Comparison of 2 PCR assays on environmental samples cultured for Mycobacterium avium subsp. paratuberculosis.

Mycobacterium avium subsp. paratuberculosis (MAP) is the causal agent of paratuberculosis, a chronic, contagious, and incurable enteric disease of ruminants. An in-house IS900 PCR assay validated for MAP detection in sheep has been shown to have a higher sensitivity than a commercial PCR and fecal culture. We have now compared the performance of this in-house IS900 PCR assay with a commercial ISMap02 PCR assay for the detection of MAP DNA in bovine dairy farm environmental samples. We purposefully selected 30 culture-positive, 62 culture-negative, and 62 non-interpretable environmental samples. We applied the IS900 PCR assay directly to the frozen inoculum of these samples. Inocula were incubated in an automated system, and growth was confirmed by an acid-fast bacilli stain and the IS900 PCR assay. Among culture-positive samples before incubation, the IS900 PCR assay yielded significantly more positive results than the ISMap02 PCR assay; however, among culture-negative samples, the IS900 PCR assay yielded positive results both before and after incubation. The ISMap02 PCR assay did not flag positively among the culture-negative samples either before or after incubation. The IS900 PCR assay is a sensitive method that can be used to detect MAP DNA in environmental samples before incubation. The ISMap02 PCR assay is a specific method used to detect MAP DNA in environmental samples both before and after incubation.

Randomised study of the immunomodulatory effects of azithromycin in severely asthmatic horses.

Neutrophilic inflammation is believed to contribute to the airway obstruction and remodelling in equine asthma. Azithromycin, an antibiotic with immunomodulatory properties, reduces pulmonary neutrophilia and hyper-responsiveness in human asthmatics and decreases airway remodelling in rodent models of asthma. It was therefore hypothesised that azithromycin would improve lung function, mucus accumulation and central airway remodelling by decreasing luminal neutrophilia in severe equine asthma. The effects of a 10-day treatment with either azithromycin or ceftiofur, an antimicrobial without immune-modulating activity, were assessed using a blind, randomised, crossover design with six severe asthmatic horses in clinical exacerbation. Lung function, tracheal mucus accumulation, tracheal wash bacteriology, bronchial remodelling, airway neutrophilia and mRNA expression of proinflammatory cytokines (interleukin (IL)-8, IL-17A, IL-1ß, tumour necrosis factor-α) in bronchoalveolar lavage fluid were evaluated. Azithromycin decreased the expression of IL-8 (P=0.03, one-tailed) and IL-1ß (P=0.047, one-tailed) but failed to improve the other variables evaluated. Ceftiofur had no effect on any parameter. The reduction of neutrophilic chemoattractants (IL-8, IL-1ß) justifies further efforts to investigate the effects of a prolonged treatment with macrolides on airway neutrophilia and remodelling. The lack of efficacy of ceftiofur suggests that severe equine asthma should not be treated with antibiotics at first-line therapy.

Molecular characterization of Mycobacterium avium subspecies paratuberculosis C-type and S-type isolated from sheep and goats by using a combination of MIRU-VNTR loci.

Mycobacterium avium subspecies paratuberculosis (Map) is the etiological agent of paratuberculosis of domestic and wild ruminants. Map strains are segregated into 2 main groups or strain types referred to as sheep (S) type and cattle (C) type. Few small ruminant Map strains have been genetically characterized to date. The present study was undertaken to genetically characterize a panel of 30 small ruminant Map strains in the province of Quebec, Canada. Mycobacterial Interspersed Repetitive Units - Variable-Number Tandem Repeat analysis (MIRU-VNTR) were used as genetic markers in addition to IS1311 PCR-REA. S-type and C-type strains were found in both sheep and goats, although C-type strains were more frequently isolated from goats and S-type strains were more common in sheep. A total of 12 distinct Map genotypes were uncovered in the present collection of strains using these markers. Considering the genetic diversity reported here, molecular characterization of Map stains in small ruminants using MIRU-VNTR markers represent an interesting avenue for both epidemiological investigations regarding the sources of herd infection and association studies between Map strains and their virulence, persistence and host-specific adaptation characteristics.

Mycobacterium avium subspecies paratuberculosis (Map) est l'agent étiologique de la paratuberculose affectant les ruminants sauvages et domestiques. Les souches de Map se répartissent dans deux grands groupes ou types appelés 'sheep (S)' et 'cattle (C)'. Très peu de souches de Map provenant des petits ruminants ont été caractérisées génétiquement jusqu'à présent. Cette étude a été initiée afin de caractériser un ensemble de 30 souches de Map provenant de 5 troupeaux de moutons et 8 troupeaux de chèvres situés dans la province de Quebec, Canada, et d'évaluer leur diversité génétique. Une analyse répétée en tandem des unités répétitives alternées des mycobactéries (MIRU-VNTR) a été utilisée comme marqueurs génétiques en plus du marqueur IS1311 PCR-REA. Les souches de type S et C ont été retrouvées chez les isolats ovins et caprins, avec une prédominance des souches de type C chez les isolats provenant de chèvres tandis que les souches de type S étaient plus fréquentes chez les moutons. Un total de 12 génotypes distincts de Map ont été retrouvés parmi les isolats d'après les marqueurs utilisés. Considérant la diversité génétique observée, la caractérisation moléculaire des isolats de Map représente une avenue intéressante pour investiguer les sources potentielles d'infection des troupeaux et pour étudier les associations entre les caractéristiques génétiques et pathogéniques des isolats.(Traduit par les auteurs).

Estimating diagnostic accuracy of fecal culture in liquid media for the detection of Mycobacterium avium subsp. paratuberculosis infections in Québec dairy cows: A latent class model.

A latent class model fit within a Bayesian framework was used to estimate the sensitivity and specificity of individual fecal culture (IFC) in liquid medium (Para TB culture liquid medium and BACTEC MGIT 960 system) for the detection of Mycobacterium avium subsp. paratuberculosis (MAP) infections in Québec dairy cows. As a secondary objective, the within-herd paratuberculosis prevalence was estimated. A dataset including 21 commercial Québec dairy herds participating in previous research projects was retrospectively analyzed. In total, 1386 adult cows on which both IFC and serum-ELISA were available were included. The selected latent class model assumed conditional dependence between the tests. Non-informative priors for IFC accuracy and paratuberculosis prevalence were used while informative priors, obtained from the literature, were used for serum-ELISA accuracy. The WinBUGS statistical freeware was used to obtain posterior estimates (medians and 95% Bayesian credibility intervals (95% BCI)) for each parameter. The sensitivity and specificity estimates for IFC were 34.4% (95% BCI: 20.3-66.1) and 99.5% (95% BCI: 98.6-100), respectively. Sensitivity and specificity for serum-ELISA were 27.3% (95% BCI: 18.1-38.3) and 97.4% (95% BCI: 96.6-98.0). Median paratuberculosis within herd prevalence was estimated to be 0.3% (0-3.3). In conclusion, a higher sensitivity of IFC compared to serum-ELISA was observed both in the unconditional and conditional dependent models. Since the sensitivity of both IFC and serum-ELISA was relatively low, conditional dependence between the tests is more likely in the true disease positive animals. We hypothesize that conditional dependence arises because an unmeasured covariate influences the performance of both tests among disease positive animals causing both tests to incorrectly misclassify the animal as negative. One limitation of this study was the very low within herd prevalence of the participant herds.

Risk factors associated with Mycobacterium avium subsp. paratuberculosis herd status in Québec dairy herds.

Paratuberculosis is a chronic and contagious enteric disease of ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP). Control of paratuberculosis is justified given the associated economic losses and the potential role of MAP in Crohn's disease in humans. Management practices that limit exposure of susceptible animals to MAP are more effective at reducing disease prevalence than testing and culling infected cows. The objective of this retrospective case-control study was to study the association between management practices and MAP status in dairy herds in Québec, Canada. A total of 26 case herds (MAP had been isolated from at least 1 environmental sample in each herd) and 91 control herds (no clinical cases of paratuberculosis and negative on 2 consecutive yearly environmental samplings) were selected among herds enrolled in the Québec Voluntary Paratuberculosis Control Program. A risk assessment questionnaire, completed at enrolment, was available for the selected herds. Culture of MAP was achieved using liquid media and the BACTEC 960 detection system. Multivariable logistic regression was used to evaluate the association between selected risk factors and MAP herd status. Herd size (OR = 1.17; 95% CI: 1.02-1.33) and proportion of cows purchased per year in the last 5 years (OR = 5.44; 95% CI: 1.23-23.98) were significantly associated with a positive MAP herd status. The management risk factors identified in the present study are in accord with previous studies. Management practices aiming to prevent the introduction of new animals into the herd and to reduce the contact of newborn calves with adult animals or their feces are key elements to minimize MAP introduction and transmission into a herd. These elements should be prioritized in control programs.

Evaluation of a PCR assay on overgrown individual fecal samples cultured for Mycobacterium avium subsp. paratuberculosis.

Microbial overgrowth can interfere with Mycobacterium avium subsp. paratuberculosis (MAP) growth and detection. We estimated the percentage of positive samples by PCR performed on the incubated media of individual fecal samples classified as non-interpretable (NI) by bacteriologic culture of liquid media. A total of 262 liquid cultures declared NI and 88 samples declared negative were included in the study. MAP DNA was detected in 7 NI samples (2.7%; 95% CI: 1.1-5.4%) and in 1 negative sample (1.1%; 95% CI: 0.3-6.2%). The PCR allowed the detection of MAP-positive samples that had been missed in the initial bacteriologic culture. However, the benefit of these few additional positive results must be weighed against the additional costs incurred. Using PCR to classify overgrown cultures optimizes the detection process and eliminates the NI outcome.

Evaluation of a PCR assay on overgrown environmental samples cultured for Mycobacterium avium subsp. paratuberculosis.

Culture of Mycobacterium avium subsp. paratuberculosis (MAP) is the definitive antemortem test method for paratuberculosis. Microbial overgrowth is a challenge for MAP culture, as it complicates, delays, and increases the cost of the process. Additionally, herd status determination is impeded when noninterpretable (NI) results are obtained. The performance of PCR is comparable to fecal culture, thus it may be a complementary detection tool to classify NI samples. Our study aimed to determine if MAP DNA can be identified by PCR performed on NI environmental samples and to evaluate the performance of PCR before and after the culture of these samples in liquid media. A total of 154 environmental samples (62 NI, 62 negative, and 30 positive) were analyzed by PCR before being incubated in an automated system. Growth was confirmed by acid-fast bacilli stain and then the same PCR method was again applied on incubated samples, regardless of culture and stain results. Change in MAP DNA after incubation was assessed by converting the PCR quantification cycle (Cq) values into fold change using the 2-ΔCq method (ΔCq = Cq after culture - Cq before culture). A total of 1.6% (standard error [SE] = 1.6) of the NI environmental samples had detectable MAP DNA. The PCR had a significantly better performance when applied after culture than before culture (p = 0.004). After culture, a 66-fold change (SE = 17.1) in MAP DNA was observed on average. Performing a PCR on NI samples improves MAP culturing. The PCR method used in our study is a reliable and consistent method to classify NI environmental samples.

Detection of Mycobacterium avium subspecies paratuberculosis in tie-stall dairy herds using a standardized environmental sampling technique and targeted pooled samples.

Mycobacterium avium ssp. paratuberculosis (MAP) is the etiologic agent of Johne's disease, a chronic contagious enteritis of ruminants that causes major economic losses. Several studies, most involving large free-stall herds, have found environmental sampling to be a suitable method for detecting MAP-infected herds. In eastern Canada, where small tie-stall herds are predominant, certain conditions and management practices may influence the survival and transmission of MAP and recovery (isolation). Our objective was to estimate the performance of a standardized environmental and targeted pooled sampling technique for the detection of MAP-infected tie-stall dairy herds. Twenty-four farms (19 MAP-infected and 5 non-infected) were enrolled, but only 20 were visited twice in the same year, to collect 7 environmental samples and 2 pooled samples (sick cows and cows with poor body condition). Concurrent individual sampling of all adult cows in the herds was also carried out. Isolation of MAP was achieved using the MGIT Para TB culture media and the BACTEC 960 detection system. Overall, MAP was isolated in 7% of the environmental cultures. The sensitivity of the environmental culture was 44% [95% confidence interval (CI): 20% to 70%] when combining results from 2 different herd visits and 32% (95% CI: 13% to 57%) when results from only 1 random herd visit were used. The best sampling strategy was to combine samples from the manure pit, gutter, sick cows, and cows with poor body condition. The standardized environmental sampling technique and the targeted pooled samples presented in this study is an alternative sampling strategy to costly individual cultures for detecting MAP-infected tie-stall dairies. Repeated samplings may improve the detection of MAP-infected herds.

Mycobacterium avium ssp. paratuberculosis (MAP) est l'agent étiologique de la maladie de Johne, une entérite chronique contagieuse des ruminants et responsable d'importantes pertes économiques. Plusieurs études, la plupart réalisées dans des grands troupeaux en stabulation libre, ont démontré que la technique de culture de prélèvements de l'environnement est appropriée pour la détection des troupeaux infectés par MAP. Dans l'est du Canada où prédominent les petits troupeaux en stabulation entravée, certaines conditions et pratiques de régie pourraient avoir un impact sur la survie, la transmission et l'isolement de MAP. Notre objectif était d'estimer la performance d'une technique standardisée de culture de prélèvements de l'environnement combinée à des échantillons groupés ciblés pour la détection des troupeaux laitiers en stabulation entravée infectés par MAP. Vingt-quatre troupeaux (19 infectés et 5 non infectés) ont été enrôlés, mais seulement 20 troupeaux ont été visités 2 fois dans la même année pour y prélever 7 échantillons de l'environnement et 2 échantillons groupés (vaches malades et vaches maigres). Des échantillons individuels de toutes les vaches dans le troupeau ont été également prélevés. L'isolement de MAP a été réalisé en utilisant le milieu de culture MGIT ParaTB et le système de détection BACTEC 960. Globalement, MAP a été isolée dans 7 % des cultures de l'environnement. La sensibilité de la technique était de 44 % (IC 95 % : 20 % à 70 %) en combinant le résultat des 2 visites et de 32 % (IC 95 % : 13 % à 57 %) en utilisant aléatoirement le résultat d'une seule visite. La meilleure stratégie d'échantillonnage était la combinaison des échantillons de la fosse, de l'écureur, du groupe de vaches malades et du groupe de vaches maigres. La technique standardisée de prélèvements de l'environnement combinée aux échantillons groupés ciblés présentée dans cette étude est une alternative économique à la culture individuelle pour détecter des troupeaux laitiers infectés par MAP. La répétition des prélèvements pourrait contribuer à améliorer la détection des troupeaux infectés par MAP.(Traduit par les auteurs).

Influence of short distance transportation on tracheal bacterial content and lower airway cytology in horses.

The aim of this study was to determine the effects of short distance transportation on airway mucus, cytology and bacterial culture to identify potential biases in the diagnosis of airway diseases in referral centres. Eight healthy adult horses were studied using a prospective cross-over design. Mucus scores, tracheal wash (cytology, bacterial culture) and bronchoalveolar lavage fluid (BALF; cytology) were obtained while stabled and following 2.5 h transportation (with and without hay). Neutrophil counts, percentages and BALF neutrophilia frequency increased following transport without hay (P <0.05). No effect was observed on tracheal cytology and bacterial count (P > 0.05). BALF neutrophilia could develop solely as a result of transportation or due to interactions between repeated transports, ambient temperature, head position or other environmental factors.

Evolution of in vitro antimicrobial resistance in an equine hospital over 3 decades.

This study identified antimicrobial resistance patterns of commonly isolated bacteria at the Equine Hospital of the Université de Montréal between 2007 and 2013, and compared the results with the resistance patterns observed in tests performed in previous decades in the same hospital. A total of 396 antimicrobial susceptibility tests were analyzed by the Kirby-Bauer method during the period 2007 to 2013 and compared to 233 and 255 tests completed in 1986 to 1988 and 1996 to 1998, respectively. The most common bacteria were Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) and Escherichia coli. Except for resistance of coagulase-positive staphylococci to trimethoprim-sulfamethoxazole, there was no overall increase in resistance observed between 1986 to 1988 and 2007 to 2013 for antimicrobials reported for all 3 periods. However, between 1996 to 1998 and 2007 to 2013, there was an increase in in vitro resistance to enrofloxacin for E. coli and Enterobacter spp., and to ceftiofur for Enterobacter spp. and coagulase-positive staphylococci. No increase in resistance was observed for S. zooepidemicus and no isolate was resistant to penicillin.

Évolution de l'antibiorésistancein vitrodans un hôpital équin pendant 3 décennies. L'objectif était d'identifier les patrons de résistance aux antimicrobiens des bactéries fréquemment isolées à l'Hôpital Équin de l'Université de Montréal de 2007 à 2013, pour ensuite les comparer aux données observées au cours des dernières décennies dans le même hôpital. Trois cent quatre-vingt-seize antibiogrammes faits à l'aide de la méthode Kirby-Bauer ont été analysés et comparés aux 233 et 255 ayant été effectués en 1986­1988 et 1996­1998, respectivement. Les bactéries les plus fréquentes étaient Streptococcus equi subsp. zooepidemicus (S. zooepidemicus) et Escherichia coli. Pour les antibiotiques testés pendant les 3 périodes de l'étude, il n'y pas eu d'augmentation de la résistance observée entre 1986­1988 et 2007­2013, à exception de celle des staphylocoques à coagulase positive au triméthoprime-sulfaméthoxazole. Cependant, entre 1996­1998 et 2007­2013, une augmentation de la résistance à l'enrofloxacin a été observée pour E. coli et Enterobacter spp., ainsi qu'une augmentation de la résistance au ceftiofur pour Enterobacter spp. et les staphylocoques à coagulase positive. Aucune augmentation de résistance n'a été observée pour S. zooepidemicus et aucun isolat n'était résistant à la pénicilline.(Traduit par les auteurs).

Genome-Wide Diversity and Phylogeography of Mycobacterium avium subsp. paratuberculosis in Canadian Dairy Cattle.

Mycobacterium avium subsp. paratuberculosis (MAP) is the causative bacterium of Johne's disease (JD) in ruminants. The control of JD in the dairy industry is challenging, but can be improved with a better understanding of the diversity and distribution of MAP subtypes. Previously established molecular typing techniques used to differentiate MAP have not been sufficiently discriminatory and/or reliable to accurately assess the population structure. In this study, the genetic diversity of 182 MAP isolates representing all Canadian provinces was compared to the known global diversity, using single nucleotide polymorphisms identified through whole genome sequencing. MAP isolates from Canada represented a subset of the known global diversity, as there were global isolates intermingled with Canadian isolates, as well as multiple global subtypes that were not found in Canada. One Type III and six "Bison type" isolates were found in Canada as well as one Type II subtype that represented 86% of all Canadian isolates. Rarefaction estimated larger subtype richness in Québec than in other Canadian provinces using a strict definition of MAP subtypes and lower subtype richness in the Atlantic region using a relaxed definition. Significant phylogeographic clustering was observed at the inter-provincial but not at the intra-provincial level, although most major clades were found in all provinces. The large number of shared subtypes among provinces suggests that cattle movement is a major driver of MAP transmission at the herd level, which is further supported by the lack of spatial clustering on an intra-provincial scale.

Limitations of variable number of tandem repeat typing identified through whole genome sequencing of Mycobacterium avium subsp. paratuberculosis on a national and herd level.

BACKGROUND: Mycobacterium avium subsp. paratuberculosis (MAP), the causative bacterium of Johne's disease in dairy cattle, is widespread in the Canadian dairy industry and has significant economic and animal welfare implications. An understanding of the population dynamics of MAP can be used to identify introduction events, improve control efforts and target transmission pathways, although this requires an adequate understanding of MAP diversity and distribution between herds and across the country. Whole genome sequencing (WGS) offers a detailed assessment of the SNP-level diversity and genetic relationship of isolates, whereas several molecular typing techniques used to investigate the molecular epidemiology of MAP, such as variable number of tandem repeat (VNTR) typing, target relatively unstable repetitive elements in the genome that may be too unpredictable to draw accurate conclusions. The objective of this study was to evaluate the diversity of bovine MAP isolates in Canadian dairy herds using WGS and then determine if VNTR typing can distinguish truly related and unrelated isolates. RESULTS: Phylogenetic analysis based on 3,039 SNPs identified through WGS of 124 MAP isolates identified eight genetically distinct subtypes in dairy herds from seven Canadian provinces, with the dominant type including over 80% of MAP isolates. VNTR typing of 527 MAP isolates identified 12 types, including "bison type" isolates, from seven different herds. At a national level, MAP isolates differed from each other by 1-2 to 239-240 SNPs, regardless of whether they belonged to the same or different VNTR types. A herd-level analysis of MAP isolates demonstrated that VNTR typing may both over-estimate and under-estimate the relatedness of MAP isolates found within a single herd. CONCLUSIONS: The presence of multiple MAP subtypes in Canada suggests multiple introductions into the country including what has now become one dominant type, an important finding for Johne's disease control. VNTR typing often failed to identify closely and distantly related isolates, limiting the applicability of using this typing scheme to study the molecular epidemiology of MAP at a national and herd-level.

A systematic review of risk factors associated with the introduction of Mycobacterium avium spp. paratuberculosis (MAP) into dairy herds.

The objective of this study was to systematically collect and appraise the scientific evidence related to risk factors associated with the introduction of Mycobacterium avium spp. paratuberculosis (MAP) into a herd of cattle. An electronic search was conducted to collect relevant references addressing 2 specific questions: are i) purchasing/introduction of cattle into a herd, and ii) presence of wildlife or domestic animals, risk factors for the introduction of MAP into a herd? The screening was based on titles and abstracts and selected studies were fully analyzed. Seventeen manuscripts published between 1996 and 2011 were ultimately analyzed. Unit of interest was mainly the herd (n = 17). The specific description of the risk factors studied varied between studies. The principal study design was cross-sectional (n = 15). The review indicated that purchase/introduction of animals was an important risk factor and that the importance of wildlife or other domestic species as a mechanism for transmission into a cattle herd was not measurable.

Revue systématique sur les facteurs de risque associés à l'introduction deMycobacterium aviumspp.paratuberculosis(MAP) dans les troupeaux laitiers. L'objectif était la collecte et l'évaluation systématique des preuves scientifiques liées à des facteurs de risque associés à l'introduction de Mycobacterium avium spp. paratuberculosis (MAP) dans un troupeau.Une recherche électronique a été réalisée pour trier des publications afin de répondre à 2 questions : Est-ce que i) l'achat/introduction de bovins dans un troupeau, et ii) la présence d'animaux sauvages et domestiques sont des facteurs de risque pour l'introduction de MAP dans un troupeau. La sélection a été effectuée en se basant sur les titres, les résumés, et les études choisies ont été entièrement analysés.Dix-sept manuscrits publiés entre 1996 et 2011 ont été sélectionnés. L'unité d'intérêt a principalement été le troupeau (n = 17). Les descriptions des facteurs de risque étudiés variaient d'une étude à l'autre. La majorité des études étaient de type transversal (n = 15).L'achat d'animaux est un facteur de risque bien documenté confirmant l'introduction de la paratuberculose dans le troupeau. Toutefois, malgré la présence de MAP dans les populations domestiques et sauvages à proximité du bétail, l'importance de ce facteur de risque dans la transmission à un troupeau n'était pas mesurable.(Traduit par les auteurs).

Genetic structure of Mycobacterium avium subsp. paratuberculosis population in cattle herds in Quebec as revealed by using a combination of multilocus genomic analyses.

Mycobacterium avium subsp. paratuberculosis is the etiological agent of paratuberculosis, a granulomatous enteritis affecting a wide range of domestic and wild ruminants worldwide. A variety of molecular typing tools are used to distinguish M. avium subsp. paratuberculosis strains, contributing to a better understanding of M. avium subsp. paratuberculosis epidemiology. In the present study, PCR-based typing methods, including mycobacterial interspersed repetitive units/variable-number tandem repeats (MIRU-VNTR) and small sequence repeats (SSR) in addition to IS1311 PCR-restriction enzyme analysis (PCR-REA), were used to investigate the genetic heterogeneity of 200 M. avium subsp. paratuberculosis strains from dairy herds located in the province of Quebec, Canada. The majority of strains were of the "cattle type," or type II, although 3 strains were of the "bison type." A total of 38 genotypes, including a novel one, were identified using a combination of 17 genetic markers, which generated a Simpson's index of genetic diversity of 0.876. Additional analyses revealed no differences in genetic diversity between environmental and individual strains. Of note, a spatial and spatiotemporal cluster was evidenced regarding the distribution of one of the most common genotypes. The population had an overall homogeneous genetic structure, although a few strains stemmed out of the consensus cluster, including the bison-type strains. The genetic structure of M. avium subsp. paratuberculosis populations within most herds suggested intraherd dissemination and microevolution, although evidence of interherd contamination was also revealed. The level of genetic diversity obtained by combining MIRU-VNTR and SSR markers shows a promising avenue for molecular epidemiology investigations of M. avium subsp. paratuberculosis transmission patterns.

Characterization of H1N1 swine influenza viruses circulating in Canadian pigs in 2009.

The 2009 pandemic H1N1 (pH1N1), of apparent swine origin, may have evolved in pigs unnoticed because of insufficient surveillance. Consequently, the need for surveillance of influenza viruses circulating in pigs has received added attention. In this study we characterized H1N1 viruses isolated from Canadian pigs in 2009. Isolates from May 2009 were comprised of hemagglutinin and neuraminidase (NA) genes of classical SIV origin in combination with the North American triple-reassortant internal gene (TRIG) cassette, here termed contemporary SIV (conSIV) H1N1. These conSIV H1N1 viruses were contiguous with the North American αH1 cluster, which was distinct from the pH1N1 isolates that were antigenically more related to the γH1 cluster. After the initial isolation of pH1N1 from an Alberta pig farm in early May 2009, pH1N1 was found several times in Canadian pigs. These pH1N1 isolates were genetically and antigenically homogeneous. In addition, H1N1 viruses bearing seasonal human H1 and N1 genes together with the TRIG cassette and an NA encoding an oseltamivir-resistance marker were isolated from pigs. The NS gene of one of these seasonal human-like SIV (shSIV) H1N1 isolates was homologous to pH1N1 NS, implicating reassortment between the two strains. Antigenic cross-reactivity was observed between pH1N1 and conSIV but not with shSIV H1N1. In summary, although there was cocirculation of pH1N1 with conSIV and shSIV H1N1 in Canadian pigs after May 2009, there was no evidence supporting the presence of pH1N1 in pigs prior to May 2009. The possibility for further reassortants being generated exists and should be closely monitored.

Epidemiological, biochemical and antimicrobial susceptibility characteristics of Streptococcus pseudoporcinus isolated in Quebec, Canada, from 1997 to 2006.

From 1997 to 2006, in the province of Quebec, Canada, 15 isolates of Streptococcus pseudoporcinus from 1 urine and 14 vaginorectal cultures were recovered from the genitourinary tract of pregnant women. All these women originated from the Caribbean or sub-Saharan Africa (P=0.00045 compared with a suitable control group). The S. pseudoporcinus isolates were compared to eight isolates of Streptococcus porcinus identified in Quebec from 1995 to 2006, all from animals, of which five were swine. 16S rRNA gene sequencing was required to differentiate between S. pseudoporcinus and S. porcinus animal isolates.

Clinical and in vitro efficacy of amoxicillin against bacteria associated with feline skin wounds and abscesses.

A clinical trial involving 122 cats with infected skin wounds or abscesses presented to 10 veterinary clinics was conducted to evaluate the efficacy of 2 oral amoxicillin drug products (a paste and a suspension). A 2nd objective of the study was to identify bacteria involved in such infections and verify their in vitro sensitivity to amoxicillin. Samples of wound exudate were harvested at the time of presentation and submitted for aerobic and anaerobic culture. The sensitivity to amoxicillin of isolates thought to be infecting agents was tested, using a standard minimum inhibitory concentration method. Pasteuralla multocida and obligate anaerobes of the genera Prevotella, Fusobacterium, and Porphyromonas were the most frequently isolated pathogens. Overall, their in vitro susceptibility to amoxicillin was very good. Both drug products were clinically efficacious with a global success rate of 95.1% for cats administered oral amoxicillin at 11-22 mg/kg bodyweight (mean 13.8 mg/kg bodyweight) twice daily for 7 to 10 days.

Comparison of susceptibility to antimicrobials of bacterial isolates from companion animals in a veterinary diagnostic laboratory in Canada between 2 time points 10 years apart.

The susceptibility to antimicrobials of bacterial species most frequently isolated from companion animals in a veterinary teaching diagnostic laboratory was evaluated retrospectively. A significant decrease between 1990-1992 and 2002-2003 was noted in the susceptibility of dog isolates to the following antimicrobials: Escherichia coli to cephalothin (86% to 61%, P < 0.001); E. coli to ampicillin (85% to 67%, P < 0.001); Proteus spp. to ampicillin (92% to 71%, P < 0.01); coagulase-positive staphylococci (Staphylococcus aureus and Staphylococcus intermedius) to enrofloxacin (99% to 95%, P < 0.01). Significantly increased susceptibilities were also noted as follows: coagulase-positive staphylococci to erythromycin (78% to 90%, P < 0.001) and tetracycline (61% to 77%, P < 0.001). Despite a limited number of results available for cats, a significant increase in susceptibility was noted for Pseudomonas spp. to gentamicin (40% to 100%, P < 0.05) and for E. coli to tetracycline (59% to 80%, P < 0.05). Regular updates on the resistance to antimicrobials used in veterinary medicine are required.

Systemic Cryptococcus albidus infection in a Doberman Pinscher.

Cryptococcus albidus is a saprophytic, encapsulated yeast usually found in air, both outdoor and indoor, and sometimes on human skin. It is not usually considered to be a primary pathogen. Most cryptococcal infections of humans and animals are caused by Cryptococcus neoformans. Several cases of C. albidus infection have been reported in humans over the past 20 years. In the veterinary literature, 2 equine cases have been described: genital infection and mycotic keratitis. The present report is the first documented case of C. albidus systemic infection in a dog. Veterinarians and diagnosticians should be aware that C. albidus may be a potential canine pathogen.