scholarly journals A Comparative Study on the Efficiency of Two Mycobacterium avium subsp. paratuberculosis (MAP)-Derived Lipopeptides of L3P and L5P as Capture Antigens in an In-House Milk ELISA Test

Vaccines ◽  
2021 ◽  
Vol 9 (9) ◽  
pp. 997
Author(s):  
Sepideh Hosseiniporgham ◽  
Franck Biet ◽  
Christelle Ganneau ◽  
John P. Bannantine ◽  
Sylvie Bay ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Elisa Test ◽  
Cross Reactivity ◽  
Mycobacterium Avium Subsp ◽  
Enzyme Analysis ◽  
Restriction Enzyme Analysis ◽  
Individual Level ◽  
Milk Samples ◽  
Milk Elisa ◽  
Mycobacterium Avium Subsp Paratuberculosis

Mycobacterium avium subsp. paratuberculosis (MAP) surface-exposed lipopeptides could be specific capture-antigen molecules targeting antibodies against MAP, in milk, through ELISA. Previous studies have revealed that MAP strains, isolated from sheep (S) or cow (C), could produce specific lipopeptides, L3P or L5P, respectively. In this study, we used L3P and L5P as capture antigens in an in-house milk ELISA (H-MELISA) to assess how these antigens perform, in comparison with other ELISA tests, on well-defined milk samples from MAP-infected sheep. The overall positivity rates of H-MELISA via L3P and L5P varied by the source of milk samples, in which, at bulk-tank-milk (BTM) level, the majority of positive cases (63.83%) reacted more against L5P, whereas a predominant number (69.14%) of milk samples were more responsive against L3P at the individual level. To clarify whether the positivity status of milk samples in H-MELISA L3P/L5P were predictive of MAP strain-types (S/C), strain-typing was carried out using PCR IS1311-restriction enzyme analysis. Although the presence of three MAP strains (S/C/bison types) was detected among the milk samples, the C-type (46.67%) and S-type (75%) MAP strains were detected with higher incidence among BTMs and individual milk samples, respectively. However, further examination on the H-MELISA L3P/L5P-positivity pattern of each C/S-type-MAP sample revealed that some samples had a reverse reactivity against both L3P and L5P. These results could be the consequence of either cross-reactivity between L3P and L5P (due to the similarity in the structures of the two epitopes) or simply a within-herd mixed infection with MAP strains of C and S types. These findings suggest that lipopeptide antigens could contribute a diagnostic test with optimal performance, considering the diversity of MAP strains.

scholarly journals Anti-Chlamydia abortus and Anti-Mycobacterium avium subsp. paratuberculosis Antibodies in Buffaloes in the State of Pernambuco, Brazil

2019 ◽  
Vol 47 (1) ◽  
Author(s):  
Amanda De Noronha Xavier ◽  
Sérgio Alves Do Nascimento ◽  
Tania Alexandra Ortega Sierra ◽  
Pollyane Raysa Fernandes de Oliveira ◽  
Rinaldo Aparecido Mota ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Clinical Signs ◽  
Epidemiological Studies ◽  
Elisa Test ◽  
The State ◽  
Mycobacterium Avium Subsp ◽  
Economic Losses ◽  
Water Buffaloes ◽  
Chlamydia Abortus ◽  
Mycobacterium Avium Subsp Paratuberculosis

Background: The occurrence of economic losses in buffaloes may be related to reproductive problems such as chlamydiosis caused by the bacteria Chlamydia abortus considered as a zoonotic agent; and digestive problems highlighting the infection by Mycobacterium avium subsp. paratuberculosis (Map), responsible for paratuberculosis. There is a little information about these diseases in buffaloes, therefore the aim of this study was to determine the occurrence of anti-Chlamydia abortus and anti- Mycobacterium avium subsp. paratuberculosis (Map) antibodies in water buffaloes in the state of Pernambuco, Brazil.Materials, Methods & Results: The 262 bubaline sera belonging to the serum bank of the Infectious Diseases Laboratory (LIDIC) of the Federal Rural University of Pernambuco (UFRPE) were analyzed. The samples were from nine properties distributed in the municipalities of Agreste and Zona da Mata of the state of Pernambuco. For the detection of anti-Chlamydia abortus and anti-Map antibodies was used a technique of Enzyme immunoabsorption Assay (ELISA) of the IDEXX® by following the manufacturer's instructions. Regarding the detection of anti-Chlamydia abortus antibodies, it was observed that 47.70% (125/262) of the samples were positive. All properties showed at least one positive animal for the investigation of anti-Chlamydia abortus antibodies. It was also verified the occurrence of 7.25% of suspected animals for the investigation of anti-C. abortus antibodies. No positives animals were observed for Map in the bubaline analyzed.Discussion: The occurrence of anti-C. abortus antibodies in buffaloes in the region can be explained by the fact that properties with buffalo breeding has sanitary management that allows the contact between animals, thus increasing the risk of transmission of the agent. In addition, the variation found from 35% to 68.75% may be associated with divergences to the type of management and breeding system used in each property. The percentage of suspected animals may suggest that the number of positive animals is higher or that there were non-specific reactions with other species of Chlamydia, but it is not possible to determine the seroconversion without the accomplishment of paired serology. Because it is a zoonosis, C. abortus may be a risk to the health of the population involved, since the transmission of the bacteria to humans can occur by contact with secretions and excretions of these animals. No positive animals were found for the occurrence of anti-Map antibodies, however, there are reports of the infection identifying animals with clinical signs or properties with a history of the disease, and this may be related to the differences in each property in relation to the history and sanitary management. Moreover, the ELISA test may not be sensitive to the agent depending on the phase of the infection, because if the response is predominantly cellular, the number of antibodies is diminished, making the serological diagnosis difficult. The use of more sensitive tests for the bubaline species can also favor the diagnosis of the infection. Although the occurrence of anti-Map antibodies in the analyzed samples was not detected, it is important to carry out routine epidemiological studies, since the disease has already been registered in water buffaloes in the state of Pernambuco. Considering the occurrence of anti-Chlamydia abortus antibodies it is suggested to conduct studies in order to isolate the agent and verify its real importance in the reproductive disorders of the bubaline species, since this agent causes reproductive losses and has a zoonotic character.

scholarly journals Effects of freezing on ability to detect Mycobacterium avium subsp. paratuberculosis from bovine tissues following culture

2018 ◽  
Vol 30 (5) ◽  
pp. 743-746
Author(s):  
Caroline S. Corbett ◽  
Jeroen De Buck ◽  
Herman W. Barkema
Keyword(s):  
Mycobacterium Avium ◽  
Gold Standard ◽  
Frozen Storage ◽  
Mycobacterium Avium Subsp ◽  
Freezing And Thawing ◽  
Tissue Samples ◽  
Milk Samples ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
Autopsy Tissues ◽  
Numerical Change

Mycobacterium avium subsp. paratuberculosis (MAP) is the bacterium that causes Johne’s disease in cattle. Although infected cattle can be identified by examining fecal, blood, or milk samples, the gold standard is identification of MAP in tissue samples postmortem. Although tissue samples are commonly frozen, the ability to detect MAP in frozen–thawed tissue samples has apparently not been reported. We therefore determined the ability to detect MAP in tissue samples following freezing. Tissue samples were collected from calves that were either inoculated (IN) 3 mo prior, or contact-exposed (CE) for 3 mo. Following autopsy, tissues were immediately processed for culture, followed by DNA extraction and detection by qPCR. Samples were categorized as positive or negative based on the cycle threshold (Ct) value. The remaining unprocessed tissue samples were frozen at −80°C. After 18 mo, 50 tissue samples designated MAP-positive were thawed and processed for detection of MAP. Four (8%) samples were qPCR-negative, and Ct values of the remaining 46 samples were higher after freezing. Given the small numerical change in Ct values for MAP-positive samples after 18 mo of frozen storage, freezing and thawing may have had some deleterious effects on MAP detection in tissues. Although the decrease in ability to detect MAP-positive samples was minor for IN calves, there may be a greater effect for CE calves that should be considered when freezing tissue samples.

scholarly journals Mycobacterium avium subsp. paratuberculosis ELISA Responses in Milk Samples from Vaccinated and Nonvaccinated Dairy Goat Herds in The Netherlands

2019 ◽  
Vol 6 (2) ◽  
pp. 58 ◽  
Author(s):  
Saskia Luttikholt ◽  
Karianne Lievaart-Peterson ◽  
Maaike Gonggrijp ◽  
Marian Aalberts ◽  
Gerdien van Schaik ◽  
...  
Keyword(s):  
The Netherlands ◽  
Mycobacterium Avium ◽  
Relative Sensitivity ◽  
Herd Size ◽  
Mycobacterium Avium Subsp ◽  
Dairy Goat ◽  
Dairy Goats ◽  
Milk Samples ◽  
Goat Population ◽  
Mycobacterium Avium Subsp Paratuberculosis

The aims of our study were to calculate the most appropriate cut-off value for milk samples in a serum-validated Mycobacterium avium subsp. paratuberculosis (MAP) ELISA and to analyze MAP ELISA responses in milk samples from vaccinated and nonvaccinated dairy goats in the Netherlands. Analyzed herds were representative for location and herd size of dairy goat herds in the Netherlands. A significantly higher proportion of the analyzed 49 herds were organic as compared with the total Dutch dairy goat population. First, the MAP ELISA was optimized using 992 paired serum and milk samples. At a cut-off of 25 S/P%, the relative sensitivity (Se) was 58.4% (n = 992, 95% CI: 48.8%−67.6%) and relative specificity (Sp) was 98.5% (n = 992, 95% CI: 97.5%−99.2%), as compared to serum ELISA results. The percentage of positively tested herds was 78.2% (n = 49, 95% CI: 63.4%−88.1%). The percentage of positive milk samples per herd (n = 22) was on average 4.6% (median, min, and max of 4.7%, 0.0%, and 10.7%, respectively). Average age of ELISA-positive (3.2 years) and -negative goats (3.2 years) was not different. Significantly more vaccinated goats tested positive (6.7%) as compared with nonvaccinated goats (1.1%). This study shows that a high number of vaccinated and nonvaccinated commercial dairy goat herds in the Netherlands have MAP-ELISA-positive goats.

scholarly journals Longitudinal Study of Mycobacterium avium Subsp. paratuberculosis Antibody Kinetics in Dairy Cattle Using Sera and Milk throughout the Lactation Period

2020 ◽  
Vol 7 (3) ◽  
pp. 81 ◽  
Author(s):  
Md. Shohel Al Faruk ◽  
Young-hoon Jung ◽  
Tai-young Hur ◽  
Sang-suk Lee ◽  
Yong-il Cho
Keyword(s):  
Dairy Cattle ◽  
Antibody Titer ◽  
Mycobacterium Avium ◽  
Elisa Test ◽  
Mycobacterium Avium Subsp ◽  
Control Group ◽  
Lactation Period ◽  
Antibody Titers ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
Antibody Kinetics

Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of Johne’s disease in dairy cattle populations around the world. The objective of this study was to evaluate MAP antibody kinetics in serum and milk samples throughout the lactation period in dairy cattle. The samples were collected simultaneously from eight MAP-positive and two healthy MAP-negative (control group) cows. The MAP antibody was detected by using serum and milk ELISA. The serum and milk MAP antibody titers fluctuated between the positive and negative cut-off values in this study. Specifically, cattle with low MAP antibody titer (<100) showed fluctuation between the cut-off values. Variable changes of MAP antibody titer were also observed after parturition. Between the serum and milk MAP antibody titers, there was a positive correlation (R2 = 0.5358) observed throughout the assessment period. The milk MAP ELISA test had low diagnostic performance in cows with low MAP titer due to its weak correlation (R2 = 0.0198). Finally, this study suggest that the periodic MAP ELISA test is recommended for the application of Johne’s eradication program due to the fluctuating nature of MAP antibody kinetics.

scholarly journals Identification of Mycobacterium avium subsp. paratuberculosis (MAP) in Sheep Milk, a Zoonotic Problem

2020 ◽  
Vol 8 (9) ◽  
pp. 1264
Author(s):  
Sepideh Hosseiniporgham ◽  
Tiziana Cubeddu ◽  
Stefano Rocca ◽  
Leonardo A. Sechi
Keyword(s):  
Mycobacterium Avium ◽  
Gold Standard ◽  
Dairy Product ◽  
Mycobacterium Avium Subsp ◽  
Individual Level ◽  
Reference Models ◽  
Sheep Milk ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
The Individual ◽  
Serum Elisa

Johne’s disease (JD) is a life-threatening gastrointestinal disease affecting ruminants, which causes crucial economical losses globally. This ailment is caused by Mycobacterium avium subsp. paratuberculosis (MAP), a fastidious intracellular pathogen that belongs to the Mycobacteriaceae family. This acid-fast, hard-to-detect bacterium can resist milk pasteurization and be conveyed to dairy product consumers. Many studies have emphasized the zoonotic nature of MAP, suggesting an association between MAP and some gastroenteric conditions such as Crohn’s disease in humans. This underlines the importance of utilizing efficient pasteurization alongside a state-of-the-art diagnostic system in order to minimize the possible ways this pathogen can be conveyed to humans. Until now, no confirmatory MAP screening technique has been developed that can reveal the stages of JD in infected animals. This is partially due to the lack of an efficient gold-standard reference method that can properly evaluate the performance of diagnostic assays. Therefore, the following research aimed to compare the merits of qPCR and ELISA assessments of milk for the detection of MAP in a total of 201 Sardinian unpasteurized sheep milk samples including 73 bulk tank milk (BTM) and 128 individual samples from a MAP-infected flock (MIF) applying various reference models. Accordingly, milk qPCR and ELISA assessments, together and individually, were used as reference models in the herd-level study, while serum ELISA and fecal PCR were similarly (together and in isolation) considered as the gold standards in the individual-level diagnosis. This study showed that the type of gold-standard test affects the sensitivity and specificity of milk qPCR and ELISA significantly. At the individual level in the MAP-infected flock, serum ELISA in isolation and together with fecal PCR were recognized as the best references; however, the best correlation was seen between milk and serum ELISA (p < 0.0001). Regarding the detection of MAP in BTM, qPCR IS900 was recognized as the most sensitive and specific diagnostic test (p < 0.0001) for monitoring the MAP shedders and animals with clinically developed symptoms within herds, under the condition that both milk qPCR and milk ELISA tests formed a binary reference model. The BTM analyses (qPCR and ELISA) revealed that MAP positivity has a seasonal pattern. This hypothesis was proven through a longitudinal study on 14 sheep herds.

scholarly journals Mycobacterium avium subsp. paratuberculosis antibodies in cows of low-tropic dairy herds in Colombia

2020 ◽  
pp. e1782
Author(s):  
Nathalia María del Pilar Correa-Valencia ◽  
Ferney Arango-Lezcano ◽  
Jorge Arturo Fernández-Silva
Keyword(s):  
Mycobacterium Avium ◽  
Positive Outcome ◽  
Cross Sectional Study ◽  
Elisa Test ◽  
Mycobacterium Avium Subsp ◽  
Dairy Herds ◽  
Cross Sectional ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
Level Variable ◽  
Associated Variables

Objective. To report the frequency of seropositivity against Mycobacterium avium subsp. paratuberculosis (MAP) and to explore the factors associated with the positive outcome, both at herd and animal level. Materials and methods. A cross-sectional study was carried out on 204 dairy cows, result of the crossing of different breeds, from five low-tropic dairy herds, located in three municipalities of the Provinces of Sucre and Córdoba (Colombia) in 2018. The animals were randomly selected, and blood samples were collected from each one. A commercial ELISA kit was used to analyze the sera, and the association between these variables and the result to ELISA (p<0.05) was explored. Results. The 17.2% (35/204; 95% CI: 12.0-22.3%) of the cows were positive for MAP by the ELISA test, and the five herds had seropositive animals. The herd-level variable presence of other ruminants in co-grazing with cattle in the last 2 years and the animal-level variables age and parity were associated with the positive ELISA results. Conclusions. The present study found that 17.2% of the cows and 100% of the herds were MAP positive by the ELISA test. Additionally, associated variables were identified that may be of interest to both producers and veterinarians and guide their approach to disease management.

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