MRSA and multidrug-resistant Staphylococcus aureus in U.S. retail meats, 2010-2011.

Methicillin-resistant Staphylococcus aureus (MRSA) has been detected in retail meats, although large-scale studies are scarce. We conducted a one-year survey in 2010-2011 within the framework of the National Antimicrobial Resistance Monitoring System. Among 3520 retail meats collected from eight U.S. states, 982 (27.9%) contained S. aureus and 66 (1.9%) were positive for MRSA. Approximately 10.4% (107/1032) of S. aureus isolates, including 37.2% (29/78) of MRSA, were multidrug-resistant (MDRSA). Turkey had the highest MRSA prevalence (3.5%), followed by pork (1.9%), beef (1.7%), and chicken (0.3%). Whole-genome sequencing was performed for all 66 non-redundant MRSA. Among five multilocus sequence types identified, ST8 (72.7%) and ST5 (22.7%) were most common and livestock-associated MRSA ST398 was assigned to one pork isolate. Eleven spa types were represented, predominately t008 (43.9%) and t2031 (22.7%). All four types of meats harbored t008, whereas t2031 was recovered from turkey only. The majority of MRSA (84.8%) possessed SCCmec IV and 62.1% harbored Panton-Valentine leukocidin. Pulsed-field gel electrophoresis showed that all ST8 MRSA belonged to the predominant human epidemic clone USA300, and others included USA100 and USA200. We conclude that a diverse MRSA population was present in U.S. retail meats, albeit at low prevalence.

Prevalence and characterization of methicillin-resistant Staphylococcus aureus isolates from retail meat and humans in Georgia.

There is increasing interest in the presence of Staphylococcus aureus, specifically methicillin-resistant S. aureus (MRSA), on retail meat products. In this study, staphylococci were isolated from retail pork and retail beef in Georgia, and MRSA from the products was compared to human MRSA from the same geographic area using broth microdilution antimicrobial susceptibility testing, multilocus sequence typing (MLST), spa typing, SCCmec typing, and pulsed-field gel electrophoresis (PFGE). S. aureus was isolated from 45% (45/100) of pork products and 63% (63/100) of beef products; mecA was detected in S. aureus from both pork (3/100; 3%) and beef (4/100; 4%). Fifty percent (50/100) of human S. aureus also contained mecA. Multidrug resistance was detected among MRSA from all sources. All MRSA (n = 57) was SCCmec type IV, and nine different spa types were present among the isolates (t002, t008, t012, t024, t179, t337, t548, t681, and t1062). Four sequence types (ST5, ST8, ST9, and ST30) were detected using MLST; the majority of MRSA isolates belonged to ST8, followed by ST5. One retail beef MRSA isolate belonged to ST8, while the remaining three were ST5. In retail pork MRSA, ST5, ST9, and ST30 were observed. The majority of human MRSA isolates belonged to ST8. Thirty-seven MRSA isolates, one of which was a retail beef MRSA isolate, were pvl(+). Using PFGE, MLST, and spa typing, three retail beef MRSA isolates were found to be identical in PFGE pattern, ST, and spa type to two human clonal MRSA isolates (USA100 and USA300). One additional retail beef MRSA isolate had a PFGE pattern similar to that of a human MRSA isolate, whereas none of the retail pork MRSA isolates had PFGE patterns similar to those of human MRSA isolates. These data suggest that the retail beef samples were contaminated by a human source, possibly during processing of the meat, and may present a source of MRSA for consumers and others who handle raw meat.

Clonally related methicillin-resistant Staphylococcus aureus isolated from short-finned pilot whales (Globicephala macrorhynchus), human volunteers, and a bayfront cetacean rehabilitation facility.

In May of 2011, a live mass stranding of 26 short-finned pilot whales (Globicephala macrorhynchus) occurred in the lower Florida Keys. Five surviving whales were transferred from the original stranding site to a nearby marine mammal rehabilitation facility where they were constantly attended to by a team of volunteers. Bacteria cultured during the routine clinical care of the whales and necropsy of a deceased whale included methicillin-sensitive and methicillin-resistant Staphylococcus aureus (MSSA and MRSA). In order to investigate potential sources or reservoirs of MSSA and MRSA, samples were obtained from human volunteers, whales, seawater, and sand from multiple sites at the facility, nearby recreational beaches, and a canal. Samples were collected on 3 days. The second collection day was 2 weeks after the first, and the third collection day was 2 months after the last animal was removed from the facility. MRSA and MSSA were isolated on each day from the facility when animals and volunteers were present. MSSA was found at an adjacent beach on all three collection days. Isolates were characterized by utilizing a combination of quantitative real-time PCR to determine the presence of mecA and genes associated with virulence, staphylococcal protein A typing, staphylococcal cassette chromosome mec typing, multilocus sequence typing, and pulsed field gel electrophoresis (PFGE). Using these methods, clonally related MRSA were isolated from multiple environmental locations as well as from humans and animals. Non-identical but genetically similar MSSA and MRSA were also identified from distinct sources within this sample pool. PFGE indicated that the majority of MRSA isolates were clonally related to the prototype human strain USA300. These studies support the notion that S. aureus may be shed into an environment by humans or pilot whales and subsequently colonize or infect exposed new hosts.

Human-associated methicillin-resistant Staphylococcus aureus from a subtropical recreational marine beach.

Reports of Staphylococcus aureus including methicillin-resistant S. aureus (MRSA) detected in marine environments have occurred since the early 1990 s. This investigation sought to isolate and characterize S. aureus from marine waters and sand at a subtropical recreational beach, with and without bathers present, in order to investigate possible sources and to identify the risks to bathers of exposure to these organisms. During 40 days over 17 months, 1,001 water and 36 intertidal sand samples were collected by either bathers or investigators at a subtropical recreational beach. Methicillin-sensitive S. aureus (MSSA) and MRSA were isolated and identified using selective growth media and an organism-specific molecular marker. Antimicrobial susceptibility, staphylococcal cassette chromosome mec (SCCmec) type, pulsed-field gel electrophoresis (PFGE) pattern, multi-locus sequence type (MLST), and staphylococcal protein A (spa) type were characterized for all MRSA. S. aureus was isolated from 248 (37 %) bather nearby water samples at a concentration range of <2-780 colony forming units per ml, 102 (31 %) ambient water samples at a concentration range of <2-260 colony forming units per ml, and 9 (25 %) sand samples. Within the sand environment, S. aureus was isolated more often from above the intertidal zone than from intermittently wet or inundated sand. A total of 1334 MSSA were isolated from 37 sampling days and 22 MRSA were isolated from ten sampling days. Seventeen of the 22 MRSA were identified by PFGE as the community-associated MRSA USA300. MRSA isolates were all SCCmec type IVa, encompassed five spa types (t008, t064, t622, t688, and t723), two MLST types (ST8 and ST5), and 21 of 22 isolates carried the genes for Panton-Valentine leukocidin. There was a correlation (r = 0.45; p = 0.05) between the daily average number of bathers and S. aureus in the water; however, no association between exposure to S. aureus in these waters and reported illness was found. This report supports the concept that humans are a potential direct source for S. aureus in marine waters.

Methicillin-resistant Staphylococcus aureus (MRSA) detected at four U.S. wastewater treatment plants.

BACKGROUND: The incidence of community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) infections is increasing in the United States, and it is possible that municipal wastewater could be a reservoir of this microorganism. To date, no U.S. studies have evaluated the occurrence of MRSA in wastewater. OBJECTIVE: We examined the occurrence of MRSA and methicillin-susceptible S. aureus (MSSA) at U.S. wastewater treatment plants. METHODS: We collected wastewater samples from two Mid-Atlantic and two Midwest wastewater treatment plants between October 2009 and October 2010. Samples were analyzed for MRSA and MSSA using membrane filtration. Isolates were confirmed using biochemical tests and PCR (polymerase chain reaction). Antimicrobial susceptibility testing was performed by Sensititre® microbroth dilution. Staphylococcal cassette chromosome mec (SCCmec) typing, Panton-Valentine leucocidin (PVL) screening, and pulsed field gel electrophoresis (PFGE) were performed to further characterize the strains. Data were analyzed by two-sample proportion tests and analysis of variance. RESULTS: We detected MRSA (n = 240) and MSSA (n = 119) in 22 of 44 (50%) and 24 of 44 (55%) wastewater samples, respectively. The odds of samples being MRSA-positive decreased as treatment progressed: 10 of 12 (83%) influent samples were MRSA-positive, while only one of 12 (8%) effluent samples was MRSA-positive. Ninety-three percent and 29% of unique MRSA and MSSA isolates, respectively, were multidrug resistant. SCCmec types II and IV, the pvl gene, and USA types 100, 300, and 700 (PFGE strain types commonly found in the United States) were identified among the MRSA isolates. CONCLUSIONS: Our findings raise potential public health concerns for wastewater treatment plant workers and individuals exposed to reclaimed wastewater. Because of increasing use of reclaimed wastewater, further study is needed to evaluate the risk of exposure to antibiotic-resistant bacteria in treated wastewater.

Development of a DNA microarray to detect antimicrobial resistance genes identified in the National Center for Biotechnology Information database.

To understand the mechanisms and epidemiology of antimicrobial resistance (AR), the genetic elements responsible must be identified. Due to the myriad of possible genes, a high-density genotyping technique is needed for initial screening. To achieve this, AR genes in the National Center for Biotechnology Information GenBank database were identified by their annotations and compiled into a nonredundant list of 775 genes. A DNA microarray was constructed of 70mer oligonucelotide probes designed to detect these genes encoding resistances to aminoglycosides, beta-lactams, chloramphenicols, glycopeptides, heavy metals, lincosamides, macrolides, metronidazoles, polyketides, quaternary ammonium compounds, streptogramins, sulfonamides, tetracyclines, and trimethoprims as well as resistance transfer genes. The microarray was validated with two fully sequenced control strains of Salmonella enterica: Typhimurium LT2 (sensitive) and Typhi CT18 (multidrug resistance [MDR]). All resistance genes encoded on the MDR plasmid, pHCM1, harbored by CT18 were detected in that strain, whereas no resistance genes were detected in LT2. The microarray was also tested with a variety of bacteria, including MDR Salmonella enterica serovars, Escherichia coli, Campylobacter spp., Enterococcus spp., methicillin-resistant Staphylococcus aureus, Listeria spp., and Clostridium difficile. The results presented here demonstrate that a microarray can be designed to detect virtually all AR genes found in the National Center for Biotechnology Information database, thus reducing the subsequent assays necessary to identify specific resistance gene alleles.

Comparative antimicrobial susceptibility of Listeria monocytogenes, L. innocua, and L. welshimeri.

The current study compared antimicrobial susceptibility of Listeria innocua, L. welshimeri, and L. monocytogenes isolated from various sources. Antimicrobial susceptibility testing was performed using a microbroth procedure with Sensititre minimum inhibitory concentration plates containing 18 antimicrobials. Resistant isolates were analyzed for the presence of antimicrobial resistance genes using PCR. The majority of L. monocytogenes isolates were resistant to oxacillin (99%, 89/90) and ceftriaxone (72%, 65/90), while few isolates were resistant to clindamycin (21%, 19/90) and ciprofloxacin (2%, 2/90). When selected sources of L. monocytogenes are compared, resistance to ceftriaxone, clindamycin, and oxacillin ranged from 27% to 86%, 7% to 43%, and 96% to 100%, respectively. Resistance to ciprofloxacin (6%, 2/34), quinupristin/dalfopristin (7%, 1/14), and tetracycline (7%, 1/15) was observed with L. monocytogenes isolated from food, animal, and environmental sources, respectively. All L. welshimeri isolates (6/6) were resistant to streptomycin, quinupristin/dalfopristin, ciprofloxacin, rifampin, oxacillin, penicillin, and clindamycin, while most isolates (67%, 4/6) were resistant to trimethoprim/sulfamethoxazole. All L. innocua isolates (4/4) were resistant to oxacillin and penicillin, whereas 75% (3/4) of isolates were resistant to tetracycline, ceftriaxone, and clindamycin. Resistant isolates were negative for aadA, strA-B, sul I-II, penA, vat(A-E), vga(A-B), and vgb(A-B). However, tetM was detected among tetracycline-resistant isolates. L. welshimeri was resistant to more of the tested antimicrobials than the other two Listeria species tested, but resistance was not attributed to selected resistance genes. These data demonstrate the variability in resistance among Listeria species. However, the human pathogen L. monocytogenes appears to be the least resistant among the tested species.