BackgroundCarbapenem-resistant Enterobacteriaceae pose a serious threat to public health worldwide, and the role of companion animals as a reservoir is still unclear.AimsThis 4-month prospective observational study evaluated carriage of carbapenem-resistant Enterobacteriaceae at admission and after hospitalisation in a large referral hospital for companion animals in Switzerland.MethodsRectal swabs of dogs and cats expected to be hospitalised for at least 48 h were taken from May to August 2018 and analysed for the presence of carbapenem-resistant Enterobacteriaceae using selective agar plates. Resistant isolates were further characterised analysing whole genome sequences for resistance gene and plasmid identification, and ad hoc core genome multilocus sequence typing.ResultsThis study revealed nosocomial acquisition of Escherichia coli harbouring the carbapenemase gene bla OXA-181, the pAmpC cephalosporinase gene bla CMY-42 as well as quinolone resistance associated with qnrS1 and mutations in the topoisomerases II (GyrA) and IV (ParC). The bla OXA-181 and qnrS1 genes were identified on a 51 kb IncX3 plasmid and bla CMY-42 on a 47 kb IncI1 plasmid. All isolates belonged to sequence type ST410 and were genetically highly related. This E. coli clone was detected in 17 of 100 dogs and four of 34 cats after hospitalisation (21.6%), only one of the tested animals having tested positive at admission (0.75%). Two positive animals were still carriers 4 months after hospital discharge, but were negative after 6 months.ConclusionsCompanion animals may acquire carbapenemase-producing E. coli during hospitalisation, posing the risk of further dissemination to the animal and human population and to the environment.

The blaCMY -2/4-carrying IncB/O/K-like plasmids of seven Escherichia coli strains from poultry, poultry meat and human urine samples were examined using comparative analysis of whole plasmid sequences. The incompatibility group was determined by analysis of the incRNAI region and conjugation assays with strains containing the IncK and IncB/O reference plasmids. Strains were additionally characterized using MLST and MIC determination. The complete DNA sequences of all plasmids showed an average nucleotide identity of 91.3%. Plasmids were detected in E. coli sequence type (ST) 131, ST38, ST420, ST1431, ST1564 and belonged to a new plasmid variant (IncK2) within the IncK and IncB/O groups. Notably, one E. coli from poultry meat and one from human contained the same plasmid. The presence of a common recently recognized IncK2 plasmid in diverse E. coli from human urine isolates and poultry meat production suggests that the IncK2 plasmids originated from a common progenitor and have the capability to spread to genetically diverse E. coli in different reservoirs. This discovery is alarming and stresses the need of rapidly introducing strict hygiene measures throughout the food chain, limiting the spread of such plasmids in the human settings.

Since the first report of the plasmid-mediated colistin resistance gene mcr-1 in November 2015 in China (1), several studies have confirmed its worldwide spread in different animal and human environments (2, 3). The routes of dissemination of mcr-1 have been associated with travelers and trading of food animals (4, 5). Nevertheless, the presence of this gene and of the type of mediating plasmids in sub-Saharan Africa in avian-pathogenic Escherichia coli (APEC) causing airsacculitis in broiler chickens has not yet been reported.In South Africa, antimicrobial treatment against APEC has become challenging due to the increased number of strains resistant to multiple antimicrobials. In order to provide targeted therapy, MICs of colistin have been determined for a total of 4,934 E. coli isolates from air sac lesions in different broiler operations since 2008 using the Müller-Hinton broth microdilution method (Deltamune [Pty] Ltd., Lyttelton, South Africa). The number of E. coli isolates with a MIC of colistin of Ն4 g/ml has significantly increased from an average of 4.5% for the years 2008 to 2014 to 13.6% in 2015 (P Ͻ 0.05, Dunn's test) (Fig. 1).The mcr-1 gene has been sought by PCR in strains isolated in the last quarter of 2015 (1). Out of 20 of 108 colistin-resistant E. coli isolates among 797 strains of 2015, 19 contained mcr-1. These strains originated from 6 geographically distant broiler operations and exhibited 18 different repetitive element palindromic PCR (rep-PCR) profiles (6) (see Table S1 in the supplemental material). Whole-plasmid sequence of strain VT55363 was obtained using the MiSeq Reagent kit v2 (Illumina Inc.) (Labormedizinisches Zentrum Dr Risch, Bern-Liebefeld, Switzerland), and the circular form was closed by PCR and subsequent Sanger sequencing. While the mcr-1-containing element (ISApl1-mcr-1) of the 62,218-bp plasmid pVT553 was identical to that present on pHNSHP45 (accession no. KP347127) originally reported in a pig from China, the two plasmids were not structurally related. Plasmid pVT553 shared 96% identity to plasmid pSH146_65 originally detected in Salmonella enterica subsp. enterica serovar Heidelberg sequenced in the United States (accession no. JN983044). Plasmids pVT553 and pSH146_65 belonged to incompatibility group IncI2 and shared the same backbone except for the insertion of the ISApl1-mcr-1 element in pVT553 and a bla CMY-2 -containing element (ISEcp1-bla CMY-2 ) in pSH146_65 (Fig. 2). Insertion of ISApl1-mcr-1 occurred within the pil region of pVT553, deleting pilS, essential for pilus formation (7), explaining the absence of conjugative transfer into E. coli strain JF33 (Rif r ) as tested by filter mating and selection on Mül-ler-Hinton agar containing 50 g/ml rifampin and 4 g/ml colistin. Strain VT55363 belonged to sequence type (ST) 3640 (mlst .warwick.ac.uk) and also contained genes conferring resistance to tetracycline [tet(A)], sulfonamides (sul3), trimethoprim (dfrA12), and streptomycin (aadA1, strA, and strB), which were not located on pVT553. The MICs of 15 additio...

Prevalence and genetic relatedness were determined for third-generation cephalosporin-resistant Escherichia coli (3GC-R-Ec) detected in Swiss beef, veal, pork, and poultry retail meat. Samples from meat-packing plants (MPPs) processing 70% of the slaughtered animals in Switzerland were purchased at different intervals between April and June 2013 and analyzed. Sixty-nine 3GC-R-Ec isolates were obtained and characterized by microarray, PCR/DNA sequencing, Multi Locus Sequence Typing (MLST), and plasmid replicon typing. Plasmids of selected strains were transformed by electroporation into E. coli TOP10 cells and analyzed by plasmid MLST. The prevalence of 3GC-R-Ec was 73.3% in chicken and 2% in beef meat. No 3GC-R-Ec were found in pork and veal. Overall, the bla(CTX-M-1) (79.4%), bla(CMY-2) (17.6%), bla(CMY-4) (1.5%), and bla(SHV-12) (1.5%) β-lactamase genes were detected, as well as other genes conferring resistance to chloramphenicol (cmlA1-like), sulfonamides (sul), tetracycline (tet), and trimethoprim (dfrA). The 3GC-R-Ec from chicken meat often harbored virulence genes associated with avian pathogens. Plasmid incompatibility (Inc) groups IncI1, IncFIB, IncFII, and IncB/O were the most frequent. A high rate of clonality (e.g., ST1304, ST38, and ST93) among isolates from the same MPPs suggests that strains persist at the plant and spread to meat at the carcass-processing stage. Additionally, the presence of the blaCTX-M-1 gene on an IncI1 plasmid sequence type 3 (IncI1/pST3) in genetically diverse strains indicates interstrain spread of an epidemic plasmid. The bla(CMY-2) and bla(CMY-4) genes were located on IncB/O plasmids. This study represents the first comprehensive assessment of 3GC-R-Ec in meat in Switzerland. It demonstrates the need for monitoring contaminants and for the adaptation of the Hazard Analysis and Critical Control Point concept to avoid the spread of multidrug-resistant bacteria through the food chain.

Gram-stain-positive cocci were isolated from miscellaneous sites of the skin of healthy dogs as well as from infection sites in dogs. The closest relative by sequencing of the 16S rRNA gene was Macrococcus caseolyticus with 99.7 % sequence identity, but compared with M. caseolyticus, the novel strains shared only 90.8 to 93.5 % DNA sequence identity with cpn60, dnaJ, rpoB and sodA partial genes, respectively. The novel strains also exhibited differential phenotypic characteristics from M. caseolyticus, and the majority displayed a visible haemolysis on sheep blood agar, while M. caseolyticus did not have any haemolytic activity. They generated different matrix-assisted laser-desorption/ionization time-of-flight (MALDI-TOF) MS spectral profiles compared with the other species of the genus Macrococcus. Furthermore, strain KM 45013 shared only 53.7 % DNA-DNA relatedness with the type strain of M. caseolyticus, confirming that they do not belong to the same species. The DNA G+C content of strain KM 45013 was 36.9 mol%. The most abundant fatty acids were C14 : 0, C18 : 3ω6c (6, 9, 12) and C16 : 0 n alcohol. MK-6 was the menaquinone type of KM 45013. Cell-wall structure analysis revealed that the peptidoglycan type was A3α l-Lys-Gly2-l-Ser. Based on genotypic and chemotaxonomic characteristics, we propose to classify these strains within a novel species of the genus Macrococcus for which the name Macrococcus canis sp. nov. is proposed. The type strain is KM 45013 (=DSM 101690=CCOS 969=CCUG 68920).

We characterized the genetic environment of mcr-1 in colistin-resistant Escherichia coli strains isolated in Switzerland during 2014 to 2016 from humans (n ϭ 3) and chicken meat (n ϭ 6). Whole-genome and plasmid sequencing identified the mcr-1 gene integrated in IncX4 (of which, one strain carried the mcr-1.2 variant), IncI2, IncHI2, and novel IncK2 plasmids (overall, n ϭ 7), as well as in the bacterial chromosome (n ϭ 2) in single or duplicate copies. Our study supports the easy mobilization of mcr-1 across diverse genetic locations.

Macrococcus caseolyticus belongs to the normal bacterial flora of dairy cows and does not usually cause disease. However, methicillin-resistant M. caseolyticus strains were isolated from bovine mastitis milk. These bacteria had acquired a chromosomal island (McRImecD-1 or McRImecD-2) carrying the methicillin resistance gene mecD. To gain insight into the distribution of McRImecD types in M. caseolyticus from cattle, 33 mecD-containing strains from Switzerland were characterized using molecular techniques, including multilocus sequence typing, antibiotic resistance gene identification, and PCR-based McRImecD typing. In addition, the same genetic features were analyzed in 27 mecD-containing M. caseolyticus strains isolated from bovine bulk milk in England/Wales using publicly available whole-genome sequences. The 60 strains belonged to 24 different sequence types (STs), with strains belonging to ST5, ST6, ST21, and ST26 observed in both Switzerland and England/Wales. McRImecD-1 was found in different STs from Switzerland (n = 19) and England/Wales (n = 4). McRImecD-2 was only found in 7 strains from Switzerland, all of which belonged to ST6. A novel island, McRImecD-3, which contains a complete mecD operon (mecD-mecR1m-mecIm [where the subscript m indicates Macrococcus]) combined with the left part of McRImecD-2 and the right part of McRImecD-1, was found in heterogeneous STs from both collections (Switzerland, n = 7; England/Wales, n = 21). Two strains from England/Wales carried a truncated McRImecD-3. Phylogenetic analyses revealed no clustering of strains according to geographical origin or carriage of McRImecD-1 and McRImecD-3. Circular excisions were also detected for McRImecD-1 and McRImecD-3 by PCR. The analyses indicate that these islands are mobile and may spread by horizontal gene transfer between genetically diverse M. caseolyticus strains. IMPORTANCE Since its first description in 2017, the methicillin resistance gene mecD has been detected in M. caseolyticus strains from different cattle sources and countries. Our study provides new insights into the molecular diversity of mecD-carrying M. caseolyticus strains by using two approaches to characterize mecD elements: (i) multiplex PCR for molecular typing of McRImecD and (ii) read mapping against reference sequences to identify McRImecD types in silico. In combination with multilocus sequence typing, this approach can be used for molecular characterization and surveillance of M. caseolyticus carrying mecD.

We screened a total of 340 veterinarians (including general practitioners, small animal practitioners, large animal practitioners, veterinarians working in different veterinary services or industry), and 29 veterinary assistants for nasal carriage of methicillin-resistant Staphylococcus aureus (MRSA) and Staphylococcus pseudintermedius (MRSP) at the 2012 Swiss veterinary annual meeting. MRSA isolates (n = 14) were detected in 3.8 % (95 % CI 2.1 - 6.3 %) of the participants whereas MRSP was not detected. Large animal practitioners were carriers of livestock-associated MRSA (LA-MRSA) ST398-t011-V (n = 2), ST398-t011-IV (n = 4), and ST398-t034-V (n = 1). On the other hand, participants working with small animals harbored human healthcare-associated MRSA (HCA-MRSA) which belonged to epidemic lineages ST225-t003-II (n = 2), ST225-t014-II (n = 1), ST5-t002-II (n = 2), ST5-t283-IV (n = 1), and ST88-t186-IV (n = 1). HCA-MRSA harbored virulence factors such as enterotoxins, β-hemolysin converting phage and leukocidins. None of the MRSA isolates carried Panton-Valentine leukocidin (PVL). In addition to the methicillin resistance gene mecA, LA-MRSA ST398 isolates generally contained additional antibiotic resistance genes conferring resistance to tetracycline [tet(M) and tet(K)], trimethoprim [dfrK, dfrG], and the aminoglycosides gentamicin and kanamycin [aac(6')-Ie - aph(2')-Ia]. On the other hand, HCA-MRSA ST5 and ST225 mainly contained genes conferring resistance to the macrolide, lincosamide and streptogramin B antibiotics [erm(A)], to spectinomycin [ant(9)-Ia], amikacin and tobramycin [ant(4')-Ia], and to fluoroquinolones [amino acid substitutions in GrlA (S84L) and GyrA (S80F and S81P)]. MRSA carriage may represent an occupational risk and veterinarians should be aware of possible MRSA colonization and potential for developing infection or for transmitting these strains. Professional exposure to animals should be reported upon hospitalization and before medical intervention to allow for preventive measures. Infection prevention measures are also indicated in veterinary medicine to avoid MRSA transmission between humans and animals, and to limit the spread of MRSA both in the community, and to animal and human hospitals.