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Objective: To identify the multilocus sequence typing (MLST) sequence types of Borrelia burgdorferi from Ixodes scapularis in Ontario, Canada., Results: One hundred and eighty-five I. scapularis ticks were submitted from 134 dogs via participating clinics from April 1, 2019, to March 31, 2020. Seventeen MLST sequence types of B. burgdorferi were detected from fifty-eight cultured isolates from 21 ticks. The most common MLST sequence types were 12 and 16. Mixed infections of two MLST sequence types were detected in four ticks. Three sequence types (48, 317, 639) were new detections in Ontario., (© 2023. The Author(s).)

Antimicrobial resistance is a complex One Health issue that exists in both human and veterinary medicine. To mitigate this ever-growing problem, efforts have been made to develop guidelines for appropriate antimicrobial use (AMU) across sectors. In veterinary medicine, there are notable literature gaps for proper AMU in minor species. We conducted a structured narrative review covering the years of July 2006 - July 2021 to find antimicrobial treatments for common bacterial infections in exotic (birds, rodents, reptiles, and others), small flock (chickens, turkeys, and other fowl), and backyard small ruminant (sheep and goats) species. We retrieved a total of 4728 articles, of which 21 articles met the criteria for our review. Studies were grouped according to species, syndrome, and body system affected. Other data extracted included the bacterial pathogen(s), treatment (active ingredient), and geographical origin. Body systems reported included: intra-oral (n = 4), gastrointestinal (n = 1), respiratory (n = 2), reproductive (n = 1), skin (n = 3), aural (n = 1), ocular (n = 4), and other/multisystem (n = 5). By species, our search resulted in: rabbit (n = 5), rat (n = 2), guinea pig (n = 1), chinchilla (n = 1), guinea pig and chinchilla (n = 1), avian species (n = 1), psittacine birds (n = 2), loris and lorikeets (n = 1), turtles (n = 2), lizards (n = 1), goats (n = 2) and sheep (n = 2). The results of our findings identified a distinct gap in consistent antimicrobial treatment information for commonly encountered bacterial conditions within these species. There is a persisting need for clinical trials that focus on antibacterial treatment to strengthen the evidence base for AMU within exotic, small flock, and backyard small ruminant species., (© 2022. The Author(s).)

Objective: Our objective was to apply next-generation sequence-based DNA barcoding to identify the remnant larval bloodmeals in wild-caught host-seeking (unengorged) Ixodes scapularis nymphs (n = 216). To infer host species identification, vertebrate DNA was amplified using universal primers for cytochrome c oxidase subunit I (COI) and sequenced using next-generation sequencing (NGS) for comparison against known barcode references., Results: Bloodmeal identification was unsuccessful in most samples (99% of 216 specimens) demonstrating a very low detection rate of this assay. Sequences that surpassed quality thresholds were obtained for 41.7% of nymphs (n = 90) and of those, confident species identification was obtained for 15.6% of nymphs (n = 14). Wild host identifications were only obtained from 2 specimens, where DNA from the eastern grey squirrel (Sciurus carolinensis) was identified. Human and bovine DNA was identified in remaining nymphs and considered to be contaminants. Further optimization of the technique is required to improve detection of remnant bloodmeals in host-seeking nymphs.

Background: The heightened prevalence of Salmonella Typhimurium remains a public health and food safety concern. Studies have reported antibiotic, flavophospholipol, may have the ability to reduce Salmonella in swine, as well as alter the gut microbiota in favour of beneficial bacteria by inhibiting pathogenic bacteria. Thus, the objective of this study was to investigate the fecal microbiota of weaned pigs receiving in-feed flavophospholipol and challenged with Salmonella Typhimurium., Results: Twenty-one weaned pigs were fed either a diet containing 4 ppm of flavophospholipol (treatment group) or a non-medicated feed (control group) for 36 days post-weaning (Day 1 to Day 36). The pigs were orally challenged with a 2 mL dose of 10 8  CFU/mL of S. Typhimurium at Day 7 and Day 8. Community bacterial DNA extracted from fecal samples collected at Day 6 (before challenge) and Day 36 (28 days after challenge) were used to assess the fecal microbiota using the V4 region of the 16S rRNA gene with Illumina MiSeq next-generation sequencing. Sequencing data were visualized using mothur and analyzed in JMP and R software. The fecal microbiota of pigs in the treatment group had differences in abundance of phyla (Firmicutes, Proteobacteria) and genera ( Lactobacillus, Roseburia , Treponema, unclassified Ruminococcaceae, Blautia , Streptococcus , Megasphaera , Dorea , Sporobacter , Peptococcus , unclassified Firmicutes, Clostridium IV and Campylobacter) when compared to pigs that were controls, 28 days after challenge with Salmonella ( P  < 0.05). Specifically, results demonstrated a significant increase in phylum Proteobacteria ( P =  0.001) and decrease in Firmicutes ( P  = 0.012) and genus Roseburia ( P  = 0.003) in the treated pigs suggestive of possible microbial dysbiosis. An increased abundance of genera Lactobacillus ( P  = 0.012) was also noted in the treated group in comparison to the control., Conclusion: Based on these findings, it is difficult to conclude whether treatment with 4 ppm of flavophospholipol is promoting favorable indigenous bacteria in the pig microbiota as previous literature has suggested., Competing Interests: Competing interestsThe authors declare that they have no competing interests., (© The Author(s) 2020.)

Background: Research in humans and mice suggests that obesity influences the abundance and diversity of gastrointestinal (GI) microbiota, and that an "obese microbiome" influences energy metabolism and fat storage in the host. Microbiota membership and composition have been previously assessed in healthy cats. However, research investigating the effects of obesity and weight loss on the cat's fecal microbiota is limited. Therefore, this study's objective was to evaluate differences in fecal microbial abundance and biodiversity, as well as serum cobalamin and folate concentrations in obese cats, before and after weight loss, and compare to lean cats. Fourteen lean and 17 obese healthy client-owned cats were fed a veterinary therapeutic weight loss food at maintenance energy requirement for 4 weeks. At the end of week 4, lean cats finished the study, whereas obese cats continued with a 10-week weight loss period on the same food, fed at individually-tailored weight loss energy requirements. Body weight and body condition score were recorded every 2 weeks throughout the study. At the end of each period, a fecal sample and food-consumption records were obtained from the owners, and serum cobalamin and folate concentrations were analysed. DNA was extracted from fecal samples, polymerase chain reaction (PCR) was performed, and products were sequenced using next-generation sequencing (Illumina MiSeq)., Results: No significant differences in the relative abundance of taxa and in biodiversity indices were observed between cats in either group (P > 0.05 for all tests). Nevertheless, some significantly enriched taxa, mainly belonging to Firmicutes, were noted in linear discriminant analysis effect size test in obese cats before weight loss compared to lean cats. Serum cobalamin concentrations were significantly higher in lean compared to obese cats both before and after weight loss. Serum folate concentrations were higher in obese cats before weight loss compared to after., Conclusions: The association between feline obesity and the fecal bacterial microbiota was demonstrated in enriched taxa in obese cats compared to lean cats, which may be related to enhanced efficiency of energy-harvesting. However, in obese cats, the fecal microbial abundance and biodiversity were only minimally affected during the early phase of a standardized weight loss plan.

Background: Canine leptospirosis is a reemerging zoonotic disease concern in North America, and a better understanding of its epidemiology is needed. Wide-scale use and subsequent analyses of polymerase chain reaction (PCR) testing may provide additional insight into leptospirosis. This study aimed to describe temporal trends, to descriptively map, and to identify environmental, dog, and human-level factors associated with positive canine leptospirosis PCR test results in the United States., Results: Data obtained from IDEXX Laboratories, Inc. on 40,118 canine leptospirosis PCR tests run in the United States between 2009 and 2016 were evaluated. Climate and socioeconomic (e.g. urban influence, income) data were obtained from public databases. Choropleth maps were created to identify high test-positive proportion areas and a cross-sectional analysis was completed with generalized (univariable, followed by multivariable) mixed logistic regression models accounting for county within state to identify significant predictors for a positive test. Overall test-positive proportion was 5.4% across the United States, with the regional point estimate highest in the southwest (8.1%). In the final multivariable model, the odds of a positive test were greater for male dogs (Odds Ratio [OR] = 1.28) and dogs 0-4 years of age (ORs ranged from 0.35-0.71 for the other age groups). The odds of a positive test were greater for dogs living in areas with wet environmental conditions (OR = 1.24). Season and temperature, as well as the interaction between them, were significant predictors of a positive test. Dogs had a greater probability of testing positive during cool temperatures (< 4 °C) compared to the other temperature categories in the fall season., Conclusions: These findings based on PCR testing allow for an improved understanding of factors influencing a positive canine leptospirosis PCR test and will assist targeted education and prevention efforts.

Background: Ulcerative colitis (UC) is an inflammatory bowel disease (IBD) that is widely identified worldwide. This study aimed to investigate the phenotypic characterization and molecular typing of Clostridium difficile isolates among patients with UC at an inflammatory bowel disease clinic in Iran., Methods: In this cross-sectional study, conducted from April 2015 to December 2015, 85 UC patients were assessed for C.difficile infection (CDI). C. difficile isolates were characterized based on their toxin profile and antimicrobial resistance pattern. Multi-locus sequence typing analysis (MLST) and PCR ribotyping were performed to define the genetic relationships between different lineages of toxigenic strains., Results: The prevalence of C. difficile isolates was 31.8% (27/85) in patients, of those 15 patients (17.6%) had CDI. Three different sequence types (STs) identified based on MLST among the toxigenic isolates, that is ST54 (33.3%), ST2 (53.3%), and ST37 (13.6%). C. difficile strains were divided into four different PCR-ribotypes (012, 014, 017 and IR1). The most common ribotype was 014 accounting for 48.3% (7/15) of all strains. The strains isolated during the first episode and recurrence of CDI usually belonged to PCR ribotype 014 (ST2). A high rate of CDI recurrence (14.1%, 12/85) experienced in UC patients. Colonization of the gastrointestinal tract with non-toxigenic C. difficile strains was frequent among patients with mild disease. All C. difficile isolates were susceptible to metronidazole, and vancomycin, 86 and 67% of isolates were resistant to clindamycin and erythromycin respectively. There was no correlation between the toxin type and antibiotic resistance (p > 0.05)., Conclusion: Overall CDI is rather prevalent in UC patients. All patients with CDI experienced moderate to severe disease and exposed to different antimicrobial and anti-inflammatory agents. Close monitoring and appropriate management including early detection and fast treatment of CDI will improve UC outcomes.

Background: Vector-borne pathogens are emerging concerns in multiple regions of Canada. Determining regional prevalence of canine vector-borne pathogens and documenting change will improve clinician awareness, enable targeted prevention, enhance diagnosis and ideally reduce the risk of disease. Study objectives were to: (i) estimate the prevalence of positive canine vector-borne test results from samples submitted in Canada; (ii) assess change in prevalence over time, from baseline (2008) to 2015; and (iii) estimate the prevalence of pathogen co-infections., Methods: This repeat cross-sectional study evaluated 753,468 test results for D. immitis antigen and B. burgdorferi, Ehrlichia canis/ewingii/muris serology, and 753,208 test results for Anaplasma phagocytophilum/platys serology using the SNAP® 4Dx®Test and SNAP 4Dx® Plus Test., Results: Based on all submitted samples from Canada (2008-2015), the period seroprevalence of B. burgdorferi, Ehrlichia spp., Anaplasma spp. and D. immitis antigen were 2.0%, 0.5%, 0.4% and 0.2%, respectively. Over the 7 years (2008 compared to 2015) we observed a significant increase in seroprevalence for B. burgdorferi (144.4%) and Ehrlichia spp. (150%). Co-infections (positive for two or more pathogens on a single 4 pathogen test kit) were estimated at 5.4% (1162/21,612) of total positive tests., Conclusions: The temporal rise and geographical differences in prevalence detected for these pathogens (notably B. burgdorferi) are consistent with anecdotal information on canine illness related to tick-borne pathogen exposure in multiple regions of Canada, particularly canine Lyme disease.

Background: Clostridium difficile infection (CDI) is known as one of the most important causes of nosocomial infections. The main objective of this study was to evaluate the presence of Clostridium difficile in the stool of hospitalized patients with diarrhea as well as in their environments., Methods: C. difficile isolates were characterized according to the presence of toxin genes and antibiotic resistance. Multilocus Sequence Typing Analysis (MLST) was applied for finding the genetic polymorphism and relationship among strain lineages., Results: A total of 821 samples (574 stools and 247 swabs) were collected between April 2015 and May 2017. The prevalence of C. difficile isolates was 28.6% (164/574) in patients and 19% (47/247) in swabs taken from medical devices, hands of healthcare workers and skin patient sites. Finally, 11.5% (66/574) toxigenic C. difficile strains isolated from stool samples of inpatients and 4.4% (11/247) from hands of healthcare workers and skin patient sites. All the toxigenic isolates were inhibited by a low concentration of vancomycin (MIC < 0.5 μg/ml). About 43% (33/77) and 39% of isolates were resistant to Clindamycin and moxifloxacin respectively. All isolates were susceptible to metronidazole. Toxigenic C. difficile strains were analyzed by MLST and were divided into 4 different STs. The detected types were ST-54 (57.9%), followed by ST-2 (31.6. %), ST-15 (5.3%) and ST-37 (5.3%), while none of the isolates were identified as ST-1 or ST-11. Significant risk factors for CDI appear to be advanced age, undergoing chemotherapy, previous surgery, and residence in the nursing home., Conclusions: CDI is common in Iran and further studies are recommended to monitor its epidemiological variations. Moreover, greater attempts must be made to encourage antibiotic stewardship by healthcare workers and the public., Competing Interests: The study was approved by the human research ethics committee at Isfahan University of Medical Sciences and the study was carried out in accordance with the approved guidelines.Not applicable.The authors declare that they have no competing interests.Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Background: Rabbits are cecotrophic, hindgut-fermenters that rely heavily on their gastrointestinal microbiota for optimal digestion of plant-based diets. Dysbiosis, caused by disruption of the gastrointestinal microbiota, is known to predispose rabbits to rabbit enteritis complex (REC), a major cause of morbidity and mortality. The objectives of this study were to describe the fecal microbiota of domestic rabbits from a variety of settings (commercial meat, companion, laboratory, and shelter) and to identify how factors such as age, season, and routine antimicrobial use affect the fecal microbiota composition., Results: A total of 86 pooled commercial meat, 54 companion, 14 pooled laboratory, and 14 shelter rabbit fecal samples were evaluated using 16S rRNA gene sequencing of the V4 region. In all sample types, the predominant bacterial phylum was Firmicutes. Other commonly identified phyla (composing ≥ 1% of the total microbiota composition) were Verrucomicrobia, Proteobacteria, and Bacteroidetes. Significant differences in composition were noted between commercial, companion, laboratory, and shelter rabbit samples for proportions of Verrucomicrobia (P < 0.01), Proteobacteria (P < 0.01), and Lentisphaerae (P = 0.01) within the total microbiota. Within the commercial meat rabbit samples, significant differences between the microbiota composition of growers (n = 42) and does (n = 44) were limited to one unclassified Firmicutes (P = 0.03) and no differences were identified at the phylum level. Significant differences were present between fecal samples taken from rabbits during the summer (n = 44) compared to the winter (n = 42), with Firmicutes (P = 0.04), Verrucomicrobia (P = 0.03), Proteobacteria (P = 0.02), Deinococcus-Thermus (P = 0.04), Armatimonadates (P = 0.003), and Actinobacteria (P = 0.03) forming significantly different proportions of the microbiota. The only significant difference in composition between those farms that routinely reported antimicrobial use and those that did not was in one unclassified Bacteroidetes (P < 0.05) and no differences were identified at the phylum level., Conclusions: Rabbit husbandry and diet, in addition to season, significantly influence the fecal microbiota composition of domestic rabbits, while age of the rabbit post-weaning has minimal impact.

Background: Feline fecal microbiota analyses can potentially be impacted by a variety of factors such as sample preparation, sequencing method and bioinformatics analyses. Another potential influence is changes in the microbiota from storage of samples prior to processing. This study examined the effect of ambient temperature exposure on the feline fecal microbiota composition. Fecal samples were collected from 12 healthy cats, within 15 min after defecation. Samples were aliquoted and the first aliquot was frozen at -80 °C within 1 hour of defecation. Remaining aliquots were maintained at ambient temperature (20 to 23 °C) and frozen at -80 °C at 6, 12, 24, 36, 48, 72 and 96 h after collection. DNA was extracted from all aliquots, and polymerase chain reaction (PCR). The PCR products were sequenced with next-generation sequencing (Illumina MiSeq)., Results: No significant differences were observed in alpha and beta biodiversity indexes, as well as relative abundance of different taxa over time (P > 0.05 for all tests between time points). Principal coordinate analyses demonstrated that samples cluster mainly by cat, with no significant differences between time points (AMOVA, P > 0.05; HOMOVA, P > 0.05). Linear discriminant analysis effect size method was performed and failed to detect any enriched taxa, between time points. Random forest algorithm analysis indicated homogeneity across time points., Conclusions: Although existing evidence from human fecal storage studies is contradictory, a recent study in companion animals agreed with the current study, demonstrating that maintenance of feline fecal samples at ambient temperature for up to 4 days has no effect on the bacterial membership and structure.

Background: Diarrhea in foals affects up to 60% of foals during the first six months of life. The effect of diarrhea on the fecal bacterial microbiota in foals has not been investigated. Little is known on the fecal bacterial microbial richness and diversity of foals at a young age. The objective was to compare the fecal bacterial microbiota of healthy foals to foals with diarrhea at two and four weeks of life., Methods: Fecal samples were collected from foals (n = 20) at 1-14 (T1) and 15-28 (T2) days of age and analyzed using high throughput sequencing. Differences in relative abundance of bacterial taxa, alpha diversity and beta diversity indices were assessed between age-matched foals with diarrhea (n = 9) and healthy foals (n = 11), and between time points., Results: Differences in microbial community composition based on time point and health status were observed on all taxonomic levels. Of 117 enriched species in healthy foals at T2, 50 (48%) were Lachnospiraceae or Ruminococcaceae. The Chao richness index was increased in healthy foals at T2 compared to T1 (p = 0.02). Foals with diarrhea had a significantly lower richness index than non-diarrheic foals at T2 (p = 0.04). Diarrhea had an inconsistent effect, while time point had a consistent effect on microbial community structure., Conclusions: Preventative and therapeutic measures for diarrhea should focus on maintaining bacterial microbiota richness. Lachnospiraceae and Ruminococcaceae were underrepresented in foals with diarrhea. These should be evaluated further as potential therapeutic options.

Background: Accurate diagnosis of Clostridium difficile infection (CDI) is paramount for patient management. The wrong diagnosis places patients at risk, delays treatment, and/ or contributes to transmission of infection in the healthcare setting. Although amplification of the toxin B gene by polymerase chain reaction (PCR) is a sensitive method for detecting toxigenic C. difficile, false negative results still occur and could impact the diagnosis and treatment of this infection., Methods: This study investigated 48 patients that tested negative for toxigenic C. difficile via GeneXpert C. difficile epi test, while simultaneously testing positive for toxigenic C. difficile via stool culture. Fifty discrepant samples were collected over a 15-month period and all C. difficile isolates were characterized by ribotype. Patient charts were reviewed to assess whether discrepant results impacted the treatment course or clinical outcome of affected patients., Results: Fifty samples of a total of 2308 samples tested in an acute healthcare facility over a 15-month period had negative PCR and positive stool culture for toxigenic C. difficile. C. difficile isolated from the discrepant samples resulted in diverse ribotyping patterns suggesting they were derived from different strains. The samples belonged to patients who were distributed evenly between age groups and wards in the hospital. In the majority of cases, the false negative C. difficile test results did not seem to impact the clinical outcome in these patients., Conclusions: The PCR limit of detection may impact the results of molecular methods for C. difficile detection. Both clinical and analytical sensitivity of C. difficile tests should be considered when deciding which diagnostic assay to use, and clinical correlates should be examined carefully before excluding CDI as a cause of disease.

Background: Clostridium difficile is an important enteropathogen affecting humans, domestic animals, and wildlife. The objectives of this study were to 1) compare the prevalence and characteristics of C. difficile isolated from the feces of raccoons trapped on swine farms and conservation sites, and 2) investigate the role of raccoons as potential reservoirs for host-adapted strains of C. difficile using a longitudinal study. Fecal swabs were collected from raccoons at 5 conservation sites and 5 swine farms, once every five weeks, from May to November, 2012., Results: Clostridium difficile was isolated from 9 % (38/444) of samples, from 12 % (37/302) of raccoons, from all 10 sites. A total of 19 different ribotypes were identified, including 5 ribotypes that matched recognized international designations and which are also found in humans (001, 014, 056, 078, and 103). Location type (farm or conservation area) was not associated with C. difficile status (P = 0.448) but only 3 ribotypes (014, 056, and 078) were found in both location types. The prevalence of ribotype 078 was significantly higher on farms (4 %; 9/220) compared to conservation sites (1 %; 2/225) (P = 0.034). Only one of 108 raccoons caught in multiple sessions was positive on more than one occasion., Conclusions: We found no evidence to support the hypothesis that raccoons harbour host-adapted strains of C. difficile; rather, it appears that raccoons transiently acquire C. difficile from the environment. Raccoons are unlikely to be maintaining C. difficile, but because we detected C. difficile strains that have the potential to cause illness in humans and livestock, and because raccoons can move relatively large distances, they may play a role in the dissemination of pathogenic ribotypes of C. difficile throughout the environment.

Background: The mammalian microbiota plays a key role in host health and disease susceptibility. However, knowledge of the early-age microbiota of pigs is lacking. The purpose of this study was to use high-throughput next-generation sequencing to characterize the fecal and nasal microbiotas of pigs during early life., Results: Ten commercially-raised pigs were randomly enrolled at birth and sampled throughout the first 7 weeks of life. DNA was extracted from fecal and nasal samples and the hypervariable region V4 of the 16S rRNA gene was amplified. The product was sequenced using the Illumina MiSeq platform and 2 × 250 chemistry. Sequencing data was processed and analyzed with the mothur algorithms using an operational taxonomic unit approach. In total, 4.7 million and 5.4 million high-quality sequences were recovered from fecal and nasal samples, respectively. Analysis revealed that these microbiotas contain a very rich and diverse population of bacteria that display a remarkable evolution during the first 7 weeks of life. During this developmental period, a pig was exposed to an average of 1,976 and 6,257 species of bacteria by way of the gastrointestinal and respiratory tracts, respectively. Aging was significantly associated with an increasing measure of richness and diversity as well as with distinct changes to the core microbiota. At 2-3 weeks post-weaning, the rapidly developing microbiotas appeared to reach a developmental milestone as a relative degree of stability was evident., Conclusions: Pigs are exposed to an incredibly rich and diverse mixture of bacteria during early-life as demonstrated by next-generation sequencing methodology. These findings expand the knowledge of the developing porcine microbiota which is important for understanding susceptibility to disease, particularly for vulnerable neonatal pigs.

Background: Infection control in veterinary clinics is important for preventing pathogen spread between patients, staff and the public. There has been no direct evaluation of the use of many basic infection control practices, including sharps handling, environmental cleaning, and personal protective clothing (PPC), in companion animal clinics. The objective of this study was to describe these and other infection control practices associated with routine companion animal appointments in veterinary clinics in Ontario., Results: Video observation of practices was performed in 51 clinics for approximately 3 weeks each as part of another study evaluating the effect of a poster campaign on hand hygiene compliance. Two small wireless surveillance cameras were used: one in an exam room, one in what was considered the most likely location for hand hygiene to be performed outside the exam room following an appointment. Video footage was coded and analyzed for 47 clinics, including 2713 appointments and 4903 individual staff-animal contacts. Recapping of a needle was seen in 84% (1137/1353) of appointments in which use was observed. Only one apparent needlestick injury (NSI) was seen, during recapping. Exam tables were cleaned and floors were mopped following 76% (2015/2646) and 7% (174/2643) of appointments, respectively. Contact time with spray used to clean the exam table ranged from 0-4611 s (mean 39 s, median 9 s). Appropriate PPC was worn for 72% (3518/4903) of staff-animal contacts., Conclusions: Although there was significant room for improvement in sharps handling behaviours in participating clinics, the number of observed NSIs was low. Contact time with environmental disinfectants and use of PPC could also be improved, as well as other basic infection control practices. Education and motivation of veterinary staff to use these simple measures more effectively could potentially have a significant impact on infection control in veterinary clinics for relatively little cost.

Background: The intestinal tract is a rich and complex environment and its microbiota has been shown to have an important role in health and disease in the host. Several factors can cause disruption of the normal intestinal microbiota, including antimicrobial therapy, which is an important cause of diarrhea in horses. This study aimed to characterize changes in the fecal bacterial populations of healthy horses associated with the administration of frequently used antimicrobial drugs., Results: Twenty-four adult mares were assigned to receive procaine penicillin intramuscularly (IM), ceftiofur sodium IM, trimethoprim sulfadiazine (TMS) orally or to a control group. Treatment was given for 5 consecutive days and fecal samples were collected before drug administration (Day 1), at the end of treatment (Days 5), and on Days 14 and 30 of the trial. High throughput sequencing of the V4 region of the 16S rRNA gene was performed using an Illumina MiSeq sequencer. Significant changes of population structure and community membership were observed after the use of all drugs. TMS caused the most marked changes on fecal microbiota even at higher taxonomic levels including a significant decrease of richness and diversity. Those changes were mainly due to a drastic decrease of Verrucomicrobia, specifically the "5 genus incertae sedis". Changes in structure and membership caused by antimicrobial administration were specific for each drug and may be predictable. Twenty-five days after the end of treatment, bacterial profiles were more similar to pre-treatment patterns indicating a recovery from changes caused by antimicrobial administration, but differences were still evident, especially regarding community membership., Conclusions: The use of systemic antimicrobials leads to changes in the intestinal microbiota, with different and specific responses to different antimicrobials. All antimicrobials tested here had some impact on the microbiota, but TMS significantly reduced bacterial species richness and diversity and had the greatest apparent impact on population structure, specifically targeting members of the Verrucomicrobia phylum.

Background: Evaluation of factors that might impact microbiota assessment is important to avoid spurious results, particularly in field and multicenter studies where sample collection may occur distant from the laboratory. This study evaluated the impact of refrigeration on next generation sequence-based assessment of the canine and feline fecal microbiota. Fecal samples were collected from seven dogs and ten cats, and analysed at baseline and after 3, 7, 10 and 14 days of storage at 4°C., Results: There were no differences in community membership or population structure between timepoints for either dogs or cats, nor were there any differences in richness, diversity and evenness. There were few differences in relative abundance of phyla or predominant genera, with the only differences being significant increases in Actinobacteria between Days 0-14 (P = 0.0184) and 1-14 (P = 0.0182) for canine samples, and a decrease in Erysipelotrichaceae incertae sedis between Day 0 and Day 7 (median 4.9 vs 2.2%, P = 0.046) in feline samples., Conclusions: Storage for at least 14 days at 4°C has limited impact on culture-independent assessment of the canine and feline fecal microbiota, although changes in some individual groups may occur.

Background: Wounds can easily become chronically infected, leading to secondary health complications, which occur more frequently in individuals with diabetes, compromised immune systems, and those that have suffered severe burns. When wounds become chronically infected, biofilm producing microbes are often isolated from these sites. The presence of a biofilm at a wound site has significant negative impact on the treatment outcomes, as biofilms are characteristically recalcitrant to removal, in part due to the formation of a protective matrix that shield residents organisms from inimical forces. Pseudomonas aeruginosa and methicillin-resistant Staphylococcus aureus (MRSA) are two of the organisms most prevalently isolated from wound sites, and are of particular concern due to their elevated levels of antibiotic resistance, rapid growth, and exotoxin production. In order to understand the biofilm forming abilities of these microbes in a simulated wound environment we used a microtiter plate assay to assess the ability of these two organisms to bind to proteins that are typically found at wound sites: collagen and hyaluronan., Results: Collagen and hyaluronan were used to coat the wells of 96-well plates in collagen:hyaluronan ratios of 0:1, 3:1, 1:1, 1:3, and 1:0 . P. aeruginosa and MRSA were inoculated as mono- and co-cultures (1:1 and a 3:1 MRSA: P. aeruginosa). We determined that coating the wells with collagen and/or hyaluronan significantly increased the biofilm biomass of attached cells compared to an uncoated control, although no one coating formulation showed a significant increase compared to any other combination. We also noted that the fold-change increase for MRSA upon coating was greater than for P. aeruginosa., Conclusions: Our study suggests that the presence of collagen and/or hyaluronan at wound sites may be an important factor that influences the attachment and subsequent biofilm formation of notorious biofilm-formers, such as MRSA and P. aeruginosa. Understanding the kinetics of binding may aid in our comprehension of recalcitrant wound infection development, better enabling our ability to design therapies that would prevent or mitigate the negative outcomes associated with such infections.

Background: In healthcare facilities, conventional surveillance techniques using rule-based guidelines may result in under- or over-reporting of methicillin-resistant Staphylococcus aureus (MRSA) outbreaks, as these guidelines are generally unvalidated. The objectives of this study were to investigate the utility of the temporal scan statistic for detecting MRSA clusters, validate clusters using molecular techniques and hospital records, and determine significant differences in the rate of MRSA cases using regression models., Methods: Patients admitted to a community hospital between August 2006 and February 2011, and identified with MRSA>48 hours following hospital admission, were included in this study. Between March 2010 and February 2011, MRSA specimens were obtained for spa typing. MRSA clusters were investigated using a retrospective temporal scan statistic. Tests were conducted on a monthly scale and significant clusters were compared to MRSA outbreaks identified by hospital personnel. Associations between the rate of MRSA cases and the variables year, month, and season were investigated using a negative binomial regression model., Results: During the study period, 735 MRSA cases were identified and 167 MRSA isolates were spa typed. Nine different spa types were identified with spa type 2/t002 (88.6%) the most prevalent. The temporal scan statistic identified significant MRSA clusters at the hospital (n=2), service (n=16), and ward (n=10) levels (P ≤ 0.05). Seven clusters were concordant with nine MRSA outbreaks identified by hospital staff. For the remaining clusters, seven events may have been equivalent to true outbreaks and six clusters demonstrated possible transmission events. The regression analysis indicated years 2009-2011, compared to 2006, and months March and April, compared to January, were associated with an increase in the rate of MRSA cases (P ≤ 0.05)., Conclusions: The application of the temporal scan statistic identified several MRSA clusters that were not detected by hospital personnel. The identification of specific years and months with increased MRSA rates may be attributable to several hospital level factors including the presence of other pathogens. Within hospitals, the incorporation of the temporal scan statistic to standard surveillance techniques is a valuable tool for healthcare workers to evaluate surveillance strategies and aid in the identification of MRSA clusters.

Background: Clostridium difficile is an important cause of intestinal infections in some animal species and animals might be a reservoir for community associated human infections. Here we describe a collection of animal associated C. difficile strains from 12 countries based on inclusion criteria of one strain (PCR ribotype) per animal species per laboratory., Results: Altogether 112 isolates were collected and distributed into 38 PCR ribotypes with agarose based approach and 50 PCR ribotypes with sequencer based approach. Four PCR ribotypes were most prevalent in terms of number of isolates as well as in terms of number of different host species: 078 (14.3% of isolates; 4 hosts), 014/020 (11.6%; 8 hosts); 002 (5.4%; 4 hosts) and 012 (5.4%; 5 hosts). Two animal hosts were best represented; cattle with 31 isolates (20 PCR ribotypes; 7 countries) and pigs with 31 isolates (16 PCR ribotypes; 10 countries)., Conclusions: This results show that although PCR ribotype 078 is often reported as the major animal C. difficile type, especially in pigs, the variability of strains in pigs and other animal hosts is substantial. Most common human PCR ribotypes (014/020 and 002) are also among most prevalent animal associated C. difficile strains worldwide. The widespread dissemination of toxigenic C. difficile and the considerable overlap in strain distribution between species furthers concerns about interspecies, including zoonotic, transmission of this critically important pathogen.

Background: The intestinal tract harbours a complex and diverse microbial population that is important for health, yet has been poorly described in many species. This study explored the fecal microbiota of semi-free-ranging Wood bison (Bison bison athabascae)., Results: A total of 2081936 16S rRNA (V4) sequences from 40 bison were evaluated. CatchAll analysis of richness predicted a mean of 10685 species per sample (range 5428-24764, SD 4136). Diversity was high, with an average inverse Simpson's index of 31.78 (SD 15.3, range 8.55-86.7). Twenty-one different phyla were identified; however, only Firmicutes and Proteobacteria, Actinobacteria accounted for >1% of sequences. Two distinct population clusters (Group A, n = 19 and Group B, n = 21) were evident based on both community membership and population structure. Group A had a significantly lower relative abundance of Actinobacteria (6.4 vs 11.8%, P = 0.002), Chloroflexi (0.002 vs 0.013%, P = 0.014), Gemmatimonadetes (0.007 vs 0.15%, P = 0.038) and Proteobacteria (18.7 vs 42.5%, P = <0.0001) and a greater relative abundance of Firmicutes (70.9 vs 39.3%, P < 0.0001) than Group B. Within Group B, Alphaproteobacteria was the most common class of Proteobacteria (28% of all sequences), while Caulobacteraceae (18.5%), Pseudomonadaceae (3.5%), Hyphomicrobiaceae (3.5%), Alcaligenaceae (3.1%) and Xanthomonadaceae (2.6%) were the most abundant families. The twenty (3.1%) most abundant genera accounted for 71% of sequences. No operational taxon units (OTUs) were found in all samples at a relative abundance of 1% or greater. One OTU (Clostridium cluster XI) was present at 1% or more in all Group A samples, with two other Clostridium cluster XI OTUs in 18/19 (95%) samples. No OTUs were found at that abundance in all Group B sample, but an unclassified Lachnospiraceae was present in 20/21 (95%) and Clostridium cluster XI and Brevundimonas were found in 19 (90%) samples., Conclusions: The fecal microbiota of Wood bison is rich and diverse. The presence of two distinct populations not associated with housing, age or gender suggest that enterotypes, distinctly different microbial population compositions that can achieve the same ultimate function, might be present in bison, as has been suggested in humans.

Background: Bacterial biofilms are of tremendous concern for clinicians, as they can compromise the ability of the immune system and antimicrobial therapy to resolve chronic and recurrent infections. Novel antimicrobial therapies or combinations targeted against biofilm establishment and growth subsequently represent a promising new option for the treatment of chronic infectious diseases. In this study, we treated bacterial biofilms produced by methicillin-resistant Staphylococcus pseudintermedius (MRSP) with a combination of fosfomycin and clarithromycin. We selected these agents, because they prevent biofilm formation and induce antimicrobial synergism that may also target other staphylococci., Results: We determined that the combination of fosfomycin and clarithromycin better impairs S. pseudintermedius biofilm formation compared to treatment with either therapy alone (P < 0.05). Morphological examination of these biofilms via scanning electron microscopy demonstrated that fosfomycin alone does impact biofilm formation on orthopaedic implants. However, this activity is enhanced in the presence of clarithromycin. We propose that the bacteriostatic activity of clarithromycin is accentuated when fosfoymcin is present, as it may allow better penetration into the biofilm matrix, allowing fosfomycin access to sessile bacteria near the surface of attachment., Conclusions: Here, we demonstrate that the combination of fosfomycin and clarithromycin may be a useful therapy that could improve the clinical outcomes of treating antimicrobial resistant MRSP biofilms.

Background: In hospitals, Clostridium difficile infection (CDI) surveillance relies on unvalidated guidelines or threshold criteria to identify outbreaks. This can result in false-positive and -negative cluster alarms. The application of statistical methods to identify and understand CDI clusters may be a useful alternative or complement to standard surveillance techniques. The objectives of this study were to investigate the utility of the temporal scan statistic for detecting CDI clusters and determine if there are significant differences in the rate of CDI cases by month, season, and year in a community hospital., Methods: Bacteriology reports of patients identified with a CDI from August 2006 to February 2011 were collected. For patients detected with CDI from March 2010 to February 2011, stool specimens were obtained. Clostridium difficile isolates were characterized by ribotyping and investigated for the presence of toxin genes by PCR. CDI clusters were investigated using a retrospective temporal scan test statistic. Statistically significant clusters were compared to known CDI outbreaks within the hospital. A negative binomial regression model was used to identify associations between year, season, month and the rate of CDI cases., Results: Overall, 86 CDI cases were identified. Eighteen specimens were analyzed and nine ribotypes were classified with ribotype 027 (n = 6) the most prevalent. The temporal scan statistic identified significant CDI clusters at the hospital (n = 5), service (n = 6), and ward (n = 4) levels (P ≤ 0.05). Three clusters were concordant with the one C. difficile outbreak identified by hospital personnel. Two clusters were identified as potential outbreaks. The negative binomial model indicated years 2007-2010 (P ≤ 0.05) had decreased CDI rates compared to 2006 and spring had an increased CDI rate compared to the fall (P = 0.023)., Conclusions: Application of the temporal scan statistic identified several clusters, including potential outbreaks not detected by hospital personnel. The identification of time periods with decreased or increased CDI rates may have been a result of specific hospital events. Understanding the clustering of CDIs can aid in the interpretation of surveillance data and lead to the development of better early detection systems.

Background: Hand hygiene is considered one of the most important infection control measures in human healthcare settings, but there is little information available regarding hand hygiene frequency and technique used in veterinary clinics. The objectives of this study were to describe hand hygiene practices associated with routine appointments in companion animal clinics in Ontario, and the effectiveness of a poster campaign to improve hand hygiene compliance., Results: Observation of hand hygiene practices was performed in 51 clinics for approximately 3 weeks each using 2 small wireless surveillance cameras: one in an exam room, and one in the most likely location for hand hygiene to be performed outside the exam room following an appointment. Data from 38 clinics were included in the final analysis, including 449 individuals, 1139 appointments before and after the poster intervention, and 10894 hand hygiene opportunities. Overall hand hygiene compliance was 14% (1473/10894), while before and after patient contact compliance was 3% (123/4377) and 26% (1145/4377), respectively. Soap and water was used for 87% (1182/1353) of observed hand hygiene attempts with a mean contact time of 4 s (median 2 s, range 1-49 s), while alcohol-based hand rub (ABHR) was used for 7% (98/1353) of attempts with a mean contact time of 8 s (median 7 s, range 1-30 s). The presence of the posters had no significant effect on compliance, although some staff reported that they felt the posters did increase their personal awareness of the need to perform hand hygiene, and the posters had some effect on product contact times., Conclusions: Overall hand hygiene compliance in veterinary clinics in this study was low, and contact time with hand hygiene products was frequently below current recommendations. Use of ABHR was low despite its advantages over hand washing and availability in the majority of clinics. The poster campaign had a limited effect on its own, but could still be used as a component of a multimodal hand hygiene campaign. Improving the infection control culture in veterinary medicine would facilitate future campaigns and studies in this area, as well as overall patient and staff safety.

Background: The nasal microbiota of pigs has been poorly assessed but could play a role in carriage of important microorganisms such as methicillin-resistant Staphylococcus aureus (MRSA). The objectives of this study were to describe the nasal microbiota in slaughter age pigs, to evaluate the impact of farm management on the nasal microbiota and to provide a preliminary assessment of the influence of the microbiota on MRSA carriage., Results: Nasal swabs were collected from five MRSA positive and eight MRSA negative pigs on one farm that used a liquid feeding system and routine tylosin treatment, and seven MRSA negative pigs from an antibiotic-free farm that used conventional feeding. A total of 946310 sequences passed all quality control filters. The number of sequences per sample ranged from 4307 to 165656 (mean 56092, SD 40007). CatchAll analysis of richness predicted a mean of 1749 OTUs (range 213-3736, SD 996). Overall, 6291 OTUs were identified, yet 5125 (81%) were identified less than 10 times and the 12 most abundant OTUs accounted for 80.7% of sequences. Proteobacteria predominated in all but two samples. Liquid-fed/tylosin-exposed pigs had significantly lower relative abundances of Verrucomicrobia (P = 0.004), Fibrobacteres (P = <0.0001) and sequences unclassified at the phylum level (P = 0.028). When comparing only liquid-fed pigs, MRSA carriers had significantly more Bacteroidetes (P = 0.037) than MRSA negative pigs. 124 genera were identified, with Moraxella accounting for 35.4% of sequences. In the Jaccard index tree, five of eight MRSA positive pigs clustered closely together, as did six of the seven conventionally-fed pigs. A significant difference was identified between conventional and liquid-fed pigs using parsimony test with the Jaccard (P < 0.001) but not the Yue&Clayton (P = 0.26) index. There were no significant differences between MRSA positive and negative pigs (P = 0.133 and 0.175). OTUs belonging to Firmicutes were the main indicators of MRSA negative pigs, including Lactobacillus and another Lactobacillaceae and Staphylococcus., Conclusions: Farm management can influence the nasal microbiota in pigs, but no impact of the microbiota on MRSA carriage was identified. Studies that further define the impact of management on the microbiota, and the impact of the microbiota on pathogen carriage are indicated.

Background: Analgesia is often indicated in rabbits undergoing surgical procedures or suffering from various painful conditions and the most common adverse effects associated with NSAIDs occur in the gastrointestinal tract (GIT). The objective of this study was to determine the potential effect of long-term (21 days) meloxicam administration on the fecal bacterial microbiota in healthy rabbits.Samples of hard feces were collected from six rabbits treated with meloxicam (1 mg/kg orally once every 24 h) on days 0,6,14 and 21. Next generation sequencing of V4 16S rRNA gene products was performed., Results: A total of 2589912 V4 rRNA gene sequences passed all quality control filters. Firmicutes predominated (82.0 ± 6.2%). Sixteen other phyla were also identified but other than Verrucomicrobia (4.4 ± 4.9%), all accounted for less than 1% of the identified sequences. Within Firmicutes, Clostridia was the dominant class, accounting for 76% of operational taxon units (OTUs). In general, there were only few differences observed between time points and different rabbits at the phylum level. A significant change was observed in the relative abundance of Proteobacteria over the 4 time points (P = 0.02)., Conclusions: The gastrointestinal tract of rabbits harbors dense and diverse microbiota. Significant alteration of the hard fecal microbiota does not appear to be a considerable adverse effect expected in rabbits treated for 21 days with oral meloxicam at a dose of 1 mg/kg.

Background: Clostridium difficile is an important bacterial pathogen of humans and a variety of animal species. Birds, especially migratory passerine species, can play a role in the spread of many pathogens, including Clostridium difficile. Barn Swallows (Hirundo rustica) nest in close proximity to human habitats and their biology is closely associated with cattle farming. Therefore, we hypothesized that Barn Swallows can be the reservoir of Clostridium difficile., Results: Barn Swallows (n = 175) were captured on their autumn migration across Europe to sub-Saharan Africa. Droppings were collected from juvenile (n = 152) and adult birds (n = 23). Overall prevalence of Clostridium difficile was 4% (7/175); 4.6% (7/152) in juvenile birds and 0/23 in adults. Clostridium difficile ribotypes 078, 002 and 014 were identified, which are commonly found in farm animals and humans. Three new Clostridium difficile ribotypes were also identified: SB3, SB159 and SB166, one of which was toxigenic, harbouring genes for toxins A and B., Conclusions: Results of this study indicate that Barn Swallows might play a role in national and international dissemination of Clostridium difficile and could serve as a source for human and animal infection. Clostridium difficile ribotype 078 was identified, which has been reported as an emerging cause of community-associated Clostridium difficile infection in humans. Based on this and other studies, however, it is more likely that Barn Swallows have a more indicative than perpetuating role in Clostridium difficile epidemiology.

Background: Clostridium difficile has been shown to be a nosocomial pathogen associated with diarrhoea and pseudomembranous colitis in hospitalised patients and the infection is believed to be acquired nosocomially. Recent studies have shown the occurrence of C. difficile in food animals which may act as a source of infection to humans.The aim of this study was to determine the occurrence of C. difficile in retail raw beef, cow, sheep, goat, camel and buffalo meat in Iran., Method: From April to October 2012, a total of 660 raw meat samples from beef, cow, sheep, goat, camel and buffalo were purchased from 49 butcheries in Isfahan and Khuzestan provinces, Iran, and were evaluated for the presence of C. difficile using a method including selective enrichment in C. difficile broth, subsequent alcohol shock-treatment and plating onto C. difficile selective medium. C. difficile isolates were tested for the presence of toxin genes and were typed using PCR ribotyping., Results: In this study, 13 of 660 meat samples (2%) were contaminated with C. difficile. The highest prevalence of C. difficile was found in buffalo meat (9%), followed by goat meat (3.3%), beef meat (1.7%), cow (0.94%) and sheep meat (0.9%). Seven of the 13C. difficile strains (53.9%) were positive for tcdA, tcdB and cdtB toxin genes and were classified as ribotype 078. Four strains (30.8%) were positive tcdA, and tcdB, and one strain (7.7%) was possessed only tcdB. The remaining isolate was non-toxigenic. Susceptibilities of 13C. difficile isolates were determined for 11 antimicrobial drugs using the disk diffusion assay. Resistance to clindamycin, gentamycin, and nalidixic acid was the most common finding., Conclusions: To our knowledge, the present study is the first report of the isolation of C. difficile from raw buffalo meat. This study indicates the potential importance of food, including buffalo meat, as a source of transmission of C. difficile to humans.

Background: A high proportion of staphylococci isolated from pigs affected with exudative epidermitis were found to be resistant to β-lactam antimicrobials. The primary objective of this research was to investigate and characterize β-lactam resistance in Staphylococcus hyicus, Staphylococcus aureus and other staphylococci isolated from these pigs., Results: The antimicrobial resistance patterns of 240 staphylococci isolates were determined by disk diffusion, of which 176 (73.3%) of the isolates were resistant to 3 β-lactams (penicillin G, ampicillin, and ceftiofur). The presence of mecA gene was identified in 63 staphylococci isolates from skin samples by PCR. The mecA gene was identified in 19 S. aureus, 31 S. hyicus, 9 Staphylococcus chromogenes, 2 Staphylococcus pseudintermedius isolates, and in 1 isolate each of Staphylococcus arlettae, and Staphylococcus cohnii subspecies urealyticus. From SCCmec typing results, the majority (45/63, 71.4%) were shown to be SCCmec type V. One isolate was SCCmec III. Fourteen isolates were detected as mec class A, mec class C or ccr type 5. The ccr complex and mec complex was not detected in 3 isolates of methicillin resistant S. hyicus (MRSH) based on multiplex PCR. Of the 30 isolates of MRSA identified from nasal samples of the pigs, 29 isolates were SCCmec type V and 1 isolate was SCCmec type II. Staphyloccoci isolates that were mecA negative but resistant to β-lactam antimicrobials were further examined by screening for mecC, however all were negative. Furthermore, the majority of mecA negative β-lactam resistant staphylococci isolates were susceptible to oxacillin and amoxicillin-clavulanic acid in a double disk diffusion test., Conclusions: Methicillin resistance can be identified in a variety of staphylococcal species isolated from pigs. In this study there was a great deal of similarity in the SCCmec types between staphylococcal species, suggesting that resistance may be passed from one species of staphylococci to another species of staphylococci. While this has been reported for acquisition of methicillin-resistance from coagulase negative staphylococci to S. aureus, these data suggest that transmission to or from the porcine pathogen S. hyicus may also occur. The identification of methicillin resistance in a variety of staphylococcal species in pigs does raise concerns about the spread of serious multi-drug resistance in food producing animals and warrants further study.

Background: Surgical site infections (SSIs) are a recognized risk of any surgical procedure in veterinary medicine. One of the keys to prevention of SSIs is reducing exposure of the surgical site to endogenous and exogenous microbes, beginning in the preoperative period. While guidelines are available for preoperative preparation procedures, there has been no objective investigation of compliance with these recommendations in veterinary practices. The objectives of this pilot study were to describe preoperative patient and surgeon preparation practices in a sample of non-equine companion animal veterinary clinics, and to determine if there were any areas that consistently did not meet current guidelines., Results: Observation of preparation practices was performed in 10 clinics over 9-14 days each using up to 3 small wireless surveillance cameras. Data were coded for 148 surgical patients, and 31 surgeons performing 190 preoperative preparations. When patient hair removal was observed, it was most commonly done using clippers (117/133, 88%), and in only one case was it performed prior to anesthetic induction. Patient contact time with soap ranged from 10-462 s (average of clinic means 75 s, average of clinic medians 67 s), and with alcohol from 3-220 s (average of clinic means 44 s, average of clinic medians 37 s). Alcohol-based hand rub (AHR) was used preoperatively in 2/10 facilities, but soap-and-water hand scrub was most commonly used at all clinics. Proximal-to-distal scrubbing was noted in 95/142 (67%) of soap-and-water scrubs. Contact time during surgeon hand preparation ranged from 7-529 s (average mean 121 s, average median 122 s) for soap-and-water and from 4-123 s (average mean 25 s, average median 19 s) for AHR. No significant changes in practices were identified over time during the observation period. Practices that did not conform to guidelines available in major companion animal surgical textbooks were commonly observed., Conclusions: Some preoperative preparation practices were relatively consistent between clinics in this study, while others were quite variable. Contact times with preoperative preparatory solutions for both patients and surgeons were often shorter than recommended. Evidence-based guidelines for these procedures in veterinary medicine should be established and implemented in order to help reduce preventable SSIs, while maintaining efficiency and cost-effectiveness.

Background: Research has indicated that the environment may play an important role in the transmission of meticillin-resistant Staphylococcus aureus (MRSA) and Clostridium difficile in healthcare facilities. Despite the significance of this finding, few data exist from longitudinal studies investigating MRSA and C. difficile contamination, concurrently, in both patient rooms and the general ward environment. The objectives of this study were to determine the prevalence of MRSA and C. difficile contamination in patient rooms and the ward environment and identify risk factors associated with a surface being contaminated with these pathogens., Methods: Environmental surfaces in patient rooms and the general environment in the medical and surgical wards of a community hospital were sampled six times over a 15 week period. Sterile electrostatic cloths were used for sampling and information pertaining to the surface sampled was recorded. MRSA isolates and C. difficile specimens were obtained from hospitalized patients.Enrichment culture was performed and spa typing or ribotyping was conducted for MRSA or C. difficile, respectively. Exact logistic regression models were constructed to examine risk factors associated with MRSA and C. difficile contamination., Results: Sixteen (41%) patient rooms had ≥ 1 surfaces contaminated with MRSA and/or C. difficile. For 218 surfaces investigated, 3.2% and 6.4% were contaminated with MRSA or C. difficile, respectively. Regression models indicated that surfaces in rooms exposed to a C. difficile patient had significantly increased odds of being contaminated with C. difficile, compared to surfaces in unexposed patient rooms. Additionally, compared to plastic surfaces, cork surfaces had significantly increased odds of being contaminated with C. difficile. For 236 samples collected from the ward environment, MRSA and C. difficile were recovered from 2.5% and 5.9% of samples, respectively. Overall, the majority of MRSA and C. difficile strains were molecularly identified as spa type 2/t002 (84.6%, n = 11) and ribotype 078 (50%, n = 14), respectively., Conclusions: In patient rooms and the ward environment, specific materials and locations were identified as being contaminated with MRSA or C. difficile. These sites should be cleaned and disinfected with increased vigilance to help limit the transmission and dissemination of MRSA and C. difficile within the hospital.

Background: Many human infections are transmitted through contact with animals (zoonoses), including household pets. Despite this concern, there is limited knowledge of the public's pet husbandry and infection control practices. The objective of this study was to characterize zoonotic disease related-husbandry and infection preventive practices in pet-owning households in Ontario, Canada., Methods: A self-administered questionnaire was distributed to individuals at two multi-physician clinics in Waterloo, Ontario, Canada during 2010. One adult from each household was invited to participate in the study., Results: Four hundred one pet-owners completed the questionnaire. Households reported ownership of dogs (68%), cats (48%), fish (13%), exotic mammals (7%), such as hamsters, and reptiles and birds (each 6%). Across all species, individuals at higher risk of infections (i.e. < 5 yrs, ≥ 65 yrs, immunocompromised) were often (46-57%) present in households. Children < 16 yrs of age had close pet contact, as households reported dogs (13%) and cats (30%) usually slept in a child's bed and dogs often licked a child's face (24%). Household husbandry practices that increase zoonotic disease risk were frequently identified; some fed high-risk foods (i.e. raw eggs, raw meat, or raw animal product treats) to their dogs (28%) or cats (3%); 14% of reptile-owning households allowed the pet to roam through the kitchen or washed it in the kitchen sink. Reported hand washing by children was high for all species (> 76% washed hands sometimes or greater after touching the pet, its feces, or housing), although fewer reported children always washed their hands (3-57%; by species). With a few exceptions, practices were not associated with the presence of higher risk members in the household or recall of having previously received zoonotic disease education., Conclusions: The results suggest there is a need for education on zoonotic disease prevention practices for pet-owning households with individuals at higher risk of infection and those with high-risk species (e.g., reptiles). Further research is needed to determine the role of education in altering higher risk pet practices.

Background: Staphylococcus pseudintermedius is an opportunistic pathogen of dogs and has emerged as a leading cause of skin, wound and surgical site infections worldwide. Methicillin resistance is common and clinical infections as a result of methicillin-resistant S. pseudintermedius (MRSP) pose a clinical challenge. In other staphylococci, biofilm formation has been shown to be a virulence factor for infection, however, it has received little attention in S. pseudintermedius. The objectives of this study were to evaluate the biofilm forming ability of clinical isolates of S. pseudintermedius obtained from dogs using phenotypic and genotypic techniques., Results: 96% (136/140) of S. pseudintermedius isolates were classified as strong or moderate biofilm producers, with the majority of isolates being able to produce biofilm. There was no difference in biofilm formation between MRSP and MSSP (p=0.8), amongst isolates from clinical infections compared with isolates obtained from colonized dogs (p=0.08), and between isolates from sequence type (ST) 71 and ST 68 (P=0.09). icaA was detected in 77.9% (109/140) of isolates and icaD was detected in 75.7% (106/140) of isolates. Scanning electron microscopic evaluation of S. pseudintermedius biofilm production revealed aggregates of cocci and irregularly produced extracellular polymeric matrix., Conclusion: The majority of S. pseudintermedius isolates evaluated in this study were able to produce biofilm and this may be an important virulence factor in the rapid emergence of this bacterium in veterinary hospitals worldwide. Further study into the mechanisms of biofilm formation by S. pseudintermedius is warranted.

Background: The composition of the microbiota of the equine intestinal tract is complex. Determining whether the microbial composition of fecal samples is representative of proximal compartments of the digestive tract could greatly simplify future studies. The objectives of this study were to compare the microbial populations of the duodenum, ileum, cecum, colon and rectum (feces) within and between healthy horses, and to determine whether rectal (fecal) samples are representative of proximal segments of the gastrointestinal tract. Intestinal samples were collected from ten euthanized horses. 16S rRNA gene PCR-based TRFLP was used to investigate microbiota richness in various segments of the gastrointestinal tract, and dice similarity indices were calculated to compare the samples., Results: Within horses large variations of microbial populations along the gastrointestinal tract were seen. The microbiota in rectal samples was only partially representative of other intestinal compartments. The highest similarity was obtained when feces were compared to the cecum. Large compartmental variations were also seen when microbial populations were compared between six horses with similar dietary and housing management., Conclusion: Rectal samples were not entirely representative of intestinal compartments in the small or large intestine. This should be taken into account when designing studies using fecal sampling to assess other intestinal compartments. Similarity between horses with similar dietary and husbandry management was also limited, suggesting that parts of the intestinal microbiota were unique to each animal in this study.

Background: Surgical site infections (SSIs) caused by biofilm-forming methicillin-resistant Staphylococcus pseudintermedius (MRSP) have emerged as the most common hospital-acquired infections in companion animals. No methods currently exist for the therapeutic remediation of SSIs caused by MRSP in biofilms. Clarithromycin (CLA) has been shown to prevent biofilm formation by Staphylococcus aureus. This study aims to assess the in vitro activity of CLA in eradicating MRSP biofilm formation on various materials., Results: Quantitative assay results (P = 0.5126) suggest that CLA does not eradicate MRSP biofilm formation on polystyrene after 4 - 24 h growth periods. Scanning electron micrographs confirmed that CLA did not eradicate MRSP biofilm formed on orthopaedic implants., Conclusions: By determining the in vitro characteristics and activities of MRSP isolates alone and against antibiotics, in vitro models of biofilm related infections can be made. In vitro data suggests that CLA does not effectively eradicate S. pseudintermedius biofilms in therapeutic doses.

Background: The hospital environment has been suggested as playing an important role in the transmission of hospital-associated (HA) pathogens. However, studies investigating the contamination of the hospital environment with methicillin-resistant Staphylococcus aureus (MRSA) or Clostridium difficile have generally focused on point prevalence studies of only a single pathogen. Research evaluating the roles of these two pathogens, concurrently, in the general hospital environment has not been conducted. The objectives of this study were to determine the prevalence and identify risk factors associated with MRSA and C. difficile contamination in the general environment of three community hospitals, prospectively., Methods: Sampling of environmental surfaces distributed over the medicine and surgical wards at each hospital was conducted once a week for four consecutive weeks. Sterile electrostatic cloths were used for environmental sampling and information regarding the surface sampled was recorded. For MRSA, air sampling was also conducted. Enrichment culture was performed and spa typing was performed for all MRSA isolates. For C. difficile, isolates were characterized by ribotyping and investigated for the presence of toxin genes by PCR. Using logistic regression, the following risk factors were examined for MRSA or C. difficile contamination: type of surface sampled, surface material, surface location, and the presence/absence of the other HA pathogen under investigation., Results: Overall, 11.8% (n=612) and 2.4% (n=552) of surfaces were positive for MRSA and C. difficile, respectively. Based on molecular typing, five different MRSA strains and eight different C. difficile ribotypes, including ribotypes 027 (15.4%) and 078 (7.7%), were identified in the hospital environment. Results from the logistic regression model indicate that compared to computer keyboards, the following surfaces had increased odds of being contaminated with MRSA: chair backs, hand rails, isolation carts, and sofas., Conclusions: MRSA and C. difficile were identified from a variety of surfaces in the general hospital environment.Several surfaces had an increased risk of being contaminated with MRSA but further studies regarding contact rates, type of surface material, and the populations using these surfaces are warranted.

Background: Many human infections are transmitted through contact with animals (zoonoses), including household pets. Although pet ownership is common in most countries and non-pet owners may have frequent contact with pets, there is limited knowledge of the public's pet contact practices and awareness of zoonotic disease risks from pets. The objective of this study was to characterize the general public's knowledge, attitudes and risks related to pet ownership and animal contact in southern Ontario, Canada., Methods: A self-administered questionnaire was distributed to individuals at two multi-physician clinics in Waterloo, Ontario, Canada during 2010. A single adult from each household was invited to participate in the study., Results: Seventy five percent (641/853) of individuals approached completed the questionnaire. Pet ownership and contact were common; 64% of participants had a pet in their household and 37% of non-pet owning households had a member with at least weekly animal contact outside the home. Pet ownership was high (55%) for households with individuals at higher risk for infections (i.e., < 5 yrs, ≥ 65 yrs, immunocompromised). Most respondents (64%) indicated that they had never received information regarding pet-associated disease risks. When given a list of 11 infectious pathogens, respondents were only able to correctly classify just over half on their potential to be transmitted from pets to people (mean 6.4); independently, pet owners and those who recalled receiving information in the past about this topic were able to make significantly more correct identifications. Pet (36%) and non-pet owning households (10%) reported dog or cat bites or scratches during the preceding year. Households with individuals at higher risk for an infection did not differ from the remaining households regarding their perceived disease risk of pets, zoonotic disease knowledge, recall of being asked by their medical provider if they owned any pets, or recall of having received information regarding pet-associated disease risks and preventive measures., Conclusions: These results suggest that there is a need for accessible zoonotic disease information for both pet and non-owning households, with additional efforts made by veterinary, human and public health personnel. Immediate educational efforts directed toward households with individuals at higher risk to infections are especially needed.

Background: Hospital-associated infections are an increasing cause of morbidity and mortality in veterinary patients. With the emergence of multi-drug resistant bacteria, these infections can be particularly difficult to eradicate. Sources of hospital-associated infections can include the patients own flora, medical staff and inanimate hospital objects. Cellular phones are becoming an invaluable feature of communication within hospitals, and since they are frequently handled by healthcare personnel, there may be a potential for contamination with various pathogens. The objective of this study was to determine the prevalence of contamination of cellular phones (hospital issued and personal) carried by personnel at the Ontario Veterinary College Health Sciences Centre with methicillin-resistant Staphylococcus pseudintermedius (MRSP) and methicillin-resistant Staphylococcus aureus (MRSA)., Results: MRSP was isolated from 1.6% (2/123) and MRSA was isolated from 0.8% (1/123) of cellular phones. Only 21.9% (27/123) of participants in the study indicated that they routinely cleaned their cellular phone., Conclusions: Cellular phones in a veterinary teaching hospital can harbour MRSP and MRSA, two opportunistic pathogens of significant concern. While the contamination rate was low, cellular phones could represent a potential source for infection of patients as well as infection of veterinary personnel and other people that might have contact with them. Regardless of the low incidence of contamination of cellular phones found in this study, a disinfection protocol for hospital-issued and personal cellular phones used in veterinary teaching hospitals should be in place to reduce the potential of cross-contamination.

Background: Clostridium difficile and Clostridium perfringens are commonly associated with colitis in equids, but healthy carriers exist. Scarce information is available on the prevalence of Clostridium spp. in gastrointestinal compartments other than faeces in healthy horses, and it is unknown whether faecal samples are representative of proximal compartments. The objectives were to investigate the prevalence of C. difficile and C. perfringens in different intestinal compartments of healthy adult horses and to determine whether faecal samples are representative of colonization in proximal sites and overall carrier status., Results: Toxigenic C. difficile was isolated from 14/135 (10.3%) samples from 8/15 (53.3%) horses. Between zero and three sites were positive per horse, and multiple sites were positive in four horses. Isolates were recovered from duodenum, jejunum, ileum, right dorsal colon, small colon and rectum. When multiple compartments were positive in a single horse, two different C. difficile ribotypes were always present. Clostridium perfringens Type A (CPE, β2 toxin gene negative) was recovered from the left ventral colon of one horse (0.74%, 1/135 samples). Agreement between faeces and overall C. difficile carrier status was good., Conclusions: Clostridium difficile can be found in different compartments of the gastrointestinal tract of healthy horses, and multiple strains can be present in an individual horse. The prevalence of C. perfringens in healthy adult hoses was low, consistent with previous reports. Faecal samples were representative for presence of C. difficile in proximal compartments in 5/8 horses (63%) but were not representative for the specific strain.

Background: The presence of indistinguishable strains of Clostridium difficile in humans, food animals and food, as well as the apparent emergence of the food-animal-associated ribotype 078/toxinotype V as a cause of community-associated C. difficile infection have created concerns about the potential for foodborne infection. While studies have reported C. difficile in calves, studies of cattle closer to the age of harvest are required. Four commercial feedlots in Alberta (Canada) were enrolled for this study. Fecal samples were collected at the time of arrival and after acclimation (< 62, 62-71 or > 71 days on feed). Selective culture for Clostridium difficile was performed, and isolates were characterized by ribotyping and pulsed-field gel electrophoresis. A logistic regression model was built to investigate the effect of exposure to antimicrobial drugs on the presence of C. difficile., Results: Clostridium difficile was isolated from 18 of 539 animals at the time of feedlot arrival (CI = 2.3-6.1) and from 18 of 335 cattle at mid-feeding period (CI = 2.9-13.1). Overall, there was no significant difference in the prevalence of C. difficile shedding on arrival versus mid-feeding period (P = 0.47). No association between shedding of the bacterium and antimicrobial administration was found (P = 0.33). All the isolates recovered were ribotype 078, a toxinotype V strain with genes encoding toxins A, B and CDT. In addition, all strains were classified as NAP7 by pulsed field gel electrophoresis (PFGE) and had the characteristic 39 base pairs deletion and upstream truncating mutation on the tcdC gene., Conclusions: It is apparent that C. difficile is carried in the intestinal tracts of a small percentage of feedlot cattle arriving and later in the feeding period and that ribotype 078/NAP7 is the dominant strain in these animals. Herd management practices associated with C. difficile shedding were not identified, however further studies of the potential role of antimicrobials on C. difficile acquisition and shedding are required.

Background: Clostridium difficile infection (CDI) is an important cause of morbidity and mortality internationally, yet there are important regional differences in the epidemiology and microbiology of disease. Most reports have come from North America and Europe, with limited information from other regions, including the Middle East. Given the changes in the epidemiology of CDI in developed countries, particularly associated with the dissemination of hypervirulent epidemic clones, an understanding of the epidemiology and microbiology of CDI in diverse regions is warranted. This study involved collection of stool samples from individuals with diarrhea at the Isfahan University of Medical Sciences Teaching Hospital, Isfahan, Iran, between October 2010 and March 2011. Selective enrichment culture for C. difficile was performed and isolates were characterised using ribotyping, PCR for the detection of tcdA, tcdB and cdtB genes, and tcdC sequence analysis., Findings: Clostridium difficile was isolated from 19/89 (21%) stool samples of 17/86 (20%) patients. 13/17 (77%) cases of CDI were hospital-associated. Patients with CDI were significantly older (43 ± 28y) than those with non-CDI diarrhea (24, ± 26y)(P = 0.018). All isolates were toxigenic, and possessed genes encoding for toxins A and B. Six (32%) of 19 isolates also possessed cdtB. Twelve ribotypes were identified. Ribotype 078/toxinotype V was most common, accounting for 4 (21%) of isolates. A single isolate of a different toxinotype V ribotype was identified, as was a toxinotype XXIV isolate. The remaining isolates consisted of 9 different toxinotype 0 ribotypes., Conclusions: CDI is an important cause of diarrhea in patients in this hospital. The diversity of ribotypes was striking, and the number of different types suggests the presence of a broad range of strains in the community, the hospital or both. The predominance of toxinotype V strains, which have been associated with community-associated disease and food animals, was unexpected and possible sources of this type require further investigation.

Background: Clostridium difficile and methicillin-resistant Staphylococcus aureus are critical human pathogens and of increasing concern in food animals. Because of the apparent impact of age on prevalence of these organisms, studies of slaughter age pigs are important when considering the potential for contamination of food. This study evaluated C. difficile and MRSA shedding by slaughter age pigs from farms across Canada., Results: Clostridium difficile was isolated from 30/436 (6.9%) samples from 15/45 (33%) farms. After adjusting for clustering at the herd level, the prevalence was 3.4%. Ribotype 078 (toxinotype V, North American Pulsotype 7) was the most common strain, accounting for 67% of isolates. MRSA was isolated from 21/460 (4.6%) pigs from 5/46 (11%) farms. The prevalence in pigs after adjusting for clustering at the herd level was 0.2%. Seven different spa types were identified, with 3 related spa types (t011, t034, new) accounting for 16 (76%) consistent with ST398 predominating. Both MRSA and C. difficile samples were collected from 45 farms. Both MRSA and C. difficile were detected on 2 (4.4%), with C. difficile only on 13 (29%), MRSA only on 3 (6.7%) and neither on 27 (60%)., Conclusions: The prevalence of C. difficile and MRSA in slaughter age pigs was relatively low, particularly in comparison with studies involving younger pigs. The predominance of C. difficile ribotype 078 and MRSA ST398 was not surprising, but there was diversity in strain types and the majority of isolates of both organisms were strains that can be found in humans. While the prevalence of C. difficile and MRSA in slaughter age pigs was relatively low, there is clearly potential for contamination of meat from healthy pigs carrying this pathogen into slaughterhouses.