Melanie Abley

Methicillin-resistant Staphylococcus aureus in pigs and farm workers on conventional and antibiotic-free swine farms in the USA.

Much uncertainty remains about the origin and public health implications of livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA). This study aimed to investigate the occurrence and prevalence of MRSA in general and LA-MRSA in particular in pigs and farm workers in five states. We collected nasal swabs from pigs and farm workers at 45 swine herds (21 antibiotic-free herds; 24 conventional herds) in Illinois, Iowa, Minnesota, North Carolina and Ohio. MRSA was isolated from 50 of 1085 pigs (4.6%) and 31 of 148 (20.9%) of farm workers. MRSA-positive pigs and people were clustered in four conventional swine farms in Iowa and Illinois. Based on genotyping, spa type t034, a common livestock associated variant, was predominant among both human and swine isolates. These results confirm the presence of LA-MRSA in pigs and swine farm workers in the USA, but the prevalence found is relatively low compared with European studies.

Salmonella enterica in Commercial Swine Feed and Subsequent Isolation of Phenotypically and Genotypically Related Strains from Fecal Samples

ABSTRACT The purpose of this study was to determine the occurrence and genotypic relatedness of Salmonella enterica isolates recovered from feed and fecal samples in commercial swine production units. Of 275 feed samples, Salmonella was detected in 10 feed samples that originated from 8 of 36 (22.2%) barns, with a prevalence of 3.6% (10/275 samples). In fecal samples, a prevalence of 17.2% was found at the early finishing stage (1,180/6,880 samples), with a significant reduction in prevalence (7.4%) when pigs reached market age (392/5,321 samples). Of the 280 Salmonella isolates systematically selected for further characterization, 50% of the feed isolates and 55.3% of the isolates of fecal origin showed similar phenotypes based on antimicrobial resistance patterns and serogrouping. About 44% of the isolates were multidrug resistant. Pulsed-field gel electrophoresis (PFGE) genotyping grouped the 46 representative isolates into five genotypic clusters, of which four of the clusters consisted of genotypically related isolates recovered from feed and fecal samples. The occurrence of genotypically related and, in some cases, clonal strains, including multidrug-resistant isolates in commercially processed feed and fecal samples, suggests the high significance of commercial feed as a potential vehicle of Salmonella transmission.

CTX-M-Type Extended-Spectrum β-Lactamases Present in Escherichia coli from the Feces of Cattle in Ohio, United States

CTX-M extended-spectrum β-lactamases are enzymes produced by bacteria that are capable of inhibiting the antimicrobial effects of cephalosporin drugs. Recently, the first domestically acquired Salmonella in the United States expressing bla(CTX-M) was reported. This is a concern because expanded-spectrum cephalosporins are the treatment of choice for invasive Gram-negative infections, including salmonellosis in children. Because Salmonella transmission is primarily foodborne, there is also concern that resistant enteric bacteria from livestock can be transferred through the food supply chain to consumers. bla(CTX-M) has not been previously identified in bacterial isolates from food animal populations in the United States. We report the recovery of CTX-M-type extended-spectrum β-lactamases from fecal Escherichia coli of sick and healthy dairy cattle in Ohio. Four individual fecal samples yielded E. coli isolates representing three clonal strains that carried bla(CTX-M) on transferable plasmids. Two distinguishable plasmids were identified, each encoding bla(CTX-M-1) or bla(CTX-M-79). Transferrable bla(CTX-M) genes in bovine E. coli have the potential to serve as a reservoir of resistance for pathogens and may represent a public health concern.

Variable within- and between-Herd Diversity of CTX-M Cephalosporinase-Bearing Escherichia coli Isolates from Dairy Cattle

ABSTRACT bla CTX-M beta-lactamases confer resistance to critically important cephalosporin drugs. Recovered from both hospital- and community-acquired infections, bla CTX-M was first reported in U.S. livestock in 2010. It has been hypothesized that veterinary use of cephalosporins in livestock populations may lead to the dissemination of beta-lactamase-encoding genes. Therefore, our objectives were to estimate the frequency and distribution of coliform bacteria harboring bla CTX-M in the fecal flora of Ohio dairy cattle populations. In addition, we characterized the CTX-M alleles carried by the isolates, their plasmidic contexts, and the genetic diversity of the bacterial isolates themselves. We also evaluated the association between ceftiofur use and the likelihood of recovering cephalosporinase-producing bacteria. Thirty fresh fecal samples and owner-reported ceftiofur use data were collected from each of 25 Ohio dairy farms. Fecal samples (n = 747) yielded 70 bla CTX-M-positive Escherichia coli isolates from 5/25 herds, 715 bla CMY-2 E. coli isolates from 25/25 herds, and 274 Salmonella spp. from 20/25 herds. The within-herd prevalence among bla CTX-M-positive herds ranged from 3.3 to 100% of samples. Multiple pulsed-field gel electrophoresis (PFGE) patterns, plasmid replicon types, and CTX-M genes were detected. Plasmids with CTX-M-1, -15, and -14 alleles were clonal by restriction fragment length polymorphism (RFLP) within herds, and specific plasmid incompatibility group markers were consistently associated with each bla CTX-M allele. PFGE of total bacterial DNA showed similar within-herd clustering, with the exception of one herd, which revealed at least 6 different PFGE signatures. We were unable to detect an association between owner-reported ceftiofur use and the probability of recovering E. coli carrying bla CTX-M or bla CMY-2.

Genotypic and Phenotypic Properties of Cattle-Associated Campylobacter and Their Implications to Public Health in the USA

Since cattle are a major source of food and the cattle industry engages people from farms to processing plants and meat markets, it is conceivable that beef-products contaminated with Campylobacter spp. would pose a significant public health concern. To better understand the epidemiology of cattle-associated Campylobacter spp. in the USA, we characterized the prevalence, genotypic and phenotypic properties of these pathogens. Campylobacter were detected in 181 (19.2%) out of 944 fecal samples. Specifically, 71 C. jejuni, 132 C. coli, and 10 other Campylobacter spp. were identified. The prevalence of Campylobacter varied regionally and was significantly (P<0.05) higher in fecal samples collected from the South (32.8%) as compared to those from the North (14.8%), Midwest (15.83%), and East (12%). Pulsed Field Gel Electrophoresis (PFGE) analysis showed that C. jejuni and C. coli isolates were genotypically diverse and certain genotypes were shared across two or more of the geographic locations. In addition, 13 new C. jejuni and two C. coli sequence types (STs) were detected by Multi Locus Sequence Typing (MLST). C. jejuni associated with clinically human health important sequence type, ST-61 which was not previously reported in the USA, was identified in the present study. Most frequently observed clonal complexes (CC) were CC ST-21, CC ST-42, and CC ST-61, which are also common in humans. Further, the cattle associated C. jejuni strains showed varying invasion and intracellular survival capacity; however, C. coli strains showed a lower invasion and intracellular survival potential compared to C. jejuni strains. Furthermore, many cattle associated Campylobacter isolates showed resistance to several antimicrobials including ciprofloxacin, erythromycin, and gentamicin. Taken together, our results highlight the importance of cattle as a potential reservoir for clinically important Campylobacter.

Prevalence of antimicrobial resistance and association with toxin genes in Clostridium difficile in commercial swine.

OBJECTIVE To estimate prevalence and determine association between antimicrobia resistance and toxin gene profile of Clostridium difficile in commercial pigs at the preharvest food-safety level. ANIMALS 68 sows and 251 young pigs from 5 farms in North Carolina and 3 in Ohio. PROCEDURES Fecal samples were collected from sows (8/farm) and matched young pigs (32/farm) at farrowing and again at the nursery and finishing stages. Clostridium difficile isolates were tested for susceptibility to 6 antimicrobials. A PCR assay was used to detect genes coding for enterotoxin A (tcdA), cytotoxin B (tcdB), and binary toxin (cdtB). RESULTS C difficile prevalence in young pigs at farrowing was 73% (n=183) with significantly higher prevalence in Ohio (875%) than in North Carolina (64%). Clostridium difficile was isolated from 32 (47%) sows with no significant difference between the 2 regions. A single pig had a positive test result at the nursery, and no isolate was recovered at the finishing farms. Resistance to ciprofloxacin was predominant in young pigs (91.3% of isolates) and sows (94%). The antimicrobial resistance profile ciprofloxacin-erythromycin-tetracycline was detected in 21.4% and 11.7% of isolates from young pigs and sows, respectively. Most isolates had positive results for tcdA (65%), tcdB (84%), and the binary toxin cdtB (77%) genes. Erythromycin resistance and tetracycline resistance were significantly associated with toxin gene profiles. CONCLUSIONS AND CLINICAL RELEVANCE The common occurrence of antimicrobial-resistant C difficile and the significant association of toxigenic strains with antimicrobial resistance could contribute to high morbidity in farms with farrowing pigs.

Antimicrobial Resistance, Toxinotype, and Genotypic Profiling of Clostridium difficile Isolates of Swine Origin

ABSTRACT The occurrence of Clostridium difficile infections in patients that do not fulfill the classical risk factors prompted us to investigate new risk factors of disease. The goal of this study was to characterize strains and associated antimicrobial resistance determinants of C. difficile isolated from swine raised in Ohio and North Carolina. Genotypic approaches used include PCR detection, toxinotyping, DNA sequencing, and pulsed-field gel electrophoresis (PFGE) DNA fingerprinting. Thirty-one percent (37/119) of isolates carried both tetM and tetW genes. The ermB gene was found in 91% of isolates that were resistant to erythromycin (68/75). Eighty-five percent (521/609) of isolates were toxin gene tcdB and tcdA positive. A total of 81% (494/609) of isolates were positive for cdtB and carry a tcdC gene (a toxin gene negative regulator) with a 39-bp deletion. Overall, 88% (196/223) of pigs carry a single C. difficile strain, while 12% (27/223) of pigs carried multiple strains. To the best of our knowledge, this is the first report of individual pigs found to carry more than one strain type of C. difficile. A significant difference in toxinotype profiles in the two geographic locations was noted, with a significantly (P < 0.001) higher prevalence of toxinotype V found in North Carolina (84%; 189/224) than in Ohio (55%; 99/181). Overall, the study findings indicate that significant proportions of C. difficile in swine are toxigenic and often are associated with antimicrobial resistance genes, although they are not resistant to drugs that are used to treat C. difficile infections.

Yersinia enterocolitica of Porcine Origin: Carriage of Virulence Genes and Genotypic Diversity

Yersinia enterocolitica is an important foodborne pathogen, and pigs are recognized as a major reservoir and potential source of pathogenic strains to humans. A total of 172 Y. enterocolitica recovered from conventional and antimicrobial-free pig production systems from different geographic regions (North Carolina, Ohio, Michigan, Wisconsin, and Iowa) were investigated to determine their pathogenic significance to humans. Phenotypic and genotypic diversity of the isolates was assessed using antibiogram, serogrouping, and amplified fragment length polymorphism (AFLP). Carriage of chromosomal and plasmid-borne virulence genes were investigated using polymerase chain reaction. A total of 12 antimicrobial resistance patterns were identified. More than two-thirds (67.4%) of Y. enterocolitica were pan-susceptible, and 27.9% were resistant against β-lactams. The most predominant serogroup was O:3 (43%), followed by O:5 (25.6%) and O:9 (4.1%). Twenty-two of 172 (12.8%) isolates were found to carry Yersinia adhesion A (yadA), a virulence gene encoded on the Yersinia virulence plasmid. Sixty-nine (40.1%) isolates were found to carry ail gene. The ystA and ystB genes were detected in 77% and 26.2% of the strains, respectively. AFLP genotyping of isolates showed wide genotypic diversity and were grouped into nine clades with an overall genotypic similarity of 66.8-99.3%. AFLP analysis revealed that isolates from the same production system showed clonal relatedness, while more than one genotype of Y. enterocolitica circulates within a farm.

Comparison of Swiffer wipes and conventional drag swab methods for the recovery of Salmonella in swine production systems.

The main goal of this study was to assess the efficacy of Swiffer wipes in comparison to conventional drag swabs for the recovery of Salmonella. A total of 800 samples (400 Swiffer wipes and 400 drag swabs) were aseptically collected from randomly selected swine barns before disinfection with specific biocides and within 2 h after disinfection. From each barn, 10 samples of each swab type and negative controls were collected. Salmonellae were isolated from 43 (10.8%) of 400 drag swabs and 34 (8.5%) of 400 Swiffer wipes. There was a significant reduction in Salmonella postdisinfection as identified with both sampling procedures irrespective of the type of biocide used (P < 0.05). With the drag swabs, salmonellae were detected in 15% of the samples before disinfection versus 6.5% after disinfection, whereas with the Swiffer wipes, 13 and 4% of the samples were positive pre- and postdisinfection, respectively. Of the total 720 fecal samples collected from pigs placed in the disinfected barns, 132 (18.3%) were Salmonella positive. About 65 and 98% of the Salmonella isolates from swine barns and fecal samples, respectively, were resistant to one or more of the antimicrobials tested. Multidrug resistance was found in 35.7% of the isolates from barn swabs and 56.4% of the isolates from fecal samples. Results of this study suggest that the conventional drag swab method results in better recovery of Salmonella than does the Swiffer wipe method and thus could be a useful sampling method in monitoring Salmonella. Pentaresistant Salmonella (mainly R-type ACSSuT) was more common in fecal samples than in environmental samples.

Quantification of Campylobacter and Salmonella in Cattle Before, During, and After the Slaughter Process

Salmonella and Campylobacter cause a significant number of human illnesses globally, most of which are food related. Cattle can be asymptomatic carriers of both of these pathogens. The objective of this study was to determine the association between the concentration of Salmonella and Campylobacter pre- and postharvest in cattle. Samples were collected from each of 98 individually identified cattle during the periharvest and postharvest period. For each animal, four different phases were sampled: on farm (fecal sample), poststunning and exsanguination (hide sponge and rectal content sample [lairage]), prechilling (carcass sponge), and final product (ground meat). Salmonella and Campylobacter were cultured and quantified at each stage by using the direct dilution and most probable number (MPN) method. Salmonella was not isolated from any sample. The proportion (%) of samples that were Campylobacter positive was 77, 82, 97, 55, and 12 for farm, rectal content, hide, carcass, and meat samples respectively. The mean Campylobacter concentration for each sample was as follows: fecal sample from farm, 3.7×10(4) cfu/g; rectal content sample from lairage, 1.6×10(5) cfu/g; hide sponge, 0.9 cfu/cm(2); carcass sponge, 8.7 cfu/half carcass; and meat, 1.1 cfu/g. There were no associations between Campylobacter concentrations for any two sample types. This lack of association could indicate that there is an environmental reservoir that can contaminate the final meat product, or since the majority of animals were positive entering the slaughter process, that the process itself reduces the load of Campylobacter regardless of the initial concentration. In addition, contamination of beef may be more strongly associated with periharvest practices than animal carriage rates.