Marisa Haenni

Epidemic spread of Escherichia coli ST744 isolates carrying mcr-3 and blaCTX-M-55 in cattle in France

1 Montezzi LF, Campana EH, Correa LL et al. Occurrence of carbapenemaseproducing bacteria in coastal recreational waters. Int J Antimicrob Agents 2015; 45: 174–7. 2 Oliveira S, Moura RA, Silva KC et al. Isolation of KPC-2-producing Klebsiella pneumoniae strains belonging to the high-risk multiresistant clonal complex 11 (ST437 and ST340) in urban rivers. J Antimicrob Chemother 2014; 69: 849–52. 3 Yang F, Huang L, Li L et al. Discharge of KPC-2 genes from the WWTPs contributed to their enriched abundance in the receiving river. Sci Total Environ 2017; 581–582: 136–43. 4 Ahammad ZS, Sreekrishnan TR, Hands CL et al. Increased waterborne blaNDM-1 resistance gene abundances associated with seasonal human pilgrimages to the Upper Ganges River. Environ Sci Technol 2014; 48: 3014–20. 5 Giacobbe DR, Del Bono V, Trecarichi EM et al. Risk factors for bloodstream infections due to colistin-resistant KPC-producing Klebsiella pneumoniae: results from a multicenter case-control-control study. Clin Microbiol Infect 2015; 21: 1106.e1–8. 6 Wang J, Yuan M, Chen H et al. First report of Klebsiella oxytoca strain simultaneously producing NDM-1, IMP-4 and KPC-2 carbapenemases. Antimicrob Agents Chemother 2017; 61: e00877-17. 7 Feng J, Qiu Y, Yin Z et al. Coexistence of a novel KPC-2-encoding MDR plasmid and an NDM-1-encoding pNDM-HN380-like plasmid in a clinical isolate of Citrobacter freundii. J Antimicrob Chemother 2015; 70: 2987–91. 8 Hocquet D, Llanes C, Thouverez M et al. Evidence for induction of integronbased antibiotic resistance by the SOS response in a clinical setting. PLoS Pathog 2012; 8: e1002778. 9 Shen P, Wei Z, Jiang Y et al. Novel genetic environment of the carbapenem-hydrolyzing b-lactamase KPC-2 among Enterobacteriaceae in China. Antimicrob Agents Chemother 2009; 53: 4333–8. 10 Wang L, Fang H, Feng J et al. Complete sequences of KPC-2-encoding plasmid p628-KPC and CTX-M-55-encoding p628-CTXM coexisted in Klebsiella pneumoniae. Front Microbiol 2015; 6: 838. J Antimicrob Chemother 2018; 73: 533–536 doi:10.1093/jac/dkx418 Advance Access publication 22 November 2017

Multidrug resistance Salmonella genomic island 1 in a Morganella morganii subsp. morganii human clinical isolate from France

Since its initial identification in epidemic multidrug-resistant Salmonella enterica serovar Typhimurium DT104 strains, several SGI1 variants, SGI1 lineages, and SGI1-related elements (SGI2, PGI1, and AGI1) have been described in many bacterial genera (Salmonella, Proteus, Morganella, Vibrio, Shewanella, etc.). They constitute a family of multidrug resistance site-specific integrative elements acquired by horizontal gene transfer, SGI1 being the best-characterized element. The horizontal transfer of SGI1/PGI1 elements into other genera is of public health concern, notably with regard to the spread of critically important resistance genes such as ESBL and carbapenemase genes. The identification of SGI1 in Morganella morganii raises the issue of (i) the potential for SGI1 to emerge in other human pathogens and (ii) its bacterial host range. Further surveillance and research are needed to understand the epidemiology, the spread, and the importance of the members of this SGI1 family of integrative elements in contributing to antibiotic resistance development. ABSTRACT Salmonella genomic island 1 (SGI1) is a multidrug resistance integrative mobilizable element that harbors a great diversity of antimicrobial resistance gene clusters described in numerous Salmonella enterica serovars and also in Proteus mirabilis. A serious threat to public health was revealed in the recent description in P. mirabilis of a SGI1-derivative multidrug resistance island named PGI1 (Proteus genomic island 1) carrying extended-spectrum-β-lactamase (ESBL) and metallo-β-lactamase resistance genes, blaVEB-6 and blaNDM-1, respectively. Here, we report the first description of Salmonella genomic island 1 (SGI1) in a multidrug-resistant clinical Morganella morganii subsp. morganii strain isolated from a patient in France in 2013. Complete-genome sequencing of the strain revealed SGI1 variant SGI1-L carrying resistance genes dfrA15, floR, tetA(G), blaPSE-1 (now referred to as blaCARB-2), and sul1, conferring resistance to trimethoprim, phenicols, tetracyclines, amoxicillin, and sulfonamides, respectively. The SGI1-L variant was integrated into the usual chromosome-specific integration site at the 3′ end of the trmE gene. Beyond Salmonella enterica and Proteus mirabilis, the SGI1 integrative mobilizable element may thus also disseminate its multidrug resistance phenotype in another genus belonging to the Proteae tribe of the family Enterobacteriaceae. IMPORTANCE Since its initial identification in epidemic multidrug-resistant Salmonella enterica serovar Typhimurium DT104 strains, several SGI1 variants, SGI1 lineages, and SGI1-related elements (SGI2, PGI1, and AGI1) have been described in many bacterial genera (Salmonella, Proteus, Morganella, Vibrio, Shewanella, etc.). They constitute a family of multidrug resistance site-specific integrative elements acquired by horizontal gene transfer, SGI1 being the best-characterized element. The horizontal transfer of SGI1/PGI1 elements into other genera is of public health concern, notably with regard to the spread of critically important resistance genes such as ESBL and carbapenemase genes. The identification of SGI1 in Morganella morganii raises the issue of (i) the potential for SGI1 to emerge in other human pathogens and (ii) its bacterial host range. Further surveillance and research are needed to understand the epidemiology, the spread, and the importance of the members of this SGI1 family of integrative elements in contributing to antibiotic resistance development.

High prevalence of ESBLs in retail chicken meat despite reduced use of antimicrobials in chicken production, France

Extended-spectrum cephalosporins (ESCs) are critically important antibiotics for humans and their use in animals poses a potential threat for public health. Chicken represents an increasing part of the human diet and has also been regarded as a source of ESC-resistant Enterobacteriaceae because of the worldwide off-label use of ceftiofur, a broad-spectrum cephalosporin. Thus, numerous studies pointed out chicken as a reservoir of ESBL/pAmpC genes, plasmids and/or clones at risk for humans. In France, late 2011, strong political pressure led to a drastic reduction of ceftiofur use and all other antibiotics in chicken production. Here, we ascertained the potential impact of those efforts on the prevalence of ESC-resistant E. coli in retail chicken. From October 2015 to January 2016, of 48 unrelated pieces of meat (chicken legs) belonging to four different brands, 44 (91.7%) were positive for ESC-resistant E. coli. The blaCTX-M-1 gene was highly prevalent (68/74, 91.9%), mostly located on IncI1/ST3 plasmids (65/68, 95.6%). Other ESBL/pAmpC genes (blaTEM-52, blaSHV-12, blaCMY-2) were carried by IncX1, IncI1/ST36, IncI1/ST95, IncA/C or IncK plasmids. The positive isolates were non-clonal, suggesting a horizontal spread of the ESBL/pAmpC genes. Obviously, the strong decrease of antimicrobial use in chicken farms had no impact yet on the ESBL/pAmpC prevalence in retail chicken meat in France. A human source of these ESBL/pAmpC genes is unlikely as blaCTX-M-1 IncI1/ST3 plasmids are dominant in animals and rare in humans. Our data question the real impact of the decrease of antimicrobial use in chicken production on ESBL contamination of chicken meat and point out the risk of ESBL/AmpCs human transfer through the food chain.

Detection of SGI1/PGI1 Elements and Resistance to Extended-Spectrum Cephalosporins in Proteae of Animal Origin in France

Proteae, and especially Proteus mirabilis, are often the cause of urinary tract infections (UTIs) in humans. They were reported as carriers of extended-spectrum β-lactamase (ESBL) genes, and recently of carbapenemases, mostly carried by the Salmonella genomic island 1 (SGI1) and Proteus genomic island 1 (PGI1). Proteae have also lately become an increasing cause of UTIs in companion animals, but antimicrobial susceptibility data in animals are still scarce. Here, we report the characterization of 468 clinical epidemiologically unrelated Proteae strains from animals collected between 2013 and 2015 in France. Seventeen P. mirabilis strains (3.6%) were positive for SGI1/PGI1 and 18 Proteae (3.8%) were resistant to extended-spectrum cephalosporins (ESC). The 28 isolates carrying SGI1/PGI1 and/or ESC-resistance genes were isolated from cats, dogs, and horses. ESBL genes were detected in six genetically related P. mirabilis harboring blaV EB-6 on the SGI1-V variant, but also independently of the SGI1-V, in 3 P. mirabilis strains (blaVEB-6 and blaCTX-M-15) and 1 Providencia rettgeri strain (blaCTX-M-1). The AmpC resistance genes blaCMY -2 and/or blaDHA-16 were detected in 9 P. mirabilis strains. One strain presented both an ESBL and AmpC gene. Interestingly, the majority of the ESBL/AmpC resistance genes were located on the chromosome. In conclusion, multiple ESC-resistance genetic determinants are circulating in French animals, even though SGI1-V-carrying P. mirabilis seems to be mainly responsible for the spread of the ESBL gene blaVEB-6 in dogs and horses. These results are of public health relevance and show that companion animals in close contact with humans should be regarded as a potential reservoir of ESC-resistant bacteria as well as a reservoir of ESC-resistance genes that could further disseminate to human pathogens.

CHROMagar COL-APSE: a selective bacterial culture medium for the isolation and differentiation of colistin-resistant Gram-negative pathogens.

Purpose. A selective chromogenic culture medium for the laboratory isolation and differentiation of colistin resistant Acinetobacter, Pseudomonas, Stenotrophomonas and Enterobacteriaceae spp. (CHROMagar COL‐APSE) was developed, evaluated and compared to an existing selective bacterial culture medium (SuperPolymyxin). Methodology. The medium was challenged with 84 isolates, including polymyxin B (POL B)‐susceptible and ‐resistant type strains and colistin (COL)‐resistant organisms recovered from human and animal samples. Susceptibility to COL and POL B was determined by agar dilution and broth microtitre dilution. The lower limit for the detection of COL‐resistant organisms was also calculated for both CHROMagar COL‐APSE and SuperPolymyxin media. The ability to isolate and correctly differentiate COL‐resistant organisms within mixed cultures was also assessed and compared using both media. Results. Using CHROMagar COL‐APSE, Gram‐negative pathogens (n=71) with intrinsic (n=8) or acquired COL (n=63) resistance were recovered with 100% specificity down to the lower limit of detection of 101 colony‐forming units (c.f.u.). The growth on SuperPolymyxin was similar, but notably weaker for COL‐resistant non‐fermentative bacteria (Acinetobacter, Pseudomonas and Stenotrophomonas). CHROMagar COL‐APSE was also more sensitive in supporting the growth of Enterobacteriaceae with COL resistance associated with the carriage of mcr‐1. Conclusion. CHROMagar COL‐APSE is a sensitive and specific medium for the growth of COL‐resistant bacterial pathogens. Due to the low limit of detection (101 c.f.u.), it may be useful as a primary isolation medium in the surveillance and recovery of COL‐resistant bacteria from complex human, veterinary and environmental samples, especially those with plasmid‐mediated MCR‐1 or novel mechanisms of polymyxin resistance.

Antimicrobial resistance trends in Escherichia coli isolated from diseased food‐producing animals in France: A 14‐year period time‐series study

Antimicrobial resistance (AMR) among bacteria isolated from food‐producing animals is a growing concern with implications for public health. AMR surveillance is essential to identify resistance trends and help in the design of effective and efficient control strategies. The aim of the study was to describe the antimicrobial susceptibility of pathogenic Escherichia coli isolated from three livestock productions in France (cattle, swine and poultry). The trend in resistance to the most commonly prescribed antibiotics in animal health was analysed as follows: amoxicillin (penicillin), spectinomycin or streptomycin (aminoglycoside), tetracycline and trimethoprim‐sulfamethoxazole/Enrofloxacin and ceftiofur were also taken into account as members of critically important antimicrobial families in human and veterinary medicine, that is fluoroquinolones and third‐generation cephalosporins, respectively. Data collected between 2002 and 2015 by the French national surveillance network of AMR referred to as RESAPATH were analysed. Resistance trends were investigated using non‐linear analysis (generalized additive models) applied to time‐series stratified by livestock production and antibiotic. Irrespective of the species and the antibiotic considered, resistance signals over time showed no significant annual cycle. Resistance to third‐generation cephalosporins emerged during the period of the study, with a peak at 22% [20.5; 24.0] in poultry in 2010, decreasing afterwards, while it remained consistently below 10% for the other species. The proportion of resistance to fluoroquinolones was broadly similar between species and remained under 30%, with a slight decreasing trend after 2009. Resistances to tetracycline and amoxicillin remained high, between 90% and 40% over time in cattle and swine. After 2010, there was a decrease in resistance to these antibiotics for all species, especially to tetracycline for poultry with a drop from 84% in 2009 to 43% in 2015. These results contribute to risk assessment and constitute objective evidence on which to evaluate the efficacy of control measures implemented to limit AMR occurrence.

OXA-48 and CTX-M-15 extended-spectrum beta-lactamases in raw milk in Lebanon: epidemic spread of dominant Klebsiella pneumoniae clones

Raw milk has recently been reported as a source of extended-spectrum beta-lactamase (ESBL) and carbapenemase genes. We thus investigated the prevalence of ESBL- and carbapenemase-producing Enterobacteriaceae in raw milk in Lebanon in order to assess the risk of transfer of these bacteria to humans. A high prevalence (30.2 %) of CTX-M-15-producing K. pneumoniae was detected in raw bovine milk. Three main K. pneumoniae clones were identified by PFGE and MLST typing. Southern blot experiments revealed that one of these clones carried the blaCTX-M-15 gene chromosomally. Moreover, one OXA-48-producing K. pneumoniae ST530 and seven CTX-M-15-producing Escherichia coli sharing the same ST were also detected. These findings highlight the spread of dominant CTX-M-15-producing K. pneumoniae clones and OXA-48-producing isolates in the food chain. Milk, which is mostly consumed raw in Lebanon, may be a source of human exposure to ESBLs and carbapenemases.

KPC-3-producing ST167 Escherichia coli from mussels bought at a retail market in Tunisia

Unité de recherche: Résistances bactériennes émergentes et sécurité des soins ‘UR12SP37’, Laboratoire de Microbiologie, Hôpital Universitaire Sahloul, Sousse, Tunisia; Institut Supérieur des Sciences Appliquées et de Technologie de Mahdia, Université de Monastir, Mahdia, Tunisia; Service de Bactériologie, Centre Hospitalo-Universitaire de Bichat, APHP, Paris, France; INSERM, IAME, UMR 1137, Paris, France; Université Paris Diderot, IAME, UMR 1137, Sorbonne Paris Cité, Paris, France; Laboratoire de Biochimie, APHP, Hôpitaux Universitaires Paris Nord Val-de-Seine, Site Bichat Claude-Bernard, Paris, France; Unité Antibiorésistance et Virulence Bactériennes, ANSES – Université de Lyon, Lyon, France; Faculté de Pharmacie de Monastir, Université de Monastir, Monastir, Tunisia; Faculté de Médecine de Sousse, Université de Sousse, Sousse, Tunisia

Resistance of Animal Strains of Pseudomonas aeruginosa to Carbapenems

Carbapenems are major antibiotics reserved to human medicine. This study aimed to investigate the mechanisms of carbapenem resistance of a selection of Pseudomonas aeruginosa veterinary strains from the French network Resapath. Thirty (5.7%) imipenem and/or meropenem non-susceptible P. aeruginosa of canine (n = 24), feline (n = 5), or bovine (n = 1) origin were identified in a large collection of 527 veterinary strains gathered by the Resapath. These resistant isolates belonged to 25 MultiLocus Sequence Types (MLST), of which 17 (68%) are shared with clinical (human) strains, such as high risk clones ST233 and ST395. Interestingly, none of the veterinary strains produced a carbapenemase, and only six of them (20%) harbored deletions or insertion sequence (IS) disrupting the porin OprD gene. The remaining 24 strains contained mutations or IS in various loci resulting in down-regulation of gene oprD coupled with upregulation of efflux system CzcCBA (n = 3; activation of sensor kinase CzcS ± CopS), MexEF-OprN (n = 4; alteration of oxido reductase MexS), MexXY (n = 8; activation of two-component system ParRS), or MexAB-OprM (n = 12; alteration of regulator MexR, NalC ± NalD). Two efflux pumps were co-produced simultaneously in three mutants. Finally, in 11 out of 12 strains displaying an intact porin OprD, derepression of MexAB-OprM accounted for a decreased susceptibility to meropenem relative to imipenem. Though not treated by carbapenems, animals thus represent a reservoir of multidrug resistant P. aeruginosa strains potentially able to contaminate fragile outpatients.

Extended-spectrum beta-lactamase (ESBL)- and carbapenemase-producing Enterobacteriaceae in water sources in Lebanon

Extended-spectrum beta-lactamases (ESBLs) have been recurrently reported in both human and veterinary medicine, and carbapenemases have also emerged in these two sectors. Such resistance phenotypes were increasingly reported in the environment, which both receives and further disseminates multidrug-resistant (MDR) bacteria. Here, we report the high contamination of water samples (68.2%; 15/22) collected in estuaries in Lebanon. From these 15 contaminated sites, a total of 21 ESBL-producing (mostly harbouring the blaCTX-M-15 gene) and four carbapenemase-producing (two blaOXA-48 and two blaOXA-244) Enterobacteriaceae were recovered. ESBL contamination was also identified in water samples collected from rural wells and spring water, although at a lower frequency. Indeed, 1.9% (3/155) and 6.1% (7/115) of the wells and springs were contaminated, respectively, and all identified isolates were CTX-M-15-producing E. coli. Interestingly, sequence types (STs) previously associated both with animal and human reservoirs were detected (ST38, ST10 and ST131), suggesting a complex source of contamination. This situation is alarming since water drawn from wells or springs is directly intended for human consumption in Lebanon without any further treatment. Moreover, even though water from estuaries is not intended for human consumption, it is used to water animals and irrigate crops. Consequently, water contamination by ESBLs and carbapenemases in Lebanon is potentially a major risk to public health. Part of this work was presented at the 7th Symposium on Antimicrobial Resistance in Animals and the Environment (ARAE).