A cross-sectional study of environmental, dog, and human-related risk factors for positive canine leptospirosis PCR test results in the United States, 2009 to 2016.

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.

Evaluation of a multispecies probiotic as a supportive treatment for diarrhea in dairy calves: A randomized clinical trial.

The objectives of this randomized clinical trial were to determine whether the utilization of a multispecies probiotic bolus (MSP) in dairy calves with diarrhea led to a rapid resolution of diarrhea and improved average daily gain (ADG). Calves, from a convenience sample of dairy farms with diarrhea challenges, having fecal scores of ≥2 were randomly assigned to receive MSP or a placebo (PLB). The MSP bolus contained Pediococcus acidilactici, Enterococcus faecium, Lactobacillus acidophilus, Lactobacillus casei, Bifidobacterium bifidum, peptide extract, an enzyme blend, killed yeast extract, dried whey, and natural flavors (Revive, Partnar Animal Health, Ilderton, ON, Canada). The enrolled calves were fecal scored daily for 7 consecutive days and resolution of diarrhea was defined as having 2 consecutive days with a fecal score ≤1. Calves were also weighed at enrollment, 7, and 14 d following enrollment and ADG was calculated. A Cox proportional hazards model was built to investigate time to resolution of an abnormal fecal score. Two mixed linear regression models were created to evaluate the effect of treatment group on ADG in the first and second weeks following enrollment. A total of 148 calves were enrolled in the experiment and no differences were observed between the groups with respect to the age or weight at enrollment. The mean time to resolution of abnormal fecal score was 5.1 and 5.9 d in the MSP and PLB groups, respectively. In the Cox proportional hazards model, the calves in the MSP group had faster resolution of diarrhea when compared with the PLB group; however, an interaction between time from enrollment of the first calf and treatment group was present. No differences were found between the 2 groups with respect to ADG. This study demonstrates a multispecies probiotic and yeast bolus administered to calves at the onset of diarrhea reduced the duration of diarrhea; however, the clinical and economic relevance of this reduction requires further exploration.

Zoonotic Transfer of Clostridium difficile Harboring Antimicrobial Resistance between Farm Animals and Humans.

The emergence of Clostridium difficile as a significant human diarrheal pathogen is associated with the production of highly transmissible spores and the acquisition of antimicrobial resistance genes (ARGs) and virulence factors. Unlike the hospital-associated C. difficile RT027 lineage, the community-associated C. difficile RT078 lineage is isolated from both humans and farm animals; however, the geographical population structure and transmission networks remain unknown. Here, we applied whole-genome phylogenetic analysis of 248 C. difficile RT078 strains from 22 countries. Our results demonstrate limited geographical clustering for C. difficile RT078 and extensive coclustering of human and animal strains, thereby revealing a highly linked intercontinental transmission network between humans and animals. Comparative whole-genome analysis reveals indistinguishable accessory genomes between human and animal strains and a variety of antimicrobial resistance genes in the pangenome of C. difficile RT078. Thus, bidirectional spread of C. difficile RT078 between farm animals and humans may represent an unappreciated route disseminating antimicrobial resistance genes between humans and animals. These results highlight the importance of the "One Health" concept to monitor infectious disease emergence and the dissemination of antimicrobial resistance genes.

Equine faecal microbiota transplant: Current knowledge, proposed guidelines and future directions.

While certainly not a novel concept, faecal microbiota transplant (FMT) has recently garnered renewed interest in veterinary medicine due to its remarkable success in treating recurrent Clostridium difficile infection (CDI) in man. There is a dearth of information on indications and efficacy of FMT for the treatment of gastrointestinal disorders in the horse; however, based on evidence in man and other veterinary species, and anecdotal reports in horses, FMT may be a useful treatment for selected cases of acute and chronic diarrhoea and inflammatory bowel disease (IBD) in the horse. In the absence of evidence, expert opinion is offered on case selection and FMT procedure. More research is needed to explore the efficacy, indications and optimal preparation, storage and delivery of FMT to horses.

Comparative Genome Analysis and Global Phylogeny of the Toxin Variant Clostridium difficile PCR Ribotype 017 Reveals the Evolution of Two Independent Sublineages.

The diarrheal pathogen Clostridium difficile consists of at least six distinct evolutionary lineages. The RT017 lineage is anomalous, as strains only express toxin B, compared to strains from other lineages that produce toxins A and B and, occasionally, binary toxin. Historically, RT017 initially was reported in Asia but now has been reported worldwide. We used whole-genome sequencing and phylogenetic analysis to investigate the patterns of global spread and population structure of 277 RT017 isolates from animal and human origins from six continents, isolated between 1990 and 2013. We reveal two distinct evenly split sublineages (SL1 and SL2) of C. difficile RT017 that contain multiple independent clonal expansions. All 24 animal isolates were contained within SL1 along with human isolates, suggesting potential transmission between animals and humans. Genetic analyses revealed an overrepresentation of antibiotic resistance genes. Phylogeographic analyses show a North American origin for RT017, as has been found for the recently emerged epidemic RT027 lineage. Despite having only one toxin, RT017 strains have evolved in parallel from at least two independent sources and can readily transmit between continents.

Comparison of the fecal bacterial microbiota of healthy and diarrheic foals at two and four weeks of life.

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.

Fate of Clostridium difficile during wastewater treatment and incidence in Southern Ontario watersheds.

AIMS: To investigate the prevalence of Clostridium difficile encountered during sewage treatment and in water sources into which treated effluent was directly or indirectly discharged. METHODS AND RESULTS: Samples from wastewater treatment plants (WWTPs) and rivers were collected and then enriched for Cl. difficile. Each of the isolates was subjected to toxinotyping and DNA typing using ribotyping, in addition to pulse-field gel electrophoresis. Cl. difficile was isolated from 92% (108/117) of the raw sludge and 96% (106/110) of the anaerobic digested sludge samples from two Ontario WWTPs. The pathogen was recovered from 73% (43/59) of dewatered biosolids and effluent discharge, in addition to river sediments 39% (25/64). Ribotype 078 (commonly associated with Community Acquired infections) was recovered from raw sewage (19%; 21/108), digested sludge (8%; 8/106), biosolids (35%; 15/43) and river sediments (60%; 15/25). CONCLUSIONS: Clostridium difficile is commonly encountered in raw sewage and survives the wastewater treatment process. The pathogen can then be disseminated into the wider environment via effluent and land application of biosolids. SIGNIFICANCE AND IMPACT OF THE STUDY: The study has illustrated the wide distribution of toxigenic Cl. difficile in WWTPs and river sediments although the clinical significance still requires to be elucidated.

Efficacy of antimicrobial and nutraceutical treatment for canine acute diarrhoea: A systematic review and meta-analysis for European Network for Optimization of Antimicrobial Therapy (ENOVAT) guidelines.

Systemic antimicrobial treatments are commonly prescribed to dogs with acute diarrhoea, while nutraceuticals (prebiotics, probiotics, and synbiotics) are frequently administered as an alternative treatment. The aim of this systematic review and meta-analysis was to assess the effectiveness of antimicrobials and nutraceutical preparations for treatment of canine acute diarrhoea (CAD). The results of this study will be used to create evidence-based treatment guidelines. PICOs (population, intervention, comparator, and outcome) were generated by a multidisciplinary expert panel taking into account opinions from stakeholders (general practitioners and dog owners). The Grading of Recommendations Assessment, Development and Evaluation (GRADE) methodology was used to evaluate the certainty of the evidence. The systematic search yielded six randomised controlled trials (RCT) for antimicrobial treatment and six RCTs for nutraceutical treatment meeting the eligibility criteria. Categories of disease severity (mild, moderate, and severe) were created based on the presence of systemic signs and response to fluid therapy. Outcomes included duration of diarrhoea, duration of hospitalization, progression of disease, mortality, and adverse effects. High certainty evidence showed that antimicrobial treatment did not have a clinically relevant effect on any outcome in dogs with mild or moderate disease. Certainty of evidence was low for dogs with severe disease. Nutraceutical products did not show a clinically significant effect in shortening the duration of diarrhoea (based on very low to moderate certainty evidence). No adverse effects were reported in any of the studies.

European Network for Optimization of Veterinary Antimicrobial Therapy (ENOVAT) guidelines for antimicrobial use in canine acute diarrhoea.

Acute diarrhoea is a common presentation in dogs, and a common reason for antimicrobial prescription and nutraceutical use. This evidence-based guideline provides recommendations for antimicrobial and probiotic treatment of canine acute diarrhoea (CAD). A multidisciplinary panel developed the recommendations by adhering to the Grading of Recommendations Assessment, Development and Evaluation (GRADE) framework. The opinions of stakeholders (general veterinary practitioners and dog owners) were collected and incorporated to ensure the applicability of this guideline. Four strong recommendations informed by high certainty evidence, and three conditional recommendations informed by very low or low certainty evidence, were drafted by the panel, along with an ungraded section on diagnostic work-up of dogs with acute diarrhoea. The ENOVAT guidelines initiative encourages national or regional guideline makers to use the evidence presented in this document, and the supporting systematic review, to draft national or local guidance documents.

Antimicrobial prophylaxis in companion animal surgery: A scoping review for European Network for Optimization of Antimicrobial Therapy (ENOVAT) guidelines.

Surgical antimicrobial prophylaxis (SAP) is widely used to reduce the risk of surgical site infections (SSI), but there is uncertainty as to what the proportion of SSI reduction is. Therefore, it is difficult for surgeons to properly weigh the costs, risks and benefits for individual patients when deciding on the use of SAP, making it challenging to promote antimicrobial stewardship in primary practice settings. The objective of this study was to map the veterinary evidence focused on assessing the effect of SAP on SSI development and in order to identify surgical procedures with some research evidence and possible knowledge gaps. In October 2021 and December 2022, Scopus, CAB Abstracts, Web of Science Core Collection, Embase and MEDLINE were systematically searched. Double blinded screening of records was performed to identify studies in companion animals that reported on the use of SAP and SSI rates. Comparative data were available from 34 out of 39123 records screened including: eight randomised controlled trials (RCT), 23 cohort studies (seven prospective and 16 retrospective) and three retrospective case series representing 12476 dogs and cats in total. Extracted data described peri- or post-operative SAP in nine, and 25 studies, respectively. In the eight RCTs evaluating SAP in companion animals, surgical procedure coverage was skewed towards orthopaedic stifle surgeries in referral settings and there was large variation in SAP protocols, SSI definitions and follow-up periods. More standardized data collection and agreement of SSI definitions is needed to build stronger evidence for optimized patient care.

Video observation of hand hygiene practices at a petting zoo and the impact of hand hygiene interventions.

Petting zoos are popular attractions, but can also be associated with zoonotic disease outbreaks. Hand hygiene is critical to reducing disease risks; however, compliance can be poor. Video observation of petting zoo visitors was used to assess animal and environmental contact and hand hygiene compliance. Compliance was also compared over five hand hygiene intervention periods. Descriptive statistics and multivariable logistic regression were used for analysis. Overall hand hygiene compliance was 58% (340/583). Two interventions had a significant positive association with hand hygiene compliance [improved signage with offering hand sanitizer, odds ratio (OR) 3·38, P<0·001; verbal hand hygiene reminders, OR 1·73, P=0·037]. There is clearly a need to improve hand hygiene compliance at this and other animal exhibits. This preliminary study was the first to demonstrate a positive impact of a hand hygiene intervention at a petting zoo. The findings suggest that active, rather than passive, interventions are more effective for increasing compliance.

Feline panleukopenia virus DNA shedding following modified live virus vaccination in a shelter setting.

This study assessed the frequency and timing of feline panleukopenia virus (FPV) shedding in feces following administration of a modified live FPV vaccine. Feces were collected from 37 shelter cats that did not meet clinical criteria for panleukopenia on the day of vaccination or on days 3, 7, 14, and 21 post-vaccination (NCL group). A commercial quantitative PCR (qPCR) fecal pathogen panel and a canine parvovirus point-of-care antigen test were performed. FPV DNA copy numbers from a concurrent study of 39 cats with panleukopenia (CL group) were compared with the NCL group. Of the 165 samples from the NCL group, one had a weak positive antigen test result on day 7, while nine samples (5.5%) from eight cats (21.6%) produced positive FPV qPCR test results, one on day 3 and eight on day 7. There were no day 21-positive qPCR results in the 11 cats that were revaccinated on day 14. There was no association between the number of additional fecal pathogens identified and a positive FPV qPCR result. Of the cats with positive results, FPV DNA copy numbers differed between NCL group and CL group (median 1.13 × 107 and 5.01 × 108 copies/g feces, respectively; P < 0.001). The FPV qPCR cannot differentiate subclinical infection from vaccine virus shedding. To avoid unnecessary isolation and euthanasia, shelters should therefore limit FPV PCR testing to cats with a high index of suspicion of panleukopenia. The timing of recent vaccination should also be considered when interpreting test results.

Factors related to Campylobacter spp. carriage in client-owned dogs visiting veterinary clinics in a region of Ontario, Canada.

From July 2008 until May 2009, 240 client-owned pet dogs from seven veterinary clinics in the Region of Waterloo, Ontario, Canada participated in a study to determine pet-related management factors that may be associated with the presence of Campylobacter spp. in dogs. The prevalence of Campylobacter spp. carriage in our study population of pet dogs was 22%, with 19% of the dogs positive for C. upsaliensis, and 3% positive for C. jejuni. A significant risk factor from multivariable logistic regression models for both Campylobacter spp. and C. upsaliensis carriage was having homemade cooked food as the dog's diet or added to its diet, and a significant sparing factor for both models was treatment with antibiotics in the previous month. Increasing age of the dog decreased the odds of Campylobacter spp. and C. upsaliensis carriage. Based on the high prevalence of Campylobacter, and specifically C. upsaliensis, further research concerning pet dogs as a risk factor for campylobacteriosis in humans is warranted.

Knowledge, attitudes and influencers of North American dog-owners surrounding antimicrobials and antimicrobial stewardship.

OBJECTIVES: To quantify the individual influences of antimicrobial cost, method of administration and drug importance in human medicine on dog-owner antimicrobial preference, and determine knowledge, attitudes and influencers of dog-owners surrounding antimicrobials and antimicrobial stewardship. MATERIALS AND METHODS: Data were collected through an online survey targeting three dog-owner participant groups. These consisted of individuals residing in: (1) Canada, (2) USA and (3) any country recruited through an educational social media site. USA and Canadian participants were financially compensated. Conjoint analysis was used to quantify the influence of antimicrobial cost, method of administration and drug importance in human medicine. Descriptive and analytical statistics were used for data evaluation. RESULTS: A total of 809 surveys were completed. Antimicrobial cost accounted for 47% of dog-owner preferences, followed by method of administration (31%) and drug importance in human medicine (22%). All groups preferred lower cost drugs that were administered once by injection. Participants were more likely to prefer drugs considered "very important" in human medicine, except for the social media participants, who preferred drugs that were "not at all important." Most respondents (86%) reported antimicrobial resistance as important in human medicine and 29% believed antimicrobial use in pets posed a risk for antimicrobial resistance in humans. Participants recruited through social media, and those in the highest education category, were significantly more likely to report antimicrobial use in pets as a risk to people. CLINICAL SIGNIFICANCE: Cost was the most important factor in dog-owner antimicrobial preferences. There is a need for dog-owner antimicrobial stewardship education.

A multidisciplinary approach to reduce and refine antimicrobial drugs use for diarrhoea in dairy calves.

This study evaluated the impact of a multidisciplinary intervention to reduce and refine antimicrobial drug use for treatment of diarrhoea in dairy calves. The intervention consisted of modification of management practices for disease prevention, health evaluation training of farm staff, implementation of an algorithm directed at antimicrobial therapy for diarrhoeic calves, and monthly farm visits. A combination of retrospective (before intervention period [BP], 1 year) and prospective (immediately after intervention period [AP], 1 year) cohorts were used. Health outcomes measured included incidence of diarrhoea and overall mortality. Antimicrobial treatment rates for diarrhoea and total mass of antimicrobial drugs used at the calf and farm level were also evaluated. Outcomes were assessed using the χ2 or Fisher's exact test, and odds ratios and 95% confidence intervals (95% CI) were calculated. Records of 2049 and 2251 calves from 10 farms were available in the BP and AP, respectively. Reduced antimicrobial treatment rates occurred on seven farms (P < 0.01), but not on three farms in the AP. A total of 85% (1303/1573) and 18% (310/1698) of diarrhoeic calves were treated with antimicrobial drugs in the BP and AP (P < 0.001), respectively. There were no differences in the incidence of diarrhoea or mortality between periods. In the AP, the use sulfamethazine and trimethoprim decreased on eight farms, one farm discontinued use of lincomycin and spectinomycin, while two farms discontinued use and one reduced use of cephalosporins. This multidisciplinary approach was effective in reducing antimicrobial drug use for calf diarrhoea on dairy farms without negative impacts on calf health.

Effects of parity, blood progesterone, and non-steroidal anti-inflammatory treatment on the dynamics of the uterine microbiota of healthy postpartum dairy cows.

This study evaluated the effects of treatment with meloxicam (a non-steroidal anti-inflammatory drug), parity, and blood progesterone concentration on the dynamics of the uterine microbiota of 16 clinically healthy postpartum dairy cows. Seven primiparous and 9 multiparous postpartum Holstein cows either received meloxicam (0.5 mg/kg SC, n = 7 cows) once daily for 4 days (10 to 13 days in milk (DIM)) or were untreated (n = 9 cows). Endometrial cytology samples were collected by cytobrush at 10, 21, and 35 DIM, from which the microbiota analysis was conducted using 16S rRNA gene sequence analysis. A radioimmunoassay was used to measure progesterone concentration in blood serum samples at 35 DIM and cows were classified as ˃ 1 ng/mL (n = 10) or ≤ 1 ng/mL (n = 6). Alpha diversity for bacterial genera (Chao1, Shannon-Weiner, and Camargo's evenness indices) were not affected by DIM, meloxicam treatment, parity, or progesterone category. For beta diversity (genera level), principal coordinate analysis (Bray-Curtis) showed differences in microbiota between parity groups. At the phylum level, the relative abundance of Actinobacteria was greater in primiparous than multiparous cows. At the genus level, there was lesser relative abundance of Bifidobacterium, Lactobacillus, Neisseriaceae, Paracoccus, Staphylococcus, and Streptococcus and greater relative abundance of Bacillus and Fusobacterium in primiparous than multiparous cows. Bray-Curtis dissimilarity did not differ by DIM at sampling, meloxicam treatment, or progesterone category at 35 DIM. In conclusion, uterine bacterial composition was not different at 10, 21, or 35 DIM, and meloxicam treatment or progesterone category did not affect the uterine microbiota in clinically healthy postpartum dairy cows. Primiparous cows presented a different composition of uterine bacteria than multiparous cows. The differences in microbiota associated with parity might be attributable to changes that occur consequent to the first calving, but this hypothesis should be investigated further.

The application of next-generation sequence-based DNA barcoding for bloodmeal detection in host-seeking wild-caught Ixodes scapularis nymphs.

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.

Evaluation of Clostridium difficile in dogs and the household environment.

Clostridium difficile may be an emerging community-associated pathogen but little is known about its sources of exposure. This study evaluated C. difficile contamination in households and colonization of pets. C. difficile was isolated from 44/836 (5.3%) sites in 26/84 (31%) households. Ribotype 027 was the most common (25%) environmental strain. C. difficile was isolated from 14/139 (10%) dogs. Living with an immunocompromised individual was associated with C. difficile colonization in dogs. All toxigenic strains identified in pets have been isolated from humans in Ontario. C. difficile was isolated concurrently from dogs and the environment in four households, but in all cases canine and environmental ribotypes were different. C. difficile was relatively common in households, suggesting that exposure to this pathogen may be a regular event. There was no evidence that dogs are a significant source of household C. difficile contamination.