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).

Validation of an in-house real-time PCR fecal assay and comparison with two commercial assays for the antemortem detection of Mycobacterium avium subsp. paratuberculosis infection in culled sheep.

Paratuberculosis is a chronic infectious enteritis of ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP). In sheep, the antemortem detection of the infection is challenging given the slow progression of the disease and the lack of sensitive, specific, and cost-effective validated tests. We adapted an in-house real-time PCR (rtPCR) assay targeting the multi-copy IS 900 element of MAP. The sensitivity and specificity of this essay for the detection of MAP infection were estimated in a convenience sample of culled ewes from 7 infected flocks and compared to a commercial fecal rtPCR, a commercial ELISA, and fecal culture. An infected ewe was defined as a ewe with a positive culture of the ileum and/or mesenteric lymph node. A non-infected ewe was defined as a ewe negative in intestinal tissue culture, negative in fecal culture, and with no lesions consistent with paratuberculosis. The in-house rtPCR had a sensitivity estimate of 84% (95% confidence interval [CI]: 59%, 97%) among the 44 infected ewes, which was significantly higher ( p ⩽ 0.05) than the sensitivity of a commercial fecal rtPCR (52%, 95% CI: 27%, 76%; or 63%, 95% CI: 35%, 87% depending on the cutoff used), an ELISA (14%, 95% CI:2.0%, 41%), and fecal culture (21%, 95% CI: 2.7%, 59%). No statistical difference in assay specificities was observed for the 30 non-infected ewes. The in-house rtPCR is a promising tool that could be used advantageously for the antemortem detection of MAP infection in sheep.

Validation of an immunohistochemical assay for bovine cysticercosis, with comparison to a standard histological method.

The larval stage (syn Cysticercus bovis) of the human tapeworm Taenia saginata causes cysticercosis in cattle, which has both aesthetic and food safety implications to consumers of beef. A monoclonal antibody-based immunohistochemical (IHC) assay developed to improve postmortem diagnosis of this parasite and a standard histological method were assessed to determine their fitness for intended use. Sections from 169 known-positive specimens of T. saginata from experimentally or naturally infected cattle, and from 30 known-negative specimens and lesions of various etiologies from non-infected cattle, were tested. The IHC assay identified significantly more known positive bovine cysticerci than the histological method (91.7% and 38.5%, respectively). Positive IHC staining occurred on sections from other cestode species, but should not affect the diagnostic specificity of this assay for bovine cysticercosis, due to the different host and/or tissue preferences amongst these parasites. Use of the IHC assay should improve the reliability of diagnosing lesions caused by degenerated cysticerci, facilitating more effective and efficient control of bovine cysticercosis.