blaCMY-2 Gene Research Articles

Molecular Detection Of Multidrug-Resistant Salmonella Isolated From Livestock Production Systems In South Africa.

BackgroundAntibiotic-resistant bacterial pathogens associated with livestock remain a major concern worldwide as they get transmitted from animals to humans and cause foodborne and zoonotic diseases.MethodsAntimicrobial resistance in livestock-associated Salmonella spp in South Africa was investigated using molecular DNA methods. Three hundred and sixty-one environmental faecal samples were randomly collected from avian (chicken and ducks), cows, pigs, goats, and sheep. Salmonella spp. were isolated on selective media and were confirmed using the polymerase chain reaction. Antimicrobial susceptibility testing against ampicillin, chloramphenicol, ciprofloxacin, ceftriaxone, azithromycin, tetracycline, amoxicillin-clavulanate and trimethoprim-sulfamethoxazole was determined using the Kirby–Bauer disk diffusion method. Isolates were screened for the presence of blaTEM-1, blaCMY-2, tetA, tetC, sul2 and dfrA7 resistance genes by PCR.ResultsMost of the isolates were resistant to ampicillin (64%), tetracycline (63%), amoxicillin-clavulanate (49%), trimethoprim-sulfamethoxazole (38%), and ceftriaxone (20%). Eight percent of the tested isolates were ciprofloxacin-resistant Salmonella spp. Multidrug resistance was observed with the mean multiple antibiotic resistance (MAR) index of 0.31. The study demonstrated that 43% of the isolates were multiple drug resistant. The prevalence rates of resistance genes were 44% for blaTEM-1, 35% for blaCMY-2, 21% for sul2, 18% for tetC, 14% for dfrA7 and 8% for tetA.ConclusionResistance to ceftriaxone, detection of blaCMY-2 gene and the high level of intermediate susceptibility (33%) against ciprofloxacin suggested that livestock carry problematic Salmonella spp. This study used the global one-health initiative to report the potential public health risks of livestock-associated pathogens and highlights the importance of monitoring the trends of antimicrobial resistance for sustainability of antibiotics.

623. Antimicrobial Resistance in Non-Typhoidal Salmonella from Retail Poultry Meat by Antibiotic Usage-related Production Claims—Pennsylvania, 2008–2017

BackgroundAntimicrobial-resistant (AMR) nontyphoidal Salmonella infections are a public health concern. Injudicious use of antimicrobials fuels emergence of resistance. The National Antimicrobial Resistance Monitoring System (NARMS) tracks AMR in Salmonella from humans, animals and foods. There is limited evidence regarding antimicrobial use in food animals and AMR bacteria in retail meat.MethodsWe reviewed antimicrobial susceptibility and whole-genome sequencing data from 320 Salmonella isolated from poultry meat in Pennsylvania as part of NARMS activities. Salmonella strains were isolated from 3,481 samples purchased from randomly selected retail outlets during 2008–2017. Antibiotic usage claims on meat packages were used to compare AMR Salmonella from conventional and antibiotic-free/organic (Abx-free) samples. Genetic mechanisms for AMR were investigated in a subset of isolates.ResultsThe prevalence of Salmonella in conventional poultry meat 10.2% (280/2,733) was significantly higher than the prevalence in poultry meat labeled as Abx-free (5.3%, 40/748; P < 0.0001). Salmonella from conventional poultry meat was more likely to be resistant to 3 or more drugs (55.0%, 154/280) compared with poultry meat labeled as Abx-free (27.5%, 11/40; P = 0.0011). Salmonella from conventional poultry exhibited significantly higher resistance to 4 drug classes including β-lactams (P = 0.006) (figure). One hundred isolates from conventional poultry meat and 8 isolates from antibiotic-free/organic samples harbored a gene conferring resistance to the β-lactam class; 24.3% (68/280) of isolates from conventional and 7.5% (3/40) of isolates from Abx-free samples (P = 0.0145) contained the extended-spectrum β-lactamase (ESBL) gene blaCMY-2.ConclusionMeat samples from conventionally-raised poultry were more likely to be contaminated with AMR Salmonella strains and have genes that reduce the effectiveness of antimicrobial drugs recommended for treatment of severe infections. Contamination of poultry with AMR Salmonella strains is concerning as is the presence of genes that decrease the power of critical antibiotics such as β-lactams. These findings highlight the importance of judicious use of antibiotics in food-producing animals. Disclosures All authors: No reported disclosures.

Distribution of fosfomycin and AmpC β-lactamase resistance genes in urinary Escherichia coli isolates obtained from patients admitted to an educational hospital in Ahvaz, southwest Iran

BackgroundEscherichia coli is the most important etiologic agent of urinary tract infections (UTIs). Today, the emergence of antibiotic resistance in UTI-causing E. coli isolates is increasing. This study aimed to investigate the extended-spectrum β-lactamases (ESBLs) production, AmpC enzymes, and fosfomycin resistance genes in E. coli clinical isolates obtained from patients with UTIs. MethodsFrom December 2018 to June 2019, the E. coli isolates were collected from patients with UTI who were referred to Golestan teaching hospital in Ahvaz, Iran. Then, the antimicrobial susceptibility testing, ESBL and AmpC β-lactamase production were performed by disk diffusion method. Finally, the polymerase chain reaction was done to investigate the presence of fosfomycin resistance (fosC2, fosA3) and AmpC (blaCMY-2, blaACC) genes. ResultsIn total, 90 E. coli isolates were collected. The most effective antibiotics were fosfomycin, nitrofurantoin and imipenem with 86 (95.5%), 82 (91.1%), and 78 (86.7%) susceptibility, respectively. AmpC beta-lactamase was confirmed in 18 (20%) isolates. Moreover, 63 (90%) isolates were found as ESBL producers and 6 (6.6%) had both AmpC and ESBL β-lactamases. The frequency rates of blaCMY-2 and blaACC resistance genes in AmpC-producing isolates was 12 (13.3%) and 15 (83.3%), respectively. The fosC2 and fosA3 genes were not detected in E. coli isolates. ConclusionCurrent study revealed the emergence of E. coli isolates harboring ESBL and AmpC enzymes in southwest region of Iran. This study highlighted the need of our laboratories to adopt an appropriate method for continuous screening of ESBL and AmpC in clinical isolates on a daily basis.

Multidrug-Resistant Escherichia coli, Klebsiella pneumoniae and Staphylococcus spp. in Houseflies and Blowflies from Farms and Their Environmental Settings.

Background: Antimicrobial resistance is rising globally at an alarming rate. While multiple active surveillance programs have been established to monitor the antimicrobial resistance, studies on the environmental link to antimicrobial spread are lacking. Methods: A total of 493 flies were trapped from a dairy unit, a dog kennel, a poultry farm, a beef cattle unit, an urban trash facility and an urban downtown area to isolate Escherichia coli, Klebsiella pneumoniae and Staphylococcus spp. for antimicrobial susceptibility testing and molecular characterization. Results: E. coli, K. pneumoniae and coagulase-negative Staphylococcus were recovered from 43.9%, 15.5% and 66.2% of the houseflies, and 26.0%, 19.2%, 37.0% of the blowflies, respectively. In total, 35.3% of flies were found to harbor antimicrobial-resistant bacteria and 9.0% contained multidrug-resistant isolates. Three Staphylococcus aureus isolates were recovered from blowflies while three extended spectrum beta lactamase (ESBL)-carrying E. coli and one ESBL-carrying K. pneumoniae were isolated from houseflies. Whole genome sequencing identified the antimicrobial resistance genes blaCMY-2 and blaCTXM-1 as ESBLs. Conclusion: Taken together, our data indicate that flies can be used as indicators for environmental contamination of antimicrobial resistance. More extensive studies are warranted to explore the sentinel role of flies for antimicrobial resistance.

Plasmid Distribution among Escherichia coli from Livestock and Associated Wastewater: Unraveling Factors That Shape the Presence of Genes Conferring Third-Generation Cephalosporin Resistance.

A key concern with agricultural wastewater storage ponds is that they may provide an environment conducive for horizontal exchange of antibiotic resistance genes (ARGs), thereby facilitating the emergence of antibiotic resistant pathogens. Central to this exchange are mobile genetic elements like plasmids; yet, the factors shaping their presence in agricultural environments remain poorly understood. Here, using Escherichia coli as a model bacterium, we examined genetic backgrounds and plasmid profiles of generic fecal and wastewater isolates and those possessing blaCTX-M and blaCMY-2 genes (which confer resistance to third-generation cephalosporins) to delineate factors shaping the environmental persistence of plasmid-associated ARGs in beef cattle feedlots. The wastewater environment exerted minimal influence on plasmid repertoires, as the number of plasmids and distribution of different incompatibility groups did not differ between generic fecal and wastewater isolates. The blaCTX-M and blaCMY-2 genes were associated with IncF and IncA/C plasmids, respectively, and host isolates possessing these ARGs had fewer plasmids than generic isolates, suggesting ARG-bearing plasmids may associate predominantly with such hosts to compensate for the metabolic burden imposed by these plasmids. Phylogeny also appeared to be a factor for blaCTX-M genes, as their bacterial hosts were restricted to particular genetic lineages, including the environmentally adapted ET-1 clade, as noted previously for these genes. Ultimately, these findings have important implications for evaluating human health risks of agricultural wastewater with respect to environmental persistence of ARGs and may help identify options for improving wastewater treatment.

CTX-M- and pAmpC-Encoding Genes Are Associated with Similar Mobile Genetic Elements in Escherichia coli Isolated from Different Brands of Brazilian Chicken Meat.

In this study we characterized the genetic environment of blaCTX-M and blaCMY-2 genes carried by 46 Escherichia coli isolates obtained from 20 chicken carcasses produced by five different brands in Brazil, including exporters and antibiotic-free-certified producers, purchased between 2010 and 2014. Similar plasmids characterized according to size and incompatibility group (Inc) were identified in E. coli belonging to different MLST-ST collected, regardless of carcass brand or production system. Hybridization assays with transconjugant strains revealed that blaCMY-2 gene (n = 19) was located on 85 kb plasmids of IncB/O, IncI1, IncFIB, or nontypeable groups. blaCTX-M-8 (n = 9) was located on 90 kb IncI1 plasmids. blaCTX-M-2 (n = 14) was inserted in class 1 integrons and conjugated only by one isolate in a 125 kb IncP plasmid. blaCTX-M-15 (n = 1), rarely described in isolates from food-producing animals in South America, was characterized by whole genome sequencing of transconjugant; the gene was carried in a 49.3 kb IncX1 plasmid. Sequencing of bla gene-flanking regions indicated the association of these genes with previously described insertion sequences. These results suggest that conserved genetic environments are related to ESBL and pAmpC genes in the Brazilian chicken production chain.

High proportion of carbapenemase-producing Escherichia coli and Klebsiella pneumoniae among extended-spectrum β-lactamase-producers in Nigerian hospitals

ObjectivesCarbapenem-resistant Enterobacterales are a global problem, however little is known about the burden and origin of carbapenem resistance in Africa. The objectives of this study were to determine the proportion of carbapenem-resistant isolates among extended-spectrum β-lactamase-producing Enterobacterales (ESBL-E), to identify the underlying mechanisms of resistance and to assess the population structure of carbapenem-resistant isolates from Nigeria. MethodsESBL-E isolates (n = 175) from infections were collected at four hospitals in Lagos, Nigeria, from July 2016 to January 2018 and were screened for carbapenem resistance using a VITEK®2 automated system. All carbapenem-resistant ESBL-E (CRE) were screened for blaKPC, blaCTX-M, blaCMY-2, blaNDM, blaVIM, blaIMP, blaOXA-181 and blaOXA-48 genes. Genotyping of randomly selected isolates was performed by whole-genome sequencing. ResultsThe isolates included Escherichia coli (n = 113; 64.6%) and Klebsiella pneumoniae (n = 62; 35.4%). Of the 175 ESBL-E isolates, 48 (27.4%) were resistant to carbapenems (15 E. coli and 33 K. pneumoniae). CRE isolates carried blaNDM (n = 30; 62.5%), blaNDM + blaOXA-181 (n = 10; 20.8%), blaOXA-181 (n = 2; 4.2%) and blaNDM + blaOXA-48 (n = 1; 2.1%); no carbapenemase gene was detected in 5 isolates (10.4%). The isolates showed low diversity and were mainly associated with multilocus sequence typing (MLST) sequence types ST410 for E. coli and ST395 and ST147 for K. pneumoniae. ConclusionCarbapenem resistance is frequent among ESBL-E in Nigeria and is mainly associated with blaNDM. Genotyping suggested that the observed clones possibly originated from Southeast Asia.

Characterization of CMY-2-type beta-lactamase-producing Escherichia coli isolated from chicken carcasses and human infection in a city of South Brazil

BackgroundFood-producing animals, mainly poultry, have been associated with the maintenance and dissemination of antibiotic-resistant bacteria, such as plasmid-mediated AmpC (pAmpC)-producing Enterobacteriaceae, to humans, thus impacting food safety. Many studies have shown that Escherichia coli strains isolated from poultry and humans infections share identical cephalosporin resistance, suggesting that transmission of resistance from poultry meat to humans may occur. The aim of this study was to characterize pAmpC-producing E. coli strains isolated from chicken carcasses and human infection in a restrict area and to determine their antimicrobial resistance profiles, and molecular type by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE).ResultsA total of 14 pAmpC-producing E. coli strains were isolated, including eight strains from chicken carcasses and six strains from human infections (from urine, tissue and secretion). The blaCMY-2 gene was identified in all pAmpC-producing E. coli strains by polymerase chain reaction (PCR) and DNA sequencing. High percentages of strains resistant to tetracycline, nalidixic acid and sulfamethoxazole-trimethoprim (78–92%) were detected, all of which were considered multidrug-resistant. Among the non-beta-lactam resistance genes, the majority of the strains showed tetA, tetB, sulI and sulII. No strain was considered an extended-spectrum beta-lactamases (ESBL) producer, and the blaTEM-1 gene was found in 2 strains isolated from human infection. Six strains from chicken carcasses and four strains from humans infections were linked to an ISEcp1-like element. Through MLST, 11 sequence types were found. Three strains isolated from human infection and one strain isolated from chicken carcasses belonged to the same sequence type (ST354). However, considerable heterogeneity between the strains from chicken carcasses and humans was confirmed by PFGE analysis.ConclusionThis study showed the prevalence of E. coli strains producing blaCMY-2 linked to ISEcp1 that were present in both chickens and humans in a restricted area. Our results also suggest the presence of a highly diverse strains that harbor pAmpC, indicating no clonal dissemination. Therefore, continuous monitoring and comparative analyses of resistant bacteria from humans and food-producing animals are needed.

Genomic Features of High-Priority Salmonella enterica Serovars Circulating in the Food Production Chain, Brazil, 2000\u20132016

Multidrug-resistant (MDR) Salmonella enterica has been deemed a high-priority pathogen by the World Health Organization. Two hundred and sixty-four Salmonella enterica isolates recovered over a 16-year period (2000 to 2016) from the poultry and swine production chains, in Brazil, were investigated by whole-genome sequencing (WGS). Most international lineages belonging to 28 serovars, including, S. enterica serovars S. Schwarzengrund ST96, S. Typhimurium ST19, S. Minnesota ST548, S. Infantis ST32, S. Heidelberg ST15, S. Newport ST45, S. Brandenburg ST65 and S. Kentucky ST198 displayed MDR and virulent genetic backgrounds. In this regard, resistome analysis revealed presence of qnrE1 (identified for the first time in S. Typhimurium from food chain), qnrB19, qnrS1, blaCTX-M-8, blaCTX-M-2 and blaCMY-2 genes, as well as gyrA mutations; whereas ColpVC, IncHI2A, IncHI2, IncFIA, Incl1, IncA/C2, IncR, IncX1 and po111 plasmids were detected. In addition, phylogenetic analysis revealed multiple independent lineages such as S. enterica serovars S. Infantis, S. Schwarzengrund, S. Minnesota, S. Kentucky and S. Brandenburg. In brief, ocurrence and persistence of international lineages of S. enterica serovars in food production chain is supported by conserved genomes and wide virulome and resistome.

High Genetic Diversity of Enterobacteriaceae Clones and Plasmids Disseminating Resistance to Extended-Spectrum Cephalosporins and Colistin in Healthy Chicken in Tunisia.

Enterobacteriaceae resistant to extended-spectrum cephalosporins (ESC-R) are listed as "priority pathogens" by the World Health Organization, and the Agri-food sector has regularly been pointed out as a potential source of ESC-R for humans through food consumption and animal handling. Chicken industry and chicken meat have recurrently been under specific scrutiny due to the high proportions of ESC-R reported worldwide in this sector. In Tunisia, recent studies suggested that the plasmidic AmpC blaCMY-2 gene may have emerged in chicken. We thus collected 258 cloacal swabs from five different farms and selected ESC-R isolates to determine the current ESC-R prevalence and epidemiology. All five farms were ESC-R positive with proportions ranging from 4% to 67.3%. blaCTX-M-1/IncI1/ST3 was the dominant gene/plasmid association in chicken, but several other CTX-M genes and plasmid backgrounds were shown to spread ESC-R. Surprisingly, the CMY-2 enzyme was only identified in one isolate. In addition, we also reported the sporadic presence of the mcr-1 gene carried by an IncHI2 plasmid. Our data suggest that the high diversity of Enterobacteriaceae clones and plasmids circulating in healthy chicken in Tunisia maintains a high ESC-R proportion in flocks and constitutes a major source of ESC-R determinants further disseminating in the food chain.

IDENTIFICATION OF VIRULENCE GENES, β-LACTAMS AND QUINOLONES RESISTANCE-ASSOCIATED GENES AND INTEGRONS IN SALMONELLA ISOLATED FROM RETAIL CHICKEN MEAT AND GIBLETS IN EGYPT

The study investigated the prevalence of virulent and antibiotic resistant Salmonella (S) in retail chicken meat (CM) and pooled giblets (PG) in Egypt. Seventy-two and 64 % of examined CM and PG samples harbored Salmonellae, respectively. When S. Infantis was the most predominant serovar, followed by S. Kentucky, and then S. Virchow. The antibiotic resistance experiment showed a high rate of multiple drug resistant (MDR) Salmonellae. Remarkably, 64.7% of the isolates were resistant to ciprofloxacin (the drug of choice for Salmonella infection). Molecularly, a noteworthy correlation between the MDR and the presence of integrons class 1 and 3. When, integrons 1 and 3 were detected in 100 and 92 % of identified MDR isolates, respectively, while class 2 integrons were not found. Surprisingly, 53.8 and 100% of the molecularly identified isolates were resistant to ciprofloxacin and nalidixic acid, despite the absence of plasmid-mediated quinolone resistance (PMQR)-associated genes (qnrA, qnrB and qnrS, aac(6′)-Ib-cr and qepA) from all MDR isolates. This indicates that other genetic factors could be incriminated in this pattern. Beta-lactam resistance was explored by the existence of blaTEM, blaCTX, and blaCMY-2 genes in most MDR isolates. Additionally, 4 virulence genes were detected in almost all serovars. Concerning virulence genes, invA, hilA, sopB, and stn were detected in 100, 100, 100 and 94.7 % of MDR serovars, respectively. While, spvC was detected in only 7.6 %, and pef was absent. The high prevalence of MDR virulent Salmonella strains in retail chicken meat and giblets in Egypt should be faced with a great worry, as consumers could be under a great public health risk.

Antimicrobial Resistance and Potential Pathogenicity of Escherichia coli Isolates from Healthy Broilers in Québec, Canada.

Antimicrobial resistance (AMR) is a global health issue, particularly when it affects critically important antimicrobials such as third-generation cephalosporins (3GC). The objective of this study was to characterize Escherichia coli isolates from healthy chickens in Québec in farms where ceftiofur has been administered to chickens in ovo over a long period with regard to their AMR, multidrug resistance (MDR), potential virulence, clonality, and possession of plasmids of the incompatibility groups carrying extended-spectrum beta-lactamases (ESBLs)/AmpC genes. More than 62% of indicator isolates were MDR with resistance observed for each of the nine classes of antimicrobials tested by disk diffusion. 3GC resistance was encoded by the blaCMY-2 gene (26.7% in indicator isolates), whereas blaCTX-M was only detected in isolates selected after supplementation with ceftriaxone (3 blaCTX-M-1 isolates). Examination of blaCMY-2-positive isolates by pulsed-field gel electrophoresis showed clustering of isolates originating from different floors of the livestock building within farms. The blaCMY-2 gene was carried on replicon plasmids FIB, I1, K/B, and B/O, whereas blaCTX-M-1 gene was located on I1 as demonstrated by transformation experiments; some of these plasmids cotransferred nonsusceptibility against tetracycline or sulfonamides. In addition, six isolates, of which three were AmpC-producers, were defined as potential human extraintestinal pathogenic E. coli. In summary, this study showed that ESBLs/AmpC-producing E. coli isolates from apparently healthy chickens in Québec, Canada predominantly possess blaCMY-2 rather than blaCTX-M maybe because of the in ovo use of ceftiofur to prevent omphalitis and may be spread through clones or plasmids, and that some of these isolates could be capable of infecting humans.

Multidrug-resistant Escherichia coli isolated from canine faeces in a public park in Quito, Ecuador.

This study focused on estimating the prevalence of extended-spectrum β-lactamases (ESBLs), plasmid-mediated AmpC β-lactamases, carbapenemases and MCR-1-producing Escherichia coli in canine faeces from a public park in Quito, Ecuador. Phenotypic and genotypic characterisation of E. coli isolated from 50 canine faecal samples recovered from a city park in Quito was performed. In addition, a multiple choice survey was conducted among 50 dog owners. Of the 50 faecal samples, 20 (40.0%) presented E. coli resistant to ceftriaxone. Moreover, 23 E. coli isolates were recovered for further analysis. All of the isolates showed as multidrug-resistant (MDR) phenotype (resistant to three or more antibiotic families). Resistance to carbapenems, tigecycline and amikacin was not observed. No major clonal relatedness was observed among the resistant isolates. The ESBL genes blaCTX-M-15, blaCTX-M-55 and blaCTX-M-65 were the most common. Two isolates harboured the blaCMY-2 gene and one isolate harboured both mcr-1 and blaCTX-M-65. Statistical analysis showed that older people were more conscious of collecting and disposing of dog faeces than subjects aged <35 years (P < 0.05). The finding of MDR E. coli in dog faeces in a city park in Ecuador illustrates the importance of analysing canine faeces in public settings (e.g. parks, playgrounds) as part of surveillance programmes for MDR E. coli. In addition, this research might be a sentinel sampling method to gain a better understanding of community sources of antibiotic-resistant Enterobacteriaceae at human-animal-environment interfaces.

Characterization of cefotaxime resistant Escherichia coli isolated from broiler farms in Ecuador.

Antimicrobial resistance (AR) is a worldwide concern. Up to a 160% increase in antibiotic usage in food animals is expected in Latin American countries. The poultry industry is an increasingly important segment of food production and contributor to AR. The objective of this study was to evaluate the prevalence, AR patterns and the characterization of relevant resistance genes in Extended Spectrum β-lactamases (ESBL) and AmpC-producing E. coli from large poultry farms in Ecuador. Sampling was performed from June 2013 to July 2014 in 6 slaughterhouses that slaughter broilers from 115 farms totaling 384 flocks. Each sample of collected caeca was streaked onto TBX agar supplemented with cefotaxime (3 mg/l). In total, 176 isolates were analyzed for AR patterns by the disk diffusion method and for blaCTX-M, blaTEM, blaCMY, blaSHV, blaKPC, and mcr-1 by PCR and sequencing. ESBL and AmpC E. coli were found in 362 flocks (94.3%) from 112 farms (97.4%). We found that 98.3% of the cefotaxime-resistant isolates were multi-resistant to antibiotics. Low resistance was observed for ertapenem and nitrofurantoin. The most prevalent ESBL genes were the ones belonging to the blaCTX-M group (90.9%), specifically the blaCTX-M-65, blaCTX-M-55 and blaCTX-M-3 alleles. Most of the AmpC strains presented the blaCMY-2 gene. Three isolates showed the mcr-1 gene. Poultry production systems represent a hotspot for AR in Ecuador, possibly mediated by the extensive use of antibiotics. Monitoring this sector in national and regional plans of AR surveillance should therefore be considered.

Pathogenic potential and the role of clones and plasmids in beta-lactamase-producing E. coli from chicken faeces in Vietnam

BackgroundAntimicrobial resistance (AMR) in food-producing animals is a global public health issue. This study investigated AMR and virulence profiles of E. coli isolated from healthy chickens in Vietnam. E. coli were isolated from fecal samples collected in five chicken farms located in the provinces of Hoa Binh, Thai Nguyen and Bac Giang in the North of Vietnam. These isolates were examined by disk diffusion for their AMR, PCR for virulence and AMR genes, pulsed-field gel electrophoresis for relatedness between blaCMY-2/blaCTX-M–positive isolates, electroporation for transferability of blaCMY-2 or blaCTX-M genes and sequencing for mutations responsible for ciprofloxacin resistance.ResultsUp to 99% of indicator isolates were multidrug resistant. Resistance to third-generation cephalosporins (3GC) was encoded by both blaCTX-M and blaCMY-2 genes; blaCTX-M genes being of genotypes blaCTX-M-1, − 14, − 15, − 17, − 57 and − 87, whereas ciprofloxacin resistance was due to mutations in the gyrA and parC genes. Some isolates originating from farms located in different provinces of Vietnam were found to be closely related, suggesting they may have been disseminated from a same source of contamination. Plasmids may also have played a role in the diffusion of 3GC-resistance as the blaCMY-2 gene was located on plasmids A/C and I1, and the blaCTX-M gene variants were carried by I1, FIB, R and HI1. Plasmids carrying the blaCMY-2/blaCTX-M genes also co-transferred resistance to other antimicrobials. In addition, isolates potentially capable of infecting humans, of which some produced blaCMY-2/blaCTX-M, were identified in this study.ConclusionsBoth clones and plasmids could be involved in the dissemination of 3GC-resistant E. coli within and between chicken farms in Vietnam. These results demonstrate the necessity to monitor AMR and control antimicrobial use in poultry in Vietnam.

Multidrug-resistant Escherichia coli harbouring mcr-1 and blaCTX-M genes isolated from swine in Argentina.

Multidrug-resistant Escherichia coli isolates recovered from food-producing animals are a global public-health concern, especially those with transferable mechanisms of antimicrobial resistance such as extended-spectrum β-lactamase (ESBL) and mcr-1 genes. Here we report for the first time E. coli recovered from diarrhoeic and healthy pigs harbouring blaCTX-M and/or mcr-1 from Argentinean farms. During 2017, a total of 34 E. coli were recovered from 31 faecal samples from diarrhoeic piglets and healthy fattening pigs from five pig farms in three Argentinean provinces. Antimicrobial susceptibility was evaluated by agar diffusion and resistance genes were identified by PCR. Multiplex PCR was applied to screen for ST69, ST73, ST95 and ST131 clones. Genetic relationships were evaluated by XbaI-PFGE. A high diversity of resistance profiles was observed (20 profiles among 34 isolates), and 71% of isolates were multidrug-resistant. Resistance to third-generation cephalosporins (3GCs) was observed in 28 isolates and was associated with blaCTX-M (24), blaCMY (3) and blaPER-2 (1) genes. blaCTX-M alleles were grouped by specific PCR as follow: 17 blaCTX-M-8/25; 4 blaCTX-M-1/15; 2 blaCTX-M-2; and 1 blaCTX-M-9/14. Twelve isolates were positive for mcr-1, of which six were also resistant to 3GCs and were positive for blaCTX-M-8/25 (4), blaCTX-M-1/15 (1) or blaCMY (1). High genetic diversity was observed, discriminating 29 profiles. One ST131 and two ST95 human-associated clones were detected. Here we describe E. coli isolates recovered from diarrhoeic piglets and healthy fattening pigs harbouring ESBL and/or mcr-1 genes. 3GC resistance was mainly associated with CTX-M, in particular with blaCTX-M-8/25 alleles.

Prevalence of antimicrobial resistance and potential pathogenicity, and possible spread of third generation cephalosporin resistance, in Escherichia coli isolated from healthy chicken farms in the region of Dakar, Senegal.

Escherichia coli is a normal inhabitant of the intestinal microbiota of chickens, a small proportion of which may be avian pathogenic E. coli (APEC) or potential extraintestinal pathogenic E. coli (ExPEC), capable of causing disease in humans. These E. coli may also be resistant to antimicrobials of critical importance in human or veterinary health. This study aims to 1) determine the prevalence of antimicrobial resistance (AMR) and resistance genes, multidrug resistance (MDR), chromosomal mechanisms of quinolone-resistance and virulence profiles of E. coli isolated from healthy chicken farms in the region of Dakar, Senegal, 2) investigate the spread of third-generation cephalosporins (3GC) resistance in E. coli isolated from healthy chicken farms with respect to virulence and resistance genes, serogroups, Pulsed-Field Gel Electrophoresis (PFGE), phylogenetic groups, plasmid types and transferability and 3) determine whether nonsusceptibility against 3GC on farms could be linked to risk factors. More than 68% of isolates from environmental faecal and drinking water samples, carcasses and carcass washes collected on 32 healthy chicken farms were multidrug resistant (MDR), resistance to antimicrobials critical in human health (3GC or ciprofloxacin) being found in all types of samples. Ciprofloxacin resistance was due to mutations in the gyrA and parC genes, 95% of tested farms harboring isolates carrying three mutations, in gyrA (Ser83Ile and Asp87Asn) and parC (Ser80Ile). Nine of the 32 farms (28.1%) demonstrated the presence of one or more 3GC-nonsusceptible indicator isolates but none of the potential risk factors were significantly associated with this presence on farms. Following ceftriaxone enrichment, presumptive extended-spectrum beta-lactamase/AmpC-beta-lactamase (ESBL/AmpC)-producer isolates were found in 17 of the 32 farms. 3GC resistance was mediated by blaCMY-2 or blaCTX-M genes, blaCTX-M being of genotypes blaCTX-M-1, blaCTX-M-8 and for the first time in chickens in Senegal, the genotype blaCTX-M-15. Clonally related ESBL/AmpC-producer isolates were found on different farms. In addition, blaCTX-M genes were identified on replicon plasmids I1 and K/B and blaCMY-2 on K/B, I1 and B/O. These plasmids were found in isolates of different clusters. In addition, 18 isolates, some of which were ESBL/AmpC-producers, were defined as potential human ExPEC. In conclusion, E. coli isolates potentially pathogenic for humans and demonstrating MDR, with resistance expressed against antimicrobials of critical importance in human health were found in healthy chickens in Senegal. Our results suggest that both clonal spreading and horizontal gene transfer play a role in the spread of 3GC-resistance and that chickens in Senegal could be a reservoir for AMR and ExPEC for humans. These results highlight the importance of raising awareness about compliance with biosecurity measures and prudent use of antimicrobials.

Comparative genetic characterization of third-generation cephalosporin-resistant Escherichia coliisolated from a layer operation system in Korea

The layer operation system has a pyramidal structure in which the grandparent stock (GPS) is at the top, followed by the parent stock (PS), and commercial chickens (CC) that produce eggs at the bottom of the pyramid. Unfortunately, this vertical integration of the layer industry allows the transfer of Escherichia coli(E. coli) to the next step. The objective of this study was to genetically characterize and investigate the prevalence of third-generation cephalosporin-resistant E. coli at all levels of the layer operation system in Korea. A total of 561 E. coli samples were tested in this study, and antimicrobial resistance to third-generation cephalosporin, cefotaxime, and ceftazidime gradually increased from GPS to CC. Fifty-one (9.1%) isolates produced β-lactamase genes: GPS harbored blaTEM-1 (1 isolate) or blaCMY-2 (2 isolates) genes; PS harbored blaTEM-1 (7 isolates) or blaCMY-2 (6 isolates) genes; CC harbored blaCTX-M-1 (1 isolate), blaCTX-M-14 (1 isolate), blaTEM-1 (13 isolates), blaCTX-M-14+TEM-1 (7 isolates), or blaCMY-2 (4 isolates) genes; and eggs harbored blaCTX-M-1 (2 isolates), blaCTX-M-14 (1 isolate), blaTEM-1 (3 isolates), or blaCMY-2 (3 isolates) genes. Pulsed-field gel electrophoresis (PFGE) analysis was performed on the 51 β-lactamase-producing E. coli isolates. In PFGE, E. coli included 7 PFGE patterns showing the same production stage and exhibited both β-lactamase genes and the antimicrobial resistance pattern. Our findings suggest that there is a critical need for comprehensive surveillance of third-generation cephalosporin-resistant E. coli at all levels of the layer-production pyramid and provide important considerations for the control of infection in large-scale layer operations in Korea.

Co-occurrence of clinically relevant β-lactamases and MCR-1 encoding genes in Escherichia coli from companion animals in Argentina

Extended-spectrum β-lactamase (ESBL), plasmid-mediated AmpC (pAmpC) and MCR-1 phosphoethanolamine transferase enzymes have been pointed out as the main plasmid-mediated mechanisms of resistance to third generation cephalosporins (TGC) and colistin, respectively, and are currently considered a major concern both in human and veterinary medicine. Little data on these resistance determinants prevalence in companion animal infections is available. The aim of this study was to determine the resistance profile of Escherichia coli isolated from pet infections, in Argentina, and to characterize the resistance mechanisms to TGC, as well as the presence of the plasmid-borne colistin resistance gene, mcr-1. A total of 54 E. coli isolates were collected from clinical samples in dogs and cats; from them, 20/54 (37%, CI95: [24%; 51%]) displayed resistance to TGC. In this regard, thirteen pAmpC-producing isolates were positive for blaCMY-2 genes, whereas seven ESBL- producers harboured blaCTX-M-2 (n = 4), blaCTX-M-15 (n = 2) and blaCTX-M-14 (n = 1) genes. One E. coli strain (V80), isolated from a canine urinary tract infection, showed resistance to colistin (MIC = 8 μg/ml) and whole-genome sequencing analysis revealed co-occurrence of mcr-1.1, blaCTX-M-2, aadA1, ant(2′')-Ia, catA1 and sul1 genes; the former being carried by a 60,587-bp IncI2 plasmid, previously reported in human colistin-resistant E. coli. E. coli V80 belonged to ST770 and the highly virulent phylogenetic group B2. In general, most of these multidrug-resistant isolates belonged to the phylogenetic group F (11/20) and to a lesser extent B2 (5/20), B1 (2/20), D (1/20) and E (1/20). In summary, CMY- and CTX-M-type β-lactamases may constitute the main TGC resistance mechanism in E. coli isolated from pet infections in Argentina, whereas dissemination of colistin resistance mechanism MCR-1 in the human-animal interface has been mediated by IncI2 plasmids.

Characteristics of cephalosporin-resistant Salmonella isolates from poultry in Korea, 2010-2017.

Cephalosporins are the key drugs for the treatment of salmonellosis. Resistance to cephalosporins in Salmonella spp. has become a serious public health concern worldwide. Although the sales of cephalosporins have increased by five times from 2008 to 2016 in Korea, limited information is available on cephalosporin resistance in Korea. Therefore, we aimed to investigate the trends in prevalence and characteristics of cephalosporin-resistant Salmonella isolates from poultry in Korea in the period between 2010 and 2017. A total of 141 Salmonella isolates were collected from various poultry industry sources, and 38 (27.0%) among them showed resistance to cephalosporins. In particular, resistance to the following cephalosporins increased significantly over the seven-year period cephalothin (from 5.0% to 29.2%), cephalexin, cefuroxime, cefotaxime, and ceftazidime (from 0% to 25.0%), and cefepime (from 0% to 12.5%). In addition, 12 isolates carried a β-lactamase gene. A non-extended-spectrum β-lactamase (ESBL) and plasmid-mediated AmpC β-lactamase (pAmpC) gene, blaTEM-1, was found in three isolates in the periods of 2010 to 2011 and 2012 to 2013, respectively. The blaCTX-M-79 (n = 4) and blaCTX-M-15 (n = 1) for ESBL genes and blaCMY-2 (n = 1) for pAmpC genes were only present in the 2016-2017 period. All ESBL/pAmpC-positive isolates had high minimum inhibitory concentrations for most cephalosporins and showed multi-drug resistance. In a conjugation experiment, the transfer of blaCTX-M-79 and blaCMY-2 genes was confirmed in transconjugants, which showed similar pattern of antibiotic resistance. This demonstrates that ESBL/pAmpC-producing Salmonella isolates might be transmitted to humans through contaminated poultry products. These findings suggest the need for the development of monitoring program and the guidelines for prudent use of antimicrobial agents in the poultry industry in Korea.