Unexpected role of pig nostrils in the clonal and plasmidic dissemination of extended-spectrum beta-lactamase-producing Escherichia coli at farm level.

The presence of methicillin-resistant or -susceptible S. aureus in pig nostrils has been known for a long time, but the occurrence of extended-spectrum beta-lactamase (ESBL)-producing E. coli has hardly been investigated. Here, we collected 25 E. coli recovered from nasal samples of 40 pigs/10 farmers of four farms. Nine ESBL-producing isolates belonging to ST48, ST117, ST847, ST5440, ST14914 and ST10 were retrieved from seven pigs. All blaESBL genes (blaCTX-M-32,blaCTX-M-14,blaCTX-M-1,blaCTX-M-65, and blaSHV-12) were horizontally transferable by conjugation through plasmids belonging to IncI1 (n=3), IncX1 (n=3) and IncHI2 (n=1) types. IncI1-plasmids displayed different genetic environments: i) IS26-blaSHV-12-deoR-IS26, ii) wbuC-blaCTX-M-32-ISKpn26 (IS5), and iii) IS930-blaCTX-M-14-IS26. The IncHI2-plasmid contained the genetic environment IS903-blaCTX-M-65-fipA with multiple resistance genes associated either to: a) Tn21-like transposon harbouring genes conferring aminoglycosides/beta-lactams/chloramphenicol/macrolides resistance located on two atypical class 1 integrons with an embedded ΔTn5393; or b) Tn1721-derived transposon displaying an atypical class 1 integron harbouring aadA2-arr3-cmlA5-blaOXA-10-aadA24-dfrA14, preceding the genetic platform IS26-blaTEM-95-tet(A)-lysR-floR-virD2-ISVsa3-IS3075-IS26-qnrS1, as well as the tellurite resistance module. Other plasmids harbouring clinically relevant genes were detected, such as a ColE-type plasmid carrying the mcr-4.5 gene. Chromosomally encoded genes (fosA7) or integrons (intI1-dfrA1-aadA1-qacE-sul1/intI1-IS15-dfrA1-aadA2) were also identified. Finally, an IncY plasmid harbouring a class 2 integron (intI2-dfrA1-sat2-aadA1-qacL-IS406-sul3) was detected but not associated with a blaESBL gene. Our results evidence that pig nostrils might favour the spread of ESBL-E. coli and mcr-mediated colistin-resistance. Therefore, enhanced monitoring should be considered, especially in a sector where close contact between animals in intensive farming increases the risk of spreading antimicrobial resistance.

Enterobacterales high-risk clones and plasmids spreading blaESBL/AmpC and blaOXA-48 genes within and between hospitalized dogs and their environment.

BACKGROUND: Compared with healthcare settings, the role of veterinary hospitals in the spread of extended-spectrum cephalosporin- and carbapenem-resistant (ESC-R/CP-R) bacteria has been overlooked. OBJECTIVES: To investigate using genome-based approaches the dynamics of ESC-R and CP-R Enterobacterales among 125 dogs admitted to the same veterinary hospital over a 4 month period. METHODS: Dogs (n = 125) were sampled within 48 h of admission and at discharge. ESC-R/CP-R were phenotypically characterized and whole-genome sequenced using short- and long-read technologies. Phylogenetic analyses were performed using appropriate pipelines. RESULTS: ESC-R/CP-R prevalence in dogs was 4.8% (6/125) upon admission and reached 24.8% (31/125) at discharge, reflecting multiple acquisitions of ESBL/AmpC and OXA-48-positive Enterobacterales during hospitalization. Indistinguishable or closely related isolates were found within dogs, shared between dogs, and shared between dogs and their environment, suggesting numerous clonal and plasmid spreads. Even though carbapenems are not licensed for use in companion animals, a wide distribution of the blaOXA-48/IncL plasmid was evidenced across different bacterial species and dogs. CONCLUSIONS: This study highlights nosocomial acquisitions of ESBL/AmpC and carbapenemase-producing Enterobacterales by companion animals and the risk of further transmission within the community in a One Health perspective. Reinforced infection prevention and control measures and screening procedures are urgently needed in small animal veterinary settings where advanced therapeutics and intensive care is provided.

Comparative phylogenomics of ESBL-, AmpC- and carbapenemase-producing Klebsiella pneumoniae originating from companion animals and humans.

BACKGROUND: WHO considers ESBL- and carbapenemase-producing Klebsiella pneumoniae a major global concern. In animals, ESBL- and carbapenemase-producing K. pneumoniae of human-related ST11, ST15 and ST307 have been reported, but not in the context of large WGS-based One Health investigations. OBJECTIVES: To perform comparative phylogenomics on a large collection of multidrug-resistant (MDR) K. pneumoniae recovered from diseased companion animals and humans. METHODS: MDR K. pneumoniae (n = 105) recovered from companion animals in France during 2010-18 were phenotypically characterized. All isolates were whole-genome sequenced using the NovaSeq technology and phylogenomic analysis across animal and human K. pneumoniae was performed using appropriate pipelines. RESULTS: bla CTX-M-15, blaDHA-1 and blaOXA-48 were strongly associated with IncFIIk, IncR and IncL plasmids, respectively. When compared with human K. pneumoniae genomes, four groups of closely related French human and animal isolates belonging to ST11, ST15 and ST307 were detected, suggesting the circulation of clones between the human and animal sectors at country level. A large cluster of 31 ST11-KL105 animal isolates from France and Switzerland suggested it corresponds to a sub-lineage that is particularly well-adapted to the animal host. CONCLUSIONS: This study demonstrates the spread of blaCTX-M-15-carrying ST15 and ST307, and blaDHA-1-carrying ST11 K. pneumoniae clones in animal populations. ST11 was the main vector of blaOXA-48/IncL, despite the absence of carbapenem use in French animals. Comparative phylogenomics suggests cross-transmission of K. pneumoniae sub-lineages more prone than others to colonize/infect the animal host. Our data also evidenced the emergence of convergent hypervirulent and MDR K. pneumoniae in animals.

Defining the scope of the European Antimicrobial Resistance Surveillance network in Veterinary medicine (EARS-Vet): a bottom-up and One Health approach.

BACKGROUND: Building the European Antimicrobial Resistance Surveillance network in Veterinary medicine (EARS-Vet) was proposed to strengthen the European One Health antimicrobial resistance (AMR) surveillance approach. OBJECTIVES: To define the combinations of animal species/production types/age categories/bacterial species/specimens/antimicrobials to be monitored in EARS-Vet. METHODS: The EARS-Vet scope was defined by consensus between 26 European experts. Decisions were guided by a survey of the combinations that are relevant and feasible to monitor in diseased animals in 13 European countries (bottom-up approach). Experts also considered the One Health approach and the need for EARS-Vet to complement existing European AMR monitoring systems coordinated by the ECDC and the European Food Safety Authority (EFSA). RESULTS: EARS-Vet plans to monitor AMR in six animal species [cattle, swine, chickens (broilers and laying hens), turkeys, cats and dogs], for 11 bacterial species (Escherichia coli, Klebsiella pneumoniae, Mannheimia haemolytica, Pasteurella multocida, Actinobacillus pleuropneumoniae, Staphylococcus aureus, Staphylococcus pseudintermedius, Staphylococcus hyicus, Streptococcus uberis, Streptococcus dysgalactiae and Streptococcus suis). Relevant antimicrobials for their treatment were selected (e.g. tetracyclines) and complemented with antimicrobials of more specific public health interest (e.g. carbapenems). Molecular data detecting the presence of ESBLs, AmpC cephalosporinases and methicillin resistance shall be collected too. CONCLUSIONS: A preliminary EARS-Vet scope was defined, with the potential to fill important AMR monitoring gaps in the animal sector in Europe. It should be reviewed and expanded as the epidemiology of AMR changes, more countries participate and national monitoring capacities improve.

Multicentre evaluation of a selective isolation protocol for detection of mcr-positive E. coli and Salmonella spp. in food-producing animals and meat.

This study was conducted to evaluate the performance of a screening protocol to detect and isolate mcr-positive Escherichia coli and Salmonella spp. from animal caecal content and meat samples. We used a multicentre approach involving 12 laboratories from nine European countries. All participants applied the same methodology combining a multiplex PCR performed on DNA extracted from a pre-enrichment step, followed by a selective culture step on three commercially available chromogenic agar plates. The test panel was composed of two negative samples and four samples artificially contaminated with E. coli and Salmonella spp. respectively harbouring mcr-1 or mcr-3 and mcr-4 or mcr-5 genes. PCR screening resulted in a specificity of 100% and a sensitivity of 83%. Sensitivity of each agar medium to detect mcr-positive colistin-resistant E. coli or Salmonella spp. strains was 86% for CHROMID® Colistin R, 75% for CHROMagarTM COL-APSE and 70% for COLISTIGRAM. This combined method was effective to detect and isolate most of the E. coli or Salmonella spp. strains harbouring different mcr genes from food-producing animals and food products and might thus be used as a harmonized protocol for the screening of mcr genes in food-producing animals and food products in Europe.

Spread of ESC-, carbapenem- and colistin-resistant Escherichia coli clones and plasmids within and between food workers in Lebanon.

OBJECTIVES: Knowledge on the dynamic of MDR Escherichia coli in the human community is still limited, especially in low- and middle-income countries. Our goal was to decipher the dynamics of E. coli lineages and plasmids resistant to ESC, carbapenem and colistin within and between food workers in Lebanon using genomic-based approaches. METHODS: Eighty-four healthy adults working in three bakeries were sampled twice at a 6 monthly interval. E. coli resistant to ESC (ESC-E), carbapenem (CP-E) and colistin (CO-E) were collected on selective plates. Non-duplicate isolates were whole-genome sequenced using the Illumina technology and plasmid transmission was assessed by long-read sequencing. Data were analysed using bioinformatics tools and SNP-based phylogeny. RESULTS: ESC-E carriage rate reached 34.5% (t0) and 52.9% (t6), and 15 workers were positive at both t0 and t6. Carbapenem resistance (blaOXA-181, blaOXA-204, blaNDM-5) was found in five workers at t0 and two at t6, while colistin resistance (mcr-1.1) was found in five workers at t0 and one at t6. Forty-seven different STs were identified, of which three STs were predominant (ST131, n = 9; ST10, n = 5; ST69, n = 5). One worker presented the same ESC-E clone at t0 and t6. Twelve different events of clonal transmission among individuals were exemplified while plasmid transmission was only shown once. CONCLUSIONS: Our study revealed a high carriage rate of MDR E. coli (60.7%) and the emergence of CP and colistin resistance in the Lebanese community. Incidental and long-term ESC-E carriage was observed in 41.7% and 17.9% of the workers, respectively. The high clonal diversity suggests an important dynamic of acquisition and loss of MDR E. coli and limited plasmid spread.

Interplay between Bacterial Clones and Plasmids in the Spread of Antibiotic Resistance Genes in the Gut: Lessons from a Temporal Study in Veal Calves.

Intestinal carriage of extended spectrum ß-lactamase (ESBL)-producing Escherichia coli is a frequent, increasing, and worrying phenomenon, but little is known about the molecular scenario and the evolutionary forces at play. We screened 45 veal calves, known to have high prevalence of carriage, for ESBL-producing E. coli on 514 rectal swabs (one randomly selected colony per sample) collected over 6 months. We characterized the bacterial clones and plasmids carrying blaESBL genes with a combination of genotyping methods, whole genome sequencing, and conjugation assays. One hundred and seventy-three ESBL-producing E. coli isolates [blaCTX-M-1 (64.7%), blaCTX-M-14 (33.5%), or blaCTX-M-15 (1.8%)] were detected, belonging to 32 bacterial clones, mostly of phylogroup A. Calves were colonized successively by different clones with a trend in decreasing carriage. The persistence of a clone in a farm was significantly associated with the number of calves colonized. Despite a high diversity of E. coli clones and blaCTX-M-carrying plasmids, few blaCTX-M gene/plasmid/chromosomal background combinations dominated, due to (i) efficient colonization of bacterial clones and/or (ii) successful plasmid spread in various bacterial clones. The scenario "clone versus plasmid spread" depended on the farm. Thus, epistatic interactions between resistance genes, plasmids, and bacterial clones contribute to optimize fitness in specific environments. IMPORTANCE The gut microbiota is the epicenter of the emergence of resistance. Considerable amount of knowledge on the molecular mechanisms of resistance has been accumulated, but the ecological and evolutionary forces at play in nature are less studied. In this context, we performed a field work on temporal intestinal carriage of extended spectrum ß-lactamase (ESBL)-producing Escherichia coli in veal farms. Veal calves are animals with one of the highest levels of ESBL producing E. coli fecal carriage, due to early high antibiotic exposure. We were able to show that calves were colonized successively by different ESBL-producing E. coli clones, and that two main scenarios were at play in the spread of blaCTX-M genes among calves: efficient colonization of several calves by a few bacterial clones and successful plasmid spread in various bacterial clones. Such knowledge should help develop new strategies to fight the emergence of antibiotic-resistance.

Genomic Insights into Methicillin-Resistant Staphylococcus aureus spa Type t899 Isolates Belonging to Different Sequence Types.

Methicillin-resistant Staphylococcus aureus (MRSA) presenting spa type t899 is commonly associated with sequence type 9 (ST9) but is also increasingly linked to ST398. This study provides genomic insight into the diversity of t899 isolates using core genome multilocus sequence typing (cgMLST), single nucleotide polymorphism (SNP)-based phylogeny, and the description of selected antimicrobial resistance and virulence markers. The SNP-based phylogenic tree showed that isolates sharing the same spa type (t899) but different STs highly diverged in their core and accessory genomes, revealing discriminant antimicrobial resistance (AMR) and virulence markers. Our results highlighted the idea that in a surveillance context where only spa typing is used, an additional multiplex PCR for the detection of the tet(M), sak, and seg genes would be valuable in helping distinguish ST9 from ST398 isolates on a routine basis.IMPORTANCE This study showed the genetic diversity and population structure of S. aureus presenting the same spa type, t899, but belonging to different STs. Our findings revealed that these isolates vary deeply in their core and accessory genomes, contrary to what is regularly inferred from studies using spa typing only. Given that identical spa types can be associated with different STs and that spa typing only is not appropriate for S. aureus isolates that have undergone major recombination events which include the passage of the spa gene (such as in t899-positive MRSA), the combination of both MLST and spa typing methods is recommended. However, spa typing alone is still largely used in surveillance studies and basic characterization. Our data suggest that additional markers, such as tet(M), sak, and seg genes, could be implemented in an easy and inexpensive manner in order to identify S. aureus lineages with a higher accuracy.

Scarless Removal of Large Resistance Island AbaR Results in Antibiotic Susceptibility and Increased Natural Transformability in Acinetobacter baumannii.

With a great diversity in gene composition, including multiple putative antibiotic resistance genes, AbaR islands are potential contributors to multidrug resistance in Acinetobacter baumannii However, the effective contribution of AbaR to antibiotic resistance and bacterial physiology remains elusive. To address this, we sought to accurately remove AbaR islands and restore the integrity of their insertion site. To this end, we devised a versatile scarless genome editing strategy. We performed this genetic modification in two recent A. baumannii clinical strains: the strain AB5075 and the nosocomial strain AYE, which carry AbaR11 and AbaR1 islands of 19.7 kbp and 86.2 kbp, respectively. Antibiotic susceptibilities were then compared between the parental strains and their AbaR-cured derivatives. As anticipated by the predicted function of the open reading frame (ORF) of this island, the antibiotic resistance profiles were identical between the wild type and the AbaR11-cured AB5075 strains. In contrast, AbaR1 carries 25 ORFs, with predicted resistance to several classes of antibiotics, and the AYE AbaR1-cured derivative showed restored susceptibility to multiple classes of antibiotics. Moreover, curing of AbaRs restored high levels of natural transformability. Indeed, most AbaR islands are inserted into the comM gene involved in natural transformation. Our data indicate that AbaR insertion effectively inactivates comM and that the restored comM is functional. Curing of AbaR consistently resulted in highly transformable and therefore easily genetically tractable strains. Emendation of AbaR provides insight into the functional consequences of AbaR acquisition.

Trends in antimicrobial resistance among Escherichia coli from defined infections in humans and animals.

OBJECTIVES: To characterize and compare resistance trends in clinical Escherichia coli isolates from humans, food-producing animals (poultry, cattle and swine) and pets (dogs and cats). METHODS: Antibiogram results collected between January 2014 and December 2017 by MedQual [the French surveillance network for antimicrobial resistance (AMR) in bacteria isolated from the community] and RESAPATH (the French surveillance network for AMR in bacteria from diseased animals) were analysed, focusing on resistance to antibiotics of common interest to human and veterinary medicine. Resistance dynamics were investigated using generalized additive models. RESULTS: In total, 743 637 antibiograms from humans, 48 170 from food-producing animals and 7750 from pets were analysed. For each antibiotic investigated, the resistance proportions of isolates collected from humans were of the same order of magnitude as those from food-producing animals or pets. However, resistance trends in humans differed from those observed in pets and food-producing animals over the period studied. For example, resistance to third-generation cephalosporins and fluoroquinolones was almost always below 10% for both humans and animals. However, in contrast to the notable decreases in resistance observed in both food-producing animals and pets, resistance in humans decreased only slightly. CONCLUSIONS: Despite several potential biases in the data, the resistance trends remain meaningful. The strength of the parallel is based on similar data collection in humans and animals and on a similar statistical methodology. Resistance dynamics seemed specific to each species, reflecting different antibiotic-use practices. These results advocate applying the efforts already being made to reduce antibiotic use to all sectors and all species, both in human and veterinary medicine.

Development and Multicentric Validation of a Lateral Flow Immunoassay for Rapid Detection of MCR-1-Producing Enterobacteriaceae.

Colistin has become a last-resort antibiotic for the treatment of infections caused by highly drug-resistant Gram-negative bacteria. Moreover, it has been widely used in the livestock sector. As a consequence, colistin resistance is emerging worldwide. Among the colistin resistance mechanisms, the spread of the plasmid-encoded colistin resistance gene mcr-1 (mostly in Escherichia coli) is of particular concern due to its increased transferability compared to that of chromosome-encoded resistance. The early detection of MCR-1-producing bacteria is essential to prevent further spread and provide appropriate antimicrobial therapy. Lateral flow immunoassays (LFIAs) were manufactured with selected monoclonal antibodies. A collection of 177 human and 121 animal enterobacterial isolates was tested in a multicentric study. One bacterial colony grown on agar plates was suspended in extraction buffer and dispensed on the cassette. Migration was allowed for 15 min, and the results were monitored by the appearance of a specific band. The positive results showed a pink line resulting in an unambiguous interpretation. All MCR-1-producing isolates were found to be positive by the LFIA, and no false-negative results were observed. Three out of four MCR-2-producing isolates were also found to be positive. Our test does not detect MCR-3-, MCR-4-, or MCR-5-producing isolates. LFIA allows the detection of MCR-1 with 100% sensitivity and 98% specificity. This test is fast, sensitive, specific, easy to use, and cost-effective and can therefore be implemented in any microbiology laboratory worldwide. LFIA is a major tool for the rapid detection and monitoring of MCR-1 producers in humans and animals.

Fluorescence-Based Detection of Natural Transformation in Drug-Resistant Acinetobacter baumannii.

Acinetobacter baumannii is a nosocomial agent with a high propensity for developing resistance to antibiotics. This ability relies on horizontal gene transfer mechanisms occurring in the Acinetobacter genus, including natural transformation. To study natural transformation in bacteria, the most prevalent method uses selection for the acquisition of an antibiotic resistance marker in a target chromosomal locus by the recipient cell. Most clinical isolates of A. baumannii are resistant to multiple antibiotics, limiting the use of such selection-based methods. Here, we report the development of a phenotypic and selection-free method based on flow cytometry to detect transformation events in multidrug-resistant (MDR) clinical A. baumannii isolates. To this end, we engineered a translational fusion between the abundant and conserved A. baumannii nucleoprotein (HU) and the superfolder green fluorescent protein (sfGFP). The new method was benchmarked against the conventional antibiotic selection-based method. Using this new method, we investigated several parameters affecting transformation efficiencies and identified conditions of transformability one hundred times higher than those previously reported. Using optimized transformation conditions, we probed natural transformation in a set of MDR clinical and nonclinical animal A. baumannii isolates. Regardless of their origin, the majority of the isolates displayed natural transformability, indicative of a conserved trait in the species. Overall, this new method and optimized protocol will greatly facilitate the study of natural transformation in the opportunistic pathogen A. baumanniiIMPORTANCE Antibiotic resistance is a pressing global health concern with the rise of multiple and panresistant pathogens. The rapid and unfailing resistance to multiple antibiotics of the nosocomial agent Acinetobacter baumannii, notably to carbapenems, prompt to understand the mechanisms behind acquisition of new antibiotic resistance genes. Natural transformation, one of the horizontal gene transfer mechanisms in bacteria, was only recently described in A. baumannii and could explain its ability to acquire resistance genes. We developed a reliable method to probe and study natural transformation mechanism in A. baumannii More broadly, this new method based on flow cytometry will allow experimental detection and quantification of horizontal gene transfer events in multidrug-resistant A. baumannii.

Spread of blaCTX-M-15-Producing Enterobacteriaceae and OXA-23-Producing Acinetobacter baumannii Sequence Type 2 in Tunisian Seafood.

Bivalves are filter-feeding animals and markers of bacterial pollution. We report a massive spread of blaCTX-M-15 through dominant Escherichia coli and Klebsiella pneumoniae lineages and/or plasmid subtypes (F31:A4:B1) as well as the presence of OXA-23-producing Acinetobacter baumannii sequence type 2 (ST2) in seafood, highlighting a direct risk for the consumer. These findings should urge authorities to consider hospital effluents, and also farm and urban effluents, as important sources of extended-spectrum-beta-lactamase (ESBL)/carbapenemase producers that filter-feeding animals can concentrate and further spread to humans.

Emergence of blaCTX-M-55 associated with fosA, rmtB and mcr gene variants in Escherichia coli from various animal species in France.

Objectives: In Asian countries, blaCTX-M-55 is the second most common ESBL-encoding gene. blaCTX-M-55 frequently co-localizes with fosA and rmtB genes on epidemic plasmids, which remain sporadic outside Asia. During 2010-13, we investigated CTX-M-55-producing Escherichia coli isolates and their co-resistance to fosfomycin, aminoglycosides, fluoroquinolones and colistin as part of a global survey of ESBLs in animals in France. Methods: blaCTX-M-55, fosA, rmtB and plasmidic quinolone and colistin resistance genes were characterized by PCR, sequencing and hybridization experiments. Plasmids were classified according to their incompatibility groups and subtypes. Genotyping was performed by MLST and repetitive extragenic palindromic sequence-based PCR. Results: Twenty-one E. coli isolates from bovines (n = 16), dogs (n = 2), horses (n = 2) and a monkey harboured blaCTX-M-55, were MDR and belonged to ST744 (n = 9) and 10 other clones. blaCTX-M-55 was mostly located on IncF (n = 19), but also on IncI1 (n = 2) plasmids. On IncF33:A1:B1 plasmids, blaCTX-M-55 co-localized with the rmtB and aac(6')-Ib genes and in one isolate with the fosA3 allele. Ten IncF46:A-:B20 plasmids, which were found in different clones from unrelated animals, also carried the mcr-3 gene. blaCTX-M-55-carrying IncF18:A-:B1 plasmids were found in different animal species from distinct locations and periods, and one additionally carried the fosA4 gene. One isolate harboured the mcr-1 gene, which did not co-localize with blaCTX-M-55. Conclusions: A large diversity of E. coli clones and plasmid types supported the spread of blaCTX-M-55, together with atypical resistance genes, in various animal species in France. fosA and rmtB genes are emerging among animals in Europe and this issue is of concern for public health.

Extended-spectrum cephalosporin-resistant Escherichia coli isolated from chickens and chicken meat in Brazil is associated with rare and complex resistance plasmids and pandemic ST lineages.

Objectives: Brazil is the greatest exporter of chicken meat (CM) in the world. It is of utmost importance to monitor resistance to extended-spectrum cephalosporins (ESCs) in this sector because resistance to ESCs in Escherichia coli isolated from food-producing animals may contaminate humans through the food chain. Thus, the aim of this study was to characterize and compare ESC-resistant E. coli isolated from chickens and retail CM produced in south-eastern Brazil. Methods: Five CM samples and 117 chicken cloacal swabs (CCSs) were inoculated on MacConkey agar supplemented with cefotaxime. Presumptive E. coli colonies were identified and antimicrobial susceptibility was tested. Virulence and acquired blaESBL and blaAmpC genes were sought and genetic environments characterized. Isolates were typed by phylogenetic grouping, XbaI-PFGE and MLST. Results: All five CM samples and 36 CCSs (30.8%) were positive for the presence of ESC-resistant E. coli, leading to the selection of 58 resistant isolates. ESC resistance was mostly due to the presence of the chromosome-encoded blaCTX-M-2 gene, but plasmid-mediated blaCTX-M-2, blaCTX-M-8, blaCTX-M-15, blaCTX-M-55 and blaCMY-2 were also detected. Multireplicon plasmids were sporadically identified, such as IncHI2/P-blaCTX-M-2 and IncFII/N-blaCTX-M-55. Phylogroup D predominated, while PFGE and MLST revealed a high genetic diversity. Conclusions: Live Brazilian chickens and CM act as reservoirs of ESC-resistant E. coli and resistance genes are located on highly diverse genetic determinants. Potentially pathogenic strains, which may represent a threat to human health and a source of environmental contamination, were also identified. Active surveillance is therefore essential in Brazil's chicken production line.

Antimicrobial resistance plasmid reservoir in food and food-producing animals.

Antimicrobial resistance (AMR) plasmids have been recognized as important vectors for efficient spread of AMR phenotypes. The food reservoir includes both food-producing animals and food products, and a huge diversity of AMR plasmids have been reported in this sector. Based on molecular typing methods and/or whole genome sequencing approaches, certain AMR genes/plasmids combinations were found more frequently in food compared to other settings. However, the food source of a definite AMR plasmid is highly complex to confirm due to cross-sectorial transfers and international spread of AMR plasmids. For risk assessment purposes related to human health, AMR plasmids found in food and bearing genes conferring resistances to critically important antibiotics in human medicine - such as to extended-spectrum cephalosporins, carbapenems or colistin - have been under specific scrutiny these last years. Those plasmids are often multidrug resistant and their dissemination can be driven by the selective pressure exerted by any of the antibiotics concerned. Also, AMR plasmids carry numerous other genes conferring vital properties to the bacterial cell and are recurrently subjected to evolutionary steps such as hybrid plasmids, making the epidemiology of AMR plasmids in food a moving picture.

Are animals a source of Stenotrophomonas maltophilia in human infections? Contributions of a nationwide molecular study.

Stenotrophomonas maltophilia (Sm) is an archetypal environmental opportunistic bacterium responsible for health care-associated infections. The role of animals in human Sm infections is unknown. This study aims to reveal the genetic and phylogenetic relationships between pathogenic strains of Sm, both animal and human, and identify a putative role for animals as a reservoir in human infection. We phenotypically and genotypically characterized 61 Sm strains responsible for animal infections (mainly respiratory tract infections in horses) from a French nationwide veterinary laboratory network. We tested antimicrobial susceptibility and performed MLST and genogrouping using the concatenation of the seven housekeeping genes from the original MLST scheme. Excluding the eight untypeable strains owing to the lack of gene amplification, only 10 out of the 53 strains yielded a known ST (ST5, ST39, ST162, ST8, ST27, ST126, ST131). The genogroup distribution highlighted not only genogroups (genogroups 5 and 9) comprised exclusively of animal strains but also genogroups shared by human and animal strains. Interestingly, these shared genogroups were primarily groups 2 and 6, which have previously been identified as the two most frequent genogroups among human-pathogenic Sm strains, especially among respiratory pathogens. The antimicrobial susceptibility testing underlined the presence of acquired resistance: 18.8 and 7.5% of the tested isolates were resistant to the sulfonamide-trimethoprim combination and ciprofloxacin, respectively. Animal strains of Sm shared phylogenetic traits with some of the most successful human strains. The exact relationships between the human and animal strains, and the genetic support of these common traits, need to be determined.

Antimicrobial resistance in bacteria isolated from mastitis in dairy cattle in France, 2006-2016.

In dairy cattle, mastitis is the most frequent bacterial disease, and the routine use of antibiotics for treatment and prevention can drive antimicrobial resistance (AMR). The aim of our study was to estimate the levels of AMR of the 3 main bacteria isolated from dairy cattle with mastitis in France (Streptococcus uberis, Escherichia coli, and coagulase-positive staphylococci) and to investigate their changes over time. Data collected between 2006 and 2016 by the French surveillance network for AMR in pathogenic bacteria of animal origin (called RESAPATH) were analyzed. The proportions of mono- and multidrug resistance were calculated and the trends were investigated using nonlinear analyses applied to time series. Over the whole period, the lowest proportions of resistance in S. uberis isolates were observed for oxacillin (2.2%) and gentamicin (2.4%) and most resistance levels were below 20%. The trends in resistance showed some significant variation, mainly for S. uberis, but without a common pattern across the various antibiotics examined. For only 2 combinations of bacteria-antibiotic the trend in resistance showed a continuous increase from 2006 to 2016: tetracycline resistance in S. uberis isolates and third-generation cephalosporin resistance in E. coli isolates. In E. coli, the highest proportions of resistance were observed for amoxicillin (28.1%) and tetracycline (23.1%). Resistance to third-generation cephalosporins in E. coli from dairy cattle was almost nil in 2006, but reached 2.4% in December 2016. This increase is particularly concerning because these antibiotics constitute one of the latest therapeutic alternatives to fight severe infectious diseases in humans. Except for penicillin (33.9%), the proportions of resistance in coagulase-positive staphylococci were below 11% during the whole study period. Multidrug resistance (isolates with acquired resistance to at least one antibiotic in 3 or more antibiotic classes) ranged from 2.4% for coagulase-positive staphylococci to 9.9% for S. uberis. These findings can serve as guidelines for practitioners in the choice of the most appropriate antibiotic according to the prevailing epidemiological context. Ultimately, our results contribute to risk assessment of AMR and provide a baseline for setting up and evaluating control measures and designing strategies to limit AMR.

Evidence for Human Adaptation and Foodborne Transmission of Livestock-Associated Methicillin-Resistant Staphylococcus aureus.

We investigated the evolution and epidemiology of a novel livestock-associated methicillin-resistant Staphylococcus aureus strain, which colonizes and infects urban-dwelling Danes even without a Danish animal reservoir. Genetic evidence suggests both poultry and human adaptation, with poultry meat implicated as a probable source.

Sequence Type 48 Escherichia coli Carrying the blaCTX-M-1 IncI1/ST3 Plasmid in Drinking Water in France.

Drinking water has rarely been recognized as a source of antimicrobial resistance for humans, and only in low-income countries. Here, a sequence type 48 Escherichia coli isolate carrying the blaCTX-M-1 IncI1/ST3 plasmid was recovered from drinking water in France. This plasmid was similar to other blaCTX-M-1 IncI1/ST3 plasmids found previously in animals and humans. Our findings highlight the possible human transfer of extended-spectrum ß-lactamase (ESBL) genes through drinking water in high-income countries.