Livestock-associated methicillin-resistant Staphylococcus aureus in Czech retailed ready-to-eat meat products.

This study was aimed on the detection of methicillin resistant Staphylococcus aureus (MRSA) in different categories of retailed ready-to-eat (RTE) meat products from the Czech producers and determination of their genetic properties, antimicrobial resistance and virulence. In RTE meat products, 2% (4/181) of examined samples were MRSA positive. MRSA strains were detected only in durable fermented meat products made exclusively from pork meat. Detection of livestock-associated MRSA (LA-MRSA) clonal lineages (ST398 and ST4999), SCCmec cassette type V and tetracycline resistance indicate a source of contamination from raw pork. The study confirms the ability of these strains to survive the technological process rather than contamination of meat products from the food processing environment. MRSA strains did not carry any of the tested genes encoding staphylococcal enterotoxins or virulence genes (for Panton-Valentine leukocidin, exfoliative toxins A, B and toxic shock syndrome). Our results point out the spread of LA-MRSA through the meat processing chain.

A European-wide dataset to uncover adaptive traits of Listeria monocytogenes to diverse ecological niches.

Listeria monocytogenes (Lm) is a ubiquitous bacterium that causes listeriosis, a serious foodborne illness. In the nature-to-human transmission route, Lm can prosper in various ecological niches. Soil and decaying organic matter are its primary reservoirs. Certain clonal complexes (CCs) are over-represented in food production and represent a challenge to food safety. To gain new understanding of Lm adaptation mechanisms in food, the genetic background of strains found in animals and environment should be investigated in comparison to that of food strains. Twenty-one partners, including food, environment, veterinary and public health laboratories, constructed a dataset of 1484 genomes originating from Lm strains collected in 19 European countries. This dataset encompasses a large number of CCs occurring worldwide, covers many diverse habitats and is balanced between ecological compartments and geographic regions. The dataset presented here will contribute to improve our understanding of Lm ecology and should aid in the surveillance of Lm. This dataset provides a basis for the discovery of the genetic traits underlying Lm adaptation to different ecological niches.

Staphylococcus ratti sp. nov. Isolated from a Lab Rat.

Staphylococci from the Staphylococcus intermedius-Staphylococcus hyicus species group include numerous animal pathogens and are an important reservoir of virulence and antimicrobial resistance determinants. Due to their pathogenic potential, they are possible causative agents of zoonoses in humans; therefore, it is important to address the properties of these strains. Here we used a polyphasic taxonomic approach to characterize the coagulase-negative staphylococcal strain NRL/St 03/464T, isolated from the nostrils of a healthy laboratory rat during a microbiological screening of laboratory animals. The 16S rRNA sequence, MALDI-TOF mass spectrometry and positive urea hydrolysis and beta-glucuronidase tests clearly distinguished it from closely related Staphylococcus spp. All analyses have consistently shown that the closest relative is Staphylococcus chromogenes; however, values of digital DNA-DNA hybridization <35.3% and an average nucleotide identity <81.4% confirmed that the analyzed strain is a distinct Staphylococcus species. Whole-genome sequencing and expert annotation of the genome revealed the presence of novel variable genetic elements, including two plasmids named pSR9025A and pSR9025B, prophages, genomic islands and a composite transposon that may confer selective advantages to other bacteria and enhance their survival. Based on phenotypic, phylogenetic and genomic data obtained in this study, the strain NRL/St 03/464T (= CCM 9025T = LMG 31873T = DSM 111348T) represents a novel species with the suggested name Staphylococcus ratti sp. nov.

Characterisation of Colistin -Resistant Enterobacterales and Acinetobacter Strains Carrying mcr Genes from Asian Aquaculture Products.

Aquaculture systems are widely recognised as hotspots for horizontal gene transfer, and the need for screening for bacteria carrying antimicrobial resistance genes in aquaculture systems is becoming more important. In this study, we characterised seventeen bacterial strains (Escherichia coli, Klebsiella pneumoniae, Acinetobacter baumannii, and A. nosocomialis) resistant to colistin originating from retailed aquaculture products imported from Vietnam to the Czech Republic. The mcr-1.1 gene was found located on plasmid types IncHI2, IncI2, and IncX4, as well as on the rarely described plasmid types IncFIB-FIC and IncFIB(K), phage-like plasmid p0111, and on the chromosome of E. coli. One E. coli strain carried the mcr-3.5 gene on IncFII(pCoo) plasmid in addition to the mcr-1.1 gene located on IncHI2 plasmid. K. pneumoniae was found to carry the mcr-1.1 and mcr-8.2 genes on IncFIA(HI1) plasmid. The mcr-4.3 gene was found on similar untypeable plasmids of A. baumannii and A. nosocomialis strains, pointing to the possible interspecies transfer of plasmids carrying the mcr-4 gene. Our results highlight that some aquaculture products of Asian origin can represent an important source of variable plasmids carrying mcr genes. The results showed an involvement of phages in the incorporation of the mcr-1 gene into plasmids or the chromosome in E. coli strains from aquaculture. The detection of E. coli with the mcr-1 gene in the chromosome points to the risks associated with the stabilisation of the mcr genes in the bacterial chromosome.

A Rapid Culture Method for the Detection of Campylobacter from Water Environments.

The natural environment and water are among the sources of Campylobacter jejuni and Campylobacter coli. A limited number of protocols exist for the isolation of campylobacters in poorly filterable water. Therefore, the goal of our work was to find a more efficient method of Campylobacter isolation and detection from wastewater and surface water than the ISO standard. In the novel rapid culture method presented here, samples are centrifuged at high speed, and the resuspended pellet is inoculated on a filter, which is placed on Campylobacter selective mCCDA agar. The motile bacteria pass through the filter pores, and mCCDA agar suppresses the growth of background microbiota on behalf of campylobacters. This culture-based method is more efficient for the detection and isolation of Campylobacter jejuni and Campylobacter coli from poorly filterable water than the ISO 17995 standard. It also is less time-consuming, taking only 72 h and comprising three steps, while the ISO standard method requires five or six steps and 144-192 h. This novel culture method, based on high-speed centrifugation, bacterial motility, and selective cultivation conditions, can be used for the detection and isolation of various bacteria from water samples.

Detection and molecular characterisation of methicillin-resistant Staphylococcus aureus isolated from raw meat in the retail market.

OBJECTIVES: This study aimed to detect and characterise methicillin-resistant Staphylococcus aureus (MRSA) from retail meat in the Czech Republic. METHODS: Isolates were identified by PCR detection of the S. aureus-specific fragment Sa442 and mecA gene. spa typing, MLST, detection of genes encoding staphylococcal enterotoxins, Panton-Valentine leukocidin (pvl), exfoliative toxins A and B (eta and etb), toxic shock syndrome toxin (tst) and staphylokinase (sak), detection of φSa3 prophage and antimicrobial susceptibility testing were performed. RESULTS: Of 65 raw meat samples examined (poultry, beef, pork and rabbit), 23 (35.4%) were positive for MRSA. Twelve positive samples originated from poultry (12/33; 36.4%), while the remaining eleven came from pork (9/9; 100%) and pork/beef mixed minced meat (2/5; 40.0%). Eight spa types belonging to five different sequence types (STs) were identified. ST398 was the most frequent (28/36; 77.8%), presenting spa types t011, t034, t2576, t4132, t588 and t899. Other livestock-associated MRSA STs (ST9-t899, ST5-t002, ST692-t8646 or the newly described ST4034-t899) were also sporadically identified. In seven isolates (19.4%), one or more staphylococcal enterotoxin genes were detected, with sea, seg and sei prevailing. Three isolates from turkey [ST398-t899 (n = 2) and ST398-t011] harboured the sak gene, and the latter also harboured the sea gene. Seven isolates from poultry harboured the φSa3 prophage and were resistant to tetracycline. CONCLUSION: Specific kinds of meat appear to be a possible source of MRSA, although the risk to humans is hard to define. Therefore, surveillance of MRSA in meat as well as hygienic practices should be improved.

The Intestinal Carriage of Plasmid-Mediated Colistin-Resistant Enterobacteriaceae in Tertiary Care Settings.

Background: In order to estimate the prevalence of plasmid borne colistin resistance and to characterize in detail the mcr-positive isolates, we carried out a sentinel testing survey on the intestinal carriage of plasmid-mediated colistin-resistant Enterobacteriaceae in hospitalized patients. Methods: Between June 2018 and September 2019, 1922 faecal samples from hospitalised patients were analysed by selective culture in presence of colistin (3.5 mg/L), and in parallel by direct detection of the mcr-1 to mcr-8 genes by qPCR. The mcr-positive isolates were characterised by whole-genome sequencing. Results: The prevalence of the mcr-1 gene was 0.21% (n = 4/1922); the mcr-2 to 8 genes were not detected. The mcr-1 gene was found to be localised in the IncX4 (n = 3) and IncHI2 (n = 1) plasmid type. One Escherichia coli isolate was susceptible to colistin due to the inactivation of the mcr-1 gene through the insertion of the IS2 element; however, the colistin resistance was inducible by culture in low concentrations of colistin. One human mcr-1 positive E. coli isolate was related genetically to the mcr-1 E. coli isolate derived from turkey meat of Czech origin. Conclusions:mcr-mediated colistin resistance currently poses little threat to patients hospitalised in Czech healthcare settings. The presence of the mcr-1 gene in the human population has a possible link to domestically produced, retail meat.

A cross-border outbreak of Salmonella Bareilly cases confirmed by whole genome sequencing, Czech Republic and Slovakia, 2017 to 2018.

In August 2017, an increased incidence of Salmonella Bareilly was detected in the Czech Republic. An investigation was conducted with Slovakia to confirm the outbreak and identify the source. Probable outbreak cases were defined as cases with laboratory-confirmed S. Bareilly reported in either of the national surveillance systems, and/or the Czech and Slovak National Reference Laboratory databases from July 2017. Confirmed cases had the pulsed-field gel electrophoresis (PFGE) outbreak pulsotype or up to 5 alleles difference from outbreak cluster members by core genome multilocus sequence typing (cgMLST). PFGE and whole genome sequencing were used for isolate comparison. The same trawling questionnaire was used in both countries. By the end of October 2018, 325 cases were identified. Among 88 human S. Bareilly isolates analysed by PFGE, 82 (93%) shared an identical pulsotype; cgMLST of 17 S. Bareilly human isolates showed 1-2 allele difference. The trawling questionnaire excluded consumption of unusual or imported foods. In September 2018, an isolate closely related to the outbreak isolates was identified in a powdered egg product. A spray dryer was recognised as the contamination source and the production plant was closed. Using molecular typing methods, we detected a diffuse cross-border outbreak caused by S. Bareilly.

Virulence Properties of mcr-1-Positive Escherichia coli Isolated from Retail Poultry Meat.

The great plasticity and diversity of the Escherichia coli genome, together with the ubiquitous occurrence, make E. coli a bacterium of world-wide concern. Of particular interest are pathogenic strains and strains harboring antimicrobial resistance genes. Overlapping virulence-associated traits between avian-source E. coli and human extraintestinal pathogenic E. coli (ExPEC) suggest zoonotic potential and safety threat of poultry food products. We analyzed whole-genome sequencing (WGS) data of 46 mcr-1-positive E. coli strains isolated from retail raw meat purchased in the Czech Republic. The investigated strains were characterized by their phylogroup-B1 (43%), A (30%), D (11%), E (7%), F (4%), B2 (2%), C (2%), MLST type, and serotype. A total of 30 multilocus sequence types (STs), of which ST744 was the most common (11%), were identified, with O8 and O89 as the most prevalent serogroups. Using the VirulenceFinder tool, 3 to 26 virulence genes were detected in the examined strains and a total of 7 (15%) strains met the pathogenic criteria for ExPEC. Four strains were defined as UPEC (9%) and 18 (39%) E. coli strains could be classified as APEC. The WGS methods and available on-line tools for their evaluation enable a comprehensive approach to the diagnosis of virulent properties of E. coli strains and represent a suitable and comfortable platform for their detection. Our results show that poultry meat may serve as an important reservoir of strains carrying both virulence and antibiotic resistance genes for animal and human populations.

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.

The colonisation of Czech travellers and expatriates living in the Czech Republic by colistin-resistant Enterobacteriaceae and whole genome characterisation of E. coli isolates harbouring the mcr-1 genes on a plasmid or chromosome: A cross-sectional study.

BACKGROUND: Travellers were recognized as a risk cohort that can be colonized by mcr-1-mediated colistin-resistant Enterobacteriaceae. We aimed to investigate the carriage of mcr-mediated colistin resistance in Enterobacteriaceae in Czech travellers or expatriates residing temporarily in the Czech Republic. METHODS: Between August 2018 and September 2019, the stool samples were cultured in enrichment broth. The enriched cultures were tested for the presence of the mcr-1-8 genes and inoculated onto selective agar with colistin. Colistin-resistant Enterobacteriaceae were tested for the presence of the mcr-1-8 genes; the mcr-positive isolates were characterised by whole genome sequencing. RESULTS: From the 177 stool samples, 15 colistin-resistant Enterobacteriaceae isolates were cultured (7.9%); two of the E. coli isolates carried the mcr-1 gene (1.1%). In the E. coli multilocus sequence type (ST) 156, the mcr-1 gene was located in an ISApl1-mcr-1-orf-ISApl1 (Tn6330) and incorporated into the chromosome; in the E. coli ST23 isolate, the mcr-1 gene was harboured by the plasmid IncX4. Both of the mcr-1 positive E. coli isolates were multidrug-resistant and one isolate was an extended-spectrum ß-lactamase producer (blaCTX-M-27). CONCLUSION: Patients with an international travel history should be monitored for the carriage of the mcr-1 gene in order to prevent its dissemination into healthcare settings.

Comparative Analysis of Genetic Determinants Encoding Cadmium, Arsenic, and Benzalkonium Chloride Resistance in Listeria monocytogenes of Human, Food, and Environmental Origin.

Environmental adaptation of Listeria monocytogenes is a complex process involving various mechanisms that can contribute to their survival in the environment, further spreading throughout the food chain and the development of listeriosis. The aim of this study was to analyze whole-genome sequencing data in a set of 270 strains of L. monocytogenes derived from human listeriosis cases and food and environmental sources in order to compare the prevalence and type of genetic determinants encoding cadmium, arsenic, and benzalkonium chloride resistance. Most of the detected genes of cadmium (27.8%), arsenic (15.6%), and benzalkonium chloride (7.0%) resistance were located on mobile genetic elements, even in phylogenetically distant lineages I and II, which indicates the possibility of their horizontal spread. Although no differences were found in the prevalence of these genes between human and food strains, they have been detected sporadically in strains from the environment. Regarding cadmium resistance genes, cadA1C1_Tn5422 predominated, especially in clonal complexes (CCs) 121, 8, and 3 strains. At the same time, qacH_Tn6188-encoding benzalkonium chloride resistance was most frequently detected in the genome of CC121 strains. Genes encoding arsenic resistance were detected mainly in strains CC2 (located on the chromosomal island LGI2) and CC9 (carried on Tn554). The results indicated a relationship between the spread of genes encoding resistance to cadmium, arsenic, and benzalkonium chloride in certain serotypes and CCs and showed the need for a more extensive study of L. monocytogenes strains to better understand their ability to adapt to the food production environment.

Characterization of the Complete Nucleotide Sequences of mcr-1-Encoding Plasmids From Enterobacterales Isolates in Retailed Raw Meat Products From the Czech Republic.

The aim of our study was to determine complete nucleotide sequence of mcr-1-carrying plasmids from Enterobacterales isolates recovered from domestic and imported raw retailed meat and compare them with plasmids available at the GenBank sequence database. A set of 16 plasmids originating from Escherichia coli (n = 13), Klebsiella pneumoniae (n = 2), and Citrobacter braakii (n = 1) were analyzed. In our previous study, data from whole genome sequencing showed that mcr-1 gene was located on plasmids of different incompatibility groups (IncHI2, IncI2, and IncX4). The IncI2 (n = 3) and IncX4 (n = 8) plasmids harbored mcr-1.1 gene only, whereas IncHI2 sequence type 4 plasmids (n = 5) carried large multidrug resistance (MDR) regions. MDR regions of IncHI2 plasmids included additional antimicrobial resistance genes conferring resistance to ß-lactams (bla TEM-1), aminoglycosides [aadA1, aadA2, and aph(6)-Id], macrolides [mef (B)], tetracycline (tetA, tetR), and sulphonamides (sul1, sul2, and sul3). Likewise, IncHI2 plasmids carried several insertion sequences including IS1, IS3, IS26, IS1326, and ISApl1. In conclusion, our findings confirmed the involvement of IncX4, IncI2, and IncHI2 plasmids in the dissemination of mcr-1.1 gene in several environmental niches, as in samples of retail meat originating from different geographical regions. In contrast to IncX4 and IncI2, IncHI2 plasmids were more diverse and carried additional genes for resistance to heavy metals and multiple antimicrobials.

The emergence of mcr-1-mediated colistin-resistant Escherichia coli and Klebsiella pneumoniae in domestic and imported turkey meat in the Czech Republic 2017-2018.

We investigated the occurrence of plasmid-mediated colistin resistance in Enterobacteriaceae in turkey meat produced in the Czech Republic as well as in turkey meat imported into the Czech Republic from other European Union countries. Seventeen samples of raw turkey meat from the Czech Republic (n = 4), Hungary (n = 2), Poland (n = 6) and Germany (n = 5) were cultured in peptone water at 37 °C overnight and the enriched cultures were tested for the presence of mcr-1-5 genes. PCR-positive enriched cultures were inoculated onto selective agar with colistin (3.5 mg/L). A minimal inhibitory concentration (MIC) of colistin was determined by using the broth microdilution method in PCR-positive isolates. In addition, a macrorestriction analysis was performed using XbaI endonuclease. Of 17 meat samples, 12 samples from Poland (6/6), Germany (3/5) and the Czech Republic (3/4) proved positive for the presence of the mcr-1 gene. Forty-two isolates carrying the mcr-1 gene were obtained: Escherichia coli (n = 39) revealing 32 distinct XbaI profiles and Klebsiella pneumoniae (n = 3) with 2 distinct XbaI profiles. The minimal inhibitory concentration (MIC) of the mcr-1 positive isolates was as follows: 4 mg/L (n = 28), 8 mg/L (n = 12), 32 mg/L (n = 1) and 64 mg/L (n = 1). The high prevalence (70.6%; 12/17 samples) of mcr-1-mediated colistin-resistant Enterobacteriaceae found in the turkey meat samples analysed in this study, builds on previously published evidence that poultry, and their products, represent a substantial risk for the dissemination of plasmid-mediated colistin resistance in Europe.

An investigation of Salmonella Fluntern illnesses linked to leopard geckos-United States, 2018.

Reptile contact can result in zoonotic non-typhoidal salmonellosis. In April 2018, Oregon Public Health Division contacted CDC about a cluster of four Salmonella serovar Fluntern (SF) illnesses in four states (OR, CA, IA, NY); patients reported contact with geckos, a popular reptile pet. PulseNet, the national molecular subtyping network of food-borne disease surveillance, subsequently identified additional SF clinical isolates. Twelve cases in 11 states were identified; median age was 5 years (range: <1-58 years). Three patients were hospitalized; no deaths were reported. Of those with exposure information (n = 10), all reported reptile exposure; 9 (90%) specified contact with leopard geckos. No common source of geckos was identified from reported purchase locations. Los Angeles County (LAC) health officials isolated SF from one patient's leopard gecko. Five reptile/gecko isolates were identified from the USDA National Veterinary Services Laboratories (NVSL) from 2015 to 2018. Five countries responded to an Epidemic Intelligence Information System post by PulseNet; reptile isolate sequence data were received from Czech Republic. A clinical case from England was identified through the National Center for Biotechnology Information pathogen detection pipeline; the patient did not report contact with leopard geckos. Whole genome sequencing analysis revealed substantial genetic diversity between clinical and animal isolates; however, gecko and clinical isolates from LAC were highly related (1 allele difference). This investigation linking SF illnesses to leopard geckos highlights an important public health risk from pets. A better understanding of how geckos are distributed by the pet industry in the United States could improve traceability to points of origin and mitigate Salmonella transmission at gecko breeders. Earlier NVSL reports of SF isolates from geckos suggest the risk of human SF infection from geckos is not new. This investigation demonstrates a need to educate gecko breeders, retailers and gecko owners about the continued Salmonella infection risk from pet geckos.

Prediction of Persistence of Listeria monocytogenes ST451 in a Rabbit Meat Processing Plant in the Czech Republic.

This study was focused on characterization of the genetic diversity of Listeria monocytogenes isolated from packed fresh rabbit meat obtained from one producer via retail outlets. The partial aim was to compare the characteristics of a suspect persistent strain with strains from human cases. The occurrence of L. monocytogenes in vacuum-packed rabbit meat was monitored during 2013 to 2016. All strains were characterized by serotyping, pulsed-field gel electrophoresis, and multilocus sequence typing (MLST). Selected strains, which represented each year, were analyzed using the whole genome sequencing method. L. monocytogenes was detected in 21 (38%) of 56 originally packed rabbit meat samples from one food producer during the whole monitored period. All strains showed the identical serotype (1/2a), AscI/ApaI pulsotype (735/2), and sequence type (ST451). The clonal similarity of strains from rabbit meat was also confirmed on the basis of core genome MLST (on 1,701 loci). This fact suggests the occurrence of a suspect persistent strain in the meat processing plant. Results of core genome MLST enabled us to unambiguously exclude rabbit meat as a source of listeriosis in humans caused by the indistinguishable AscI/ApaI pulsotype and sequence type, although all strains carried all genes important for the virulence of L. monocytogenes. No specific genes that may be associated with its persistence in the food processing environment were detected among the tested strains of ST451.

Complete Nucleotide Sequences of mcr-4.3-Carrying Plasmids in Acinetobacter baumannii Sequence Type 345 of Human and Food Origin from the Czech Republic, the First Case in Europe.

Here, we describe two plasmids carrying mcr-4.3 in two Acinetobacter baumannii strains isolated from imported food and a clinical sample. The comparative analysis of these plasmids, with two other plasmids reported in the NCBI database, highlighted the common origin of the plasmidic structure carrying mcr-4.3 This is the first case of the mcr-4.3 gene in a A. baumannii strain isolated from a clinical case in Europe. We hypothesize that food import is initiating the spread in Czech Republic.

[Detection of colistin-resistant Acinetobacter baumannii with the mcr-4 gene].

The emergence of plasmid-mediated colistin resistance carried by mcr genes and polymyxin resistance in carbapenem-resistant bacteria poses a threat to antibiotic therapy of bacterial infections. The worldwide spread of colistin resistance carried by mcr genes, particularly in Enterobacteriaceae, points to the possibility of the spread of this type of resistance also in non-fermenting Gram-negative bacteria such as Pseudomonas aeruginosa or Acinetobacter baumannii. This study provides information on the first described occurrence of the mcr-4 gene in A. baumannii isolated from imported turkey liver obtained in the retail market of the Czech Republic.

Genome Sequences of Livestock-Associated Methicillin-Resistant Staphylococcus aureusspa Type t899 Strains Belonging to Three Different Sequence Types (ST398, ST9, and ST4034).

Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) is an emerging MRSA lineage rapidly evolving in the community. In this report, we present the draft genome sequences of nine LA-MRSA strains. These strains were isolated from meat and a human nasal swab sample and belong to one unique spa type (t899), but to three different sequence types, ST398, ST9, and ST4034.

Detection and Molecular Characterization of Methicillin-Resistant Staphylococcus aureus Isolated from Bulk Tank Milk of Cows, Sheep, and Goats.

This study is aimed at detecting and characterizing methicillin-resistant Staphylococcus aureus (MRSA) from bulk tank milk samples of cows, sheep, and goats collected from dairy farms in the Czech Republic. All MRSA isolates were identified using PCR detection of the Staphylococcus aureus-specific fragment SA442 and mecA gene. The staphylococcal chromosomal cassettes mec (SCCmec), spa, and multilocus sequence types (MLST) were determined. The presence of genes encoding enterotoxins (ses), Panton-Valentine leukocidin (pvl), exfoliative toxins A, B (eta, etb), and toxic shock syndrome toxin (tst) were assessed. To differentiate human and animal origin, the presence of staphylokinase (sak) gene, ϕSa3 prophage, and susceptibility to tetracycline was tested. Out of 49 bulk tank milk samples examined, 14 (28.6%) were MRSA-positive. Eleven positive samples came from cow's milk (38%) and the remaining three from goat's milk (33%). All samples of ewe's milk were negative. In MRSA isolates three sequence types containing seven spa types were identified. Twelve isolates (85.7%) belonged to ST398 spa types t011/SCCmec IVa, t011/SCCmec V, t034/SCCmec V, t1456/SCCmec IVa, t1255/SCCmec V, and t2346/SCCmec V. Another two isolates belonged to ST5/t3598/SCCmec IVa and ST8/t064/SCCmec IVNT. In six isolates, one or more ses genes (seb, sed, seg, sei, and sej) were confirmed. One isolate from cow's milk harbored the tst gene. Another two isolates (ST398/t1456/SCCmec IVa and ST5/t3598/SCCmec IVa) harbored the sak gene and ϕSa3 prophage, and the latter was the only tetracycline-susceptible isolate in this study. However, none of the isolates was positive for pvl or eta, etb. These results suggest that there is the wide geographical spread of ST398 across different regions of the Czech Republic with no host preference among dairy cattle and goats. Therefore, when evaluating the occupational and foodborne risks, MRSA carriage and infection should be taken into account.