ISMap02 element targeted nested polymerase chain in the detection of Mycobacterium avium subsp. paratuberculosis in fecal samples of cattle and buffaloes
Background and Aim: Johne’s disease is chronic granulomatous enteritis which affects ruminants. There are many diagnostic approaches for the detection of Mycobacterium avium subsp. paratuberculosis (MAP) of which molecular detection methods using various elements are less time consuming and more accurate. The present study was conducted using ISMap02 element for nested polymerase chain reaction (nPCR) based detection of MAP in fecal samples. The aim was to test the sensitivity and specificity of the ISMap02 element and also to use this element for the detection of MAP in fecal samples of cattle and buffaloes. Materials and Methods: A total of 211 fecal samples of cattle and buffaloes from different herds around Ludhiana aged between 2 and 13 years were collected, and DNA extraction was done from these samples. The nPCR was carried out for the detection of MAP in fecal samples. Results: The ISMap02 element was specific for the detection of MAP only and showed a sensitivity of detection of 7.6 fg/µL of the standard genomic DNA. Among the 211 fecal samples of cattle and buffaloes tested for the ISMap02 element, 18 samples (8.5%) were positive for MAP. Conclusion: The ISMap02 element is specific and sensitive for the detection of MAP in various samples, and when used in nPCR format, it can increase the sensitivity of detection.
Mycobacterium avium subsp. paratuberculosis Antibody Response, Fecal Shedding, and Antibody Cross-Reactivity to Mycobacterium bovis in M. avium subsp. paratuberculosis-Infected Cattle Herds Vaccinated against Johne's Disease
ABSTRACT Vaccination for Johne's disease with killed inactivated vaccine in cattle herds has shown variable success. The vaccine delays the onset of disease but does not afford complete protection. Johne's disease vaccination has also been reported to interfere with measurements of cell-mediated immune responses for the detection of bovine tuberculosis. Temporal antibody responses and fecal shedding of Mycobacterium avium subsp. paratuberculosis, the causative agent of Johne's disease, were measured in 2 dairy cattle herds using Johne's disease vaccine (Mycopar) over a period of 7 years. Vaccination against Johne's disease resulted in positive serum M. avium subsp. paratuberculosis antibody responses in both herds, and the responses persisted in vaccinated cattle up to 7 years of age. Some vaccinated animals (29.4% in herd A and 36.2% in herd B) showed no serological reactivity to M. avium subsp. paratuberculosis. M. avium subsp. paratuberculosis-specific antibody responses were also detected in milk from Johne's disease-vaccinated animals, but fewer animals (39.3% in herd A and 49.4% in herd B) had positive results with milk than with serum samples. With vaccination against M. avium subsp. paratuberculosis, fecal shedding in both dairy herds was reduced significantly (P < 0.001). In addition, when selected Johne's disease-vaccinated and -infected animals were investigated for serological cross-reactivity to Mycobacterium bovis, no cross-reactivity was observed.
Antibody and skin-test responses of sheep vaccinated against Johne's Disease.
Current vaccines against Mycobacterium avium subsp. paratuberculosis (MAP, Johne's Disease) may cause animals to react positively when tested for Mycobacterium bovis (Bovis). Therefore, the effects of vaccination on MAP serum Ab and skin-test responses to MAP and Bovis PPD were compared in 25 ewes vaccinated against MAP with 24 control ewes in an infected flock 3 years post-vaccination. MAP-specific Ab levels were higher (P<0.001) in vaccinated ewes than in control ewes. All increases in skinfold-thickness from 0 to 48h were greater (P<0.0001) than zero while increases in skinfold-thickness from 48 to 72h were greater (P<0.05) than zero for Johnin but not for Bovis PPD. The Vaccine x PPD x Time interaction for skinfold-thickness was significant (P<0.001) with greater increases to Johnin than to Bovis, but with much greater increases in vaccinated ewes. These data suggest that administration of vaccines against MAP developed from whole organisms increase the likelihood that animals will be classified as "responders" to a Bovis screening test and negative by the follow-up comparative cervical tuberculin test, but they also show that vaccination initiates both humoral and cell-mediated MAP-specific responses.