Publications by authors named "Wondwossen A Gebreyes"

78 Publications

Effects of flavophospholipol on conjugation and plasmid curing of multidrug-resistant Enteritidis in broiler chickens.

JAC Antimicrob Resist 2021 Mar 11;3(1):dlab022. Epub 2021 Mar 11.

Department of Veterinary Preventive Medicine, College of Veterinary Medicine, The Ohio State University, A100R Sisson Hall, 1920 Coffey Rd, Columbus, OH 43210, USA.

Background: Early studies suggested that flavophospholipol has plasmid-curing effects and could inhibit conjugation by disrupting pilus formation between bacteria.

Objectives: This 36-day controlled-challenge study aimed to evaluate the anti-conjugative and plasmid-curing effect of flavophospholipol on plasmid-mediated antimicrobial resistance (AMR) in MDR transconjugant Enteritidis in chickens.

Methods: A total of 270-day-old chicks were randomly assigned to four control and four treatment groups with two doses of in-feed flavophospholipol (10 ppm and 64 ppm) and in the presence and absence of ampicillin in drinking water. Chicks were orally challenged with Enteritidis with known plasmid-encoded AMR factors. Cloacal swabs were collected on Day 7, 14 and 23. On Day 35, all chickens were euthanized, and caecal tissue and content were collected. Antimicrobial susceptibility testing was done with a panel of 12 antimicrobials and interpreted according to CLSI breakpoints.

Results: Flavophospholipol given in-feed at 64 ppm had an anti-conjugative effect. There was a significant reduction of acquisition of resistance to ampicillin, streptomycin and tetracycline by the recipient strains of Enteritidis in treatment groups given flavophospholipol in-feed at 64 ppm (0.05). This was not seen with flavophospholipol given in-feed at 10 ppm.

Conclusions: The results demonstrate that flavophospholipol given in-feed at 64 ppm had an anti-conjugative effect. The results also suggest that AMR is reduced through other mechanisms of action, which are yet to be determined. There is insufficient evidence that flavophospholipol at 64 ppm in feed alone or with sub-therapeutic levels of antibiotics had a plasmid-curing effect.
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http://dx.doi.org/10.1093/jacamr/dlab022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8210090PMC
March 2021

The posthatch prophylactic use of ceftiofur affects the cecal microbiota similar to the dietary sanguinarine supplementation in broilers.

Poult Sci 2020 Nov 12;99(11):6013-6021. Epub 2020 Aug 12.

Department of Animal Science, Center for Agricultural Sciences, Federal University of Paraiba (CCA/UFPB), Areia, PB, Brazil; Global One Health initiative (GOHi), The Ohio State University, Columbus, OH, USA. Electronic address:

The prophylactic administration of ceftiofur to newly hatched chicks is a common practice in some hatcheries worldwide to mitigate early gastrointestinal infections caused by Enterobacteriaceae. In spite of the crucial role of the gut microbiome for the broiler's health, there is still limited information on how the microbial composition is affected by such procedure. We investigated the effects of posthatch prophylactic application of ceftiofur on the cecal microbiota of 14-day-old broilers fed regular or sanguinarine-supplemented diets. DNA samples were extracted from cecal contents, amplified for the V3-V4 regions of the microbial 16S rRNA gene, and sequenced in a high-throughput sequencing platform (Illumina MiSeq). After downstream bioinformatics and statistical analyses, our results demonstrated that both ceftiofur and sanguinarine treatments similarly increased the proportions of the phylum Bacteroidetes and the genera Bacteroides and Megamonas, whereas reduced the relative abundances of Firmicutes and Lachnospiraceae in the ceca of the birds. Such changes are probably associated with increased carbohydrate fermentation processes favoring the production of short-chain fatty acids. This was also corroborated by the functional prediction findings, which suggest an increase in some metabolic pathways associated with digestibility in broilers receiving ceftiofur. Considering that antimicrobial stewardship in animal production systems is strongly needed to mitigate the threat of antimicrobial resistance, our findings show that supplementation with a phytogenic feed additive can lead to a similar microbial composition in the ceca of commercial broiler chickens, suggesting that the use of alternative products could lead to functional modifications without increasing pressure for antimicrobial resistance.
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http://dx.doi.org/10.1016/j.psj.2020.06.078DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7647783PMC
November 2020

Molecular Epidemiology of Infectious Zoonotic and Livestock Diseases.

Microbiol Spectr 2020 03;8(2)

Population Health and Pathobiology (PHP), College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27606.

Zoonotic and livestock diseases are very important globally both in terms of direct impact on human and animal health and in terms of their relationship to the livelihood of farming communities, as they affect income generation and food security and have other, indirect consequences on human lives. More than two-thirds of emerging infectious diseases in humans today are known to be of animal origin. Bacterial, viral, and parasitic infections that originate from animals, including hypervirulent and multidrug-resistant (MDR) bacterial pathogens, such as livestock-associated methicillin-resistant (LA-MRSA), invasive nontyphoidal of animal origin, hyperviruent , and others, are of major significance to public health. Understanding the origin, risk factors, transmission, prevention, and control of such strains has been a challenge for various reasons, particularly due to the transdisciplinary partnership between and among human, environment, and animal health sectors. MDR bacteria greatly complicate the clinical management of human infections. Food animal farms, pets in communities, and veterinary hospital environments are major sources of such infections. However, attributing such infections and pinpointing sources requires highly discriminatory molecular methods as outlined in other parts of this curated series. Genotyping methods, such as multilocus sequence typing, pulsed-field gel electrophoresis, restriction fragment length polymorphism, and several others, have been used to decipher sources of foodborne and other zoonotic infectious diseases. In recent years, whole-genome-sequence-based approaches have been increasingly used for molecular epidemiology of diseases at the interface of humans, animals, and the environment. This part of the series highlights the major zoonotic and foodborne disease issues. *This article is part of a curated collection.
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http://dx.doi.org/10.1128/microbiolspec.AME-0011-2019DOI Listing
March 2020

Accuracy of Two Point-of-Care Tests for Rapid Diagnosis of Bovine Tuberculosis at Animal Level using Non-Invasive Specimens.

Sci Rep 2020 03 25;10(1):5441. Epub 2020 Mar 25.

Texas Biomedical Research Institute, San Antonio, TX, United States.

Bovine tuberculosis (BTB) testing in cattle requires a significant investment of time, equipment, and labor. Novel, rapid, cheaper and accurate methods are needed. The Alere Determine TB lipoarabinomannan antigen (LAM-test) is a World Health Organization-endorsed point-of-care urine test designed to detect active TB disease in humans. The Lionex Animal TB Rapid Test (Lionex-test) is a novel animal specific TB diagnostic blood test. An animal level analysis was performed using urine (n = 141) and milk (n = 63) samples from depopulated BTB-suspected cattle to test the accuracy of the LAM-test when compared to results of positive TB detection by any routine BTB tests (BOVIGAM, necropsy, histology, culture, PCR) that are regularly performed by the United States Department of Agriculture (USDA). The agreement between the urine LAM-test and USDA standard tests were poor at varying testing time points. The same milk samples did not elicit statistically significant agreement with the Lionex-test, although positive trends were present. Hence, we cannot recommend the LAM-test as a valid BTB diagnostic test in cattle using either urine or milk. The Lionex-test's production of positive trends using milk samples suggests larger sample sizes may validate the Lionex-test in accurately diagnosing BTB in cattle using milk samples, potentially providing a quick and reliable field test for BTB.
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http://dx.doi.org/10.1038/s41598-020-62314-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7096388PMC
March 2020

Burden and antimicrobial resistance of S. aureus in dairy farms in Mekelle, Northern Ethiopia.

BMC Vet Res 2020 Jan 22;16(1):20. Epub 2020 Jan 22.

Tropical Veterinary Medicine, College of Veterinary Medicine, Mekelle University, Mekelle, Ethiopia.

Background: Staphylococcus aureus is a frequent colonizer of human and several animal species, including dairy cows. It is the most common cause of intramammary infections in dairy cows. Its public health importance increases inline to the continuous emergence of drug-resistant strains; such as Methicillin-resistant S. aureus (MRSA). Indeed, the recent emergence of human and veterinary adapted MRSA demands serious attention. The aim of this study was to determine the burden and drug resistance pattern of S. aureus in dairy farms in Mekelle and determine the molecular characteristics of MRSA.

Results: This study was done on 385 lactating dairy cows and 71 dairy farmers. The ages of the cows and farmworkers were between 3 and 14 and 17-63 years respectively. S. aureus was isolated from 12.5% of cows and 31% of farmworkers. Highest resistance was observed for penicillin (> 90%) followed by tetracycline (32-35%) and trimethoprim-sulphamethoxazole (10-27%). But no resistance was observed for vancomycin, daptomycin, and rifampin. Only one isolate was MRSA both phenotypically and harboring mecA. This isolate was from nasal of a farmworker and was MRSA SCCmec Iva, spa type t064 of CC8. Multi-drug resistance was observed in 6.2% of cow isolates and 13.6% of nasal isolates.

Conclusions: In this study, S. aureus infected 12.5% of dairy cows and colonized 31% of farmworkers. Except for penicillin, resistance to other drugs was rare. Although no MRSA was found from dairy cows the existence of the human and animal adapted and globally spread strain, MRSA SCCmec IVa spa t064, warrants for a coordinated action to tackle AMR in both human and veterinary in the country.
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http://dx.doi.org/10.1186/s12917-020-2235-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6977269PMC
January 2020

Clostridioides difficile on Ohio swine farms (2015): A comparison of swine and human environments and assessment of on-farm risk factors.

Zoonoses Public Health 2019 11 7;66(7):861-870. Epub 2019 Aug 7.

Department of Veterinary Preventive Medicine, The Ohio State University College of Veterinary Medicine, Columbus, OH, USA.

Swine are known reservoirs for Clostridioides difficile, formerly known as Clostridium difficile, and transmission from swine to human farm workers is strongly suggested by previous studies. This cross-sectional study evaluated the potential role of farm environmental surfaces, including those in worker breakrooms and swine housing areas, in the possible transmission of C. difficile from swine to farm workers. Environmental surfaces and piglet faeces at 13 Ohio swine farms were sampled in 2015. Typical culturing techniques were performed to isolate C. difficile from samples, and amplification of toxin genes (tcdA, tcdB and cdtB) and PCR-ribotyping were used to genetically characterize recovered isolates. In addition, sequencing of toxin regulatory gene, tcdC, was done to identify the length of identified deletions in some isolates. A survey collected farm-level management risk factor information. Clostridioides difficile was recovered from all farms, with 42% (188/445) of samples testing positive for C. difficile. Samples collected from all on-farm locations recovered C. difficile, including farrowing rooms (60%, 107/178), breakrooms (50%, 69/138) and nursery rooms (9%, 12/129). Three ribotypes recovered from both swine and human environments (078, 412 and 005) have been previously implicated in human disease. Samples taken from farrowing rooms and breakrooms were found to have greater odds of C. difficile recovery than those taken from nursery rooms (OR = 40.5, OR = 35.6, p < .001 respectively). Farms that weaned ≥23,500 pigs per year had lower odds of C. difficile recovery as compared to farms that weaned fewer pigs (OR = 0.4, p = .01) and weekly or more frequent cleaning of breakroom counters was associated with higher odds of C. difficile recovery (OR = 11.7, p < .001). This study provides important insights into the presence and characterization of C. difficile found in human environments on swine farms and highlights how these areas may be involved in transmission of C. difficile to swine farm workers and throughout the facility.
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http://dx.doi.org/10.1111/zph.12637DOI Listing
November 2019

Genomic surveillance links livestock production with the emergence and spread of multi-drug resistant non-typhoidal Salmonella in Mexico.

J Microbiol 2019 Apr 5;57(4):271-280. Epub 2019 Feb 5.

Facultad de Medicina Veterinaria y Zootecnia, Universidad Nacional Autónoma de México, Mexico City, Mexico.

Multi-drug resistant (MDR) non-typhoidal Salmonella (NTS) is increasingly common worldwide. While food animals are thought to contribute to the growing antimicrobial resistance (AMR) problem, limited data is documenting this relationship, especially in low and middle-income countries (LMIC). Herein, we aimed to assess the role of non-clinical NTS of bovine origin as reservoirs of AMR genes of human clinical significance. We evaluated the phenotypic and genotypic AMR profiles in a set of 44 bovine-associated NTS. For comparative purposes, we also included genotypic AMR data of additional isolates from Mexico (n = 1,067) that are publicly available. The most frequent AMR phenotypes in our isolates involved tetracycline (40/44), trimethoprim-sulfamethoxazole (26/44), chloramphenicol (19/44), ampicillin (18/44), streptomycin (16/44), and carbenicillin (13/44), while nearly 70% of the strains were MDR. These phenotypes were correlated with a widespread distribution of AMR genes (i.e. tetA, aadA, dfrA12, dfrA17, sul1, sul2, bla-TEM-1, blaCARB-2) against multiple antibiotic classes, with some of them contributed by plasmids and/or class-1 integrons. We observed different AMR genotypes for betalactams and tetracycline resistance, providing evidence of convergent evolution and adaptive AMR. The probability of MDR genotype occurrence was higher in meat-associated isolates than in those from other sources (odds ratio 11.2, 95% confidence interval 4.5-27.9, P < 0.0001). The study shows that beef cattle are a significant source of MDR NTS in Mexico, highlighting the role of animal production on the emergence and spread of MDR Salmonella in LMIC.
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http://dx.doi.org/10.1007/s12275-019-8421-3DOI Listing
April 2019

Off-label use of ceftiofur in one-day chicks triggers a short-term increase of ESBL-producing E. coli in the gut.

PLoS One 2018 11;13(9):e0203158. Epub 2018 Sep 11.

Department of Animal Science, Center for Agricultural Sciences, Federal University of Paraiba (CCA/UFPB), Areia, Paraíba, Brazil.

This trial was designed to evaluate the off-label use of ceftiofur with Marek's vaccine in one-day-old broiler chicks, a prophylactic treatment that has been done in some commercial hatcheries, on the emergence of extended-spectrum beta-lactamase producing Escherichia coli (ESBL-E. coli). A total of 168 chicks (Cobb500®) were used in a completely randomized design. Birds were assigned to two treatments (Marek's vaccine plus saline vs Marek's vaccine plus ceftiofur) and six repetitions, with 14 animals each. Cloacal swabs were collected from 1 to 14 days post-hatch. The majority (86%; p<0.0001) of the ESBL-producing isolates harboring blaCTX-M and blaSHV genes originated from animals receiving the antimicrobial. None of the isolates were positive for plasmid-mediated AmpC betalactamase genes (blaACC, blaCMY-2, blaDHA, blaFOX, blaMOX and blaMIR). These findings indicate that the off-label use of ceftiofur with Marek's vaccine is associated with the short-term increase in ESBL-producing Escherichia coli in the gut of chicks.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0203158PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6133352PMC
February 2019

Phenotypic and genotypic characterization of temporally related nontyphoidal Salmonella strains isolated from humans and food animals in central Ethiopia.

Zoonoses Public Health 2018 11 8;65(7):766-776. Epub 2018 Jul 8.

Department of Pharmacology and Clinical Pharmacy, College of Health Sciences, Addis Ababa University, Addis Ababa, Ethiopia.

Salmonella is one of the common causes of food-borne bacterial illnesses. The primary sources of human nontyphoidal Salmonella (NTS) infection are food animals. This study characterized temporally and spatially related Salmonella isolated during April 2013 to March 2014 from faeces of diarrhoeic human patients in Addis Ababa (n = 68) and food animals (n = 84) in Addis Ababa and surrounding districts (dairy cattle, n = 30; slaughtered cattle, n = 20; poultry, n = 26; swine n = 8). Isolates were serotyped, page typed and tested for antimicrobial susceptibility using Kirby-Bauer disc diffusion method, and genotyped by pulsed-field gel electrophoresis (PFGE). The dominant Salmonella serovars isolated from food animals were S. Saintpaul (38.1%), S. Typhimurium (17.9%) and S. Kentucky (9.5%), whereas in humans, S. Typhimurium (39.7%), S. Virchow (30.9%) and S. Kottbus (10.3%) were frequently isolated. Resistance to streptomycin, sulfisoxazole, tetracycline, ampicillin and cephalothin was higher in animal isolates than human isolates, and mean number of antimicrobials to which isolates were resistant was significantly higher in isolates from cattle and poultry compared to those from humans (p < 0.05). All S. Kentucky isolated from animals and humans were multidrug resistant (MDR) with shared resistance phenotype (AmpCfCipTeSuSNa). Although this study involved small sample size and was not able to show clear epidemiological linkage among isolates from various sources, genotyping by PFGE analysis demonstrated circulation of closely related genotypes of S. Virchow, S. Typhimurium and S. Kentucky among humans and food animals. Detection of related Salmonella isolates from humans and animals, the high MDR status of isolates from animals and close proximity of farms and human residential areas in the absence of appropriate biosecurity present major public health problem. Integrated surveillance of Salmonella serovars in humans and animals and implementation of appropriate hazard analysis and pathogen control strategies along critical points of the food chain from farm to table is recommended.
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http://dx.doi.org/10.1111/zph.12490DOI Listing
November 2018

Whole genome sequencing reveals widespread distribution of typhoidal toxin genes and VirB/D4 plasmids in bovine-associated nontyphoidal Salmonella.

Sci Rep 2018 06 29;8(1):9864. Epub 2018 Jun 29.

Facultad de Medicina Veterinaria y Zootecnia, Universidad Nacional Autónoma de México, Mexico City, 04510, Mexico.

Nontyphoidal Salmonella (NTS) is a common pathogen in food-producing animals and a public health concern worldwide. Various NTS serovars may be present in apparently healthy animals. This could result in carcass contamination during the slaughter process leading to human exposure. While most genomic research has focused on Salmonella pathogenesis, little is known on the factors associated with subclinical infections and environmental persistence. We report here the widespread distribution of typhoidal toxin genes (i. e. the cdtB islet, hlyE, taiA), among NTS strains from a beef slaughter operation (n = 39) and from epidemiologically unconnected ground beef (n = 20). These genes were present in 76% of the strains, regardless of serovar, isolation source or geographical location. Moreover, strains that predominated in the slaughterhouse carry plasmid-borne type IV secretion systems (T4SS), which have been linked to persistent infections in numerous pathogens. Population genomics supports clonal dissemination of NTS along the food production chain, highlighting its role as reservoir of genetic variability in the environment. Overall, the study provides a thorough characterization of serovar diversity and genomic features of beef-associated NTS in Mexico. Furthermore, it reveals how common genetic factors could partially explain the emergence and persistence of certain NTS serovars in the beef industry.
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http://dx.doi.org/10.1038/s41598-018-28169-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6026178PMC
June 2018

Accuracy of PCR targeting different markers for Staphylococcus aureus identification: a comparative study using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry as the gold standard.

J Vet Diagn Invest 2018 Mar 6;30(2):252-255. Epub 2017 Nov 6.

Departments of Animal Sciences (Saraiva, De Leon, Santos, Oliveira), College of Agricultural Sciences, Federal University of Paraiba, Areia, Paraíba, Brazil.

Staphylococcus aureus is considered a major pathogen in veterinary and human medicine, and the emergence of multidrug-resistant strains, such as livestock-associated methicillin-resistant S. aureus, means that reliable, inexpensive, and fast methods are required to identify S. aureus obtained from animal sources. We tested the accuracy of a PCR targeting the genes femA, nuc, and coa in identifying S. aureus from animals. A total of 157 Staphylococcus spp. isolates were examined by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry; 18 different Staphylococcus species were identified. Of 68 S. aureus isolates, the genes femA, nuc, and coa were found in 61, 53, and 32 isolates, respectively. Considering MALDI-TOF as the gold standard, the PCR assays targeting all 3 genes showed 100% specificity; the sensitivity values were 89.7, 77.9, and 47.0% for femA, nuc, and coa, respectively. Sensitivity was 100% when femA and nuc markers were targeted simultaneously. These results confirm PCR as an accurate method to identify S. aureus species from animal sources and strongly suggest the simultaneous use of primers targeting femA and nuc genes.
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http://dx.doi.org/10.1177/1040638717732370DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6505873PMC
March 2018

Genetic markers associated with resistance to beta-lactam and quinolone antimicrobials in non-typhoidal isolates from humans and animals in central Ethiopia.

Antimicrob Resist Infect Control 2017 17;6:13. Epub 2017 Jan 17.

Department of Pharmacology and Clinical Pharmacy, School of Pharmacy, College of Health Sciences, Addis Ababa University, Churchill Avenue, P.O. Box 1176, Addis Ababa, Ethiopia.

Background: Beta-lactam and quinolone antimicrobials are commonly used for treatment of infections caused by non-typhoidal (NTS) and other pathogens. Resistance to these classes of antimicrobials has increased significantly in the recent years. However, little is known on the genetic basis of resistance to these drugs in isolates from Ethiopia.

Methods: isolates with reduced susceptibility to beta-lactams ( = 43) were tested for genes encoding for beta-lactamase enzymes, and those resistant to quinolones ( = 29) for mutations in the quinolone resistance determining region (QRDR) as well as plasmid mediated quinolone resistance (PMQR) genes using PCR and sequencing.

Results: Beta-lactamase genes () were detected in 34 (79.1%) of the isolates. The dominant gene was TEM, recovered from 33 (76.7%) of the isolates, majority being TEM-1 (24, 72.7%) followed by TEM-57, (10, 30.3%). The OXA-10 and CTX-M-15 were detected only in a single Concord human isolate. Double substitutions in A (Ser83-Phe + Asp87-Gly) as well as C (Thr57-Ser + Ser80-Ile) subunits of the quinolone resistance determining region (QRDR) were detected in all Kentucky isolates with high level resistance to both nalidixic acid and ciprofloxacin. Single amino acid substitutions, Ser83-Phe ( = 4) and Ser83-Tyr ( = 1) were also detected in the A gene. An isolate of . Miami susceptible to nalidixic acid but intermediately resistant to ciprofloxacin had Thr57-Ser and an additional novel mutation (Tyr83-Phe) in the C gene. Plasmid mediated quinolone resistance (PMQR) genes investigated were not detected in any of the isolates. In some isolates with decreased susceptibility to ciprofloxacin and/or nalidixic acid, no mutations in QRDR or PMQR genes were detected. Over half of the quinolone resistant isolates in the current study 17 (58.6%) were also resistant to at least one of the beta-lactam antimicrobials.

Conclusion: Acquisition of TEM was the principal beta-lactamase resistance mechanism and mutations within QRDR of A and C were the primary mechanism for resistance to quinolones. Further study on extended spectrum beta-lactamase and quinolone resistance mechanisms in other gram negative pathogens is recommended.
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http://dx.doi.org/10.1186/s13756-017-0171-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5240271PMC
January 2017

Fecal prevalence, serotype distribution and antimicrobial resistance of Salmonellae in dairy cattle in central Ethiopia.

BMC Microbiol 2016 Feb 16;16:20. Epub 2016 Feb 16.

Department of Microbial Infection and Immunity, Center for Microbial Interface Biology, The Ohio State University, Biomedical Research Tower, 460 West 12th, Columbus, OH, 43210-1214, USA.

Background: Salmonellae are major worldwide zoonotic pathogens infecting a wide range of vertebrate species including humans. Consumption of contaminated dairy products and contact with dairy cattle represent a common source of non-typhoidal Salmonella infection in humans. Despite a large number of small-scale dairy farms in Addis Ababa and its surrounding districts, little is known about the status of Salmonella in these farms.

Results: Salmonella was recovered from the feces of at least one animal in 7.6% (10/132) of the dairy farms. Out of 1203 fecal samples examined, 30 were positive for Salmonella resulting in a weighted animal level prevalence of 2.3%. Detection of diarrhea in an animal and in a farm was significantly associated with animal level (p = 0.012) and herd level (p < 0.001) prevalence of Salmonella. Animal level prevalence of Salmonella was significantly associated with age (p = 0.023) and study location; it was highest among those under 6 months of age and in farms from Adaa district and Addis Ababa (p < 0.001). Nine different serotypes were identified using standard serological agglutination tests. The most frequently recovered serotypes were Salmonella Typhimurium (23.3%), S. Saintpaul (20%), S. Kentucky (16.7%) and S. Virchow (16.7%). All isolates were resistant or intermediately resistant to at least one of the 18 drugs tested. Twenty-six (86.7%), 19 (63.3 %), 18 (60%), 16 (53.3%) of the isolates were resistant to streptomycin, nitrofurantoin, sulfisoxazole and tetracycline , respectively. Resistance to 2 drugs was detected in 27 (90%) of the isolates. Resistance to 3 or more drugs was detected in 21 (70%) of the isolates, while resistance to 7 or more drugs was detected in 11 (36.7%) of the isolates. The rate of occurrence of multi-drug resistance (MDR) in Salmonella strains isolated from dairy farms in Addis Ababa was significantly higher than those isolated from farms outside of Addis Ababa (p = 0.009). MDR was more common in S. Kentucky, S. Virchow and S. Saintpaul.

Conclusion: Isolation of Salmonella serotypes commonly known for causing human salmonellosis that are associated with an MDR phenotype in dairy farms in close proximity with human population is a major public health concern. These findings imply the need for a strict pathogen reduction strategy.
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http://dx.doi.org/10.1186/s12866-016-0638-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4754838PMC
February 2016

Non-typhoidal Salmonella serotypes, antimicrobial resistance and co-infection with parasites among patients with diarrhea and other gastrointestinal complaints in Addis Ababa, Ethiopia.

BMC Infect Dis 2015 Nov 4;15:497. Epub 2015 Nov 4.

Department of Pharmacology and Clinical Pharmacy, School of Pharmacy, College of Health Sciences, Addis Ababa University, Churchill Avenue, P.O. Box 1176, Addis Ababa, Ethiopia.

Background: Non-typhoidal Salmonella (NTS) is an important public health problem worldwide. Consumption of animal-derived food products and direct and/or indirect contact with animals are the major routes of acquiring infection with NTS. Published information, particularly on the serotype distribution of NTS among human patients with gastroenteritis and associated risk factors, is scarce in Ethiopia. This study investigated the prevalence, risk factors, serotype distribution and antimicrobial susceptibility of Salmonella species among diarrheic out-patients attending health centers in Addis Ababa and patients with various gastrointestinal complaints at Tikur Anbessa Specialized Hospital (TASH).

Methods: Stool samples were cultured for Salmonella species according to the WHO Global Foodborne Infections Network laboratory protocol. Salmonella serotyping was conducted using slide agglutination and microplate agglutination techniques. Antibiotic susceptibility testing was performed using the disk diffusion method according to Clinical and Laboratory Standards Institute guidelines.

Results: A total of 59 (6.2 %) stool samples, out of 957 were culture positive for Salmonella species. Fifty-five (7.2 %) of 765 diarrheic patients from health centers and 4 (2.1 %) of 192 patients from TASH were culture positive for Salmonella species. Multivariable logistic regression analysis after adjusting for all other variables revealed statistically significant association of Salmonella infection with consumption of raw vegetables (OR = 1.91, 95 % CI = 1.29-2.83, χ(2) = 4.74, p = 0.025) and symptom of watery diarrhea (OR = 3.3, 95 % CI = 1.23-8.88, χ(2) = 10.54, p = 0.005). Eleven serotypes were detected, and the most prominent were S. Typhimurium (37.3 %), S. Virchow (34 %), and S. Kottbus (10.2 %). Other serotypes were S. Miami, S. Kentucky, S. Newport, S. Enteritidis, S. Braenderup, S. Saintpaul, S. Concord and S. V:ROUGH-O. Resistance to three or more antimicrobials was detected in 27 (40.3 %) of the isolates. Resistance to five or more antimicrobials was detected in 17 (25.4 %). Resistance to individual antimicrobials was found at varying proportions: streptomycin (50; 74.6 %), nitrofurantoin (27; 40.3 %), sulfisoxazole (26; 38.8 %), kanamycin (23; 34.3 %), cephalothin (12; 17.9 %), and ampicillin (11; 16.4 %) respectively. Two S. Kentucky, one S. Typhimurium and one S. Concord isolates were multi-drug resistant to more than 10 antimicrobials.

Conclusions: The study demonstrated significant association of Salmonella infection with consumption of raw vegetables. There was no significant association of Salmonella infection with co-occurring parasites. The study also showed the dominance of S. Typhimurium and S. Virchow in primary health care units. Overall, prevalence of MDR was low compared to previous studies. Although their proportion was low, S. Kentucky and S. Concord demonstrated wider spectrum of MDR. Continuous monitoring of circulating serotypes, antimicrobial resistance profile and characterization on molecular resistance determinants is essential for proper treatment of patients and for identifying potential environmental origins of antimicrobial resistance.
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http://dx.doi.org/10.1186/s12879-015-1235-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4634906PMC
November 2015

Supplementation with Quaternary Benzo(c)phenanthridine Alkaloids Decreased Salivary Cortisol and Salmonella Shedding in Pigs After Transportation to the Slaughterhouse.

Foodborne Pathog Dis 2015 Nov 22;12(11):891-7. Epub 2015 Sep 22.

1 Department of Veterinary Preventive Medicine, College of Veterinary Medicine, The Ohio State University , Columbus, Ohio.

The present study was aimed at evaluating the effect of herbal extracts supplementation, particularly quaternary-benzo(c)phenanthridine alkaloids (QBA), which have been previously demonstrated to have anti-inflammatory, antimicrobial, and immune-modulator effects. We investigated the role of QBA on stress response and Salmonella shedding in finishing pigs transported to the slaughterhouse. A total of 82 pigs were orally challenged with a Salmonella cocktail (day 0) containing Salmonella Meleagridis, Hartford, Bovismorbificans and Newport serovars and randomly assigned to three treatment groups after 2 wks (day [D] 14): T1, in-feed QBA; T2, in-feed and water-soluble QBA; CON, nonsupplemented). Pigs were transported to the slaughterhouse 2 weeks after intervention (D 28) and slaughtered after nearly 19 h (D 29). Saliva, fecal samples, and carcass swabs were collected from all pigs. Salivary cortisol, Salmonella shedding, and carcass contamination were measured. A high positive correlation (Spearman rank correlation coefficient range 0.82-0.93) between salivary cortisol and Salmonella shedding was found after transportation in all groups (p < 0.05). Only the CON group showed an increase in salivary cortisol after transportation (5.48 ng/mL; p < 0.0001) to concentrations that were higher than in T1 (2.73 ng/mL; p = 0.0002) and T2 (1.88 ng/mL; p < 0.0001). Salmonella prevalence and shedding decreased after transportation in pigs receiving the QBA intervention (p < 0.05), whereas the control group showed a significant increase in Salmonella shedding after transportation (p = 0.04). At D 28, pigs in T2 shed lower numbers of Salmonella as compared to T1 (1.3E + 02 CFU/mL versus 8E + 03 CFU/mL; p = 0.002). Additionally, carcass contamination by Salmonella was higher in the CON group than the treated groups (p = 0.01). The findings show QBA intervention was effective in reducing transportation stress of pigs, resulting in reduced Salmonella shedding and positively impacting animal welfare and pork safety.
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http://dx.doi.org/10.1089/fpd.2015.2009DOI Listing
November 2015

The use of pulsed-field gel electrophoresis for genotyping of Clostridium difficile.

Methods Mol Biol 2015 ;1301:95-101

Department of Veterinary Preventive Medicine, Infectious Diseases Molecular Epidemiology Laboratory, The Ohio State University, Columbus, OH, USA,

Genotyping approaches are important for tracking infectious agents and can be used for various purposes. Pulsed-Field Gel Electrophoresis (PFGE) is among the highly discriminatory genotyping approaches commonly used for characterizing Clostridium difficile. Other genotyping methods used for C. difficile include Ribotyping, Restriction Endonuclease Assay (REA), Multilocus Variable Number Tandem Repeats (VNTR) Assay, and others. PFGE has a high discriminatory power, high reproducibility, and typeability. We utilized PFGE for typing C. difficile isolates of porcine and human origin. We used a macrorestriction fragment analysis of an intact genomic DNA using SmaI, a rare cutting restriction endonuclease. Using a Contour-Clamped Homogeneous Electric Field (CHEF) system with running conditions of 120° angle; initial switch time of 5 s; final switch time of 40 s with a run time of 18 h in a low-melting temperature agarose (Seakem Gold); and 0.5× TBE circulated in the CHEF system at 6 V/cm [CDC (2014) Pulsenet. http://www.cdc.gov/pulsenet/index.html . Accessed 22 Aug 2014] supported by 14 °C cooling module, we were able to separate very large DNA fragments (up to 2 Mb).
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http://dx.doi.org/10.1007/978-1-4939-2599-5_9DOI Listing
December 2015

Pre-parturition staphylococcal mastitis in primiparous replacement goats: persistence over lactation and sources of infection.

Vet Res 2014 Dec 9;45:115. Epub 2014 Dec 9.

Department of Animal Science, Federal University of Paraiba, Areia, PB, 58397-000, Brazil.

This investigation reported for the first time the occurrence of intramammary infections caused by Staphylococcus in primiparous replacement goats before parturition and the persistence of clinical Staphylococcus aureus infection during the lactation period. Subclinical infections, mainly caused by coagulase negative staphylococci (CoNS), did not persist during lactation. Genotyping analysis indicated that environment seems to play a moderate role as source of intramammary infections to goats before parturition, but causative agents of mastitis in lactating animals are not genotypically related to environmental staphylococci. The occurrence and persistence of intramammary infections in replacement goats demonstrate the need to consider those animals as potential sources of infections in dairy goat herds.
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http://dx.doi.org/10.1186/s13567-014-0115-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4302595PMC
December 2014

The global one health paradigm: challenges and opportunities for tackling infectious diseases at the human, animal, and environment interface in low-resource settings.

PLoS Negl Trop Dis 2014 13;8(11):e3257. Epub 2014 Nov 13.

Global Health Programs, College of Veterinary Medicine, The Ohio State University and VPH-Biotec Global Consortium, Columbus, Ohio, United States of America.

Zoonotic infectious diseases have been an important concern to humankind for more than 10,000 years. Today, approximately 75% of newly emerging infectious diseases (EIDs) are zoonoses that result from various anthropogenic, genetic, ecologic, socioeconomic, and climatic factors. These interrelated driving forces make it difficult to predict and to prevent zoonotic EIDs. Although significant improvements in environmental and medical surveillance, clinical diagnostic methods, and medical practices have been achieved in the recent years, zoonotic EIDs remain a major global concern, and such threats are expanding, especially in less developed regions. The current Ebola epidemic in West Africa is an extreme stark reminder of the role animal reservoirs play in public health and reinforces the urgent need for globally operationalizing a One Health approach. The complex nature of zoonotic diseases and the limited resources in developing countries are a reminder that the need for implementation of Global One Health in low-resource settings is crucial. The Veterinary Public Health and Biotechnology (VPH-Biotec) Global Consortium launched the International Congress on Pathogens at the Human-Animal Interface (ICOPHAI) in order to address important challenges and needs for capacity building. The inaugural ICOPHAI (Addis Ababa, Ethiopia, 2011) and the second congress (Porto de Galinhas, Brazil, 2013) were unique opportunities to share and discuss issues related to zoonotic infectious diseases worldwide. In addition to strong scientific reports in eight thematic areas that necessitate One Health implementation, the congress identified four key capacity-building needs: (1) development of adequate science-based risk management policies, (2) skilled-personnel capacity building, (3) accredited veterinary and public health diagnostic laboratories with a shared database, and (4) improved use of existing natural resources and implementation. The aim of this review is to highlight advances in key zoonotic disease areas and the One Health capacity needs.
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http://dx.doi.org/10.1371/journal.pntd.0003257DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4230840PMC
May 2016

Association of multicellular behaviour and drug resistance in Salmonella enterica serovars isolated from animals and humans in Ethiopia.

J Appl Microbiol 2014 Oct 7;117(4):961-971. Epub 2014 Jul 7.

Department of Microbial Infection and Immunity, Center for Microbial Interface Biology, The Ohio State University, Biomedical Research Tower, 460 West 12th, Columbus, OH 43210-1214.

Aims: To determine the association between multicellular behaviour, integron status and antibiotic resistance among 87 Ethiopian Salmonella enterica isolates of animal and human origin.

Methods And Results: Isolates were characterized for their biofilm forming ability, antimicrobial susceptibility and the presence and characteristics of a class 1 integron and Salmonella genomic island 1 (SGI1). The majority of isolates grown at environmental temperatures (20°C) exhibited robust biofilm formation (72·4%) and displayed RDAR colony morphology on Congo red agar plates. The presence of a class 1 integron correlated with the extent of drug resistance and ability to exhibit multicellular behaviour.

Conclusions: Although cellulose production and RDAR morphology correlated with increased multicellular behaviour, neither was required for biofilm formation. Contrary to previous reports, colony morphology was generally consistent within a serovar. No integrons were detected in isolates deficient for multicellular behaviour, indicating a potential role of bacterial community formation in transfer of genetic elements among environmental isolates.

Significance And Impact Of Study: Infection by Salm. enterica is a major public health problem worldwide. The dominance of multidrug resistance and multicellular behaviour in Salmonella isolates of Ethiopian origin highlights a need for integrated surveillance and further detailed phenotypic and molecular studies of isolates from this region.
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http://dx.doi.org/10.1111/jam.12579DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4165716PMC
October 2014

Distribution, quantitative load and characterization of Salmonella associated with swine farms in upper-northern Thailand.

J Vet Sci 2014 21;15(2):327-34. Epub 2014 Mar 21.

Department of Food Animal Clinic, Faculty of Veterinary Medicine, Chiang Mai University, Chiang Mai 50200, Thailand.

This study was conducted to analyze the prevalence and quantitative loads of Salmonella spp. on pig farms in Chiang Mai, Lamphun, Thailand to assess loading levels before slaughtering. The serotype diversity, antimicrobial-resistance pattern and pulse-field type of Salmonella spp. were also characterized to assess the dynamic propagation of the pathogen. The Salmonella-positive prevalence was 246/805 (30.56%), and the quantitative loads varied from 1.48 ˜ 4.04 Log10MPN/g, with a mean ± standard deviation of 2.11 ± 0.57. AMP/S/TE (ampicillin/streptomycin/tetracycline) was the highest frequency antimicrobial resistance pattern found in this study. In addition, Salmonella Rissen was the primary serotype in this region. PFGE results indicated the occurrence of infection by cross contamination among pig farms. Our study showed that pork is easily contaminated with this pathogen. Farm control programs must be based on strict biosecurity and hygienic measures, which could further reduce the contamination pressure at slaughterhouses or retail shops.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4087236PMC
http://dx.doi.org/10.4142/jvs.2014.15.2.327DOI Listing
February 2015

In-feed use of heavy metal micronutrients in U.S. swine production systems and its role in persistence of multidrug-resistant salmonellae.

Appl Environ Microbiol 2014 Apr 31;80(7):2317-25. Epub 2014 Jan 31.

Department of Veterinary Preventive Medicine, College of Veterinary Medicine, The Ohio State University, Columbus, Ohio, USA.

The study aimed to characterize the role of heavy metal micronutrients in swine feed in emergence of heavy-metal-tolerant and multidrug-resistant Salmonella organisms. We conducted a longitudinal study in 36 swine barns over a 2-year period. The feed and fecal levels of Cu(2+) and Zn(2+) were measured. Salmonella was isolated at early and late finishing. MICs of copper sulfate and zinc chloride were measured using agar dilution. Antimicrobial susceptibility was tested using the Kirby-Bauer method, and 283 isolates were serotyped. We amplified pcoA and czcD genes that encode Cu(2+) and Zn(2+) tolerance, respectively. Of the 283 isolates, 113 (48%) showed Cu(2+) tolerance at 24 mM and 164 (58%) showed Zn(2+) tolerance at 8 mM. In multivariate analysis, serotype and source of isolates were significantly associated with Cu(2+) tolerance (P < 0.001). Fecal isolates were more likely to be Cu(2+) tolerant than those of feed origin (odds ratio [OR], 27.0; 95% confidence interval [CI], 2.8 to 250; P = 0.0042) or environmental origin (OR, 5.8), implying the significance of gastrointestinal selective pressure. Salmonella enterica serotypes Typhimurium and Heidelberg, highly significant for public health, had higher odds of having >20 mM MICs of Cu(2+) than did "other" serotypes. More than 60% of Salmonella isolates with resistance type (R-type) AmStTeKm (32 of 53) carried pcoA; only 5% with R-type AmClStSuTe carried this gene. czcD gene carriage was significantly associated with a higher Zn(2+) MIC (P < 0.05). The odds of having a high Zn(2+) MIC (≥8 mM) were 14.66 times higher in isolates with R-type AmClStSuTe than in those with R-type AmStTeKm (P < 0.05). The findings demonstrate strong association between heavy metal tolerance and antimicrobial resistance, particularly among Salmonella serotypes important in public health.
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http://dx.doi.org/10.1128/AEM.04283-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3993138PMC
April 2014

Effects of quaternary benzo(c)phenanthridine alkaloids on growth performance, shedding of organisms, and gastrointestinal tract integrity in pigs inoculated with multidrug-resistant Salmonella spp.

Am J Vet Res 2013 Dec;74(12):1530-5

Department of Population Health and Pathobiology, College of Veterinary Medicine, North Carolina State University, Raleigh, NC 27695.

Objective: To evaluate effects of quaternary benzo(c)phenanthridine alkaloids (QBAs) against Salmonella spp and determine effects on growth performance, organism shedding, and gastrointestinal tract integrity in pigs inoculated with Salmonella enterica serovar Typhimurium.

Sample: 36 Salmonella isolates and twenty 5-week-old pigs.

Procedures: Minimum inhibitory concentration of QBAs against the Salmonella isolates was determined. Pigs were allocated to 4 groups and inoculated with Salmonella organisms. Pigs received diets supplemented with 1.5 g of QBAs/1,000 kg of feed, 0.75 g of QBAs/1,000 kg of feed, or 59.4 g of chlortetracycline/1,000 kg of feed or a nonsupplemented (control) diet. Pigs were weighed on day 0 and then weekly for 40 days. Fecal samples were collected to quantify Salmonella organisms. Gastrointestinal tract integrity was evaluated by measuring transepithelial resistance.

Results: In vitro, 9 of 36 (25%) Salmonella isolates were inhibited at 90 μg of QBAs/mL; all 36 were inhibited at 179 μg of QBAs/mL. Diets containing QBAs significantly decreased Salmonella spp shedding; shedding was lower 40 days after inoculation for pigs fed diets containing QBAs or chlortetracycline than for pigs fed the control diet. Growth performance was similar for pigs fed diets containing QBA or chlortetracycline. Gastrointestinal tract integrity was improved in pigs fed the diet containing 1.5 g of QBAs/1,000 kg of feed.

Conclusions And Clinical Relevance: QBAs and chlortetracycline decreased Salmonella spp shedding but did not differ with regard to growth performance. Gastrointestinal tract integrity was better, albeit not significantly, in pigs fed diets containing QBAs. Further investigation into the role of QBAs and their mechanism as an immunomodulator is necessary.
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http://dx.doi.org/10.2460/ajvr.74.12.1530DOI Listing
December 2013

Comparative phenotypic and genotypic characterization of temporally related nontyphoidal Salmonella isolated from human clinical cases, pigs, and the environment in North Carolina.

Foodborne Pathog Dis 2014 Feb 16;11(2):156-64. Epub 2013 Nov 16.

1 Department of Population Health and Pathobiology, College of Veterinary Medicine, North Carolina State University , Raleigh, North Carolina.

Nontyphoidal Salmonella infections caused by antimicrobial-resistant (AMR) strains are of great public health concern. We compared the phenotypic and genotypic relationships among temporally and spatially related AMR Salmonella isolates (n=1058) representing several predominant serovars, including Salmonella Typhimurium, Salmonella Typhimurium var. 5-, Salmonella Derby, Salmonella Heidelberg, Salmonella Muenchen, Salmonella Schwarzengrund, and Salmonella Rissen of human clinical cases (n=572), pig (n=212), and farm environment (n=274) origin in North Carolina. Antimicrobial susceptibility testing was performed using the broth microdilution method, and genotypic resistance determinants, including class I and II integrons, were identified. Overall, Salmonella isolates exhibited the highest frequency of resistance to tetracycline (50%), followed by sulfisoxazole (36%) and streptomycin (27%). We identified 16 different antimicrobial resistance genes, including extended spectrum and AmpC β-lactamases-producing genes (bla(TEM), bla(PSE), and bla(CMY-2)), in all the β-lactam- and cephalosporin-resistant Salmonella isolates from humans, pigs, and the environment. Class I integrons of 1-kb and 1.2-kb size were identified from all the three sources (humans, 66%; pigs, 85%; environment, 58%), while Class II integrons of 2-kb size were identified only in pig (10%) and environmental (19%) isolates. We detected genotypic similarity between Salmonella Typhimurium isolated from humans, pigs, and the environment while serovars Derby, Heidelberg, and Muenchen exhibited genotypic diversity. Detection of AMR Salmonella isolates from humans, pigs, and the environment is a concern for clinicians and veterinarians to mitigate the dissemination of AMR Salmonella strains.
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http://dx.doi.org/10.1089/fpd.2013.1630DOI Listing
February 2014

Longitudinal study of distributions of similar antimicrobial-resistant Salmonella serovars in pigs and their environment in two distinct swine production systems.

Appl Environ Microbiol 2013 Sep 21;79(17):5167-78. Epub 2013 Jun 21.

Department of Population Health and Pathobiology, College of Veterinary Medicine, North Carolina State University, Raleigh, North Carolina, USA.

The aim of this longitudinal study was to determine and compare the prevalences and genotypic profiles of antimicrobial-resistant (AR) Salmonella isolates from pigs reared in antimicrobial-free (ABF) and conventional production systems at farm, at slaughter, and in their environment. We collected 2,889 pig fecal and 2,122 environmental (feed, water, soil, lagoon, truck, and floor swabs) samples from 10 conventional and eight ABF longitudinal cohorts at different stages of production (farrowing, nursery, finishing) and slaughter (postevisceration, postchill, and mesenteric lymph nodes [MLN]). In addition, we collected 1,363 carcass swabs and 205 lairage and truck samples at slaughter. A total of 1,090 Salmonella isolates were recovered from the samples; these were isolated with a significantly higher prevalence in conventionally reared pigs (4.0%; n = 66) and their environment (11.7%; n = 156) than in ABF pigs (0.2%; n = 2) and their environment (0.6%; n = 5) (P < 0.001). Salmonella was isolated from all stages at slaughter, including the postchill step, in the two production systems. Salmonella prevalence was significantly higher in MLN extracted from conventional carcasses than those extracted from ABF carcasses (P < 0.001). We identified a total of 24 different serotypes, with Salmonella enterica serovar Typhimurium, Salmonella enterica serovar Anatum, Salmonella enterica serovar Infantis, and Salmonella enterica serovar Derby being predominant. The highest frequencies of antimicrobial resistance (AR) were exhibited to tetracycline (71%), sulfisoxazole (42%), and streptomycin (17%). Multidrug resistance (resistance to ≥ 3 antimicrobials; MDR) was detected in 27% (n = 254) of the Salmonella isolates from the conventional system. Our study reports a low prevalence of Salmonella in both production systems in pigs on farms, while a higher prevalence was detected among the carcasses at slaughter. The dynamics of Salmonella prevalence in pigs and carcasses were reciprocated in the farm and slaughter environment, clearly indicating an exchange of this pathogen between the pigs and their surroundings. Furthermore, the phenotypic and genotypic fingerprint profile results underscore the potential role played by environmental factors in dissemination of AR Salmonella to pigs.
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http://dx.doi.org/10.1128/AEM.01419-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3753971PMC
September 2013

Prevalence of porcine noroviruses, molecular characterization of emerging porcine sapoviruses from finisher swine in the United States, and unified classification scheme for sapoviruses.

J Clin Microbiol 2013 Jul 15;51(7):2344-53. Epub 2013 May 15.

Food Animal Health Research Program, Ohio Agricultural Research and Development Center, The Ohio State University, Wooster, Ohio, USA.

Noroviruses (NoVs) and sapoviruses (SaVs) are important human pathogens. Although the involvement of porcine NoVs in disease in pigs is unclear, they are genetically and antigenically closely related to human NoVs. Human NoV-like strains have been detected in pigs, raising public health concerns of potential interspecies transmission. Porcine SaVs are highly diverse and emerging in swine populations. Recently, at least three new genogroups of porcine SaVs have been proposed. In this study, we tested 413 pooled fecal samples collected from apparently healthy finisher pigs in North Carolina swine farms during 2009. Reverse transcription (RT)-PCR coupled hybridization assays were performed to detect known porcine NoVs. The overall prevalence of porcine NoVs determined was 18.9% based on this method. Samples were then tested by RT-PCR targeting the 5' end of the capsid region for genogroup II (GII) NoVs, a group which includes human NoVs, followed by sequence analysis. All NoVs identified belonged to typical porcine NoV genotypes, and no human NoV-like strains were detected in specimens from these pigs. Porcine NoV-negative samples (n = 335) were subsequently screened using universal calicivirus primers, and 17 SaV strains were confirmed by sequencing. Based on the partial RNA-dependent RNA polymerase (RdRp) region, they clustered with GIII, GVII, and GVIII and with currently unclassified SaVs. According to analysis of the complete capsid sequences, 7 representative strains clustered with GVII, GVIII, and GIX? SaVs. We tentatively classified SaVs into 14 genogroups based on the complete capsid protein VP1. In summary, porcine NoVs and highly divergent SaVs were present in North Carolina finisher pigs.
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http://dx.doi.org/10.1128/JCM.00865-13DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3697660PMC
July 2013

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

PLoS One 2013 7;8(5):e63704. Epub 2013 May 7.

Center for Emerging Infectious Diseases, Department of Epidemiology, University of Iowa College of Public Health, University of Iowa, Iowa City, Iowa, United States of America.

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.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0063704PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3646818PMC
December 2013

Staphylococcus aureus shedding pattern throughout lactation in dairy cows with naturally occurring intramammary infection.

J Am Vet Med Assoc 2013 May;242(10):1410-8

Department of Veterinary Preventive Medicine, College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210, USA.

Objective: To evaluate shedding patterns of Staphylococcus aureus, specifically the association between clonal relatedness and shedding patterns of S aureus for cows with naturally occurring S aureus intramammary infection.

Design: Longitudinal field study.

Sample: Milk samples from 22 lactating cows (29 mammary glands) of varied numbers of lactations on 2 dairies. Procedures-Foremilk samples were collected weekly for 26 to 44 weeks during lactation from individual mammary glands. Milk samples were cultured bacteriologically with a 0.01-mL inoculum. Samples were considered culture positive for S aureus if ≥ 1 colony-forming units were obtained. Milk samples from known S aureus-positive mammary glands that were culture negative for S aureus or culture positive with a single colony of S aureus were cultured bacteriologically a second time with a 0.1-mL inoculum. Longitudinal shedding patterns of S aureus and the effect of strain type on ln(colony forming unit count) were examined.

Results: With the 0.01-mL inoculum, 914 of 1,070 (85%) samples were culture positive. After reculturing of negative samples with a 0.1-mL inoculum, 1,011 (95%) of the samples were culture positive. There was no significant difference in the detection of S aureus between genotypic clusters when either the 0.01- or 0.1-mL inoculum was used. There was no significant difference in the amount of shedding between mammary glands infected with isolates in genotypic cluster 1 or 2. No consistent shedding patterns were identified among or within cows. There was a significant difference in mammary gland linear score and test day (composite) linear score between mammary glands infected with isolates in genotypic clusters 1 and 2.

Conclusions And Clinical Relevance: S aureus was shed consistently in cows with naturally occurring intramammary infection in cows, and regardless of the pulsotype, variations in the amount of S aureus shedding had no significant effect on the ability to detect S aureus with a 0.1-mL inoculum.
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http://dx.doi.org/10.2460/javma.242.10.1410DOI Listing
May 2013

Prevalence of Yersinia enterocolitica in antimicrobial-free and conventional antimicrobial use swine production.

Foodborne Pathog Dis 2013 Jun 24;10(6):514-9. Epub 2013 Apr 24.

Department of Large Animal Clinical Sciences, College of Veterinary Medicine, Michigan State University, East Lansing, Michigan 48824, USA.

Swine are the primary reservoir for foodborne illness associated with Yersinia enterocolitica. The use of antimicrobials in animal agriculture has been hypothesized as having a potential role in the increase in prevalence of zoonotic pathogens. The objective of this study was to compare the frequency of Y. enterocolitica fecal shedding in swine reared on farms with conventional antimicrobial use policies to farms that were antimicrobial free (ABF). Swine farms were selected from three regions in the United States. In each region, farms were categorized based on antimicrobial use policy. Fecal samples were collected from pigs on-farm within 48 h of harvest. The overall proportion of Y. enterocolitica and ail-harboring Y. enterocolitica-positive pigs was 10.9% and 4.0%, respectively. There were increased odds (odds ratio [OR] 6.8, 95% confidence interval [CI] 3.46-13.28) for a pig to be Y. enterocolitica positive if it was reared on an ABF farm as compared to a conventional farm. There was no significant association between farm antimicrobial use policy and isolation of an ail-harboring Y. enterocolitica from an individual pig (OR 1.8, 95% CI 0.90-3.61). The association of antimicrobial use policy with Y. enterocolitica shedding in feces should be interpreted cautiously, as antimicrobial use cannot be separated from other management factors (e.g., confinement or outdoor housing), which may be associated with risk of Y. enterocolitica in swine.
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http://dx.doi.org/10.1089/fpd.2012.1354DOI Listing
June 2013

Enterotoxin-encoding genes in Staphylococcus spp. from bulk goat milk.

Foodborne Pathog Dis 2013 Feb;10(2):126-30

Federal Institution for Education, Science, and Technology of Alagoas (IFAL), Santana do Ipanema-AL, Brazil.

Although Staphylococcus aureus has been implicated as the main Staphylococcus species causing human food poisoning, recent studies have shown that coagulase-negative Staphylococcus could also harbor enterotoxin-encoding genes. Such organisms are often present in goat milk and are the most important mastitis-causing agents. Therefore, this study aimed to investigate the occurrence of enterotoxin-encoding genes among coagulase-positive (CoPS) and coagulase-negative (CoNS) staphylococci isolated from raw goat milk produced in the semi-arid region of Paraiba, the most important region for goat milk production in Brazil. Enterotoxin-encoding genes were screened in 74 staphylococci isolates (30 CoPS and 44 CoNS) by polymerase chain reaction targeting the genes sea, seb, sec, sed, see, seg, seh, and sei. Enterotoxin-encoding genes were found in nine (12.2%) isolates, and four different genes (sea, sec, seg, and sei) were identified amongst the isolates. The most frequent genes were seg and sei, which were often found simultaneously in 44.5% of the isolates. The gene sec was the most frequent among the classical genes, and sea was found only in one isolate. All CoPS isolates (n=7) harboring enterotoxigenic genes were identified as S. aureus. The two coagulase-negative isolates were S. haemolyticus and S. hominis subsp. hominis and they harbored sei and sec genes, respectively. A higher frequency of enterotoxin-encoding genes was observed amongst CoPS (23.3%) than CoNS (4.5%) isolates (p<0.05), reinforcing the importance of S. aureus as a potential foodborne agent. However, the potential risk posed by CoNS in goat milk should not be ignored because it has a higher occurrence in goat milk and enterotoxin-encoding genes were detected in some isolates.
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http://dx.doi.org/10.1089/fpd.2012.1256DOI Listing
February 2013

Yersinia enterocolitica of porcine origin: carriage of virulence genes and genotypic diversity.

Foodborne Pathog Dis 2013 Jan;10(1):80-6

Division of Animal and Food Microbiology, Office of Research, Center for Veterinary Medicine, U.S. Food and Drug Administration, Laurel, MD, USA.

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.
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http://dx.doi.org/10.1089/fpd.2011.1120DOI Listing
January 2013
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