK was found to be highly specific for EHEC O26:H11 and its stx-negative derivative strains isolated in European countries. E. coli O26 strains isolated in Brazil from infant diarrhea, foods, and the environment have consistently been shown to lack stx genes and are thus considered atypical EPEC. However, no further information related to their genetic background is known. Therefore, in this study, we aimed to discriminate and characterize these Brazilian O26 stx-negative strains by phenotypic, genetic, and biochemical approaches. Among 44 isolates confirmed to be O26 isolates, most displayed flagellar antigen H11 or H32. Out of the 13 nonmotile isolates, 2 tested positive for fliCH11, and 11 were fliCH8 positive. The identification of genetic markers showed that several O26:H11 and all O26:H8 strains tested positive for espK and could therefore be discriminated as EHEC derivatives. The presence of H8 among EHEC O26 and its stx-negative derivative isolates is described for the first time. The interaction of three isolates with polarized Caco-2 cells and with intestinal biopsy specimen fragments ex vivo confirmed the ability of the O26 strains analyzed to cause attaching-and-effacing (A/E) lesions. The O26:H32 strains, isolated mostly from meat, were considered nonvirulent. Knowledge of the virulence content of stx-negative O26 isolates within the same serotype helped to avoid misclassification of isolates, which certainly has important implications for public health surveillance.
Tripathi, A K; Mishra, B M
Glycine betaine relieved sodium chloride-mediated inhibition of growth in Azospirillum lipoferum ATCC 29708. 35S-methionine labelling of proteins after salinity up-shock revealed strong induction of a 30 kDa protein which cross-reacted with the anti-glycine betaine binding protein antibody from Escherichia coli. This suggested that A. lipoferum had a salinity-induced ProU-like high-affinity glycine betaine transport system. A genomic library of A. lipoferum ATCC 29708 was screened for the proU-like gene by complementation of a proU mutant of E. coli. Four recombinant cosmids, capable of restoring growth of the proU mutant on plates containing 600 mM NaCl and 1 mM glycine betaine were selected. Selected recombinant cosmids hybridized with a proU gene probe from E. coli. Complementation of E. coli proU mutant with the A. lipoferum genomic DNA was evident by the ability of proU mutant (containing selected recombinant cosmids) to grow on minimal medium supplemented with 600 mM NaCl and 1 mM glycine betaine.
Yamamoto, Shiori; Iwabuchi, Eriko; Hasegawa, Megumi; Esaki, Hidetake; Muramatsu, Masatake; Hirayama, Norio; Hirai, Katsuya
We investigated the prevalence of antimicrobial-resistant Escherichia coli in Japanese black beef cattle from the three major production regions of Japan. We collected and examined 291 fecal samples from Japanese black beef cattle in Hokkaido, Chubu, and Kyushu. Of the 3,147 E. coli isolates, 1,397 (44.4%) were resistant to one or more antibiotics; these included 553 (39.8%) of 1,388 isolates from Hokkaido, 352 (54.4%) of 647 isolates from Chubu, and 492 (44.2%) of 1,112 isolates from Kyushu. The difference in resistance rates between the three regions was significant. The antibiotics with the highest rates of resistance were oxytetracycline and dihydrostreptomycin (35.8% each), followed by ampicillin (21.4%). Further, E. coli isolates from calves had higher resistance rates than those from growing cattle and mature cattle, and the calf isolates showed high rates of resistance to gentamicin (20.2%), enrofloxacin (9.4%), and ceftiofur (4.2%). In addition, the high degrees of similarity in the genotypes of the isolates and in the resistance patterns on each farm suggest that resistance bacteria and resistance genes were horizontally transferred. Most isolates, in each of the three regions, harbored resistance genes such as blaTEM, strA, strB, aphA1, aphAI-IAB, and catI. In contrast to the isolates from Kyushu, most of which harbored aacC2, tetB, and dfrA12, the isolates from Hokkaido and Chubu harbored a variety of resistance genes. Furthermore, the prevalence of genes for resistance to dihydrostreptomycin, gentamicin, chloramphenicol, and trimethoprim differed significantly between the regions. This is the first large-scale study describing and comparing antimicrobial-resistant bacteria from different regions in Japan. The results will contribute to improving food safety and promoting careful usage of antimicrobial agents.
Zhao, Hong-Xia; Zhao, Jun-Li; Shen, Jian-Zhong; Fan, Hong-Liang; Guan, Hong; An, Xiao-Ping; Li, Pei-Feng
Fluoroquinolones are frequently used to treat infectious disease that is caused by Escherichia coli in dairy cattle. However, fluoroquinolone resistance occurs and is due either to chromosomal mutations in the bacterial topoisomerase genes and/or to plasmid-mediated resistance genes. The purpose of this study was to determine the prevalence and molecular characteristics of fluoroquinolone resistance determinants in E. coli strains (n=148) isolated from dairy cattle with bovine endometritis in Inner Mongolia (China). Analysis of the mutations in the quinolone resistance-determining regions of resistant E. coli isolates confirmed previously reported substitutions in the GyrA and ParE. However, we identified additional substitutions in the ParC and GyrB that have not been reported earlier. No plasmid-mediated quinolone resistance genes in any of the isolates were found. The number of point mutations found per isolate correlated with an increase in the minimum inhibitory concentration of ciprofloxacin. Overall, 45.5% of the isolates were positive for the class I integrase gene along with four gene cassettes that were responsible for resistance to trimethoprim (dfr1 and dfrA17) and aminoglycosides (aadA1 and aadA5), respectively. The prevalence of extended-spectrum β-lactamases (ESBLs) was 100%, and the blaTEM gene was predominant in all of the isolates. In conclusion, our results identify the mechanism of quinolone resistance for the first time and reveal the prevalence of integron and ESBLs in E. coli isolates from dairy cattle with bovine endometritis in China after 20 years of quinolone usage in cattle.
Zhao, Jingjing; Chen, Zhangliu; Chen, Sheng; Deng, Yuting; Liu, Yahong; Tian, Wei; Huang, Xianhui; Wu, Congming; Sun, Yongxu; Sun, Yan; Zeng, Zhenling; Liu, Jian-Hua
OqxAB has recently been identified as one of the mechanisms of plasmid-mediated quinolone resistance (PMQR). Compared to what is observed for other PMQR determinants, there is a paucity of data with regard to the prevalence and epidemiology of OqxAB and its contribution to resistance to different antimicrobials. In this study, the prevalence and dissemination of oqxAB and other PMQR genes in Escherichia coli isolates from animals, farmworkers, and the environment in 2002 in China were investigated. Of the 172 E. coli isolates, 39.0% carried oqxA, while only 4.1%, 2.9%, and 0.6% carried qnr (1 qnrB6 isolate, 5 qnrS1 isolates, and 1 qnrD isolate), qepA, and aac(6′)-Ib-cr, respectively. Among the 33 isolates from farmworkers, 10 (30.3%) were positive for oqxA. oqxAB was associated with IS26 and was carried on the 43- to 115-kb IncF transferable plasmid. Transconjugants carrying oqxAB showed 4- to 16-fold increases in the MICs of quinolones, 16- to 64-fold increases in the MICs of quinoxalines, 8- to 32-fold increases in the MICs of chloramphenicol and trimethoprim-sulfamethoxazole, and 4- to 8-fold increases in the MICs of florfenicol compared to the levels for the recipient. The pulsed-field gel electrophoresis (PFGE) analysis showed that the high levels of prevalence and dissemination of oqxAB in E. coli in animal farms were primarily due to the transmission of plasmids carrying oqxAB, although clonal transmission between human and swine E. coli isolates was observed. It is concluded that oqxAB was widespread in animal farms in China, which may be due to the overuse of quinoxalines in animals. This study warrants the prudent use of quinoxalines in food animals. PMID:20696876
Khoramrooz, Seyed Sajjad; Sharifi, Asghar; Yazdanpanah, Mahboubeh; Malek Hosseini, Seyed Ali Asghar; Emaneini, Mohammad; Gharibpour, Farzaneh; Parhizgari, Najmeh; Mirzaii, Mehdi; Zoladl, Mohammad; Khosravani, Seyed Abdolmajid
Background: Most urinary tract infections (UTI) are caused by Escherichia coli. Integrons have an important role in distributing antibiotic resistance genes among bacteria. Objectives: The aim of this study was to investigate the presence of class 1, 2 and 3 integrons and their association with antibiotic resistance in E. coli isolated from patient with UTI in Yasuj, Iran. Patients and Methods: In this cross-sectional study a total of 200 E. coli were collected from 1820 patients diagnosed with UTI that had been referred to two clinical laboratories between February 2013 and November 2014 in Yasuj city, southwest of Iran. Susceptibility of isolates to 11 different antibiotics was determined by the disk agar diffusion method. multiplex-polymerase chain reaction (PCR) was used for detection of class 1, 2 and 3 integrons. The data were analyzed using the SPSS software (version 16) and the chi-square test. A P value of < 0.05 was considered statistically significant. Results: The highest rate of resistance was observed toward cephalothin (99%) and amoxicillin (76%) while only two (1%) isolates showed resistance to imipenem. Overall, 79% of isolates were multi drug resistant (MDR). Class 1 and 2 integrons were detected in 104 (52%) and 5 (2.5%) isolates respectively, while none of the isolates were positive for class 3 integrons. A significant association was observed between the presence of integrons and resistance to co-trimoxazole, nalidixic acid, ciprofloxacin, amoxicillin, ceftazidime and tetracycline (P < 0.05). Conclusions: High MDR isolates of E. coli were observed in this study. The significant association between class 1 integrons and resistance to ciprofloxacin, nalidixic acid, co-trimoxazole, amoxicillin, ceftazidime and tetracycline showed that class 1 integrons have an important role in resistance to these antibiotics in this region. PMID:26889395
Daoud, Ziad; Salem Sokhn, Elie; Masri, Khalil; Cheaito, Katia; Haidar-Ahmad, Nathaline; Matar, Ghassan M.; Doron, Shira
The early treatment of urinary tract infections (UTIs) is directly related to decrease in morbidity, which makes the empirical treatment of great importance. Recently, beta lactamases of several types have emerged as significant mechanisms of resistance in Gram-negative bacilli, especially Escherichia coli. Our aim was to study the urinary E. coli isolated from Lebanese patients and to characterize their mechanisms of resistance. The study analyzed data between 2005 and 2012 of UTIs caused by E. coli. The mechanisms of resistance were characterized by phenotypic and genotypic methods and the pulsed-field gel electrophoresis (PFGE) was used to determine the different bacterial clusters. As expected, the highest incidence was observed with E. coli (60.53–73.98%) followed by K. pneumoniae (5.32–8.33%). ICU isolates were constantly associated with the lowest rates of susceptibility to extended-spectrum cephalosporins, ciprofloxacin, as well as most of the tested antibiotics. A 100% occurrence of CTX-M in extended-spectrum β-lactamase (ESBL)-producing isolates was recorded, followed by TEM, SHV, and OXA. In addition, 15.9% harbored 4 different ESBL enzymes and only 13 isolates (14.8%) harbored only one enzyme (CTX-M). Over the years, the simultaneous susceptibility of E. coli to ceftazidime and ciprofloxacin decreased from 62.5% in 2006 to 48.7% in 2012. PFGE results demonstrated that 10 clusters were 32 generated, denoting diversity among detected isolates. Understanding the epidemiology of resistance is 33 instrumental for the implementation of recommendations for the management of antimicrobials, infection 34 control measures, as well as active surveillance and antimicrobial stewardship. PMID:25984513
Sallam, Khalid Ibrahim; Mohammed, Mahmoud Ahmed; Ahdy, Asmaa Mohammed; Tamura, Tomohiro
Sorbitol-fermenting (SF) Escherichia coli O157:H- strains have emerged as important pathogens and have been associated with a higher incidence of progression to hemolytic-uremic syndrome (HUS) than non-sorbitol fermenting (NSF) E. coli O157:H7. The present study was carried out to determine the prevalence of SF E. coli O157:H- and NSF E. coli O157:H7 strains in retail beef products in Mansoura, Egypt. The contamination rates with rfbEO157-positive E. coli O157 strains were 26.7% (8/30), 10% (3/30) and 3.7% (1/27) in ground beef, beef burger, and fresh beef samples, respectively with an overall mean of 13.8% (12/87) among all meat products tested. SF E. coli O157:H- were the most dominant among the isolated O157 strains. Of the fifteen O157 strains isolated, 11 (73.3%) were SF E. coli O157:H-, while the remaining 4 (26.7%) were NSF E. coli O157:H7. The 11 SF O157H- strains were genetically positive for sfpA gene. Restriction fragment length polymorphism (RFLP) analysis for fliC gene demonstrated a similar pattern for both SF and NSF O157 isolates. PCR assays verified the existence of stx1 gene in 7 (46.7%) and stx2 gene in 13 (86.7%) of the 15 O157 strains isolated. Unexpectedly, two of the 15 O157 strains isolated were negative for Shiga toxin genes. The eae gene was identified in all of the 15 O157 strains except in one NSF O157:H7 strain. EHEC-hlyA gene was detected in 14 (93.3%) of the 15 O157 isolates, nonetheless only 11 strains showed enterohemolytic phenotype on blood agar. A combination of the four virulence genes, stx1, stx2, eae and EHEC-hlyA were detected in 7 (46.7%) strains, while six (40%) strains were positive for stx2, eae and hlyA genes. This is the first record for isolation of E. coli O157: H- in Egypt as well as in the African continent.
Bansal, Sandhya; Tandon, Vibha
DNA gyrase (GyrA and GyrB) and topoisomerase IV (ParC and ParE) are the two essential type II topoisomerases in Escherichia coli. These enzymes act via inhibition of DNA replication. Mutations in the quinolone resistance-determining region (QRDR) of the gyrA, gyrB, parC and parE genes from clinical isolates of E. coli were determined by DNA sequencing of 54 ciprofloxacin-resistant clinical isolates from a hospital in Delhi, India. The majority of the E. coli isolates were shown to carry mutations in gyrA, parC and parE. Ciprofloxacin resistance due to accumulation of such a high number of mutations in the QRDR regions of gyrA at positions Ser83 and Asp87 and parC at position Ser80 as well as outside of the QRDR region of parE at Ser458 and Glu460 confers high-level resistance of ciprofloxacin in clinical isolates. The high frequency of occurrence of mutations in the parE gene (44.4% strains) is alarming, as topoisomerase IV is a secondary target of quinolones.
Maluta, Renato P; Borges, Clarissa A; Beraldo, Lívia G; Cardozo, Marita V; Voorwald, Fabiana A; Santana, André M; Rigobelo, Everlon C; Toniollo, Gilson H; Avila, Fernando A
Escherichia coli is the most common bacterial agent isolated from canine pyometra. The frequencies of 24 virulence genes and pulsed field gel electrophoresis (PFGE) profiles were determined for 23 E. coli isolates from cases of canine pyometra in Brazil. The frequencies of virulence genes were 91.3% fimH, 91.3% irp-2, 82.6% fyuA, 56.5% iroN, 47.8% traT, 39.1% usp, 34.8% sfaD/E, 34.8% tsh, 30.4% papC, 30.4% hlyA, 26.1% papGIII, 26.1% cnf-1, 21.7% papE/F, 21.7% iss, 17.4% iutA, 17.4% ompT, 17.4% cvaC, 17.4% hlyF, 17.4% iucD, 13.0% iucC, 13.0% astA, 4.3% papGII, 0% afaB/C and 0% papGI. The high frequency of yersiniabactin (fyuA and irp2) and salmochelin (iroN) genes suggests that iron uptake systems might be important in the pathogenesis of canine pyometra. PFGE profiles of 19 isolates were heterogeneous, confirming that E. coli isolates from canine pyometra are unlikely to be epidemic clones.
Shar, A H; Kazi, Y F; Soomro, I H
A total 72 drinking water sample were collected and analyzed by membrane filtration method during 1 year study from various points in Khairpur City. Out of these 58 (80.55%) samples were found to be contaminated with thermo-tolerant Escherichia coli 2. The susceptibility of these isolates to 35 antibiotics was studied by disc diffusion method and the organism was highly sensitive to levoflaxin, cefipime, enoxobid, noroxin, tarivid, ciproxin, avelox, amikacin, kanamycin, rocifin, pipenedic acid and slightly sensitive to cravit, naladixic acid, neomycin, cefizox, fortum cefotaxime, cefizox, fortum, tobramycin and cefoperoxone. The resistance against 16 antibiotics such as meropenem, linkomycin, fusidic acid, orbenin, penicillin, streptomycin, bacitracin, minocin, zinacef, amoxil, ceclor, claracid, cephalexin, augmentin, cephradin and dalacin was shown by these isolates. We report the presence of multi-drug resistance in thermo-tolerant Escherichia coli isolated in municipal water with different levels of prevalence in Khairpur City. In this study a higher number of positive results were obtained in all sampling points indicating the more fecally polluted municipal water.
Raya, Raul R.; Varey, Peter; Oot, Rebecca A.; Dyen, Michael R.; Callaway, Todd R.; Edrington, Tom S.; Kutter, Elizabeth M.; Brabban, Andrew D.
Bacteriophage CEV1 was isolated from sheep resistant to Escherichia coli O157:H7 colonization. In vitro, CEV1 efficiently infected E. coli O157:H7 grown both aerobically and anaerobically. In vivo, sheep receiving a single oral dose of CEV1 showed a 2-log-unit reduction in intestinal E. coli O157:H7 levels within 2 days compared to levels in the controls. PMID:16957272
Kuczkowski, Maciej; Krawiec, Marta; Voslamber, Berend; Książczyk, Marta; Płoskońska-Bugla, Gabriela; Wieliczko, Alina
Affiliation to four phylogroups (A, B1, B2, and D) was examined, among 190 Escherichia coli strains, collected from five, wild waterbird species, including the following: the Greylag goose-Anser anser (61) and the Canada goose-Branta canadensis (33) obtained in the Netherlands, and the Mallard-Anas platyrhynchos (38), the Mute swan-Cygnus olor (37), and the Great cormorant-Phalacrocorax carbo (21) obtained in Poland. Moreover, the prevalence of 10 virulence factors: astA, iss, iucD, irp2, papC, tsh, vat, cva/cvi, stx2f, and bfp, as well as antimicrobial susceptibility to amoxicillin, enrofloxacin, and tetracycline (minimum inhibitory concentration [MIC] using E-tests) were investigated, in the examined E. coli strains. Results demonstrated that the greatest number of E. coli strains belonged to phylogenetic groups, B1 (86 strains-45.3%) and D (49 strains-25.8%), whereas 40 (21.0%) and only 15 (7.9%) isolates were classified as being of phylogenetic groups, A and B2, respectively. Among the 10 tested virulence-associated genes, 7 genes were detected in 61 examined strains (32.1%) with highly varying frequency. Virulence profiles showed that astA, iss, and irp2 genes were detected most frequently among all examined E. coli strains, isolated from every chosen bird species. Antimicrobial susceptibility, as detected by MIC for the examined antibiotics, is variable among strains isolated from different species of birds. The aim of this study was to examine the prevalence of E. coli strains, isolated from different species of wild waterbirds and determine their potential pathogenicity to the environment, other birds, and people.
Zhang, Song He; Lv, Xiaoyang; Han, Bing; Gu, Xiucong; Wang, Pei Fang; Wang, Chao; He, Zhenli
The rapid development of antibiotic-resistant bacteria (ARB) has been of concern worldwide. In this study, antibiotic resistance genes (ARGs) were investigated in antibiotic-resistant Escherichia coli isolated from surface water samples (rivers, n = 17; Taihu Lake, n = 16) and from human, chicken, swine, and Egretta garzetta sources in the Taihu Basin. E. coli showing resistance to at least five drugs occurred in 31, 67, 58, 27, and 18% of the isolates from surface water (n = 665), chicken (n = 27), swine (n = 29), human (n = 45), and E. garzetta (n = 15) sources, respectively. The mean multi-antibiotic resistance (MAR) index of surface water samples (0.44) was lower than that of chicken (0.64) and swine (0.57) sources but higher than that of human (0.30) and E. garzetta sources (0.15). Ten tetracycline, four sulfonamide, four quinolone, five β-lactamase, and two streptomycin resistance genes were detected in the corresponding antibiotic-resistant isolates. Most antibiotic-resistant E. coli harbored at least two similar functional ARGs. Int-I was detected in at least 57% of MAR E. coli isolates. The results of multiple correspondence analysis and Spearman correlation analysis suggest that antibiotic-resistant E. coli in water samples were mainly originated from swine, chicken, and/or human sources. Most of the ARGs detected in E. garzetta sources were prevalent in other sources. These data indicated that human activities may have contributed to the spread of ARB in the aquatic environment.
Murphy, Colleen P; Reid-Smith, Richard J; Boerlin, Patrick; Weese, J Scott; Prescott, John F; Janecko, Nicol; Hassard, Lori; McEwen, Scott A
Hospital-based infection control in veterinary medicine is emerging and the role of the environment in hospital-acquired infections (HAI) in veterinary hospitals is largely unknown. This study was initiated to determine the recovery of Escherichia coli and selected veterinary and zoonotic pathogens from the environments of 101 community veterinary hospitals. The proportion of hospitals with positive environmental swabs were: E. coli--92%, Clostridium difficile--58%, methicillin-resistant Staphylococcus aureus (MRSA)--9%, CMY-2 producing E. coli--9%, methicillin-resistant Staphylococcus pseudintermedius--7%, and Salmonella--2%. Vancomycin-resistant Enterococcus spp., canine parvovirus, and feline calicivirus were not isolated. Prevalence of antimicrobial resistance in E. coli isolates was low. Important potential veterinary and human pathogens were recovered including Canadian epidemic strains MRSA-2 and MRSA-5, and C. difficile ribotype 027. There is an environmental reservoir of pathogens in veterinary hospitals; therefore, additional studies are required to characterize risk factors associated with HAI in companion animals, including the role of the environment.
Samuel, Jastin; Paul, Madona Lien; Ravishankar, Harish; Mathur, Ankita; Saha, Dipti Priya; Natarajan, Chandrasekaran; Mukherjee, Amitava
In the current study, indigenous bacterial isolates Bacillus subtilis VITSUKMW1 and Escherichia coli VITSUKMW3 from a chromite mine were adapted to 100 mg L(-1) of Cr(VI). The phase contrast and scanning electron microscopic images showed increase in the length of adapted E. coli cells and chain formation in case of adapted B. subtilis. The presence of chromium on the surface of the bacteria was confirmed by energy dispersive X-ray spectroscopy (EDX), which was also supported by the conspicuous Cr-O peaks in FTIR spectra. The transmission electron microscopic (TEM) images of adapted E. coli and B. subtilis showed the presence of intact cells with Cr accumulated inside the bacteria. The TEM-EDX confirmed the internalization of Cr(VI) in the adapted cells. The specific growth rate and Cr(VI) reduction capacity was significantly higher in adapted B. subtilis compared to that of adapted E. coli. To study the possible role of Cr(VI) toxicity affecting the Cr(VI) reduction capacity, the definite assays for the released reactive oxygen species (ROS) and ROS scavenging enzymes (SOD and GSH) were carried out. The decreased ROS production as well as SOD and GSH release observed in adapted B. subtilis compared to the adapted E. coli corroborated well with its higher specific growth rate and increased Cr(VI) reduction capacity.
Jung, Yuna; Lim, Dongbin
We assessed heterologous protein expression in 64 strains obtained from the Escherichia coli Reference (ECOR) collection, a collection representing diverse natural E. coli populations. A plasmid generating a glutathione S-transferase and plant carbonic anhydrase fusion protein (GST-CA) under the control of the tac promoter was introduced into the ECOR strains, and the quantity of the fusion protein was determined by SDS-PAGE. The foreign protein was generated at various levels, from very high (40 strains, high producers) to very low (six strains, low producers). Immunoblotting showed that the high producers expressed approximately 250-500 times more GST-CA protein than the low producers. The results of semi-quantitative RT-PCR showed that the low producers generated mRNA levels comparable to those of the high producers, thereby suggesting that, at least in this case, inefficient translation is a major cause of the low production. We introduced a different plasmid, which expressed a maltose binding protein and plant guanylate kinase fusion protein (MBP-GK) into the six low producers. Interestingly, five of these expressed MBP-GK at very high levels. Thus, we conclude that the production of a particular protein from an expression vector can vary considerably, depending on the host strain. Strains in the ECOR collection could function as useful alternative hosts when a desired level of protein expression is not obtained from commonly used strains, such as E. coli K12 or B derivatives.
Alonso, C A; González-Barrio, D; Tenorio, Carmen; Ruiz-Fons, F; Torres, C
Eighty-nine Escherichia coli isolates recovered from faeces of red deer and small mammals, cohabiting the same area, were analyzed to determine the prevalence and mechanisms of antimicrobial resistance and molecular typing. Antimicrobial resistance was detected in 6.7% of isolates, with resistances to tetracycline and quinolones being the most common. An E. coli strain carrying blaCTX-M-1 as well as other antibiotic resistant genes included in an unusual class 1 integron (Intl1-dfrA16-blaPSE-1-aadA2-cmlA1-aadA1-qacH-IS440-sul3-orf1-mef(B)Δ-IS26) was isolated from a deer. The blaCTX-M-1 gene was transferred by conjugation and transconjugants also acquired an IncN plasmid. This strain was typed as ST224, which seems to be well adapted to both clinical and environmental settings. The phylogenetic distribution of the 89 strains varied depending on the animal host. This work reveals low antimicrobial resistance levels among faecal E. coli from wild mammals, which reflects a lower selective pressure affecting these bacteria, compared to livestock. However, it is remarkable the detection of a multi-resistant ESBL-E. coli with an integron carrying clinically relevant antibiotic-resistance genes, which can contribute to the dissemination of resistance determinants among different ecosystems.
Moser, A; Stephan, R; Corti, S; Lehner, A
Between March 2011 and February 2012 83 E. coli strains were isolated from mastitis milk samples from 83 different animals (67 farms) and tested for their sensitivity to various antibiotics by means of disk diffusion method and genotyped by determination of the phylogenetic groups as well as by pulsed field gel electrophoresis (PFGE). The antibiotics were chosen on the basis of their licenses for intramammary application in Switzerland. As many as 16.9 % of the isolates were resistant to one or more antimicrobial agents. Amoxicillin-clavulanic acid, gentamicin and third generation cephalosporins proved effective against the majority of these strains. Nevertheless, one blaCTX-M-14 harbouring extended-spectrum-beta-lactamase producing strain was found. Genetic analysis grouped most of the strains (87 %) into phylogenetic groups A and B1. PFGE genotyping demonstrated that E. coli from cows with mastitis even from the same farm were genotypically very diverse.
Godbout-DeLasalle, F; Higgins, R
Using the Analytab (API 20E) Enterobacteriaceae system of biochemical identification, a total of 506 Escherichia coli isolates from different animal species were coded numerically or biotyped. Fifty-four different biotypes were identified, 11 accounting for 83.1% of the isolates examined. Three of these profiles accounted for 65.3% of the isolates and were found in almost all animal species. Some of the biotypes were found in only one animal species: six in cattle, five in horses, 15 in pigs, two in sheep, two in birds, one in dogs and one in a porpoise. Biotypes, as determined here, could not be related to a particular pathology and more work is needed to assess the extent and significance of this relative biotype specificity among animal species. The use of other, more sophisticated, typing systems, i.e. plasmid "fingerprinting", or restriction endonuclease analysis of chromosomal DNA, would have to be investigated. PMID:3527394
Honda, M; Kodama, T; Ishida, Y; Takei, K; Matsuda, M
Crossed-field gel electrophoresis was used to analyse the chromosomal genomic DNA from 41 clinical isolates of Escherichia coli serotypes O-2, O-4, O-6 and O-75 from patients with lower urinary tract infections after digestion of the DNA with Not I and Xba I, respectively. Not I generated 28 distinctly different genomic profiles after crossed-field gel electrophoresis (CFGE) from 34 strains and Xba I generated 35 distinctly different genomic profiles from 41 strains. Not I failed to cleave the genomic DNA from one strain of E. coli serotype O-6 and from all of six strains of serotype O-75. Both enzymes were shown to have equal potential utility for detection of variability in CFGE profiles among 34 strains because both classified 34 of the 41 strains into 28 profiles in a similar manner. No examples of essentially identical restriction profiles among strains of different serotypes were obtained.
Snesrud, Erik; Ong, Ana C; Corey, Brendan; Kwak, Yoon I; Clifford, Robert; Gleeson, Todd; Wood, Shannon; Whitman, Timothy J; Lesho, Emil P; Hinkle, Mary; Mc Gann, Patrick
The emergence of the transferable colistin resistance gene mcr-1 is of global concern. The insertion sequence (IS) Apl1 is a key component in the mobilization of this gene, but its role remains poorly understood.Six Escherichia coli were cultured from the same patient over one month in Germany and the USA after a brief hospitalization in Bahrain for an unconnected illness. Four carried mcr-1 by real-time PCR, but two were negative. Two additional mcr-1-negative E. coli were collected during follow-up surveillance nine months later. All isolates were analyzed with whole genome sequencing (WGS).WGS revealed that the six initial isolates were composed of two distinct strains; an initial ST-617 E.coli harboring mcr-1and a second, unrelated mcr-1-negative ST-32 E. coli that emerged 2 weeks after hospitalization. Follow-up swabs 9 months later were negative for the ST-617 strain, but the mcr-1-negative ST-32 strain was still present. Mcr-1 was associated with a single copy of ISApl1 located on a 64.5 Kb IncI2 plasmid that shared >95% homology to other mcr-1 IncI2 plasmids. ISApl1 copy number ranged from 2 in the first isolate to 6 in the final isolate, but movement was independent of mcr-1 Some movement was accompanied by gene disruption, including the loss of genes encoding proteins involved in stress response, arginine catabolism and L-arabinose utilization.These data represent the first comprehensive analysis of ISApl1 movement in serial clinical isolates and reveals that under certain conditions ISApl1 is a highly active IS element whose movement may be detrimental to the host cell.
Sahoo, Krushna Chandra; Tamhankar, Ashok J; Sahoo, Soumyakanta; Sahu, Priyadarshi Soumyaranjan; Klintz, Senia Rosales; Lundborg, Cecilia Stålsby
Little information is available on relationships between the biophysical environment and antibiotic resistance. This study was conducted to investigate the antibiotic resistance pattern of Escherichia coli isolated from child stool samples, cow-dung and drinking water from the non-coastal (230 households) and coastal (187 households) regions of Odisha, India. Susceptibility testing of E. coli isolates (n = 696) to the following antibiotics: tetracycline, ampicillin/sulbactam, cefuroxime, cefotaxime, cefixime, cotrimoxazole, amikacin, ciprofloxacin, norfloxacin and nalidixic acid was performed by the disk diffusion method. Ciprofloxacin minimum inhibitory concentration (MIC) values were determined for ciprofloxacin-resistant isolates (n = 83). Resistance to at least one antibiotic was detected in 90% or more of the E. coli isolates. Ciprofloxacin MIC values ranged from 8 to 32 µg/mL. The odds ratio (OR) of resistance in E. coli isolates from children's stool (OR = 3.1, 95% CI 1.18-8.01), cow-dung (OR = 3.6, 95% CI 1.59-8.03, P = 0.002) and drinking water (OR = 3.8, 95% CI 1.00-14.44, P = 0.049) were higher in non-coastal compared to coastal region. Similarly, the co-resistance in cow-dung (OR = 2.5, 95% CI 1.39-4.37, P = 0.002) and drinking water (OR = 3.2, 95% CI 1.36-7.41, P = 0.008) as well as the multi-resistance in cow-dung (OR = 2.2, 95% CI 1.12-4.34, P = 0.022) and drinking water (OR = 2.7, 95% CI 1.06-7.07, P = 0.036) were also higher in the non-coastal compared to the coastal region.
Mainali, Chunu; McFall, Margaret; King, Robin; Irwin, Rebecca
This study was conducted to investigate antimicrobial resistance profiles of Escherichia coli isolates from broiler chickens in Alberta, Canada. Cecal contents of broiler chickens from 24 flocks were collected at slaughter between January and March 2005 for culture and antimicrobial susceptibility testing against a panel of 15 antimicrobials using a broth microdilution technique. Of 600 E. coli isolates tested, 475 (79.2%) were resistant to one or more antimicrobials, 326 (54.3%) were resistant to three or more antimicrobials, 65 (10.8%) were resistant to five or more antimicrobials, and 15 (2.5%) were resistant to seven or more antimicrobials. The most common resistance was to tetracycline (69.2%), followed by streptomycin (48.2%), kanamycin (40.3%), and sulfisoxazole (38.0%). None of the E. coli isolates were resistant to amikacin, ceftriaxone, or ciprofloxacin. Of the isolates that were resistant to two or more antimicrobials, the most common multidrug resistance patterns were streptomycinte-tracycline (44.0%), streptomycin-sulfisoxazole-tetracycline (30.7%), and kanamycin-streptomycin-sulfisoxazole-tetracycline (23.5%). Resistance to tetracycline and kanamycin (odds ratio = 46.7, P = 0.0001) was highly associated, followed by resistance to streptomycin and sulfisoxazole (odds ratio = 12.0, P = 0.0001), and streptomycin and tetracycline (odds ratio = 10.3, P = 0.0001). The flock level prevalence of resistance varied from 16.7% for chloramphenicol to 100.0% for ampicillin, streptomycin, sulfisoxazole, and tetracycline. The results of this study provided baseline information on antimicrobial susceptibility of E. coli isolates of broiler chickens at slaughter in Alberta, which can serve as a bench mark for future research.
Huang, Ying; Ogutu, James O.; Gu, Jiarui; Ding, Fengshu; You, Yuhong; Huo, Yan; Zhao, Hong; Li, Wenjing; Zhang, Zhiwei; Zhang, Wenli; Chen, Xiaobei; Fu, Yingmei; Zhang, Fengmin
The objective of this study was to compare quinolone resistance and gyrA mutations in clinical isolates of Klebsiella pneumoniae and Escherichia coli from Chinese adults who used quinolone in the preceding month and children without any known history of quinolone administration. The antimicrobial susceptibilities of 61 isolates from children and 79 isolates from adults were determined. The mutations in the quinolone resistance-determining regions in gyrA gene were detected by PCR and DNA sequencing. Fluoroquinolone resistance and types of gyrA mutations in isolates from children and adults were compared and statistically analyzed. No significant differences were detected in the resistance rates of ciprofloxacin and levofloxacin between children and adults among isolates of the two species (all P > 0.05). The double mutation Ser83→Leu + Asp87→Asn in the ciprofloxacin-resistant isolates occurred in 73.7% isolates from the children and 67.9% from the adults, respectively (P = 0.5444). Children with no known history of quinolone administration were found to carry fluoroquinolone-resistant Enterobacteriaceae isolates. The occurrence of ciprofloxacin resistance and the major types of gyrA mutations in the isolates from the children were similar to those from adults. The results indicate that precautions should be taken on environmental issues resulting from widespread transmission of quinolone resistance. PMID:25756041
Huang, Ying; Ogutu, James O; Gu, Jiarui; Ding, Fengshu; You, Yuhong; Huo, Yan; Zhao, Hong; Li, Wenjing; Zhang, Zhiwei; Zhang, Wenli; Chen, Xiaobei; Fu, Yingmei; Zhang, Fengmin
The objective of this study was to compare quinolone resistance and gyrA mutations in clinical isolates of Klebsiella pneumoniae and Escherichia coli from Chinese adults who used quinolone in the preceding month and children without any known history of quinolone administration. The antimicrobial susceptibilities of 61 isolates from children and 79 isolates from adults were determined. The mutations in the quinolone resistance-determining regions in gyrA gene were detected by PCR and DNA sequencing. Fluoroquinolone resistance and types of gyrA mutations in isolates from children and adults were compared and statistically analyzed. No significant differences were detected in the resistance rates of ciprofloxacin and levofloxacin between children and adults among isolates of the two species (all P > 0.05). The double mutation Ser83→Leu + Asp87→Asn in the ciprofloxacin-resistant isolates occurred in 73.7% isolates from the children and 67.9% from the adults, respectively (P = 0.5444). Children with no known history of quinolone administration were found to carry fluoroquinolone-resistant Enterobacteriaceae isolates. The occurrence of ciprofloxacin resistance and the major types of gyrA mutations in the isolates from the children were similar to those from adults. The results indicate that precautions should be taken on environmental issues resulting from widespread transmission of quinolone resistance.
Oh, Kyung-Hwan; Kim, Dong Wook; Jung, Su-Mi; Cho, Seung-Hak
Enterotoxigenic Escherichia coli (ETEC) is one of the major causes of infectious diarrhea in developing countries. In order to characterize the molecular features of human ETEC isolates from Korea, we investigated the profiles of enterotoxin and colonization factor (CF) genes by polymerase chain reaction (PCR) and performed multilocus sequence typing (MLST) with a total of 291 ETEC strains. The specimens comprised 258 domestic strains isolated from patients who had diarrhea and were from widely separated geographic regions in Korea and 33 inflow strains isolated from travelers visiting other Asian countries. Heat-stable toxin (STh)-possessing ETEC strains were more frequent than heat-labile toxin (LT)-possessing ETEC strains in the domestic isolates, while the detection rates of both enterotoxin genes were similar in the inflow isolates. The profile of CF genes of domestic isolates was similar to that of inflow isolates and the major CF types of the strains were CS3-CS21-CS1/PCF071 and CS2-CS3-CS21. Most of these 2 CF types were detected in ETEC strains that possess both lt and sth genes. The major MLSTST types of domestic isolates were ST171 and ST955. Moreover, the 2 major CF types were usually found concomitantly with the 2 major MLST STs, ST171 and ST955. In conclusion, our genotyping results may provide useful information for guiding the development of geographically specific vaccines against human ETEC isolates. PMID:24841334
Ríos, Esther; López, Maria Carmen; Rodríguez-Avial, Iciar; Pena, Irene; Picazo, Juan Jose
The aim of present work was to characterize the inhibitor-resistant TEM (IRT) β-lactamases produced by Escherichia coli in Hospital Clínico San Carlos (Madrid, Spain). Mechanisms of fluoroquinolone resistance among IRT-producing strains were also studied. Isolates with susceptibility to cephalosporins and amoxicillin-clavulanate (AMC) resistance were collected in our hospital (November 2011-July 2012) from both outpatients and hospitalized patients. Among 70 AMC-resistant E. coli strains, 28 (40%) produced IRT enzymes. Most of them were uropathogens (82.1%) and recovered from outpatients (75%). Seven different IRT enzymes were identified with TEM-30 (IRT-2) being the most prevalent, followed by TEM-40 (IRT-11). A high rate of ciprofloxacin resistance was found among IRT-producing strains (50%). Most of the ciprofloxacin-resistant isolates showed ciprofloxacin minimum inhibitory concentration >32 mg/L and contained two mutations in both gyrA and parC genes. Four IRT enzyme producers harbored the qnr gene. ST131 clone was mainly responsible for both IRT enzyme production and ciprofloxacin resistance. In conclusion, data from this study show that the frequency of IRT producers was 40% and a high rate of ciprofloxacin resistance was found among IRT-producing isolates. Current and future actions should be taken into account to avoid or reduce the development of AMC and fluoroquinolone resistance in E. coli.
Dias, Diana; Torres, Rita T; Kronvall, Göran; Fonseca, Carlos; Mendo, Sónia; Caetano, Tânia
Antibiotic resistance is an emerging global problem. Wild animals are rarely exposed to antibiotics and therefore low levels of antibiotic resistance are expected. However, the growing interactions of these animals with humans and livestock may have a huge impact on their bacterial flora. This study aimed to assess the levels of antibiotic resistance in Escherichia coli isolated from widespread wild ungulates in Portugal. The interpretation of inhibition zone diameters was performed according to clinical breakpoints and epidemiological cut-offs, determined with the normalized resistance interpretation (NRI) method. For clinical breakpoints, 16% of the isolates were resistant to at least one antibiotic, including ampicillin (10%), tetracycline (9%), streptomycin (5%) co-trimoxazole (4%), amoxicillin/clavulanic acid (1%) and cefoxitin (1%). The levels of resistance detected in E. coli strains isolated from wild boar were statistically different for ampicillin and co-trimoxasol. According to NRI cut-offs, 10% of the population showed a non-wild-type phenotype against at least one antibiotic, also including tetracycline (9%), co-trimoxazole (6%), streptomycin (4%), ampicillin (2%) and amoxicillin/clavulanic acid (1%). Considering this parameter of comparison, no statistically different levels of resistance were identified between E. coli recovered from the three wild ungulates. Screening of Salmonella spp., which can be potentially pathogenic, was also performed, revealing that its prevalence was very low (1.5%). The study demonstrated that wild ungulates from Portugal are also reservoirs of antibiotic-resistant bacteria.
Lu, Zhongjing; Breidt, Fred
A novel phage, Φ241, specific for Escherichia coli O157:H7 was isolated from an industrial cucumber fermentation where both acidity (pH ≤ 3.7) and salinity (≥5% NaCl) were high. The phage belongs to the Myoviridae family. Its latent period was 15 min and average burst size was 53 phage particles per infected cell. The phage was able to lyse 48 E. coli O157:H7 strains, but none of the 18 non-O157 strains (including E. coli O104:H7) or the 2 O antigen-negative mutants of O157:H7 strain, 43895Δper (also lacking H7 antigen) and F12 (still expressing H7 antigen). However, the phage was able to lyse a per-complemented strain (43895ΔperComp) which expresses O157 antigen. These results indicated that phage Φ241 is specific for O157 antigen, and E. coli strains lacking O157 antigen were resistant to the phage infection, regardless of the presence or absence of H7 antigen. SDS-PAGE profile revealed at least 13 structural proteins of the phage. The phage DNA was resistant to many commonly used restriction endonucleases, suggesting the presence of modified nucleotides in the phage genome. At the multiplicity of infection of 10, 3, or 0.3, the phage caused a rapid cell lysis within 1 or 2 h, resulting in 3.5- or 4.5-log-unit reduction in cell concentration. The high lytic activity, specificity and tolerance to low pH and high salinity make phage Φ241 a potentially ideal biocontrol agent of E. coli O157:H7 in various foods. To our knowledge, this is the first report on E. coli O157:H7 phage isolated from high acidity and salinity environment.
Lu, Zhongjing; Breidt, Fred
A novel phage, Φ241, specific for Escherichia coli O157:H7 was isolated from an industrial cucumber fermentation where both acidity (pH ≤ 3.7) and salinity (≥5% NaCl) were high. The phage belongs to the Myoviridae family. Its latent period was 15 min and average burst size was 53 phage particles per infected cell. The phage was able to lyse 48 E. coli O157:H7 strains, but none of the 18 non-O157 strains (including E. coli O104:H7) or the 2 O antigen-negative mutants of O157:H7 strain, 43895Δper (also lacking H7 antigen) and F12 (still expressing H7 antigen). However, the phage was able to lyse a per-complemented strain (43895ΔperComp) which expresses O157 antigen. These results indicated that phage Φ241 is specific for O157 antigen, and E. coli strains lacking O157 antigen were resistant to the phage infection, regardless of the presence or absence of H7 antigen. SDS-PAGE profile revealed at least 13 structural proteins of the phage. The phage DNA was resistant to many commonly used restriction endonucleases, suggesting the presence of modified nucleotides in the phage genome. At the multiplicity of infection of 10, 3, or 0.3, the phage caused a rapid cell lysis within 1 or 2 h, resulting in 3.5- or 4.5-log-unit reduction in cell concentration. The high lytic activity, specificity and tolerance to low pH and high salinity make phage Φ241 a potentially ideal biocontrol agent of E. coli O157:H7 in various foods. To our knowledge, this is the first report on E. coli O157:H7 phage isolated from high acidity and salinity environment. PMID:25741324
Tuckman, Margareta; Petersen, Peter J; Howe, Anita Y M; Orlowski, Mark; Mullen, Stanley; Chan, Karen; Bradford, Patricia A; Jones, C Hal
Tigecycline, a member of the glycylcycline class of antibiotics, was designed to maintain the antibacterial spectrum of the tetracyclines while overcoming the classic mechanisms of tetracycline resistance. The current study was designed to monitor the prevalence of the tet(A), tet(B), tet(C), tet(D), tet(E), and tet(M) resistance determinants in Escherichia coli isolates collected during the worldwide tigecycline phase 3 clinical trials. A subset of strains were also screened for the tet(G), tet(K), tet(L), and tet(Y) genes. Of the 1,680 E. coli clinical isolates screened for resistance to classical tetracyclines, 405 (24%) were minocycline resistant (MIC > or = 8 microg/ml) and 248 (15%) were tetracycline resistant (MIC > or = 8 microg/ml) but susceptible to minocycline (MIC < or = 4 microg/ml). A total of 452 tetracycline-resistant, nonduplicate isolates were positive by PCR for at least one of the six tetracycline resistance determinants examined. Over half of the isolates encoding a single determinant were positive for tet(A) (26%) or tet(B) (32%) with tet(C), tet(D), tet(E), and tet(M), collectively, found in 4% of isolates. Approximately 33% of the isolates were positive for more than one resistance determinant, with the tet(B) plus tet(E) combination the most highly represented, found in 11% of isolates. The susceptibilities of the tetracycline-resistant strains to tigecycline (MIC(90), 0.5 microg/ml), regardless of the encoded tet determinant(s), were comparable to the tigecycline susceptibility of tetracycline-susceptible strains (MIC(90), 0.5 microg/ml). The results provide a current (2002 to 2006) picture of the distribution of common tetracycline resistance determinants encoded in a globally sourced collection of clinical E. coli strains.
Tuckman, Margareta; Petersen, Peter J.; Howe, Anita Y. M.; Orlowski, Mark; Mullen, Stanley; Chan, Karen; Bradford, Patricia A.; Jones, C. Hal
Tigecycline, a member of the glycylcycline class of antibiotics, was designed to maintain the antibacterial spectrum of the tetracyclines while overcoming the classic mechanisms of tetracycline resistance. The current study was designed to monitor the prevalence of the tet(A), tet(B), tet(C), tet(D), tet(E), and tet(M) resistance determinants in Escherichia coli isolates collected during the worldwide tigecycline phase 3 clinical trials. A subset of strains were also screened for the tet(G), tet(K), tet(L), and tet(Y) genes. Of the 1,680 E. coli clinical isolates screened for resistance to classical tetracyclines, 405 (24%) were minocycline resistant (MIC ≥ 8 μg/ml) and 248 (15%) were tetracycline resistant (MIC ≥ 8 μg/ml) but susceptible to minocycline (MIC ≤ 4 μg/ml). A total of 452 tetracycline-resistant, nonduplicate isolates were positive by PCR for at least one of the six tetracycline resistance determinants examined. Over half of the isolates encoding a single determinant were positive for tet(A) (26%) or tet(B) (32%) with tet(C), tet(D), tet(E), and tet(M), collectively, found in 4% of isolates. Approximately 33% of the isolates were positive for more than one resistance determinant, with the tet(B) plus tet(E) combination the most highly represented, found in 11% of isolates. The susceptibilities of the tetracycline-resistant strains to tigecycline (MIC90, 0.5 μg/ml), regardless of the encoded tet determinant(s), were comparable to the tigecycline susceptibility of tetracycline-susceptible strains (MIC90, 0.5 μg/ml). The results provide a current (2002 to 2006) picture of the distribution of common tetracycline resistance determinants encoded in a globally sourced collection of clinical E. coli strains. PMID:17620376
Briñas, Laura; Moreno, Miguel Angel; Zarazaga, Myriam; Porrero, Concepción; Sáenz, Yolanda; García, María; Dominguez, Lucas; Torres, Carmen
Genes encoding the CMY-2, CTX-M-14, and SHV-12 beta-lactamases were detected in three of five Escherichia coli isolates from fecal samples from healthy chickens which showed resistance or diminished susceptibility to extended-spectrum cephalosporins. A -42 mutation at the promoter region of the ampC gene was detected in the other two isolates.
Briñas, Laura; Moreno, Miguel Angel; Zarazaga, Myriam; Porrero, Concepción; Sáenz, Yolanda; García, María; Dominguez, Lucas; Torres, Carmen
Genes encoding the CMY-2, CTX-M-14, and SHV-12 β-lactamases were detected in three of five Escherichia coli isolates from fecal samples from healthy chickens which showed resistance or diminished susceptibility to extended-spectrum cephalosporins. A −42 mutation at the promoter region of the ampC gene was detected in the other two isolates. PMID:12760899
Yanat, Betitera; Dali Yahia, Radia; Yazi, Leila; Machuca, Jesús; Díaz-De-Alba, Paula; Touati, Abdelaziz; Pascual, Álvaro; Rodríguez-Martínez, José-Manuel
QepA is a plasmid-mediated quinolone resistance determinant of low prevalence described worldwide, mainly in Enterobacteriaceae. This study describes, for the first time in Algeria, two clonally related, QepA-producing Escherichia coli clinical isolates positive for CTX-M-15. The clonal spread of these multidrug-resistant isolates is a major public health concern.
Bayles, Darrell O.; Alt, David P.; Looft, Torey
Escherichia coli O157:H7 strain 86-24 does not produce curli fimbriae, but gives rise to curli-positive isolates at a variable frequency. Here, we report the complete genome sequences of curli-negative and curli-positive isolates of strain 86-24. PMID:27979932
Van, Thi Thu Hao; Chin, James; Chapman, Toni; Tran, Linh Thuoc; Coloe, Peter J
This study was conducted to examine a current baseline profile of antimicrobial resistance and virulence of Escherichia coli isolated from foods commonly sold in the market place in Vietnam. E. coli were isolated from 180 samples of raw meat, poultry and shellfish and also isolated from 43 chicken faeces samples. Ninety-nine E. coli isolates recovered from all sources were selected for the investigation of their susceptibility to 15 antimicrobial agents by the disk diffusion method. Eighty-four percent of the isolates were resistant to one or more antibiotics, and multi-resistance, defined as resistance to at least 3 different classes of antibiotics, was detected in all sources. The rates of multi-resistance were up to 89.5% in chicken, 95% in chicken faeces and 75% in pork isolates. Resistance was most frequently observed to tetracycline (77.8%), sulfafurazole (60.6%), ampicillin (50.5%), amoxicillin (50.5%), trimethoprim (51.5%), chloramphenicol (43.4%), streptomycin (39.4%), nalidixic acid (34.3%) and gentamicin (24.2%). In addition, the isolates also displayed resistance to fluoroquinolones (ciprofloxacin 16.2%, norfloxacin 17.2%, and enrofloxacin 21.2%), with chicken isolates showing the highest rates of resistance to these antibiotics (52.6-63.2%). Thirty-eight multi-resistant isolates were selected for further the examination of antibiotic resistance genes and were also evaluated for virulence gene profiles by multiplex and uniplex polymerase chain reaction. The beta-lactam TEM gene and tetracycline resistance tetA, tetB genes were frequently detected in the tested isolates (84.2% and 89.5% respectively). Genes which are responsible for resistance to streptomycin (aadA) (68.4%), chloramphenicol (cmlA) (42.1%), sulfonamides (sulI) (39.5%), trimethoprim (dhfrV) (26.3%) and kanamycin (aphA-1) (23.7%) were also widely distributed. Plasmid-mediated ampC genes were detected in E. coli isolates from chicken and pork. The isolates were tested for the presence of 58
Canal, Natália; Meneghetti, Karine Lena; de Almeida, Clara Ponzi; da Rosa Bastos, Marina; Otton, Letícia Muner; Corção, Gertrudes
Fecal bacteria are considered to be a potential reservoir of antimicrobial resistance genes in the aquatic environment and could horizontally transfer these genes to autochthonous bacteria when carried on transferable and/or mobile genetic elements. Such circulation of resistance genes constitutes a latent public health hazard. The aim of this study was to characterize the variable region of the class 1 integron and relate its genetic content to resistance patterns observed in antimicrobial-resistant Escherichia coli isolated from the surface waters of Patos Lagoon, Southern Brazil. Genetic diversity of the isolates and presence of the qacEΔ1 gene, which confers resistance to quaternary ammonium compounds, were also investigated. A total of 27 isolates were analyzed. The variable region harbored dfrA17, dfrA1 and dfrA12 genes, which confer resistance to trimethoprim, and aadA1, aadA5 and aadA22 genes that encode resistance to streptomycin/spectinomycin. Most of the isolates were considered resistant to quaternary ammonium compounds and all of them carried the qacEΔ1 gene at the 3' conserved segment of the integron. ERIC-PCR analyses of E. coli isolates that presented the integrons showed great genetic diversity, indicating diverse sources of contamination in this environment. These results suggest that fecal bacteria with class 1 integrons in aquatic environments are potentially important reservoirs of antibiotic-resistance genes and may transfer these elements to other bacteria that are capable of infecting humans.
Zhang, Rui-Quan; Ying, Guang-Guo; Su, Hao-Chang; Zhou, Li-Jun; Liu, You-Sheng
This study investigated antibiotic resistance profiles including antibiotic resistance frequencies, resistance genes and resistance patterns in Escherichia coli strains isolated from traditional and integrated aquaculture systems in South China by using antibiotic susceptibility testing and real time polymerase chain reaction (PCR) technique. The E. coli isolates were found to be resistant to at least one antibiotic among 12 antibiotics. Higher resistance frequencies to ampicillin, sulfamethoxazole, trimethoprime, streptomycin and tetracycline were found compared to the rest antibiotics. Among the 10 tetracycline resistance genes detected in the resistant isolates, the most prevalent tetracycline resistance genes were tetA, tetW and tetB with the frequency of 69.7%, 63.5% and 21.9%, respectively. Three sulfonamide resistance genes were detected in these resistant isolates, with their detection frequencies in the following order: sul2 (55.3%) > sul3 (28.2%) > sul1 (6.2%). Four resistance genes mainly encoding extended-spectrum β-lactamases (ESBLs) were detected in these resistant isolates, with the detection frequencies of blaTEM (28.4%) > blaOXA (9.7%) > blaCTX (9.3%) > blaCARB (5.2%) > blaSHV (0.0%). It was found that the integrated aquaculture system exhibited generally higher prevalence of antibiotic resistance than the traditional aquaculture system. An integrated aquaculture system could facilitate development of bacterial resistance and spread of the antibiotic resistance genes, and consequently become an important reservoir of resistance genes.
Wang, Rong; Kalchayanand, Norasak; King, David A; Luedtke, Brandon E; Bosilevac, Joseph M; Arthur, Terrance M
In the meat industry, a "high event period" (HEP) is defined as a time period during which commercial meat plants experience a higher than usual rate of Escherichia coli O157:H7 contamination. Genetic analysis indicated that within a HEP, most of the E. coli O157:H7 strains belong to a singular dominant strain type. This was in disagreement with the current beef contamination model stating that contamination occurs when incoming pathogen load on animal hides, which consists of diverse strain types of E. coli O157:H7, exceeds the intervention capacity. Thus, we hypothesize that the HEP contamination may be due to certain in-plant colonized E. coli O157:H7 strains that are better able to survive sanitization through biofilm formation. To test our hypothesis, a collection of 45 E. coli O157:H7 strains isolated from HEP beef contamination incidents and a panel of 47 E. coli O157:H7 strains of diverse genetic backgrounds were compared for biofilm formation and sanitizer resistance. Biofilm formation was tested on 96-well polystyrene plates for 1 to 6 days. Biofilm cell survival and recovery growth after sanitization were compared between the two strain collections using common sanitizers, including quaternary ammonium chloride, chlorine, and sodium chlorite. No difference in "early stage" biofilms was observed between the two strain collections after incubation at 22 to 25°C for 1 or 2 days. However, the HEP strains demonstrated significantly higher potency of "mature" biofilm formation after incubation for 4 to 6 days. Biofilms of the HEP strains also exhibited significantly stronger resistance to sanitization. These data suggest that biofilm formation and sanitization resistance could have a role in HEP beef contamination by E. coli O157:H7, which highlights the importance of proper and complete sanitization of food contact surfaces and food processing equipment in commercial meat plants.
Caballero, Moisés; Rivera, Isabel; Jara, Luis M; Ulloa-Stanojlovic, Francisco M; Shiva, Carlos
Feral pigeons (Columbia livia) live in close contact with humans and other animals. They can transmit potentially pathogenic and zoonotic agents. The objective of this study was to isolate and detect strains of diarrheagenic Escherichia coli and Campylobacter jejuni of urban feral pigeons from an area of Lima, Peru. Fresh dropping samples from urban parks were collected for microbiological isolation of E. coli strains in selective agar, and Campylobacter by filtration method. Molecular identification of diarrheagenic pathotypes of E.coli and Campylobacter jejuni was performed by PCR. Twenty-two parks were sampled and 16 colonies of Campylobacter spp. were isolated. The 100% of isolates were identified as Campylobacter jejuni. Furthermore, 102 colonies of E. coli were isolated and the 5.88% resulted as Enteropathogenic (EPEC) type and 0.98% as Shiga toxin-producing E. coli (STEC). The urban feral pigeons of Lima in Peru can act as a reservoir or carriers of zoonotic potentially pathogenic enteric agents.
Wegner, Anna S; Wintraecken, Kathelijne; Spurio, Roberto; Woldringh, Conrad L; de Vries, Renko; Odijk, Theo
Escherichia coli nucleoids were compacted by the inert polymer polyethylene glycol (PEG) in the presence of the H-NS protein. The protein by itself appears to have little impact on the size of the nucleoids as determined by fluorescent microscopy. However, it has a significant impact on the nucleoidal collapse by PEG. This is quantitatively explained by assuming the H-NS protein enhances the effective diameter of the DNA helix leading to an increase in the depletion forces induced by the PEG. Ultimately, however, the free energy of the nucleoid itself turns out to be independent of the H-NS concentration. This is because the enhancement of the supercoil excluded volume is negligible. The experiments on the nucleoids are corroborated by dynamic light scattering and EMSA analyses performed on DNA plasmids in the presence of PEG and H-NS.
Hopkins, Katie L; Mushtaq, Shazad; Richardson, Judith F; Doumith, Michel; de Pinna, Elizabeth; Cheasty, Tom; Wain, John; Livermore, David M; Woodford, Neil
Rifaximin is licensed in the EU and USA for treating travellers' diarrhoea caused by non-invasive bacteria. Selection for resistance mechanisms of public health significance might occur if these are linked to rifamycin resistance. Rifaximin MICs were determined by agar dilution for 90 isolates each of Escherichia coli, Shigella spp., nontyphoidal Salmonella enterica, typhoidal S. enterica and Campylobacter spp., an additional 60 E. coli with CTX-M ESBLs isolated from patients with travellers' diarrhoea, and 30 non-diarrhoeal carbapenemase-producing E. coli. Comparators were rifampicin, ciprofloxacin, azithromycin, trimethoprim/sulfamethoxazole and doxycycline. Isolates with rifaximin MICs>32 mg/L were screened for arr genes, and critical rpoB regions were sequenced. Rifaximin was active at ≤32 mg/L against 436/450 (96.9%) diverse Enterobacteriaceae, whereas 81/90 (90%) Campylobacter spp. were resistant to rifaximin at ≥128 mg/L. Rifaximin MICs were ≥128 mg/L for two Shigella and five MDR E. coli producing NDM (n = 3), OXA-48 (n = 1) or CTX-M-15 (n = 1). Two of the five MDR E. coli had plasmids harbouring arr-2 together with bla(NDM), and two (one each with bla(NDM) and bla(CTX-M-15)) had His526Asn substitutions in RpoB. The rifamycin resistance mechanism remained undefined in one MDR E. coli isolate (with bla(OXA-48)) and the two Shigella isolates. Rifaximin showed good in vitro activity against diverse Enterobacteriaceae but was largely inactive against Campylobacter spp. Rifaximin has potential to co-select MDR E. coli in the gut flora, but much stronger associations were seen between ESBL and/or carbapenemase production and resistance to alternative treatments for travellers' diarrhoea, notably ciprofloxacin and azithromycin.
Cao, Xiaoli; Cavaco, Lina M.; Lv, Yuan; Li, Yun; Zheng, Bo; Wang, Pengyuan; Hasman, Henrik; Liu, Yucun; Aarestrup, Frank M.
A total of 222 urinary Escherichia coli isolates from 20 tertiary hospitals in 15 different provinces and 4 municipalities in mainland China were characterized by antimicrobial susceptibility, phylogrouping, and the presence of plasmid-mediated quinolone resistance genes. A subset of 138 suspected extended-spectrum cephalosporinase (ESC) producers were examined for genes encoding cephalosporin resistance. Forty-three isolates harboring blaCTX-M-14 or blaCTX-M-15 were analyzed by pulsed-field gel electrophoresis (PFGE), and plasmids containing these genes were typed using PCR-based replicon typing (PBRT). Thirteen phylogroup B2 blaCTX-M-14- and blaCTX-M-15-positive isolates were analyzed by multilocus sequence typing (MLST). A frequent occurrence of resistance (>46%) was observed toward cephalosporins, gentamicin, and fluoroquinolones. Among the 222 isolates, 4 qnrS1, 4 qepA, and 16 aac(6′)-Ib-cr genes were confirmed. Four major phylogroups (A, B1, B2, and D) and nontypeable isolates (NTs) were found among the isolates, with phylogroup D (54%) being the most common phylogroup. A total of 110 (80%) of the 138 screened isolates harbored blaCTX-M genes, with blaCTX-M-14 (71%) and blaCTX-M-15 (24%) being the most prevalent of these genes. Nine of the 13 CTX-M-15- or CTX-M-14-containing B2 isolates belonged to ST131. PFGE typing showed a high level of diversity, and plasmid analysis indicated a very large pool of different resistance plasmids mediating the spread of blaCTX-M genes in mainland China. An equally very high frequency of resistance and equally high levels of diversity in phylogroups, PFGE types, and plasmids were observed among community- and hospital-acquired E. coli isolates, indicating the presence of a large reservoir in the community and a long-term spread of cephalosporin resistance in China. PMID:21525216
Ode, Takashi; Saito, Ryoichi; Kumita, Wakako; Sato, Kenya; Okugawa, Shu; Moriya, Kyoji; Koike, Kazuhiko; Okamura, Noboru
This study investigated the relationship of plasmid-mediated quinolone resistance (PMQR) and aminoglycoside resistance among oxyimino-cephalosporin-resistant Escherichia coli (n=46) and Klebsiella oxytoca (n=28) clinical isolates in Japan. Seventy-three isolates appeared to produce an extended-spectrum beta-lactamase (ESBL) and one K. oxytoca isolate produced IMP-1 metallo-beta-lactamase (MBL). Polymerase chain reaction (PCR) and sequencing confirmed that eight CTX-M-9/SHV-12-producing isolates, one IMP-1-producing K. oxytoca isolate, and six ESBL-positive E. coli isolates respectively possessed PMQR genes qnrA1, qnrB6, and aac(6')-Ib-cr. All qnr-positive isolates also carried either aac(6')-Ib or aac(6')-IIc aminoglycoside acetyltransferase genes. Resistance determinants to beta-lactams, quinolones and aminoglycosides were co-transferred with a plasmid of ca. 140 kb. The qnrA1 gene was located downstream of insertion sequence ISCR1 in complex class 1 integrons. A novel qnrA1-carrying class 1 integron with the cassette arrangement aac(6')-IIc-aadA2 as well as a unique class 1 integron with bla(IMP-1)-aac(6')-IIc cassettes on the plasmid carrying qnrB6 were found in K. oxytoca isolates. We describe the identification of qnrB6 and aac(6')-Ib-cr and the close association of qnr with aac(6')-Ib and aac(6')-IIc for the first time in clinical isolates producing ESBL or MBL in Japan.
Oh, Jae-Young; Kwon, Yong-Kuk; Tamang, Migma Dorji; Jang, Hyung-Kwan; Jeong, Ok-Mi; Lee, Hee-Soo; Kang, Min-Su
A total of 2,423 nonduplicate isolates of Escherichia coli recovered from wild birds (n=793) and chickens (n=1,630) in South Korea were investigated for plasmid-mediated quinolone resistance (PMQR) genes. Altogether, 56 isolates with PMQR genes were identified, including 25 (3.2%) from wild birds and 31 (1.9%) from chickens, which were further characterized using molecular methods. Among them, qnrS, aac(6')-Ib-cr, qnrB, and qepA genes were detected in 47 (1.9%), 6 (0.24%), 2 (0.08%), and 1 (0.04%) isolates, respectively. The most prevalent gene, qnrS, was identified in 21 (0.9%) and 26 (1.1%) isolates from wild birds and chickens, respectively. The qnrB gene was identified in two chicken isolates, which included qnrB19 and a novel qnrB44 gene. Plasmid isolation and Southern hybridization revealed that qnrS1 was located on a large (>200 kbp) plasmid. The spread of the PMQR genes was attributed to a combination of horizontal dissemination and clonal expansion. The horizontal dissemination of PMQR genes was mostly mediated by IncK plasmids. Molecular typing demonstrated that the majority of the PMQR-positive isolates were genetically diverse. Only one chicken isolate belonged to ST131, which harbored an additional CMY-2 gene. Our findings suggest that the wild birds could serve as reservoirs of PMQR genes and spread them over long distances through migration. To our knowledge, this is the first report of PMQR genes in Korean wild birds. This study also reports qnrS2, qnrB19, qnrB44, and qepA genes for the first time in animal E. coli isolates from South Korea.
Dhaka, Pankaj; Vijay, Deepthi; Vergis, Jess; Negi, Mamta; Kumar, Manesh; Mohan, Vysakh; Doijad, Swapnil; Poharkar, Krupali V.; Malik, Satyaveer Singh; Barbuddhe, Sukhadeo Baliram; Rawool, Deepak B.
Introduction Infectious diarrhoea particularly due to pathogenic bacteria is a major health problem in developing countries, including India. Despite significant reports of diarrhoeagenic Escherichia coli (DEC) pathotypes around the globe, studies which address genetic relatedness, antibiogram profile and their correlation with respect to their isolation from different sources are sparse. The present study determines isolation and identification of DEC pathotypes from different sources, their genetic characterisation, antibiogram profile and their correlation if any. Materials and methods A total of 336 samples comprising diarrhoeic stool samples from infants (n=103), young animal (n=106), foods (n=68) and associated environmental sources (n=59) were collected from Bareilly region of India. All the samples were screened by using standard microbiological methods for the detection of E. coli. The identified E. coli were then confirmed as DEC pathotypes using polymerase chain reaction–based assays. Those DEC pathotypes identified as Enteroaggregative E. coli (EAEC) were further confirmed using HEp-2 adherence assay. All the isolated DEC pathotypes were studied for their genetic diversity using pulsed-field gel electrophoresis (PFGE), and antimicrobial susceptibility testing was performed by using disc diffusion method as per Clinical Laboratory Standards Institute guidelines. Results and discussion Of the four DEC pathotypes investigated, EAEC was found to be the predominant pathogen with an isolation rate of 16.5% from infants, 17.9% from young animals, 16.2% from foods and 3.4% from the associated environmental sources. These EAEC isolates, on further characterisation, revealed predominance of ‘atypical’ EAEC, with an isolation rate of 10.7% from infants, 15.1% from young animals, 16.2% from foods, and 3.4% from the associated environmental sources. On PFGE analysis, discrimination was evident within DEC pathotypes as 52 unique pulsotypes were observed for 59
Bicalho, R C; Santos, T M A; Gilbert, R O; Caixeta, L S; Teixeira, L M; Bicalho, M L S; Machado, V S
The objective of this study was to isolate bacteriophages from environmental samples of 2 large commercial dairy farms using Escherichia coli isolated from the uteri of postpartum Holstein dairy cows as hosts. A total of 11 bacteriophage preparations were isolated from manure systems of commercial dairy farms and characterized for in vitro antimicrobial activity. In addition, a total of 57 E. coli uterine isolates from 5 dairy cows were phylogenetically grouped by triplex PCR. Each E. coli bacterial host from the uterus was inoculated with their respective bacteriophage preparation at several different multiplicities of infections (MOI) to determine minimum inhibitory MOI. The effect of a single dose (MOI=10(2)) of bacteriophage on the growth curve of all 57 E. coli isolates was assessed using a microplate technique. Furthermore, genetic diversity within and between the different bacteriophage preparations was assessed by bacteriophage purification followed by DNA extraction, restriction, and agarose gel electrophoresis. Phylogenetic grouping based on triplex PCR showed that all isolates of E. coli belonged to phylogroup B1. Bacterial growth was completely inhibited at considerably low MOI, and the effect of a single dose (MOI=10(2)) of bacteriophage preparations on the growth curve of all 57 E. coli isolates showed that all bacteriophage preparations significantly decreased the growth rate of the isolates. Bacteriophage preparation 1230-10 had the greatest antimicrobial activity and completely inhibited the growth of 71.7% (n=57) of the isolates. The combined action of bacteriophage preparations 1230-10, 6375-10, 2540-4, and 6547-2, each at MOI=10(2), had the broadest spectrum of action and completely inhibited the growth (final optical density at 600 nm
Chaslus-Dancla, E; Martel, J L; Carlier, C; Lafont, J P; Courvalin, P
We studied two outbreaks of calf salmonellosis caused by apramycin and gentamicin-resistant Salmonella typhimurium strains. In both cases, the responsible strains were resistant to ampicillin, chloramphenicol, kanamycin, streptomycin, tetracycline, and trimethoprim; one strain was also resistant to nalidixic acid in one outbreak. A systematic survey of the intestinal Escherichia coli strains of calves from the two affected flocks showed that 11 of 24 animals sampled were also colonized by apramycin- and gentamicin-resistant E. coli strains. These isolates belonged to four biotypes and were resistant to ampicillin, chloramphenicol, kanamycin, streptomycin, tetracycline, trimethoprim, and nalidixic acid. All of the strains were resistant to high levels of apramycin (MICs, 512 to 1,024 micrograms/ml) and to gentamicin (MICs, 8 to 32 micrograms/ml), and these resistances were always transferred en bloc. In S. typhimurium, this coresistance was borne by plasmids that were approximately 39 kilobases long (outbreak 1) or 90 kilobases long (outbreak 2), whereas in E. coli, the coresistance was due to plasmids that were approximately 110 kilobases long in both outbreaks. The two plasmids of Salmonella and four plasmids of E. coli encoded type IV aminoglycoside 3-N-acetyltransferases. The intensive use of curative and preventive treatments in calf production could be responsible for the emergence of enzymic resistance to apramycin and gentamicin. Images PMID:3521474
Jones, C. Hal; Tuckman, Margareta; Murphy, Ellen; Bradford, Patricia A.
The presence of the tetracycline resistance determinant tet(M) in human clinical isolates of Escherichia coli is described for the first time in this report. The homologue was >99% identical to the tet(M) genes reported to occur in Lactobacillus plantarum, Neisseria meningitidis, and Streptococcus agalactiae, and 3% of the residues in its deduced amino acid sequence diverge from tet(M) of Staphylococcus aureus. Sequence analysis of the regions immediately flanking the gene revealed that sequences upstream of tet(M) in E. coli have homology to Tn916; however, a complete IS26 insertion element was present immediately upstream of the promoter element. Downstream from the termination codon is an insertion sequence that was homologous to the ISVs1 element reported to occur in a plasmid from Vibrio salmonicida that has been associated with another tetracycline resistance determinant, tet(E). Results of mating experiments demonstrated that the E. coli tet(M) gene was on a mobile element so that resistance to tetracycline and minocycline could be transferred to a susceptible strain by conjugation. Expression of the cloned tet(M) gene, under the control of its own promoter, provided tetracycline and minocycline resistance to the E. coli host. PMID:17015654
Jones, C Hal; Tuckman, Margareta; Murphy, Ellen; Bradford, Patricia A
The presence of the tetracycline resistance determinant tet(M) in human clinical isolates of Escherichia coli is described for the first time in this report. The homologue was >99% identical to the tet(M) genes reported to occur in Lactobacillus plantarum, Neisseria meningitidis, and Streptococcus agalactiae, and 3% of the residues in its deduced amino acid sequence diverge from tet(M) of Staphylococcus aureus. Sequence analysis of the regions immediately flanking the gene revealed that sequences upstream of tet(M) in E. coli have homology to Tn916; however, a complete IS26 insertion element was present immediately upstream of the promoter element. Downstream from the termination codon is an insertion sequence that was homologous to the ISVs1 element reported to occur in a plasmid from Vibrio salmonicida that has been associated with another tetracycline resistance determinant, tet(E). Results of mating experiments demonstrated that the E. coli tet(M) gene was on a mobile element so that resistance to tetracycline and minocycline could be transferred to a susceptible strain by conjugation. Expression of the cloned tet(M) gene, under the control of its own promoter, provided tetracycline and minocycline resistance to the E. coli host.
Das, Subhadip; Chatterjee, Nabanita; Bose, Dipayan; Banerjee, Somenath; Pal, Prajnamoy; Jha, Tarun; Das Saha, Krishna
Sepsis is the reflection of systemic immune response that manifests in the sequential inflammatory process in presence of infection. This may occur as a result of gram-negative bacterial sepsis including Escherichia coli infection that gives rise to excessive production of inflammatory mediators and causes severe tissue injuries. We have reported earlier that the lipid of attenuated Leishmania donovani suppresses the inflammatory responses in arthritis patients. Using heat killed E. coli stimulated macrophages, we have now investigated the effect of leishmanial total lipid (LTL) isolated from Leishmania donovani (MHO/IN/1978/UR6) for amelioration of the inflammatory mediators and transcriptional factor with suppression of TLR4-CD14 expression. To evaluate the in vivo effect, E. coli induced murine sepsis model was used focusing on the changes in different parameter(s) of lung injury caused by sepsis, namely, edema, vascular permeability, and pathophysiology, and the status of different cytokine-chemokine(s) and adhesion molecule(s). Due to the effect of LTL, E. coli induced inflammatory cytokine-chemokine(s) levels were significantly reduced in serum and bronchoalveolar lavage fluid simultaneously. LTL also improved the lung injury and suppressed the cell adhesion molecules in lung tissue. These findings indicate that LTL may prove to be a potential anti-inflammatory agent and provide protection against gram-negative bacterial sepsis with pulmonary impairment.
The present study was conducted to gain insights into the occurrence and characteristics of extended-spectrum beta-lactamase- (ESBL-) producing Escherichia coli (E. coli) from drinking well water in the rural area of Laiwu, China, and to explore the role of the nearby pit latrine as a contamination source. ESBL-producing E. coli from wells were compared with isolates from pit latrines in the vicinity. The results showed that ESBL-producing E. coli isolates, with the same antibiotic resistance profiles, ESBL genes, phylogenetic group, plasmid replicon types, and enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) fingerprints, were isolated from well water and the nearby pit latrine in the same courtyard. Therefore, ESBL-producing E. coli in the pit latrine may be a likely contributor to the presence of ESBL-producing E. coli in rural well water. PMID:27965975
Aedo, Sandra; Tse-Dinh, Yuk-Ching
DNA topoisomerases are important targets in anticancer and antibacterial therapy because drugs can initiate cell death by stabilizing the transient covalent topoisomerase-DNA complex. In this study, we employed a method that uses CsCl density gradient centrifugation to separate unbound from DNA-bound GyrA/ParC in Escherichia coli cell lysates after quinolone treatment, allowing antibody detection and quantitation of the covalent complexes on slot blots. Using these procedures modified from the in vivo complexes of enzyme (ICE) bioassay, we found a correlation between gyrase-DNA complex formation and DNA replication inhibition at bacteriostatic (1× MIC) norfloxacin concentrations. Quantitation of the number of gyrase-DNA complexes per E. coli cell permitted an association between cell death and chromosomal gyrase-DNA complex accumulation at norfloxacin concentrations greater than 1× MIC. When comparing levels of gyrase-DNA complexes to topoisomerase IV-DNA complexes in the absence of drug, we observed that the gyrase-DNA complex level was higher (∼150-fold) than that of the topoisomerase IV-DNA complex. In addition, levels of gyrase and topoisomerase IV complexes reached a significant increase after 30 min of treatment at 1× and 1.7× MIC, respectively. These results are in agreement with gyrase being the primary target for quinolones in E. coli. We further validated the utility of this method for the study of topoisomerase-drug interactions in bacteria by showing the gyrase covalent complex reversibility after removal of the drug from the medium, and the resistant effect of the Ser83Leu gyrA mutation on accumulation of gyrase covalent complexes on chromosomal DNA.
Murinda, Shelton E; Nguyen, Lien T; Landers, Tippi L; Draughon, F Ann; Mathew, Alan G; Hogan, Joseph S; Smith, K Larry; Hancock, Dale D; Oliver, Stephen P
The objective of this study was to characterize Escherichia coli isolates from dairy cows/feedlots, calves, mastitis, pigs, dogs, parrot, iguana, human disease, and food products for prevalence of Shiga toxin-producing E. coli (STEC) virulence markers. The rationale of the study was that, isolates of the same serotypes that were obtained from different sources and possessed the same marker profiles, could be cross-species transmissible. Multiplex polymerase chain reaction (PCR) was used to detect presence of genes encoding Shiga toxin 1 and 2 (stx1 and stx2), H7 flagella (flicC), enterohemolysin (hly) and intimin (eaeA) in E. coli isolates (n = 400). Shiga toxin-producing isolates were tested for production of Shiga toxins (Stx1 and Stx2 and enterohemolysin. Of the E. coli O157:H7/H- strains, 150 of 164 (mostly human, cattle, and food) isolates were stx+. Sixty-five percent of O157 STEC produced both Stx1 and Stx2; 32% and 0.7% produced Stx2 or Stx1, respectively. Ninety-eight percent of O157 STEC had sequences for genes encoding intimin and enterohemolysin. Five of 20 E. coli O111, 4 of 14 O128 and 4 of 10 O26 were stx+ . Five of 6 stx+ O26 and O111 produced Stx1, however, stx+ O128 were Stx-negative. Acid resistance (93.3%) and tellurite resistance (87.3%) were common attributes of O157 STEC, whereas, non-O157 stx+ strains exhibited 38.5% and 30.8% of the respective resistances. stx-positive isolates were mostly associated with humans and cattle, whereas, all isolates from mastitis (n = 105), and pigs, dogs, parrot and iguanas (n = 48) were stx-negative. Multiplex PCR was an effective tool for characterizing STEC pathogenic profiles and distinguished STEC O157:H7 from other STEC. Isolates from cattle and human disease shared similar toxigenic profiles, whereas isolates from other disease sources had few characteristics in common with the former isolates. These data suggest interspecies transmissibility of certain serotypes, in particular, STEC O157:H7, between
Jiménez-Belenguer, Ana; Doménech, Eva; Villagrá, Arantxa; Fenollar, Alejandro; Ferrús, Maria Antonia
The use of antimicrobials in food animals is the major determinant for the propagation of resistant bacteria in the animal reservoir. However, other factors may also play a part, and in particular vertical spread between the generations has been suggested to be an important transmission pathway. The objective of this paper was to determine the resistance patterns of Escherichia coli isolated from newly-hatched chickens as well as to study the antibiotic pressure effect when amoxicillin was administered during their growing period. With this aim, meconium from 22 one-day-old Ross chickens was analysed. In addition, during their growth period, amoxicillin treatments at days 7, 21 and 35 were carried out. Results showed a high number of E. coli-resistant strains were isolated from the treated one-day-old chickens, and were the highest for β-lactams group, followed by quinolone and tetracyclines. After treatment with amoxicillin, the highest percentage of resistances were detected for this antibiotic compared to the others analysed, with significant differences in resistance percentages between control and treated broilers detected in relation to ampicillin, cephalothin, streptomycin, kanamycin, gentamicin, chloramphenicol and tetracycline. Differences in resistances to ciprofloxacin and nalidixic acid between control and treated animals were not observed and there was lack of resistance for amikacin and ceftriaxone. These results suggest the possibility of vertical transmission of resistant strains to newly-hatched chicks from parent flocks, and seem to indicate that the treatment with amoxicillin increased the resistance of E. coli to other antibiotics.
Villegas, Maria Virginia; Correa, Adriana; Perez, Federico; Miranda, Maria Consuelo; Zuluaga, Tania; Quinn, John P
Gram-negative pathogens harboring extended-spectrum beta-lactamases (ESBL) are widely prevalent in Latin America, but little is known about their prevalence in Colombia. A network of 8 tertiary care hospitals in Bogotá, Medellín, and Cali, Colombia, was formed in January 2002 to determine the prevalence of ESBL-producing Klebsiella pneumoniae and Escherichia coli. We characterized and established the molecular epidemiology of ESBLs from these hospitals. Data from 1074 E. coli and 394 K. pneumoniae isolates were obtained from hospital laboratories during 6 months. Isolates resistant to third-generation cephalosporins or aztreonam were sent to a central laboratory. The prevalence of strains with this phenotype was 32.6% in K. pneumoniae and 11.8% in E. coli from the intensive care units, with slightly lower percentages from wards. Although TEM and SHV enzymes were present, the dominant class was CTX-M. Molecular typing of chromosomal DNA showed that most strains were not clonal.
Ciesielczuk, Holly; Betts, Jonathon; Phee, Lynnette; Doumith, Michel; Hope, Russell; Woodford, Neil; Wareham, David W
Extra-intestinal pathogenic Escherichia coli (ExPEC) are a significant cause of urinary tract infections and bacteraemia worldwide. Currently no single virulence factor or ExPEC lineage has been identified as the sole contributor to severe extra-intestinal infection and/or urosepsis. Galleria mellonella has recently been established as a simple model for studying the comparative virulence of ExPEC. In this study we investigated the virulence of 40 well-characterized ExPEC strains, in G. mellonella, by measuring mortality (larvae survival), immune recognition/response (melanin production) and cell damage (lactate dehydrogenase production). Although mortality was similar between urinary and bloodstream isolates, it was heightened for community-associated infections, complicated UTIs and urinary-source bacteraemia. Isolates of ST131 and those possessing afa/dra, ompT and serogroup O6 were also associated with heightened virulence.
Ding, Juanjuan; Ma, Xitao; Chen, Zhuochang; Feng, Keqing
A total of 52 strains were resistant to amoxicillin-clavulanate by disk diffusion method in a Chinese tertiary hospital from July 2011 to December 2011. Among these isolates, 2 isolates possessed a phenotype consistent with production of inhibitor-resistant temoniera (TEM) (IRT) β-lactamase, and the TEM-type gene was cloned into strains of Escherichia coli JM109 cells. Both had no blaTEM mutations and were identified as TEM-1 β-lactamase producers. As a result, no IRT β-lactamase was detected. Multiplex PCR detected most of these strains produced TEM-1 enzymes, and plasmid-mediated AmpC β-lactamase and oxacillinase-1 β-lactamases are important mechanisms of resistance as well.
Clemente, Lurdes; Manageiro, Vera; Jones-Dias, Daniela; Correia, Ivone; Themudo, Patricia; Albuquerque, Teresa; Geraldes, Margarida; Matos, Filipa; Almendra, Cláudia; Ferreira, Eugénia; Caniça, Manuela
The impact of extended-spectrum β-lactamases (ESBLs) and plasmid-mediated AmpC β-lactamases (PMAβs) of animal origin has been a public health concern. In this study, 562 Salmonella enterica and 598 Escherichia coli isolates recovered from different animal species and food products were tested for antimicrobial resistance. Detection of ESBL-, PMAβ-, plasmid-mediated quinolone resistance (PMQR)-encoding genes and integrons was performed in isolates showing non-wild-type phenotypes. Susceptibility profiles of Salmonella spp. isolates differed according to serotype and origin of the isolates. The occurrence of cefotaxime non-wild-type isolates was higher in pets than in other groups. In nine Salmonella isolates, blaCTX-M (n = 4), blaSHV-12 (n = 1), blaTEM-1 (n = 2) and blaCMY-2 (n = 2) were identified. No PMQR-encoding genes were found. In 47 E. coli isolates, blaCTX-M (n = 15), blaSHV-12 (n = 2), blaCMY-2 (n = 6), blaTEM-type (n = 28) and PMQR-encoding genes qnrB (n = 2), qnrS (n = 1) and aac(6')-Ib-cr (n = 6) were detected. To the best of our knowledge, this study is the first to describe the presence of blaCMY-2 (n = 2) and blaSHV-12 (n = 1) genes among S. enterica from broilers in Portugal. This study highlights the fact that animals may act as important reservoirs of isolates carrying ESBL-, PMAβ- and PMQR-encoding genes that might be transferred to humans through direct contact or via the food chain.
Gomis, S M; Riddell, C; Potter, A A; Allan, B J
The objective of this study was to characterize virulence factors of Escherichia coli isolates from broilers with simultaneous occurrence of cellulitis and other colibacillosis lesions. Thirty flocks were sampled and 237 birds with cellulitis were examined. Eighty-two (34.6%) of 237 birds condemned for cellulitis had gross lesions in the heart, air sacs, joints, or liver. In 58 chickens, E. coli was isolated from both the cellulitis and other lesions of colibacillosis, and 18.9% of the E. coli isolates from the 2 types of lesions belonged to the same O group. Escherichia coli of serogroups O78, O1, and O2 predominated. Isolates of the same serogroup that were derived from different lesions in the same birds had similar patterns of biotype, aerobactin production, serum sensitivity profile, antibiotic sensitivity, and K1 capsule production. Escherichia coli derived from cellulitis lesions produced virulence factors similar to those found in E. coli isolated from other colibacillosis lesions in poultry. PMID:11227188
Wajima, Takeaki; Sabui, Subrata; Kano, Shigeyuki; Ramamurthy, Thandavarayan; Chatterjee, Nabendu Sekhar; Hamabata, Takashi
Coli surface antigen 6 (CS6) is one of the most prevalent colonization factors among enterotoxigenic Escherichia coli (ETEC) isolated in developing countries. Although it is known that CS6 is encoded by a plasmid, there are no reports on the sequence analysis of the CS6-encoding plasmid or genes exhibiting similar behavior to CS6. Here, we report the isolation of the CS6-encoding plasmid, pCss165Kan, from 4266 ΔcssB::kanamycin (Km) and its complete nucleotide sequence. This plasmid consisted of 165,311bp and 222 predicted coding sequences. Remarkably, there were many insertion sequence (IS) elements, which comprised 24.4% of the entire sequence. Virulence-associated genes such as heat-stable enterotoxin, homologues of ATP-binding cassette transporter in enteroaggregative E. coli (EAEC), and ETEC autotransporter A were also present, although the ETEC autotransporter A gene was disrupted by the integration of IS629. We found that 2 transcriptional regulators belonging to the AraC family were not involved in CS6 expression. Interestingly, pCss165 had conjugative transfer genes, as well as 3 toxin-antitoxin systems that potentially exclude other plasmid-free host bacteria. These genes might be involved in the prevalence of CS6 among ETEC isolates.
Kim, Semi; Sung, Ji Youn; Cho, Hye Hyun; Kwon, Kye Chul; Koo, Sun Hoe
The aims of this study were to characterize the molecular epidemiological profiles of CTX-M-producing uropathogenic Escherichia coli isolates from a tertiary hospital in Daejeon, Korea, and to investigate the genetic diversity and compare the prevalence of sequence types (STs) in different areas. Extended spectrum β-lactamase-producing E. coli strains isolated from urine were analyzed for CTX-M, integrons, and insertion sequence common regions (ISCRs) by PCR and sequencing. Multilocus sequence typing (MLST), pulsed-field gel electrophoresis (PFGE), phylogenetic analysis, and rep-PCR were also used for molecular typing of the isolates. Of 80 CTX-M producers, 31 and 46 expressed CTX-M-15 and CTX-M-14, respectively. MLST analysis indicated that the most prevalent ST was ST131 (n = 34, 42.5%), followed by ST38 (n = 22, 27.5%), ST405 (n = 8, 10.0%), and ST69 (n = 6, 7.5%). Most CTX-M producers harbored class 1 integrons. ST131 strains belonged to phylogenetic group B2 and showed identical rep-PCR patterns, whereas ST69, ST38, and ST405 strains belonged to phylogenetic group D; the ST38 and ST405 strains displayed the same rep-PCR pattern, respectively. ST131 and ST38 isolates showed 21 and 19 distinct types, respectively, by PFGE. In Daejeon, D-ST38 CTX-M-14 producers were relatively more prevalent than in other countries and Korean cities. Our results indicate that CTX-M-producing E. coli isolates belonged mostly to ST131 or ST38 and were more related to hospital-onset than to community-onset infections and that the blaCTX-M gene may vary according to the ST.
Takahashi, I; Yoshida, T; Higashide, Y; Sakano, T
Susceptibilities of Escherichia coli, Salmonella and Staphylococcus aureus isolated from chickens, pigs and cattle to ofloxacin (OFLX) and commonly used antimicrobial agents were investigated. 1. E. coli (28 isolates) demonstrated the highest level of susceptibility of OFLX (MIC 0.10-0.39 micrograms/ml for all the isolates) among all the test drugs. Commonly used antimicrobial agents to which these isolates responded with relatively high susceptibilities (MIC50 0.78-6.25 micrograms/ml) included oxolinic acid (OXA), ampicillin (ABPC), kanamycin (KM) and chloramphenicol (CP) with their MIC50 values in the increasing order as above. Drugs to which these isolates responded with moderate to weak susceptibilities (MIC50 25 approximately greater than 800 micrograms/ml) were doxycycline (DOXY), streptomycin (SM), spectinomycin (SPCM) and sulfadimethoxine (SDMX) in the increasing order of MIC50. E. coli isolates with resistances to all the test drugs other than OFLX and OXA amounted to 7.1-57.1% of the isolates examined and 20 isolates (71.4%) in total. 2. Susceptibilities to OFLX and 4 existing pyridonecarboxylic acid derivatives of E. coli (48 samples) isolated recently from diarrheal pigs were compared. When evaluated in terms of MIC50, the values of OFLX and norfloxacin were both 0.10 micrograms/ml. The values increased by differences of 0.39-3.13 micrograms/ml in an order of OXA, pipemidic acid and nalidixic acid. 3. Salmonella (28 isolates) demonstrated the highest level of susceptibility to OFLX (MIC 0.20-0.39 micrograms/ml for all the isolates) among all the test drugs. The drugs to which these isolates responded with relatively high to moderate susceptibilities (MIC50 0.78-12.5 micrograms/ml) included ABPC, OXA, DOXY, KM, CP and SM with their MIC50 values increasing in this order. The drugs to which the isolates responded with low susceptibilities (MIC50 above 100 micrograms/ml) were SPCM and SDMX. Of all the 28 Salmonella isolates tested, 7.1-32.1% were resistant
Lim, Jong-Soo; Choi, Da-Som; Kim, Young-Jo; Chon, Jung-Whan; Kim, Hong-Seok; Park, Hyun-Jung; Moon, Jin-San; Wee, Sung-Hwan; Seo, Kun-Ho
In South Korea, few reports have indicated the occurrence and characteristics of extended-spectrum β-lactamase (ESBL)-producing Escherichia coli in food-producing animals, particularly in poultry slaughterhouses. In this study, we investigated the occurrence and antibiotic resistance of ESBL-producing E. coli from whole chicken carcasses (n=156) and fecal samples (n=39) of chickens obtained from 2 slaughterhouses. Each sample enriched in buffered peptone water was cultured on MacConkey agar with 2 mg/L cefotaxime and ESBL agar. ESBL production and antibiotic susceptibility were determined using the Trek Diagnostics system. The ESBL genotypes were determined by polymerase chain reaction (PCR) using the bla(SHV), bla(TEM), and bla(CTX-M) gene sequences. Subtyping using a repetitive sequence-based PCR system (DiversiLab™) and multilocus sequence typing (MLST) were used to assess the interspecific biodiversity of isolates. Sixty-two ESBL-producing E. coli isolates were obtained from 156 samples (39.7%). No bla(SHV) genes were detected in any of the isolates, whereas all contained the bla(TEM) gene. Twenty-five strains (40.3%) harbored the CTX-M group 1 gene. The most prevalent MLST sequence type (ST) was ST 93 (14.5%), followed by ST 117 (9.7%) and ST 2303 (8.1%). This study reveals a high occurrence and β-lactams resistance rate of E. coli in fecal samples and whole chickens collected from slaughterhouses in South Korea.
Charnock, Colin; Nordlie, Anne-Lise; Hjeltnes, Bjarne
The presence of enterovirulent and/or antibiotic resistant strains of Escherichia coli in recreational bathing waters would represent a clear health issue. In total, 144 E. coli isolated from 26 beaches along the inner Oslo fjord were examined for virulence determinants and resistance to clinically important antibiotics. No isolates possessed the genetic determinants associated with enterotoxigenic strains and none showed the prototypic sorbitol negative, O157:H7 phenotype. A small number (∼1 %) produced alpha-hemolysin. Occurrences and patterns of antibiotic resistances were similar to those of E. coli isolated previously from environmental samples. In total, 6 % of the strains showed one or more clinically relevant resistances and 1.4 % were multi-drug resistant. Microarray analyses suggested that the resistance determinants were generally associated with mobile genetic elements. Resistant strains were not clonally related, and were, furthermore not concentrated at one or a few beach sites. This suggests that these strains are entering the waters at a low rate but in a widespread manner. The study demonstrates that resistant E. coli are present in coastal bathing waters where they can come into contact with bathers, and that the resistance determinants are potentially transferable. Some of the resistances registered in the study are to important antibiotics used in human medicine such as fluoroquinolones. The spread of antibiotic resistant genes, from the clinical setting to the environment, has clear implications with respect to the current management of bacterial infections and the long term value of antimicrobial therapy. The present study is the first of its kind in Norway.
Evans, D J; Evans, D G; Höhne, C; Noble, M A; Haldane, E V; Lior, H; Young, L S
Escherichia coli isolated from cases of bacteremia and from a variety of urinary tract infections were characterized according to serotype (O:H antigenicity), K type (possession of K1, K2, K3, K12, or K13), hemagglutination (HA) type, and production of beta-hemolysin. Results obtained with the bacteremia and urinary tract infection isolates were similar except for more hemolytic isolated from urine than from blood (42 versus 29%) and more K1+ isolates from blood than from urine (50 versus 29%). A close correlation was found between Ha type VI (production of fimbriae which mediate mannose-resistant HA of human and African green monkey erythrocytes) and the production of hemolysin or K1 capsular antigen or both. Most (95 of 98, or 95%) of the HA type VI+ blood isolates and most (146 of 164, or 89%) of the HA type VI+ urine isolates produced hemolysin or K1 or both, in contrast to 22 and 26%, respectively, of those belonging to HA types other than HA type VI. Also, 76% of all hemolytic and 70% of all K1+ isolates belonged to HA type VI. Remarkably few of the HA type VI+ isolates (13%) and even fewer of the HA type VI- isolates (3%) produced both K1 and hemolysin; these belonged mainly to serotypes O16:H6, O18:H7 and O2:H4. Other major serogroups were usually K1+/hemolysin- (O1, O7) or K1-/hemolysin+ (O2, O4, O6). At least 74% (262 of 351) and possibly as many as 83% (293 of 351) of those isolates which produced mannose-resistant HA of human erythrocytes were classified as HA type VI+; 31 isolates produced mannose-resistant HA with all erythrocytes tested. Taking serogroup and serotype into consideration, we conclude that the E. coli fimbrial hemagglutinin(s) responsible for the HA type VI phenotype will prove to be the same as the virulence-associated mannose-resistant adhesins of uropathogenic E. coli which other investigators have characterized as unique fimbrial antigens detectable by mannose-resistant HA of human erythrocytes. PMID:7007421
Shin, Seung Won; Jung, Myunghwan; Shin, Min-Kyung
In this study, 78 isolates of Escherichia coli isolated from Korean beef cattle farms were investigated for the production of extended-spectrum β-lactamase (ESBL) and/or AmpC β-lactamase. In the disc diffusion test with ampicillin, amoxicillin, cephalothin, ceftiofur, cefotaxime, ceftazidime, and cefoxitin, 38.5% of the isolates showed resistance to all of ampicillin, amoxicillin, and cephalothin. The double disc synergy method revealed that none of the isolates produced ESBL or AmpC β-lactamases. DNA sequencing showed that all isolates encoded genes for TEM-1-type β-lactamase. Moreover, 78.2% of the isolates transferred the TEM-1-type β-lactamase gene via conjugation. In plasmid replicon typing of all donors, IncFIB and IncFIA were identified in 71.4% and 41.0% of plasmids, respectively. In transconjugants, IncFIB and IncFIA were the most frequent types detected (61.5% and 41.0%, respectively). Overall, the present study indicates that selection pressures of antimicrobials on β-lactamases in beef cattle may be low relative to other livestock animals in Korea. Moreover, to reduce selection pressure and dissemination of β-lactamase, the long-term surveillance of antimicrobial use in domestic beef cattle should be established. PMID:26119172
Medina, Anicia M.; Rivera, Fulton P.; Pons, Maria J.; Riveros, Maribel; Gomes, Cláudia; Bernal, María; Meza, Rina; Maves, Ryan C.; Huicho, Luis; Chea-Woo, Elsa; Lanata, Claudio F.; Gil, Ana I.; Ochoa, Theresa J.; Ruiz, Joaquim
Background Antibiotic resistance is increasing worldwide, being of special concern in low- and middle-income countries. The aim of this study was to determine the antimicrobial susceptibility and mechanisms of resistance in 205 enterotoxigenic Escherichia coli (ETEC) isolates from two cohort studies in children <24 months in Lima, Peru. Methods ETEC were identified by an in-house multiplex real-time PCR. Susceptibility to 13 antimicrobial agents was tested by disk diffusion; mechanisms of resistance were evaluated by PCR. Results ETEC isolates were resistant to ampicillin (64%), cotrimoxazole (52%), tetracycline (37%); 39% of the isolates were multidrug-resistant. Heat-stable toxin producing (ETEC-st) (48%) and heat-labile toxin producing ETEC (ETEC-lt) (40%) had higher rates of multidrug resistance than isolates producing both toxins (ETEC-lt-st) (21%), p<0.05. Only 10% of isolates were resistant to nalidixic acid and none to ciprofloxacin or cefotaxime. Ampicillin and sulfamethoxazole resistance were most often associated with blaTEM (69%) and sul2 genes (68%), respectively. Tetracycline resistance was associated with tet(A) (49%) and tet(B) (39%) genes. Azithromycin inhibitory diameters were ≤15 mm in 36% of isolates, with 5% of those presenting the mph(A) gene. Conclusions ETEC from Peruvian children are often resistant to older, inexpensive antibiotics, while remaining susceptible to ciprofloxacin, cephalosporins and furazolidone. Fluoroquinolones and azithromycin remain the drugs of choice for ETEC infections in Peru. However, further development of resistance should be closely monitored. PMID:26175267
Liu, Xiaoqiang; Lazzaroni, Caterina; Aly, Sherine A; Thungrat, Kamoltip; Boothe, Dawn M
This study explored and compared the mechanisms and selective concentration of resistance between a 3rd (pradofloxacin) and 2nd (ciprofloxacin) generation fluoroquinolone. Pradofloxacin- and ciprofloxacin-resistant mutants were selected by stepwise exposure of Escherichia coli (E. coli) to escalating concentrations of pradofloxacin and ciprofloxacin. The sequence of the quinolone resistance determining region (QRDR) and the transcriptional regulator soxS were analyzed, and efflux pump AcrAB-TolC activity was measured by quantitative real-time reverse transcription-PCR (qRT-PCR). First-step mutants reduced the fluoroquinolone sensitivity and one mutant bore a single substitution in gyrA. Four of six second-step mutants expressed ciprofloxacin resistance, and displayed additional mutations in gyrA and/or parC, while these mutants retained susceptibility to pradofloxacin. All the third-step mutants were fluoroquinolone resistant, and each expressed multidrug resistance (MDR) phenotypes. Further, they displayed resistance to all antibacterials tested except cefotaxime, ceftazidime and meropenem. The number of mutations in QRDR of gyrA and parC correlated with fluoroquinolone MICs. Mutations in parC were not common in pradofloxacin-associated mutants. Moreover, one second- and one third-step ciprofloxacin-associated mutants bore both mutations at position 12 (Ala12Ser) and 78 (Met78Leu) in the soxS gene, yet no mutations in the soxS gene were detected in the pradofloxacin-selected mutants. Altogether, these results demonstrated that resistance emerged relatively more rapidly in 2nd compared to 3rd generation fluoroquinolones. Point mutations in gyrA were a key mechanism of resistance to pradofloxacin, and overexpression of efflux pump gene acrB played a potential role in the emergence of MDR phenotypes identified in this study.
Tong, Panpan; Sun, Yang; Ji, Xue; Du, Xiaoli; Guo, Xuejun; Liu, Jun; Zhu, Lingwei; Zhou, Bo; Zhou, Wei; Liu, Guo; Feng, Shuzhang
Extended-spectrum β-lactamase (ESBL)-producing Escherichia coli have been frequently isolated from food-producing animals and pose a serious threat to human health. This study collected 195 ESBL-producing E. coli isolates from 20 chicken farms and 3 live-bird markets located in Northeast China (Heilongjiang, Liaoning, Jilin) and Jiangsu province from February 2011 to October 2013. ESBL genes, including blaCTX-M, blaTEM, and blaSHV, were detected and characterized, and the susceptibilities of these strains to various antimicrobial agents were determined. One hundred ninety-one of these isolates carried 1 or more bla genes. blaCTX-M, blaTEM-1, and blaSHV-5 were identified in 183, 121, and 2 isolates, respectively. The most common blaCTX-M genes were blaCTX-M-15 (68 strains), blaCTX-M-65 (41 strains), blaCTX-M-55 (35 strains), blaCTX-M-14 (32 strains), followed by blaCTX-M-3, blaCTX-M-13, blaCTX-M-79, and blaCTX-M-101, as well as the chimeric genes blaCTX-M-64, blaCTX-M-123, and blaCTX-M-132. Fifteen strains (7.7%) co-harboring CTX-M-1 group and CTX-M-9 group genes were detected in 195 ESBL-producing strains. Pulsed-field gel electrophoresis of 45 strains showed that these CTX-M-producing isolates belonged to 34 different types. To our knowledge, this is the first study to report the blaSHV-5 gene in E. coli isolated from chickens in China. Conjugation experiments demonstrated that the blaCTX-M and blaTEM genes could be transferred to E. coli strain J53, while conjugative transfer of the blaSHV-5 gene from two isolates was not detectable. blaCTX-M genes are carried by many kinds of transferable and untypable plasmids. Our findings demonstrate that the CTX-M enzymes are predominant in both type and quantity.
Balıkçı, Hilal; Açıkgöz, Z Cibali; Güvenman, Selda; Celikbilek, Nevreste; Ozdem, Birsen
This study was conducted to obtain data about prevalence of plasmid-mediated AmpC (pAmpC), efficacy of various phenotypic tests applied for the detection of pAmpC-mediated resistance and the enzyme types responsible for this resistance. A total of 1316 isolates (1030 Escherichia coli and 286 Klebsiella spp.) obtained from the clinical samples sent to our laboratory between 2008-2011 period, from various inpatient and outpatient clinics and intensive care units, were included in this study. Antimicrobial susceptibility profiles of the isolates were determined by using Kirby-Bauer disk diffusion method. Production of pAmpC was phenotypically investigated by Boronic Acid Inhibition Test (BAIT), Cefoxitin Hodge Test (CHT) and AmpC Disk Test (ACDT) in a total of 36 (2.7%) cefoxitin-resistant isolates (77% were E.coli, 23% were K.pneumoniae); and by synergy tests with or without AmpC inhibitors in a total of 165 (88% were E.coli, 12% were K.pneumoniae) cefoxitin-susceptible, third generation cephalosporin-resistant (S3R) isolates. For the detection of pAmpC genes a multiplex-PCR protocol was applied to the isolates found positive at least by one of the phenotypic tests. CHT, ACDT and BAIT were found positive in 21 (58%), 18 (50%) and 7 (19%) of the 36 cefoxitin-resistant isolates, respectively. In only 10 (27.7%) of these isolates (all were E.coli strains), pAmpC presence was detected by PCR; and the enzyme produced was assessed as CMY-2. Based on the positive PCR results; specificity rates of the phenotypic tests, BAIT, ACDT and CHT were 97%, 69% and 58%; while the sensitivity rates were 50%, 100% and 100%, respectively. Our data indicated that, pAmpC prevalence (10/1316, 0.8%) detected in this study, did not seem to be an important problem yet, in the population screened. However, since the first detection of this resistance was in 2010, it should be considered as a sign to get necessary precautions preventing its spread. In conclusion, none of the phenotypic methods
Gibbons, J F; Boland, F; Egan, J; Fanning, S; Markey, B K; Leonard, F C
Antimicrobial use and resistance in animal and food production are of concern to public health. The primary aims of this study were to determine the frequency of resistance to 12 antimicrobials in Escherichia coli isolates from 39 pig farms and to identify patterns of antimicrobial use on these farms. Further aims were to determine whether a categorization of farms based on the duration of in-feed antimicrobial use (long-term versus short-term) could predict the occurrence of resistance on these farms and to identify the usage of specific antimicrobial drugs associated with the occurrence of resistance. Escherichia coli were isolated from all production stages on these farms; susceptibility testing was carried out against a panel of antimicrobials. Antimicrobial prescribing data were collected, and farms were categorized as long term or short term based on these. Resistance frequencies and antimicrobial use were tabulated. Logistic regression models of resistance to each antimicrobial were constructed with stage of production, duration of antimicrobial use and the use of 5 antimicrobial classes included as explanatory variables in each model. The greatest frequencies of resistance were observed to tetracycline, trimethoprim/sulphamethoxazole and streptomycin with the highest levels of resistance observed in isolates from first-stage weaned pigs. Differences in the types of antimicrobial drugs used were noted between long-term and short-term use farms. Categorization of farms as long- or short-term use was sufficient to predict the likely occurrence of resistance to 3 antimicrobial classes and could provide an aid in the control of resistance in the food chain. Stage of production was a significant predictor variable in all models of resistance constructed and did not solely reflect antimicrobial use at each stage. Cross-selection and co-selection for resistance was evident in the models constructed, and the use of trimethoprim/sulphonamide drugs in particular was
Tabasi, Mohsen; Karam, Mohammad Reza Asadi; Habibi, Mehri; Mostafavi, Ehsan
Introduction Urinary Tract Infections (UTIs) caused by Uropathogenic Escherichia coli (UPEC) are among the most common infections worldwide. It is well-documented that the pathogenesis of UPEC is mediated by the production of a wide variety of Virulence Factors (VFs). Thus, detection of these VFs and evaluation of their association with different clinical types of UTIs could help to understand the role of these factors in pathogenesis of UPEC isolates. Aim To investigate the genotypic characteristics of UPEC isolates and to examine the relationship between VFs and different clinical symptoms of UTI. Materials and Methods In this cross-sectional study conducted at Pasteur Institute of Iran, a total of 156 UPEC isolated from outpatients and inpatients (symptomatic and asymptomatic UTI patients) visiting general and private hospitals in Tehran, Iran between March 2014 and February 2015 were included. Among them, 49 patients experienced at least one episode of recurrent UTI. A Polymerase Chain Reaction (PCR) assay was developed to detect the presence of different VFs in the isolates. Moreover, Pulsed-Field Gel Electrophoresis (PFGE) was used to characterize clonal relationships among UPEC isolates. Results The prevalence of virulence genes ranged from 0% for cdtB to 100% for fimH. The papEF, hlyA and aer genes were found to be significantly more frequent in UPEC isolated from patients with pyelonephritis, while the afa gene, the only indicator of recurrent UTIs, was more prevalent in UPEC isolated from patients with cystitis. In the present study, 34 PFGE clonal groups were found in the UPEC genome. Conclusion Our findings showed that from outpatients and patients with pyelonephritis, isolates were more virulent than those isolated from inpatients and cystitis patients, respectively. PFGE displayed a large diversity in the UPEC isolates that could be considered as an evolutionary strategy in the survival of the bacteria. PMID:28208853
Iovine, Renata de Oliveira; Dejuste, Catia; Miranda, Flávia; Filoni, Claudia; Bueno, Marina Galvão; de Carvalho, Vania Maria
Increasing interactions between humans, domestic animals and wildlife may result in inter-species transmission of infectious agents. To evaluate the presence of pathogenic E. coli and Salmonella spp. and to test the antimicrobial susceptibility of isolates, rectal swabs from 36 different free-ranging wild mammals were taken from two distinct natural sites in Brazil: Cantareira State Park (CSP, state of São Paulo) and Santa Isabel do Rio Negro Region (SIRNR, state of Amazonas). The swabs were randomly collected and processed for bacterial isolation, identification, characterization and antimicrobial resistance. Eighteen E. coli strains from CSP and 20 from SIRNR were recovered from 14 and 22 individuals, respectively. Strains from animals captured in CSP, the site with the greatest anthropization, exhibited a higher range and percentage of virulence genes, including an eae+/bfpA+ strain. Antimicrobial resistance was verified in strains originating from both sites; however, in strains from SIRNR, aminopenicillins were almost the exclusive antimicrobial class to which strains exhibited resistance, whereas in CSP there were strains resistant to cephalosporins, sulfonamide, aminoglycoside, tetracycline and fluoroquinolone, in addition to strains exhibiting multidrug resistance. Two strains of Salmonella enterica that are known to be associated with reptiles, serotypes Belem and 60:r:e,n,z15, were recovered only from Amazonian animals and showed susceptibility to all classes of antimicrobials that were tested. Although the potential impact of these pathogens on wildlife remains unknown, bacteria isolated from free-ranging wild animals may provide relevant information about environmental health and should therefore be more deeply studied.
Ishii, Y; Ohno, A; Taguchi, H; Imajo, S; Ishiguro, M; Matsuzawa, H
Escherichia coli TUH12191, which is resistant to piperacillin, cefazolin, cefotiam, ceftizoxime, cefuzonam, and aztreonam but is susceptible to cefoxitin, latamoxef, flomoxef, and imipenem, was isolated from the urine of a patient treated with beta-lactam antibiotics. The beta-lactamase (Toho-1) purified from the bacteria had a pI of 7.8, had a molecular weight of about 29,000, and hydrolyzed beta-lactam antibiotics such as penicillin G, ampicillin, oxacillin, carbenicillin, piperacillin, cephalothin, cefoxitin, cefotaxime, ceftazidime, and aztreonam. Toho-1 was markedly inhibited by beta-lactamase inhibitors such as clavulanic acid and tazobactam. Resistance to beta-lactams, streptomycin, spectinomycin, sulfamethoxazole, and trimethoprim was transferred by conjugational transfer from E. coli TUH12191 to E. coli ML4903, and the transferred plasmid was about 58 kbp, belonging to incompatibility group M. The cefotaxime resistance gene for Toho-1 was subcloned from the 58-kbp plasmid by transformation of E. coli MV1184. The sequence of the gene for Toho-1 was determined, and the open reading frame of the gene consisted of 873 or 876 bases (initial sequence, ATGATG). The nucleotide sequence of the gene (DDBJ accession number D37830) was found to be about 73% homologous to the sequence of the gene encoding a class A beta-lactamase produced by Klebsiella oxytoca E23004. According to the amino acid sequence deduced from the DNA sequence, the precursor consisted of 290 or 291 amino acid residues, which contained amino acid motifs common to class A beta-lactamases (70SXXK, 130SDN, and 234KTG). Toho-1 was about 83% homologous to the beta-lactamase mediated by the chromosome of K. oxytoca D488 and the beta-lactamase mediated by the plasmid of E. coli MEN-1. Therefore, the newly isolated beta-lactamase Toho-1 produced by E. coli TUH12191 is similar to beta-lactamases produced by K. oxytoca D488, K. oxytoca E23004, and E. coli MEN-1 rather than to mutants of TEM or SHV enzymes
Stirm, Stephan; Ørskov, Frits; Ørskov, Ida; Mansa, Bendt
The K88 antigen was carried by episomal transfer to D282, a nonmotile Escherichia coli strain without K antigen. D520, obtained by this episomal transfer, was used for the extraction of K88 antigen. It was shown by the agar gel precipitation technique that some K88 antigen was released from D520 into suspending aqueous medium. The amount of liberated material was increased by gentle heating (60 C) or treatment in a Waring Blendor. The antigen was obtained from the extracts in a purified form by making use of its insolubility between pH 3.5 and 5.5 and of its high sedimentation rate (S020,w = 36.7S). The homogeneity of the material was demonstrated by agar gel precipitation with D520 antiserum, by analytical ultracentrifugation, and by moving-boundary electrophoresis. Chemical analysis revealed that K88 is a pure protein containing all the common amino acids with the exception of cysteine-cystine. Purified K88 selectively precipitated the K88 antibodies from D520 antiserum and was shown to be immunogenic in rabbits. Images PMID:4960185
Shafritz, K M; Sandigursky, M; Franklin, W A
The bacteria Escherichia coli contains several exonucleases acting on both double- and single-stranded DNA and in both a 5'-->3' and 3'-->5' direction. These enzymes are involved in replicative, repair and recombination functions. We have identified a new exonuclease found in E.coli, termed exonuclease IX, that acts preferentially on single-stranded DNA as a 3'-->5' exonuclease and also functions as a 3'-phosphodiesterase on DNA containing 3'-incised apurinic/apyrimidinic (AP) sites to remove the product trans -4-hydroxy-2-pentenal 5-phosphate. The enzyme showed essentially no activity as a deoxyribophosphodiesterase acting on 5'-incised AP sites. The activity was isolated as a glutathione S-transferase fusion protein from a sequence of the E.coli genome that was 60% identical to a 260 bp region of the small fragment of the DNA polymerase I gene. The protein has a molecular weight of 28 kDa and is free of AP endonuclease and phosphatase activities. Exonuclease IX is expressed in E.coli , as demonstrated by reverse transcription-PCR, and it may function in the DNA base excision repair and other pathways. PMID:9592142
Kojima, Yasuko; Harada, Sohei; Aoki, Kotaro; Sawa, Tomoka; Hasegawa, Kei; Saji, Tsutomu; Yamaguchi, Keizo; Tateda, Kazuhiro
We document the household spread of extended-spectrum β-lactamase-producing Escherichia coli. One isolate belonged to sequence type 1193 and caused urinary tract infection in a 4-month-old female, and the other isolate belonged to sequence type 131 and colonized three family members, including the index patient. These isolates carried similar Inc-I1-Iγ plasmids, harboring blaCTX-M-15. PMID:24554749
Kojima, Yasuko; Harada, Sohei; Aoki, Kotaro; Ishii, Yoshikazu; Sawa, Tomoka; Hasegawa, Kei; Saji, Tsutomu; Yamaguchi, Keizo; Tateda, Kazuhiro
We document the household spread of extended-spectrum β-lactamase-producing Escherichia coli. One isolate belonged to sequence type 1193 and caused urinary tract infection in a 4-month-old female, and the other isolate belonged to sequence type 131 and colonized three family members, including the index patient. These isolates carried similar Inc-I1-Iγ plasmids, harboring blaCTX-M-15.
Dubreuil, J. Daniel; Isaacson, Richard E.; Schifferli, Dieter M.
Enterotoxigenic Escherichia coli (ETEC) is the most common cause of E. coli diarrhea in farm animals. ETEC are characterized by the ability to produce two types of virulence factors; adhesins that promote binding to specific enterocyte receptors for intestinal colonization and enterotoxins responsible for fluid secretion. The best-characterized adhesins are expressed in the context of fimbriae, such as the F4 (also designated K88), F5 (K99), F6 (987P), F17 and F18 fimbriae. Once established in the animal small intestine, ETEC produces enterotoxin(s) that lead to diarrhea. The enterotoxins belong to two major classes; heat-labile toxin that consist of one active and five binding subunits (LT), and heat-stable toxins that are small polypeptides (STa, STb, and EAST1). This chapter describes the disease and pathogenesis of animal ETEC, the corresponding virulence genes and protein products of these bacteria, their regulation and targets in animal hosts, as well as mechanisms of action. Furthermore, vaccines, inhibitors, probiotics and the identification of potential new targets identified by genomics are presented in the context of animal ETEC. PMID:27735786
Guimarães, Bruno; Barreto, Angela; Radhouani, Hajer; Figueiredo, Nicholas; Gaspar, Eurico; Rodrigues, Jorge; Torres, Carmen; Igrejas, Gilberto; Poeta, Patrícia
One hundred twelve fecal samples of healthy children were recovered in Portugal during October 2007 and February 2008 and were tested for extended-spectrum beta-lactamases (ESBLs) containing Escherichia coli isolates and vancomycin-resistant enterococci (VRE). Three of the 112 fecal samples (2.7%) harbored ESBL-positive E. coli isolates and the bla(CTX-M-1), bla(TEM-52), and bla(SHV-12) genes were identified in these isolates. The bla(TEM-52)-containing isolate showed a phenotype of multiresistance that included fluoroquinolones, tetracycline, trimethoprim-sulfamethoxazole, and chloramphenicol; sul1, sul3, and cmlA genes were detected in this isolate, in addition to two amino acid changes in GyrA (Ser83Leu + Asp87Asn) and one in ParC protein (Ser80Ile). The ESBL isolates corresponded to phylogroup A (one isolate), B1 (one isolate), and D (one isolate). vanA-containing Escherichia faecium isolates were detected in 13 of the 112 fecal samples (11.6%), and vanC-1 isolates were found in 2 samples. A diversity of resistance genes [(tet(M), tet(L), erm(B), aph(3')-IIIa, ant(6)-Ia, catA, and vat(E)] were found in VRE isolates. These results show that the intestinal tract of healthy children constitutes a reservoir of ESBL-containing E. coli and VRE isolates.
Ismail, Miriam D; Luo, Ting; McNamara, Sara; Lansing, Bonnie; Koo, Evonne; Mody, Lona; Foxman, Betsy
BACKGROUND Rates of multidrug-resistant gram-negative organisms are surpassing those of methicillin-resistant Staphylococcus aureus and vancomycin-resistant enterococci in nursing homes (NHs). OBJECTIVE To characterize the incidence and duration of carriage of ciprofloxacin-resistant Escherichia coli (CipREc) in NHs and identify those in the O25b-ST131 lineage. METHODS We collected 227 CipREc isolates obtained by routine and regular surveillance of high-risk NH residents with indwelling devices. Repetitive element palindromic (REP)-polymerase chain reaction assay and multiplex polymerase chain reaction amplification for O25b-ST131 E. coli detection were performed using (GTG)5-primers and O25pabBspe and trpA2 primer pairs, respectively. RESULTS We found a high period prevalence of CipREc colonization (21.5%), high rates of recolonization with the same strain following clearing (0.46 recolonizations/ person/ year), and an acquisition incidence of 1.05 cases/1,000 person-days. Almost three-quarters of colonized residents carried strains in the O25b-ST131 E. coli lineage. Compared with isolates not in the lineage, O25b-ST131 isolates were carried significantly longer (10 vs 3 months). We identified 18 different REP-types; 2 occurred in 55% of the residents colonized with CipREc, and in more than 1 NH. Duration of CipREc carriage varied by REP-type and averaged 6 months. CONCLUSION CipREc occurred frequently in NH residents and is carried for long durations, and reacquisition following clearance is common Trial registration. ClinicalTrials.gov identifier: NCT01062841.
Canizalez-Roman, Adrian; Gonzalez-Nuñez, Edgar; Vidal, Jorge E; Flores-Villaseñor, Héctor; León-Sicairos, Nidia
Diarrheogenic Escherichia coli (DEC) strains are an important cause of intestinal syndromes in the developing world mainly affecting children. DEC strains often infect tourists from developed countries traveling to Mexico, causing so-called "traveler diarrhea". DEC strains are typically transmitted by contaminated food and water; however, the prevalence of these strains in food items that are produced, consumed and sometimes exported in northwestern Mexico has not been evaluated. In this study, we conducted a large microbiological survey of DEC strains in 5162 food items and beverages consumed throughout Sinaloa state during 2008 and 2009. We developed a panel of eight sequential PCR reactions that detected the presence of all DEC categories, including typical or atypical variants. Thermotolerant coliforms (also known as fecal coliforms) and E. coli were detected by conventional bacteriology in 13.4% (692/5162) and 7.92% (409/5162) of food items, respectively. Among 409 E. coli isolates, 13.6% (56/409) belonged to DEC strains. Dairy products (2.8%) were the most contaminated with DEC, while DEC strains were not detected in beverages and ice samples. The pathogenic type that was most commonly isolated was EPEC (78.5%), followed by EAEC (10.7%), STEC (8.9%) and ETEC (1.7%). EHEC, DAEC and EIEC strains were not detected. Approximately 80% of EPEC and EAEC strains were classified as atypical variants; they did not adhere to a culture of HEp-2 cell. Of the isolated DEC strains, 66% showed resistance to at least one commonly prescribed antibiotic. In conclusion, the presence of DEC strains in food items and beverages available in northwestern Mexico is low and may not represent a threat for the general population or those traveling to tourist areas.
Horcajo, Pilar; Domínguez-Bernal, Gustavo; Carrión, Javier; De La Fuente, Ricardo; Ruiz-Santa-Quiteria, José A; Orden, José A
Differences in the pathogenicity of atypical enteropathogenic Escherichia coli (EPEC) strains may be due, at least partially, to different expression patterns of some virulence genes. To investigate this hypothesis, the virulence gene expression patterns of 6 atypical EPEC strains isolated from healthy and diarrheic ruminants were compared using quantitative real-time reverse transcription polymerase chain reaction after growing the bacteria in culture medium alone or after binding it to HeLa epithelial cells. Some virulence genes in strains from diarrheic animals were upregulated relative to their expression in strains from healthy animals. When bacteria were cultured in the presence of HeLa cells, the ehxA and efa1/lifA genes, previously associated with the production of diarrhea, were expressed at higher levels in strains from diarrheic animals than in strains from healthy animals. Thus, the expression levels of some virulence genes may help determine which atypical EPEC strains cause diarrhea in ruminants.
Horcajo, Pilar; Domínguez-Bernal, Gustavo; Carrión, Javier; De La Fuente, Ricardo; Ruiz-Santa-Quiteria, José A.; Orden, José A.
Differences in the pathogenicity of atypical enteropathogenic Escherichia coli (EPEC) strains may be due, at least partially, to different expression patterns of some virulence genes. To investigate this hypothesis, the virulence gene expression patterns of 6 atypical EPEC strains isolated from healthy and diarrheic ruminants were compared using quantitative real-time reverse transcription polymerase chain reaction after growing the bacteria in culture medium alone or after binding it to HeLa epithelial cells. Some virulence genes in strains from diarrheic animals were upregulated relative to their expression in strains from healthy animals. When bacteria were cultured in the presence of HeLa cells, the ehxA and efa1/lifA genes, previously associated with the production of diarrhea, were expressed at higher levels in strains from diarrheic animals than in strains from healthy animals. Thus, the expression levels of some virulence genes may help determine which atypical EPEC strains cause diarrhea in ruminants. PMID:24082409
Paddock, Zac D; Renter, David G; Cull, Charley A; Shi, Xiarong; Bai, Jianfa; Nagaraja, Tiruvoor G
Escherichia coli O26 is second only to O157 in causing foodborne, Shiga toxin-producing E. coli (STEC) infections. Our objectives were to determine fecal prevalence and characteristics of E. coli O26 in commercial feedlot cattle (17,148) that were enrolled in a study to evaluate an E. coli O157:H7 siderophore receptor and porin (SRP(®)) vaccine (VAC) and a direct-fed microbial (DFM; 10(6) colony-forming units [CFU]/animal/day of Lactobacillus acidophilus and 10(9) CFU/animal/day of Propionibacterium freudenreichii). Cattle were randomly allocated to 40 pens within 10 complete blocks; pens were randomly assigned to control, VAC, DFM, or VAC+DFM treatments. Vaccine was administered on days 0 and 21, and DFM was fed throughout the study. Pen-floor fecal samples (30/pen) were collected weekly for the last 4 study weeks. Samples were enriched in E. coli broth and subjected to a multiplex polymerase chain reaction (PCR) designed to detect O26-specific wzx gene and four major virulence genes (stx1, stx2, eae, and ehxA) and to a culture-based procedure that involved immunomagnetic separation and plating on MacConkey agar. Ten presumptive E. coli colonies were randomly picked, pooled, and tested by the multiplex PCR. Pooled colonies positive for O26 serogroup were streaked on sorbose MacConkey agar, and 10 randomly picked colonies per sample were tested individually by the multiplex PCR. The overall prevalence of E. coli O26 was higher (p<0.001) by the culture-based method compared to the PCR assay (22.7 versus 10.5%). The interventions (VAC and or DFM) had no impact on fecal shedding of O26. Serogroup O26 was recovered in pure culture from 23.9% (260 of 1089) of O26 PCR-positive pooled colonies. Only 7 of the 260 isolates were positive for the stx gene and 90.1% of the isolates possessed an eaeβ gene that codes for intimin subtype β, but not the bfpA gene, which codes for bundle-forming pilus. Therefore, the majority of the O26 recovered from feedlot cattle feces was
Vila, Jordi; Vargas, Martha; Casals, Climent; Urassa, Honorato; Mshinda, Hassan; Schellemberg, David; Gascon, Joaquim
Diarrhea caused by multidrug-resistant bacteria is an important public health problem among children in developing countries. The prevalence and antimicrobial susceptibility of diarrheagenic Escherichia coli in 346 children under 5 years of age in Ifakara, Tanzania, were studied. Thirty-eight percent of the cases of diarrhea were due to multiresistant enterotoxigenic E. coli, enteroaggregative E. coli, or enteropathogenic E. coli. Strains of all three E. coli categories showed high-level resistance to ampicillin, tetracycline, co-trimoxazole, and chloramphenicol but were highly susceptible to quinolones. Guidelines for appropriate use of antibiotics in developing countries need updating. PMID:10582903
Batra, Dhwani; Rowe, Lori; Loparev, Vladimir N.; Stripling, Devon; Garcia-Toledo, Lisley; Knipe, Kristen; Juieng, Phalasy; Sheth, Mili; Martin, Haley; Laufer Halpin, Alison
ABSTRACT Enterobacteriaceae carrying plasmid-mediated colistin resistance have been found around the world. We report here the high-quality whole-genome sequence of an Escherichia coli O157:H48 isolate (2016C-3936C1) from Connecticut that carried the mcr-1 resistance gene on an IncX4-type plasmid. PMID:28302788
Navarro-Gonzalez, N; Porrero, M C; Mentaberre, G; Serrano, E; Mateos, A; Domínguez, L; Lavín, S
Antimicrobial resistance was assessed in indicator Escherichia coli isolates from free-ranging livestock and sympatric wild boar (Sus scrofa) and Iberian ibex (Capra pyrenaica) in a National Game Reserve in northeastern Spain. The frequency of antimicrobial resistance was low (0% to 7.9%). However, resistance to an extended-spectrum cephalosporin and fluoroquinolones was detected.
Introduction: Shiga toxin-producing Escherichia coli (STEC) are important food-borne pathogens that cause outbreaks and serious cases of food-borne illness. Methods for detection and isolation of STEC, particularly the non-O157 STEC, are needed to prevent their transmission through contaminated fo...
We report the isolation of Campylobacter species from the same population of feral swine that was investigated in San Benito County, California during the 2006 spinach-related Escherichia coli O157:H7 outbreak. This is the first survey of Campylobacter in a free-ranging feral swine population in the...
Rincón, Giovanna; Radice, Marcela; Giovanakis, Marta; Di Conza, Jose A; Gutkind, Gabriel
This is the first report of the presence of qepA1 efflux pump gene in Escherichia coli clinical isolate from Argentina, which was associated to other plasmid-mediated quinolone resistance determinants, such as aac(6')-Ib-cr and qnrB10 and also quinolone resistance determining regions mutations.
Porrero, M. C.; Mentaberre, G.; Serrano, E.; Mateos, A.; Domínguez, L.; Lavín, S.
Antimicrobial resistance was assessed in indicator Escherichia coli isolates from free-ranging livestock and sympatric wild boar (Sus scrofa) and Iberian ibex (Capra pyrenaica) in a National Game Reserve in northeastern Spain. The frequency of antimicrobial resistance was low (0% to 7.9%). However, resistance to an extended-spectrum cephalosporin and fluoroquinolones was detected. PMID:23892753
Johnson, James R.; Davis, Gregg; Clabots, Connie; Johnston, Brian D.; Porter, Stephen; DebRoy, Chitrita; Pomputius, William; Ender, Peter T.; Cooperstock, Michael; Slater, Billie Savvas; Banerjee, Ritu; Miller, Sybille; Kisiela, Dagmara; Sokurenko, Evgeni V.; Aziz, Maliha; Price, Lance B.
Background. Within-household sharing of strains from the resistance-associated H30R1 and H30Rx subclones of Escherichia coli sequence type 131 (ST131) has been inferred based on conventional typing data, but it has been assessed minimally using whole genome sequence (WGS) analysis. Methods. Thirty-three clinical and fecal isolates of ST131-H30R1 and ST131-H30Rx, from 20 humans and pets in 6 households, underwent WGS analysis for comparison with 52 published ST131 genomes. Phylogenetic relationships were inferred using a bootstrapped maximum likelihood tree based on core genome sequence polymorphisms. Accessory traits were compared between phylogenetically similar isolates. Results. In the WGS-based phylogeny, isolates clustered strictly by household, in clades that were distributed widely across the phylogeny, interspersed between H30R1 and H30Rx comparison genomes. For only 1 household did the core genome phylogeny place epidemiologically unlinked isolates together with household isolates, but even there multiple differences in accessory genome content clearly differentiated these 2 groups. The core genome phylogeny supported within-household strain sharing, fecal-urethral urinary tract infection pathogenesis (with the entire household potentially providing the fecal reservoir), and instances of host-specific microevolution. In 1 instance, the household's index strain persisted for 6 years before causing a new infection in a different household member. Conclusions. Within-household sharing of E coli ST131 strains was confirmed extensively at the genome level, as was long-term colonization and repeated infections due to an ST131-H30Rx strain. Future efforts toward surveillance and decolonization may need to address not just the affected patient but also other human and animal household members. PMID:27703993
The background of quinolone resistance was characterized in ciprofloxacin-resistant commensal Escherichia coli selected out of 3,551 isolates from slaughtered animals in Poland between 2009 and 2012. Plasmid-mediated determinants were suspected in 6.2% of the study group, ranging from 1.1% in cattle to 9.7% in turkeys. Polymerase chain reaction and sequencing identified up to four quinolone resistance-determining substitutions in gyrA (Ser83, Asp87) and parC (Ala56, Ser80). Plasmid-mediated mechanisms were identified as qnrS1 (or qnrS3, n=70, including six isolates with chromosomal mutations), qnrB19 (or qnrB10, n=19), and qnrB17 (n=1). All tested isolates were negative for qnrA, qnrC, qnrD, qepA, and aac(6')-Ib-cr. Still, there were several E. coli suspected for both plasmid- and chromosome-mediated resistance with unrevealed genetic background of the phenomenon. Since all tested isolates showed diverse XbaI-PFGE profiles, chromosome-encoded quinolone resistance does not result from the spread of a single resistant clone, however, it is rather due to antimicrobial pressure leading to the selection of random gyr and par mutants. It also favors the selection and spread of plasmids carrying predominant qnr genes, since the same determinants were found in Salmonella, isolated from similar sources. The identification of carrier plasmids and mitigation of their spread might be essential for sustainable quinolone usage in animal husbandry and efficient protection of human health.
Ghosh, Pankaj Kumar; Ali, Arif
A total of 17 typical and atypical enteropathogenic Escherichia coli (EPEC) were isolated from 396 children with and without diarrhoea. Out of 12 EPEC isolates from patients with diarrhoea, 3 (25 %) were atypical EPEC while 9 (75 %) were typical EPEC. It was observed that atypical EPEC strains had colonized the intestines of healthy children and its isolation rates were higher in healthy children than in children with diarrhoea. Interestingly all of the atypical EPEC isolates carried a megaplasmid, mostly comparable with the size of EPEC adherence factor (EAF) encoding gene but no virulence gene was detected in this megaplasmid. Studies also indicated that multidrug resistance EPEC are emerging and all the atypical EPEC strains showed significantly less resistance to all antimicrobial agents used in this study than typical EPEC. This study also supports the opinion that Shiga toxin-producing E. coli does not pose a major threat to human health in India. Subtyping analysis reveals that eae-α1, eae-β2 and eae-λ could be common EPEC subtypes prevalent in children with diarrhoea in Delhi. The present study is believed to be the first report of the detection of atypical EPEC from children without diarrhoea and records of isolation of eae-γ1, eae-γ2 and the rare eae-λ subtype in India. The data also indicated that typical EPEC are a common cause of diarrhoea and atypical EPEC are emerging as colonizers of the intestine of children with and without diarrhoea in Delhi and the National Capital Region, India.
Heuvelink, A. E.; van de Kar, N. C.; Meis, J. F.; Monnens, L. A.; Melchers, W. J.
Fifty verocytotoxin (VT)-producing Escherichia coli (VTEC) strains of serogroup O157 were characterized by phage typing, polymerase chain reaction (PCR) for VT genes and the E. coli attaching and effacing (eae) gene, and random amplified polymorphic DNA-PCR (RAPD-PCR) fingerprinting. The collection represented isolates obtained from patients with diarrhoea-associated haemolytic-uraemic syndrome (D+ HUS) and their family contacts, isolated in the Netherlands, Belgium and Germany between 1989 and 1993. Based on isolates from separate families (n = 27) seven different phage types were identified, types 2 (44%) and 4 (33%) were predominant. Eighty-five percent of the strains contained only VT2 gene sequences and 15% both VT1 and VT2. All strains of the dominant phage types 2 and 4 carried the VT2 gene. Strains that belonged to the minor phage types 8, 14, 32 carried both VT1 and VT2 genes, with the exception of two isolates identified as phage types 49 and 54 which contained only VT2 genes. All O157 VTEC strains possessed the chromosomally-located eae gene, which indicates its usefulness as virulence marker. RAPD-PCR fingerprinting identified four distinct banding patterns, with one profile found among 79% of the strains. Based on the combined results of all typing methods used in this study, the collection of 50 O157 VTEC strains could be divided into nine distinct groups. Strains isolated from different persons within one family could not be distinguished by any of these methods. The data suggest that O157 VTEC strains are members of one clone that has become widely distributed. Images Fig. 1 Fig. 2 Fig. 3 PMID:7641823
Todd, E C; Szabo, R A; MacKenzie, J M; Martin, A; Rahn, K; Gyles, C; Gao, A; Alves, D; Yee, A J
Verotoxigenic Escherichia coli (VTEC) strains were isolated from food and animal fecal samples by using PCR to screen for the presence of VTEC after broth enrichment and then filtering VTEC-positive cultures through hydrophobic-grid membrane filters (HGMFs) which were incubated on MacConkey agar. The filters were probed with a digoxigenin-labeled PCR product generated by amplification of a conserved verotoxin gene sequence. Replication of the growth on filters allowed probe-positive colonies to be picked. When ground beef samples were inoculated with VTEC strains, 100% of the strains were recovered, and the detection limit was 0.1 CFU per g. Similar results were obtained with seven types of artificially contaminated vegetables. A survey of 32 packages of vegetables and 23 samples of apple cider obtained at the retail level did not reveal the presence of VTEC. However, the intestinal fecal contents of a moose, 1 of 35 wild mammals and birds examined, contained E. coli O157:H7. The DNA hybridization-HGMF method was also used in a prevalence survey of 327 raw and 744 ready-to-eat products; VTEC strains were recovered from 4.9% of the raw products and 0.7% of the ready-to-eat products. No serotype O157:H7 strains were detected. This method is particularly suited for surveys in which low numbers of VTEC-positive samples are expected and isolates are required.
Todd, E. C. D.; Szabo, R. A.; MacKenzie, J. M.; Martin, A.; Rahn, K.; Gyles, C.; Gao, A.; Alves, D.; Yee, A. J.
Verotoxigenic Escherichia coli (VTEC) strains were isolated from food and animal fecal samples by using PCR to screen for the presence of VTEC after broth enrichment and then filtering VTEC-positive cultures through hydrophobic-grid membrane filters (HGMFs) which were incubated on MacConkey agar. The filters were probed with a digoxigenin-labeled PCR product generated by amplification of a conserved verotoxin gene sequence. Replication of the growth on filters allowed probe-positive colonies to be picked. When ground beef samples were inoculated with VTEC strains, 100% of the strains were recovered, and the detection limit was 0.1 CFU per g. Similar results were obtained with seven types of artificially contaminated vegetables. A survey of 32 packages of vegetables and 23 samples of apple cider obtained at the retail level did not reveal the presence of VTEC. However, the intestinal fecal contents of a moose, 1 of 35 wild mammals and birds examined, contained E. coli O157:H7. The DNA hybridization-HGMF method was also used in a prevalence survey of 327 raw and 744 ready-to-eat products; VTEC strains were recovered from 4.9% of the raw products and 0.7% of the ready-to-eat products. No serotype O157:H7 strains were detected. This method is particularly suited for surveys in which low numbers of VTEC-positive samples are expected and isolates are required. PMID:10543785
Maaland, Marit; Guardabassi, Luca
Minimum inhibitory concentrations (MIC) of nitrofurantoin were determined by agar dilution in 269 canine and feline isolates of Escherichia coli and Staphylococcus pseudintermedius, two of the most common bacterial species associated with urinary tract infection (UTI) in small animals. The MIC90 for E. coli and S. pseudintermedius were 32 and 16 μg/ml, respectively. All isolates, including multidrug-resistant strains of known genetic background, displayed MICs below the drug concentrations reported in canine urine following oral administration of nitrofurantoin. Preliminary data on mutant prevention concentration (MPC) and many years of nitrofurantoin usage in human medicine suggest that emergence of resistant mutants during treatment is not a critical issue for this drug. The study provides species-specific data on nitrofurantoin MIC distribution that can be used for setting dog- and cat-specific breakpoints. Although nitrofurantoin is not an appropriate first-line agent for empirical treatment of canine UTI due to toxicity and poor pharmacokinetic properties, it may be indicated for treatment of UTI caused by multidrug-resistant bacteria, which are otherwise difficult to treat using conventional veterinary antimicrobial agents.
Chen, Yahui; Lin, Jianhan; Jiang, Qin; Chen, Qi; Zhang, Shengjun; Li, Li
The objective of this study was to evaluate the performance of a nucleic acid isolation and purification instrument using Escherichia coli O157:H7 as the model. The instrument was developed with magnetic nanoparticles for efficiently capturing nucleic acids and an intelligent mechanical unit for automatically performing the whole nucleic acid extraction process. A commercial DNA extraction kit from Huier Nano Company was used as reference. Nucleic acids in 1 ml of E. coli O157: H7 at a concentration of 5 x 10(8) CFU/mL were extracted by using this instrument and the kit in parallel and then detected by an ultraviolet spectrophotometer to obtain A260 values and A260/A280 values for the determination of the extracted DNA's quantity and purity, respectively. The A260 values for the instrument and the kit were 0.78 and 0.61, respectively, and the A260/A280 values were 1.98 and 1.93. The coefficient of variations of these parallel tests ranged from 10.5% to 16.7%. The results indicated that this nucleic acid isolation and purification instrument could extract a comparable level of nucleic acid within 50 min compared to the commercial DNA extraction kit.
Januszkiewicz, A; Wołkowicz, T; Chróst, A; Szych, J
Shiga toxin-producing Escherichia coli (STEC) O26 infections can be comparable with STEC O157 infections in severity of the acute haemolytic-uremic syndrome HUS and long-term sequelae. Among O26 STEC isolates, highly virulent clone O26:H11/H- Sequence Type 29 (ST 29) emerged in Germany in mid-1990s and spread to European countries. However, up to date, no STEC O26:H11/H- belonging to ST29 has been documented in Poland. In this study, we determined the relationship and clonal structure, stx genotypes, plasmid gene profiles and antimicrobial resistance of nine human STEC O26:H11/H- strains from human patients in Poland between 1996 and 2014. Of the 9 human STEC O26:H11/H- strains, two belonged to ST29 and were isolated from two children with HUS and renal failure with sepsis respectively. These strains showed the molecular characteristics of the emerging human-pathogenic ST29 clone (stx1-, stx2a+, eae+, ehxA+, etpD+, katP-, espP-). The remaining STEC O26:H11/H- strains examined in this study, belonged to ST21, with plasmid genes profiles frequently reported in ST21 strains in Europe. STEC O26 infections with serious human health consequences highlight the need of continuous surveillance of non-O157 STEC and implementation of the diagnostic approaches focused on their detection. Significance and impact of the study: These study provides the first data on the occurrence of emerging Shiga toxin-producing Escherichia coli O26:H11 ST 29 clone in human patients in Poland. Those strains show the molecular characteristics of highly virulent new ST29 pathotype (stx1-, stx2a+, eae+ ehxA+, etpD+, katP-, espP-). These results demonstrated prompt efforts to implement diagnostic approaches detection of those pathogen in the European countries.
Escherichia coli isolates from broiler chicken meat, broiler chickens, pork, and pigs share phylogroups and antimicrobial resistance with community-dwelling humans and patients with urinary tract infection.
Jakobsen, Lotte; Kurbasic, Azra; Skjøt-Rasmussen, Line; Ejrnaes, Karen; Porsbo, Lone J; Pedersen, Karl; Jensen, Lars B; Emborg, Hanne-Dorthe; Agersø, Yvonne; Olsen, Katharina E P; Aarestrup, Frank M; Frimodt-Møller, Niels; Hammerum, Anette M
Escherichia coli is the most common cause of urinary tract infection (UTI). Phylogroup B2 and D isolates are associated with UTI. It has been proposed that E. coli causing UTI could have an animal origin. The objective of this study was to investigate the phylogroups and antimicrobial resistance, and their possible associations in E. coli isolates from patients with UTI, community-dwelling humans, broiler chicken meat, broiler chickens, pork, and pigs in Denmark. A total of 964 geographically and temporally matched E. coli isolates from UTI patients (n = 102), community-dwelling humans (n = 109), Danish (n = 197) and imported broiler chicken meat (n = 86), Danish broiler chickens (n = 138), Danish (n = 177) and imported pork (n = 10), and Danish pigs (n = 145) were tested for phylogroups (A, B1, B2, D, and nontypeable [NT] isolates) and antimicrobial susceptibility. Phylogroup A, B1, B2, D, and NT isolates were detected among all groups of isolates except for imported pork isolates. Antimicrobial resistance to three (for B2 isolates) or five antimicrobial agents (for A, B1, D, and NT isolates) was shared among isolates regardless of origin. Using cluster analysis to investigate antimicrobial resistance data, we found that UTI isolates always grouped with isolates from meat and/or animals. We detected B2 and D isolates, that are associated to UTI, among isolates from broiler chicken meat, broiler chickens, pork, and pigs. Although B2 isolates were found in low prevalences in animals and meat, these sources could still pose a risk for acquiring uropathogenic E. coli. Further, E. coli from animals and meat were very similar to UTI isolates with respect to their antimicrobial resistance phenotype. Thus, our study provides support for the hypothesis that a food animal and meat reservoir might exist for UTI-causing E. coli.
Olesen, Bente; Hansen, Dennis S; Nilsson, Frida; Frimodt-Møller, Jakob; Leihof, Rikke Fleron; Struve, Carsten; Scheutz, Flemming; Johnston, Brian; Krogfelt, Karen A; Johnson, James R
We report the characteristics of 115 extended-spectrum beta-lactamase (ESBL)-producing Escherichia coli clinical isolates, from 115 unique Danish patients, over a 1-year study interval (1 October 2008 to 30 September 2009). Forty-four (38%) of the ESBL isolates represented sequence type 131 (ST13)1, from phylogenetic group B2. The remaining 71 isolates were from phylogenetic groups D (27%), A (22%), B1 (10%), and B2 (3%). Serogroup O25 ST131 isolates (n = 42; 95% of ST131) comprised 7 different K antigens, whereas two ST131 isolates were O16:K100:H5. Compared to non-ST131 isolates, ST131 isolates were associated positively with CTX-M-15 and negatively with CTX-M-1 and CTX-M-14. They also were associated positively with 11 virulence genes, including afa and dra (Dr family adhesins), the F10 papA allele (P fimbria variant), fimH (type 1 fimbriae), fyuA (yersiniabactin receptor), iha (adhesin siderophore), iutA (aerobactin receptor), kpsM II (group 2 capsules), malX (pathogenicity island marker), ompT (outer membrane protease), sat (secreted autotransporter toxin), and usp (uropathogenicity-specific protein) and negatively with hra (heat-resistant agglutinin) and iroN (salmochelin receptor). The consensus virulence gene profile (>90% prevalence) of the ST131 isolates included fimH, fyuA, malX, and usp (100% each), ompT and the F10 papA allele (95% each), and kpsM II and iutA (93% each). ST131 isolates were also positively associated with community acquisition, extraintestinal pathogenic E. coli (ExPEC) status, and the O25, K100, and H4 antigens. Thus, among ESBL E. coli isolates in Copenhagen, ST131 was the most prevalent clonal group, was community associated, and exhibited distinctive and comparatively extensive virulence profiles, plus a greater variety of capsular antigens than reported previously.
Krüger, Alejandra; Lucchesi, Paula M. A.; Parma, Alberto E.
In this study, we determined vt subtypes and evaluated verotoxicity in basal as well as induced conditions of verotoxin-producing Escherichia coli (VTEC) strains isolated from cattle and meat products. Most (87%) of the 186 isolates carried a vt2 gene. Moreover, the vt2 subtype, which is associated with serious disease, was present in 42% of our VTEC collection. The other vt subtypes detected were vt1, vt1d, vt2vha, vt2vhb, vt2O118, vt2d (mucus activatable), and vt2g. A total of 41 (22%) of the isolates possessed more than one vt subtype in its genome, and among them the most frequent combination was vt1/vt2, but we also observed multiple combinations among vt2 subtypes. Differences in verotoxicity titers were found among a selection of 54 isolates. Among isolates with a single vt2 variant, those carrying the vt2 subtype had high titers under both uninduced and induced conditions. However, the highest increase in cytotoxicity under mitomycin C treatment was detected among the strains carrying vt2vha or vt2hb variants. Notably, the isolates carrying the vt1 subtype showed a lesser increase than that of most of the vt2-positive VTEC strains. Furthermore, the presence of more than one vt gene variant in the same isolate was not reflected in higher titers, and generally the titers were lower than those for strains with only one gene variant. The main observation was that both basal and induced cytotoxic effects seemed to be associated with the type and number of vt variants more than with the serotype or origin of the isolate. PMID:21037301
Galland, J C; Hyatt, D R; Crupper, S S; Acheson, D W
Prevalence, antibiotic susceptibility, and genetic diversity were determined for Escherichia coli O157:H7 isolated over 11 months from four beef cattle feedlots in southwest Kansas. From the fecal pat (17,050) and environmental (7,134) samples collected, 57 isolates of E. coli O157:H7 were identified by use of bacterial culture and latex agglutination (C/LA). PCR showed that 26 isolates were eaeA gene positive. Escherichia coli O157:H7 was identified in at least one of the four feedlots in 14 of the 16 collections by C/LA and in 9 of 16 collections by PCR, but consecutive positive collections at a single feedlot were rare. Overall prevalence in fecal pat samples was low (0.26% by C/LA, and 0.08% by PCR). No detectable differences in prevalence or antibiotic resistance were found between isolates collected from home pens and those from hospital pens, where antibiotic use is high. Resistant isolates were found for six of the eight antibiotics that could be used to treat E. coli infections in food animals, but few isolates were multidrug resistant. The high diversity of isolates as measured by random amplification of polymorphic DNA and other characteristics indicates that the majority of isolates were unique and did not persist at a feedlot, but probably originated from incoming cattle. The most surprising finding was the low frequency of virulence markers among E. coli isolates identified initially by C/LA as E. coli O157:H7. These results demonstrate that better ways of screening and confirming E. coli O157:H7 isolates are required for accurate determination of prevalence.
Zweifel, C; Giezendanner, N; Corti, S; Krause, G; Beutin, L; Danuser, J; Stephan, R
Food is an important vehicle for transmission of Shiga toxin-producing Escherichia coli (STEC). To assess the potential public health impact of STEC in Swiss raw milk cheese produced from cow's, goat's, and ewe's milk, 1,422 samples from semihard or hard cheese and 80 samples from soft cheese were examined for STEC, and isolated strains were further characterized. By PCR, STEC was detected after enrichment in 5.7% of the 1,502 raw milk cheese samples collected at the producer level. STEC-positive samples comprised 76 semihard, 8 soft, and 1 hard cheese. By colony hybridization, 29 STEC strains were isolated from 24 semihard and 5 soft cheeses. Thirteen of the 24 strains typeable with O antisera belonged to the serogroups O2, O22, and O91. More than half (58.6%) of the 29 strains belonged to O:H serotypes previously isolated from humans, and STEC O22:H8, O91:H10, O91:H21, and O174:H21 have also been identified as agents of hemolytic uremic syndrome. Typing of Shiga toxin genes showed that stx(1) was only found in 2 strains, whereas 27 strains carried genes encoding for the Stx(2) group, mainly stx(2) and stx(2vh-a/b). Production of Stx(2) and Stx(2vh-a/b) subtypes might be an indicator for a severe outcome in patients. Nine strains harbored hlyA (enterohemorrhagic E. coli hemolysin), whereas none tested positive for eae (intimin). Consequently, semihard and hard raw milk cheese may be a potential source of STEC, and a notable proportion of the isolated non-O157 STEC strains belonged to serotypes or harbored Shiga toxin gene variants associated with human infections.
Sotto, Albert; De Boever, Corinne Merle; Fabbro-Peray, Pascale; Gouby, Anne; Sirot, Danielle; Jourdan, Jacques
From November 1998 to February 1999 we prospectively evaluated the prevalence of resistance to penicillins, cephalosporins, carbapenem, quinolones, aminoglycosides, and trimethoprim-sulfamethoxazole (SXT) in 320 Escherichia coli isolates isolated from hospitalized patients with acute urinary tract infections (UTIs). We also studied for these strains risk factors for resistance to amoxicillin-clavulanic acid (AMC), fluoroquinolones (FQs), and SXT. Resistance rates were consistent with those from major recent studies reported in the literature. Multivariate analyses selected the following factors as being significantly associated with E. coli resistance: (i) for resistance to AMC, prior (1 year) UTI (odds ratio [OR] = 2.71, P = 0.006), prior (1 year) urinary catheter (OR = 2.98, P = 0.0025), and prior (6 months) antibiotic exposure (OR = 2.68, P = 0.005); (ii) for resistance to FQs male sex (OR = 3.87, P = 0.03), with a trend toward significance for age >65 years (OR = 7.67, P = 0.06) and prior (1 year) UTI (OR = 2.98, P = 0.07); and (iii) for resistance to SXT, male sex (OR = 1.91, P = 0.046), hospitalization in an intermediate-term-care unit (OR = 2.18, P = 0.008), and prior (1 year) UTI (OR = 2.03, P = 0.03). Ours results suggest that prior UTI is a common risk factor for resistance to the different antibiotics tested. Although few studies on risk factors for E. coli resistance to antibiotics have been published, careful interpretation of their findings, taking into consideration the population, infection site, and period studied, should contribute to the formulation of a better strategy that can be used to overcome antibiotic resistance. PMID:11158087
Babu, Mohan; Butland, Gareth; Pogoutse, Oxana; Li, Joyce; Greenblatt, Jack F; Emili, Andrew
Biochemical purification of affinity-tagged proteins in combination with mass spectrometry methods is increasingly seen as a cornerstone of systems biology, as it allows for the systematic genome-scale characterization of macromolecular protein complexes, representing demarcated sets of stably interacting protein partners. Accurate and sensitive identification of both the specific and shared polypeptide components of distinct complexes requires purification to near homogeneity. To this end, a sequential peptide affinity (SPA) purification system was developed to enable the rapid and efficient isolation of native Escherichia coli protein complexes (J Proteome Res 3:463-468, 2004). SPA purification makes use of a dual-affinity tag, consisting of three modified FLAG sequences (3X FLAG) and a calmodulin binding peptide (CBP), spaced by a cleavage site for tobacco etch virus (TEV) protease (J Proteome Res 3:463-468, 2004). Using the lambda-phage Red homologous recombination system (PNAS 97:5978-5983, 2000), a DNA cassette, encoding the SPA-tag and a selectable marker flanked by gene-specific targeting sequences, is introduced into a selected locus in the E. coli chromosome so as to create a C-terminal fusion with the protein of interest. This procedure aims for near-endogenous levels of tagged protein production in the recombinant bacteria to avoid spurious, non-specific protein associations (J Proteome Res 3:463-468, 2004). In this chapter, we describe a detailed, optimized protocol for the tagging, purification, and subsequent mass spectrometry-based identification of the subunits of even low-abundance bacterial protein complexes isolated as part of an ongoing large-scale proteomic study in E. coli (Nature 433:531-537, 2005).
Eppinger, Mark; Mammel, Mark K.; LeClerc, Joseph E.; Ravel, Jacques; Cebula, Thomas A.
Cattle comprise a main reservoir of Shiga toxin-producing Escherichia coli O157:H7 (STEC). The significant differences in host prevalence, transmissibility, and virulence phenotypes among strains from bovine and human sources are of major interest to the public health community and livestock industry. Genomic analysis revealed divergence into three lineages: lineage I and lineage I/II strains are commonly associated with human disease, while lineage II strains are overrepresented in the asymptomatic bovine host reservoir. Growing evidence suggests that genotypic differences between these lineages, such as polymorphisms in Shiga toxin subtypes and synergistically acting virulence factors, are correlated with phenotypic differences in virulence, host ecology, and epidemiology. To assess the genomic plasticity on a genome-wide scale, we have sequenced the whole genome of strain EC869, a bovine-associated E. coli O157:H7 isolate. Comparative phylogenomic analysis of this key isolate enabled us to place accurately bovine lineage II strains within the genetically homogenous E. coli O157:H7 clade. Identification of polymorphic loci that are anchored both in the chromosomal backbone and horizontally acquired regions allowed us to associate bovine genotypes with altered virulence phenotypes and host prevalence. This study catalogued numerous novel lineage II-specific genome signatures, some of which appear to be associated intimately with the altered pathogenic potential and niche adaptation within the bovine rumen. The presented extended list of polymorphic markers is valuable in the development of a robust typing system critical for forensic, diagnostic, and epidemiological studies of this emerging human pathogen. PMID:21421787
Shehabi, A A; Odeh, J F; Fayyad, M
This study demonstrates that Escherichia coli isolates from human stools showed mostly higher minimum inhibitory concentrations (MICs) and significant rates of resistance (32%-67%, P<0.05) than Escherichia coli water isolates in Jordan, as follows: ampicillin (67% vs 28%), trimethoprim/sulfamethoxazole (67% vs 28%) nalidixic acid (63% vs 20%), cefuroxime (32% vs 4%), gentamicin (32% vs 17%), norfloxacin (32% vs 12%) and tetracycline (33% vs 16%). The prevalence of integron integrase genes (Intl1) in these isolates was also significantly higher in patients' stools (67%, P <0.05) than in water (36%), but the distribution of Sul 1/Sul 2 or both in association with postive Intl1 and resistance to ampicillin and sulfamethoxazole was not significantly higher (74% versus 62%, P <0.05) in isolates from stool and water. Plasmid profiles of representative multiresistant E. coli isolates from both sources indicated the presence of two common plasmids (49,25 kb) in 11/12 (91.6%), and all E. coli transconjugants were positive for class 1 integron markers (Intl 1, Sul 1 and Sul2) and mostly associated with three transferable drug-resistant determinants to ampicillin, sulfamethoxazole and tetracycline. These results indicate that class 1 integrons with conjugative R-plasmids are common and transferable among commensal antimicrobial multiresistant E. coli isolated from human feces and drinking water sources in Jordan.
Yamamoto, T; Nakazawa, M
Enterotoxigenic Escherichia coli (ETEC) strains isolated from piglets and calves with diarrhea were tested for the presence of the enteroaggregative E. coli enterotoxin 1 (EAST1) gene sequences by PCR and colony hybridization. The EAST1 gene was found in most porcine ETEC strains with adherence factor K88, especially in those elaborating heat-labile enterotoxin. One porcine ETEC strain with adherence factor K99 was also positive for the EAST1 gene. In contrast, 987P-positive (987P+) ETEC strains from piglets, K99+ ETEC strains from calves, and K99+ F41+ or F41+ ETEC strains from piglets and calves were negative for the EAST1 gene. The K88ab+ or K88ac+ ETEC strains tested possessed the EAST1 gene on a plasmid that was distinct from a K88-encoding plasmid. The EAST1 gene sequences of the K88+ ETEC strains were identical to each other and 99.1 and 98.3% homologous to the previously reported sequences of ETEC strains colonizing humans and enteroaggregative E. coli strains, respectively. The data indicate that the EAST1 gene is distributed among porcine ETEC strains in association with the adherence factor type.
Meireles, D; Leite-Martins, L; Bessa, L J; Cunha, S; Fernandes, R; de Matos, A; Manaia, C M; Martins da Costa, P
The increasing prevalence of antimicrobial resistances is now a worldwide problem. Investigating the mechanisms by which pets harboring resistant strains may receive and/or transfer resistance determinants is essential to better understanding how owners and pets can interact safely. Here, we characterized the genetic determinants conferring resistance to β-lactams and quinolones in 38 multidrug-resistant Escherichia coli isolated from fecal samples of dogs, through PCR and sequencing. The most frequent genotype included the β-lactamase groups TEM (n=5), and both TEM+CTX-M-1 (n=5). Within the CTX-M group, we identified the genes CTX-M-32, CTX-M-1, CTX-M-15, CTX-M-55/79, CTX-M-14 and CTX-M-2/44. Thirty isolates resistant to ciprofloxacin presented two mutations in the gyrA gene and one or two mutations in the parC gene. A mutation in gyrA (reported here for the first time), due to a transversion and transition (TCG→GTG) originating a substitution of a serine by a valine in position 83 was also detected. The plasmid-encoded quinolone resistance gene, qnrs1, was detected in three isolates. Dogs can be a reservoir of genetic determinants conferring antimicrobial resistance and thus may play an important role in the spread of antimicrobial resistance to humans and other co-habitant animals.
Luo, Xi; Wang, Peng; Cheng, Jian-guo; Luo, Yan; Dai, Lei; Zhou, Xin; Zou, Li-kou; Li, Bei; Xiao, Jiu-Jin
This study investigated genotypic diversity, 26 virulence genes, and antimicrobial susceptibility of lung pathogenic Escherichia coli (LPEC) isolated from forest musk deer. Associations between virulence factors (VFs) and phylogenetic group, between antimicrobial resistance (AMR) and phylogenetic group, and between AMR and VFs were subsequently assessed. The results showed 30 LPEC isolated were grouped into seven different clusters (A, B, C, D, E, F, and G). The detection rates of crl (90%), kpsMT II (76.67%), mat (76.67%), and ompA (80%) were over 75%. The most frequent types of resistance were to amoxicillin (100%), sulfafurazole (100%), ampicillin (96.67%), and tetracycline (96.67%), with 93.33% (n = 28) of isolates resistant to more than eight types of drugs. There were significant relationships between resistance to cefalotin and the presence of iucD(a) (P < 0.001), papC (P = 0.032), and kpsMT II (P = 0.028); between resistance to chloromycetin and the presence of irp2 (P = 0.004) and vat (P = 0.047); between resistance to nalidixic acid and the presence of crl (P = 0.002) and iucD(a) (P = 0.004); and between resistance to ampicillin/sulbactam and the presence of vat (P = 0.013). These results indicated there could be some association between resistance and VFs, and there is a great need for the prudent use of antimicrobial agents in LPEC.
Filioussis, George; Tzivara, Athanasia; Petridou, Evanthia; Giadinis, Nektarios D; Burriel, Angeliki R; Kritas, Spyridon K
The presence of ciprofloxacin resistance commensal Escherichia coli (C-R-Ec) was determined for goats in the absence of selective pressure in Northern and Central Greece. The C-R-Ec was categorized in 3 groups with respect to their phenotypic resistance to other antibiotics as well as the carriage of antibiotic resistance genes. The first group consisted of 7 C-R-Ec that were found also resistant to tetracycline. Among them tet(B) (n = 7), qnr(S) (n = 7), and qnr(B) (n = 3) producers were identified by polymerase chain reaction. The second group consisted of 10 C-R-Ec that were found sensitive to all other antibiotics, and their phenotypic resistance to ciprofloxacin was not attributed to the presence of resistance genes. Finally, the third group consisted of 2 C-R-Ec also resistant to sulfamethoxazole. These strains were not carrying any transferable elements that contribute to resistance either to ciprofloxacin or to sulfamethoxazole. This is the first report of ciprofloxacin-resistant E. coli isolated from goats in Greece.
Sato, Toyotaka; Yokota, Shin-ichi; Uchida, Ikuo; Okubo, Torahiko; Usui, Masaru; Kusumoto, Masahiro; Akiba, Masato; Fujii, Nobuhiro; Tamura, Yutaka
Fluoroquinolone resistance can cause major clinical problems. Here, we investigated fluoroquinolone resistance mechanisms in a clinical Escherichia coli isolate, HUE1, which had no mutations quinolone resistance-determining regions (QRDRs) of DNA gyrase and topoisomerase IV. HUE1 demonstrated MICs that exceeded the breakpoints for ciprofloxacin, levofloxacin, and norfloxacin. HUE1 harbored oqxAB and qnrS1 on distinct plasmids. In addition, it exhibited lower intracellular ciprofloxacin concentrations and higher mRNA expression levels of efflux pumps and their global activators than did reference strains. The genes encoding AcrR (local AcrAB repressor) and MarR (MarA repressor) were disrupted by insertion of the transposon IS3-IS629 and a frameshift mutation, respectively. A series of mutants derived from HUE1 were obtained by plasmid curing and gene knockout using homologous recombination. Compared to the MICs of the parent strain HUE1, the fluoroquinolone MICs of these mutants indicated that qnrS1, oqxAB, acrAB, acrF, acrD, mdtK, mdfA, and tolC contributed to the reduced susceptibility to fluoroquinolone in HUE1. Therefore, fluoroquinolone resistance in HUE1 is caused by concomitant acquisition of QnrS1 and OqxAB and overexpression of AcrAB–TolC and other chromosome-encoded efflux pumps. Thus, we have demonstrated that QRDR mutations are not absolutely necessary for acquiring fluoroquinolone resistance in E. coli. PMID:23745120
Mainda, Geoffrey; Bessell, Paul B.; Muma, John B.; McAteer, Sean P.; Chase-Topping, Margo E.; Gibbons, James; Stevens, Mark P.; Gally, David L.; deC. Bronsvoort, Barend M.
This study focused on the use of antibiotics on small, medium and commercial-sized dairy farms in the central region of Zambia and its relationship to antibiotic resistance in Escherichia coli. A stratified random sample of 104 farms was studied, representing approximately 20% of all dairy farms in the region. On each farm, faecal samples were collected from a random sample of animals and a standardised questionnaire on the usage of antibiotics was completed. An E. coli isolate was obtained from 98.67% (371/376) of the sampled animals and tested for resistance to six classes of antibiotics. The estimated prevalence of resistance across the different farming systems was: tetracycline (10.61; 95%CI: 7.40–13.82), ampicillin (6.02; 95%CI: 3.31–8.73), sulfamethoxazole/ trimethoprim (4.49; 95%CI: 2.42–6.56), cefpodoxime (1.91; 95%CI: 0.46–3.36), gentamicin (0.89; 95%CI: 0.06–1.84) and ciprofloxacin (0%). Univariate analyses indicated certain diseases, exotic breeds, location, farm size and certain management practices as risk factors for detection of resistance, whereas multivariate analyses showed an association with lumpy skin disease and a protective effect for older animals (>25 months). This study has provided novel insights into the drivers of antibiotic use and their association with antibiotic resistance in an under-studied region of Southern Africa. PMID:26211388
Feng, Peter C H; Keys, Christine; Lacher, David W; Beutin, Lothar; Bentancor, Adriana; Heuvelink, Annet; Afset, Jan E; Rumi, Valeria; Monday, Steven
Atypical enteropathogenic Escherichia coli (aEPEC) is comprised of a large heterogeneous group of strains and serotypes that carry the intimin gene (eae) but no other EPEC virulence factors. In a previous study, we examined a few aEPEC strains of O157:H16 serotype from the U.S. and France and found these to be nearly homologous, and speculated that the same strain had been disseminated or perhaps they are part of a large clonal group that exists worldwide. To test that hypothesis, we examined additional 45 strains isolated from various sources from 4 other countries and determined that although there are a few eae-negative O157:H16 strains, most are aEPEC that carried eae and specifically, the ε-eae allele. Analysis by pulsed field gel electrophoresis (PFGE) and multilocus sequence typing showed that as a whole, O157:H16 strains are phylogenetically diverse and have different sequence types and PFGE profiles. But the aEPEC strains within the O157:H16 serotype, regardless of the eae allele carried, are a highly conserved and homologous group of sequence type (ST)-171 strains that shared similar PFGE profiles. These aEPEC strains of O157:H16 serotype are not closely related to any of the major EPEC and enterohemorrhagic E. coli clonal lineages and appear to be part of a large clonal group that are prevalent worldwide.
Benito, Amparo; Ventoura, Georgia; Casadei, Maria; Robinson, Tobin; Mackey, Bernard
Strains of Escherichia coli O157 isolated from patients with clinical cases of food-borne illness and other sources exhibited wide differences in resistance to high hydrostatic pressure. The most pressure-resistant strains were also more resistant to mild heat than other strains. Strain C9490, a representative pressure-resistant strain, was also more resistant to acid, oxidative, and osmotic stresses than the pressure-sensitive strain NCTC 12079. Most of these differences in resistance were observed only in stationary-phase cells, the only exception being acid resistance, where differences were also apparent in the exponential phase. Membrane damage in pressure-treated cells was revealed by increased uptake of the fluorescent dyes ethidium bromide and propidium iodide. When strains were exposed to the same pressure for different lengths of time, the pressure-sensitive strains took up stain sooner than the more resistant strain, which suggested that the differences in resistance may be related to susceptibility to membrane damage. Our results emphasize the importance of including stress-resistant strains of E. coli O157 when the efficacy of a novel or mild food preservation treatment is tested. PMID:10103251
Chung, Yeon Soo; Song, Jae Won; Kim, Dae Ho; Shin, Sook; Park, Young Kyung; Yang, Soo Jin; Lim, Suk Kyung
Limited information is available regarding horse-associated antimicrobial resistant (AR) Escherichia (E.) coli. This study was designed to evaluate the frequency and characterize the pattern of AR E. coli from healthy horse-associated samples. A total of 143 E. coli (4.6%) were isolated from 3,078 samples collected from three national racetracks and 14 private horse-riding courses in Korea. Thirty of the E. coli isolates (21%) showed antimicrobial resistance to at least one antimicrobial agent, and four of the AR E. coli (13.3%) were defined as multi-drug resistance. Most of the AR E. coli harbored AR genes corresponding to their antimicrobial resistance phenotypes. Four of the AR E. coli carried class 1 integrase gene (intI1), a gene associated with multi-drug resistance. Pulsed-field gel electrophoretic analysis showed no genetic relatedness among AR E. coli isolated from different facilities; however, cross-transmissions between horses or horses and environments were detected in two facilities. Although cross-transmission of AR E. coli in horses and their environments was generally low, our study suggests a risk of transmission of AR bacteria between horses and humans. Further studies are needed to evaluate the risk of possible transmission of horse-associated AR bacteria to human communities through horse riders and horse-care workers. PMID:26645344