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Aims: This study was to investigate and to characterize methicillin-resistant coagulase-positive staphylococci (MRCoPS) harboring in dogs and people associated with dogs in Thailand. Methods and Results: Staphylococci were collected from 100 dogs, 100 dog owners, 200 small animal veterinarians and 100 people without pet association. Species of MRCoPS were identified phenotypically and genotypically. Molecular characteristics were determined by multilocus sequence typing (MLST), pulsedfield gel electrophoresis (PFGE) and SCCmec typing, and antimicrobial susceptibility was assayed by broth microdilution and by microarray analysis for resistance genes. Methicillin-resistant Staphylococcus pseudintermedius (MRSP), methicillin-resistant Staphylococcus schleiferi subsp. coagulans (MRSSc) and methicillin-resistant Staphylococcus aureus (MRSA) were isolated from dogs (45, 17 and 1%, respectively), veterinarians (8, 2 and 1Á5%, respectively) and dog owners (3, 2 and 0%, respectively). Seventeen sequence types (STs) were identified among 83 MRSP isolates which specifically carried SCCmec V, II-III, ΨSCCmec 57395 and three uncharacterized SCCmec types. MRSP ST 45, 68 and novel STs including 169, 178, 181 and 183 were shared among canine and human isolates. Most of MRSA ST398 and MRSSc carried SCCmec type V. The MRCoPS commonly displayed multiple resistances to tested antimicrobials and carried various resistance genes. Conclusion: Variety of MRCoPS, especially new MRSP clones, distributed in dogs and people in Thailand. Significance and Impact of the Study: The existence of MRCoPS circulating between dogs and humans in Thailand provides indirect evidence of interspecies transmission and represents a potential public health hazard.

Leptospirosis is an important zoonotic disease that is often associated with animal carriers and contamination of the environment via infected urine. This study aimed to assess pathogenic leptospiral carriage in Nan province, a rural area of Thailand where leptospirosis is endemic. Samples from 20 villages were obtained during the period 2013 to 2016, comprising urine samples collected from asymptomatic people (n=37) and domestic animals (n=342), and environmental water samples (n=14). Leptospira were cultured in Ellinghauson McCullough Johnson and Harris (EMJH) media. An rrs nested PCR identified 9.92% (95% confidence interval (CI) 6.96-12.88) of the urine and water samples as being positive for Leptospira spp., and phylogenetic analysis was conducted on the 443bp amplicons. Leptospira weilii, which has not previously been identified in Thailand, was recovered from 13 cattle, 9 pigs, 2 dogs, 2 water samples and 1 goat. L. interrogans was found in 4 dogs, 3 pigs, 3 cattle, 1 human and 1 water sample. Four leptospiral strains were isolated and multilocus sequence typing (MLST) analysis was performed on these. Three novel sequence types were identified, including two singletons of L. interrogans in ST26 and ST33, and one of L. weilii in ST94, with this having a close relationship to previous isolates from cases of human leptospirosis in Laos and China. Our results revealed that pathogenic Leptospira occur commonly in asymptomatic domestic animals, humans and environmental water samples in Nan Province, and emphasize the high potential for zoonotic transmission in the province.

A novel staphylococcal cassette chromosome mec (SCCmec) composite island (SCCmec AI16 -SCCczr AI16 -CI) was identified in Staphylococcus pseudintermedius. Four integration site sequences for SCC subdivided the 60,734-bp island into 41,232-bp SCCmec AI16 , 19,400-bp SCCczr AI16 , and 102-bp SCC-like AI16 elements. SCCmec AI16 represents a new combination of ccrA1B3 genes with a class A mec complex. SCCczr AI16 contains ccrA1B6 and genes related to restriction modification and heavy metal resistance. SCCmec AI16 -SCCczr AI16 -CI was found in methicillin-resistant S. pseudintermedius sequence type 112 (ST112) and ST111 isolated from dogs and veterinarians in Thailand. Methicillin-resistant Staphylococcus pseudintermedius (MRSP) has been associated with clinical manifestations in companion animals and occasionally causes diseases in humans (1, 2). MRSP contains the staphylococcal cassette chromosome mec (SCCmec) element, which is defined by the presence of the mec gene that mediates ␤-lactam resistance and by the cassette chromosome recombinase (ccr) gene(s) responsible for site-specific integration/excision of the element (3). Composite SCC structures containing two ccr gene complexes likely resulting from multiple element integrations have been increasingly reported in Staphylococcus aureus and in coagulase-negative staphylococci where such structures frequently carry heavy metal resistance gene clusters (3-9). So far, only five SCCmec elements have been completely characterized in MRSP, including SCCmec II-III, SCCmec VII-241, SCCmec V, ⌿SCCmec 57395 , and SCCmec IV, and are mainly associated with predominant clones of the following sequence types (ST): ST71, ST93, ST68, ST233, ST45,. Recently, MRSP strains of ST112 and ST111 (a single locus variant of ST112) isolated from dogs and veterinarians in Thailand were found to contain an unusual combination of the class A mec gene complex with the type I ccr gene complex, suggesting a new SCCmec type (15). We therefore aimed to characterize this novel SCCmec element and to determine whether it is conserved within other MRSP isolates of the same clonal lineage.Identification of the SCCmec AI16 -SCCczr AI16 composite element (CI). MRSP strains of ST111 and ST112 were obtained from a previous study and are listed in Table 1 (15). Phenotypic and genotypic characteristics, including those determined by antimicrobial susceptibility testing and antibiotic resistance gene detection, pulsed-field gel electrophoresis (PFGE), and multilocus sequence typing (MLST), were determined previously (15). In this study, the strains were additionally tested for mec-associated direct repeat unit (dru) and spa types (16, 17), for the presence of the novel SCCmec element, and for resistance to heavy metals. The genomic DNA of all strains was extracted using the peqGOLD bacterial DNA kit (Peqlab Biotechnologie GmBH, Jena, Germany), and that of strain AI16 was sequenced using Ion Torrent semiconductor (Life Technologies, Carlsbad, CA) and Illumina MiSeq (Illumina, San Diego, CA) technologies. S...

Objectives To analyse the genetic context of mecB in two Macrococcus canis strains from dogs, compare the mecB-containing elements with those found in other Macrococcus and Staphylococcus species, and identify possible mobilizable mecB subunits. Methods Whole genomes of the M. canis strains Epi0076A and KM0218 were sequenced using next-generation sequencing technologies. Multiple PCRs and restriction analysis confirmed structures of mecB-containing elements, circularization and recombination of mecB subunits. Results Both M. canis strains contained novel composite pseudo (Ψ) staphylococcal cassette chromosome mec (SCCmec) elements. Integration site sequences for SCC flanked and subdivided composite ΨSCCmecEpi0076A (69569 bp) into ΨSCC1Epi0076A-ΨSCCmecEpi0076A-ΨSCC2Epi0076A and composite ΨSCCmecKM0218 (24554 bp) into ΨSCCKM0218-ΨSCCmecKM0218. Putative γ-haemolysin genes (hlgB and hlgC) were found at the 3′ end of both composite elements. ΨSCCmecKM0218 contained a complete mecB gene complex (mecIm-mecR1m-mecB-blaZm) downstream of a new IS21-family member (ISMaca1). ΨSCCmecEpi0076A carried a blaZm-deleted mecB gene complex similar to that reported in ‘Macrococcus goetzii’ CCM4927T. A second mecB gene was found on the 81325 bp MDR plasmid pKM0218 in KM0218. This plasmid contained a complete Tn6045-associated mecB gene complex distinct from that of ΨSCCmecKM0218. pKM0218 was almost identical to the mecB-containing plasmid recently reported in Staphylococcus aureus (overall 99.96% nucleotide identity). Mobilization of mecB within an unconventional circularizable structure was observed in Epi0076A as well as chromosomal plasmid insertion via recombination of mecB operons in KM0218. Conclusions Our findings provide evidence of both the continuing evolution of mecB-containing elements in macrococci and M. canis as a potential source of the mecB-containing plasmid found in staphylococci.

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Resistance to extended-spectrum cephalosporins (ESC) and carbapenems in Escherichia coli (E. coli), increasingly identified in small animals, indicates a crisis of an antimicrobial resistance situation in veterinary medicine and public health. This study aimed to characterise the genetic features of ESC-resistant E. coli isolated from cats and dogs with urinary tract infections in Thailand. Of 72 ESC-resistant E. coli isolated from diagnostic samples (2016–2018), blaCTX-M including group 1 (CTX-M-55, -15 and -173) and group 9 (CTX-M-14, -27, -65 and -90) variants were detected in 47 isolates (65.28%) using PCR and DNA sequencing. Additional antimicrobial resistance genes, including plasmid-mediated AmpC (CIT and DHA), blaNDM-5, mcr-3, mph(A) and aac(6′)-Ib-cr, were detected in these isolates. Using a broth microdilution assay, all the strains exhibited multidrug-resistant phenotypes. The phylogroups were F (36.11%), A (20.83%), B1 (19.44%), B2 (19.44%) and D (4.17%), with several virulence genes, plasmid replicons and an integrase gene. The DNA fingerprinting using a repetitive extragenic palindromic sequence-PCR presented clonal relationships within phylogroups. Multiple human-associated, high-risk ExPEC clones associated with multidrug resistance, including sequence type (ST) 38, ST131, ST224, ST167, ST354, ST410, ST617 and ST648, were identified, suggesting clonal dissemination. Dogs and cats are a potential reservoir of ESC-resistant E. coli and significant antimicrobial resistance genes.