scholarly journals Development and Validation of a Novel ELISA for the Specific Detection of Antibodies against Mycobacterium avium Subspecies paratuberculosis Based on a Chimeric Polyprotein

2021 ◽  
Vol 2021 ◽  
pp. 1-10
Author(s):  
Roberto Damián Moyano ◽  
Magali Andrea Romero ◽  
María Alejandra Colombatti Olivieri ◽  
María Fiorella Alvarado Pinedo ◽  
Gabriel Eduardo Traveria ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Roc Curve ◽  
Bovine Serum ◽  
Mycobacterium Avium Subspecies Paratuberculosis ◽  
Cross Reactivity ◽  
Specific Detection ◽  
Serum Samples ◽  
Specific Antigens ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
Development And Validation

Bovine paratuberculosis (PTB) is caused by Mycobacterium avium subsp. paratuberculosis (MAP). The optimization of detection tests specific for MAP is crucial to improve PTB control. In this work, we aimed to develop and validate a diagnostic tool based on an ELISA to specifically detect anti-MAP antibodies from bovine serum samples. For that purpose, we designed a recombinant polyprotein containing four specific antigens from MAP and optimized the ELISA. The validation consisted of the assessment of 10 sera from PTB-infected and healthy bovines with different OD values. The diagnostic performance of the polyprotein-ELISA was evaluated by testing 130 bovine serum samples (47 healthy, 48 MAP-infected, and 35 M. bovis-infected bovines). The ELISA using the polyprotein yielded an area under the ROC curve (AUC) of 0.9912 (95% CI, 0.9758–1.007; P  < 0.0001). Moreover, for this ELISA, the cut-off selected from the ROC curve based on the point with a sensitivity of 95.56% (95% CI, 0.8485–0.9946) and specificity of 97.92 (95% CI, 0.8893–0.9995) was 0.3328. Similar results were obtained with an ELISA using the commercial Paratuberculosis Protoplasmatic Antigen (PPA). However, the ELISA with the polyprotein antigen showed a better performance against sera from animals infected with Mycobacterium bovis compared to the ELISA with PPA: lower cross-reactivity (2.85% versus 25.71%). These results demonstrate a very low cross-reactivity of the polyprotein with antibodies present in serum samples from animals infected with M. bovis. The designed polyprotein and the validated ELISA could be very useful for the specific identification of MAP-infected animals in herds.

scholarly journals Profiling Bovine Antibody Responses to Mycobacterium avium subsp. paratuberculosis Infection by Using Protein Arrays

2007 ◽  
Vol 76 (2) ◽  
pp. 739-749 ◽  
Author(s):  
John P. Bannantine ◽  
Michael L. Paustian ◽  
W. Ray Waters ◽  
Judith R. Stabel ◽  
Mitchell V. Palmer ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Cross Reactivity ◽  
Surface Proteins ◽  
Peptide Transport ◽  
Serum Samples ◽  
Protein Arrays ◽  
Healthy Control ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
Immunological Assays ◽  
Bovine Antibody

ABSTRACT With the genome sequence of Mycobacterium avium subsp. paratuberculosis determined, technologies are now being developed for construction of protein arrays to detect the presence of antibodies against M. avium subsp. paratuberculosis in host serum. The power of this approach is that it enables a direct comparison of M. avium subsp. paratuberculosis proteins to each other in relation to their immunostimulatory capabilities. In this study, 93 recombinant proteins, produced in Escherichia coli, were arrayed and spotted onto nitrocellulose. These proteins include unknown hypothetical proteins and cell surface proteins as well as proteins encoded by large sequence polymorphisms present uniquely in M. avium subsp. paratuberculosis. Also included were previously reported or known M. avium subsp. paratuberculosis antigens to serve as a frame of reference. Sera from healthy control cattle (n = 3) and cattle infected with either M. avium subsp. avium and Mycobacterium bovis were exposed to the array to identify nonspecific or cross-reactive epitopes. These data demonstrated a degree of cross-reactivity with the M. avium subsp. avium proteins that was higher than the degree of cross-reactivity with the more distantly related M. bovis proteins. Finally, sera from naturally infected cattle (n = 3) as well as cattle experimentally infected with M. avium subsp. paratuberculosis (n = 3) were used to probe the array to identify antigens in the context of Johne's disease. Three membrane proteins were the most strongly detected in all serum samples, and they included an invasion protein, an ABC peptide transport permease, and a putative GTPase protein. This powerful combination of genomic information, molecular tools, and immunological assays has enabled the identification of previously unknown antigens of M. avium subsp. paratuberculosis.

scholarly journals Identification of Mycobacterium avium subspecies paratuberculosis in sheep farms in Bayannaoer, Inner Mongolia, China

2021 ◽  
Author(s):  
yuandi yu ◽  
Suhui Zhang ◽  
Guoyang Xu ◽  
Dengfeng Xu ◽  
Hua Zheng ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Inner Mongolia ◽  
Clinical Symptoms ◽  
Mycobacterium Avium Subspecies Paratuberculosis ◽  
Elisa Test ◽  
Economic Losses ◽  
Serum Samples ◽  
Polymerase Chain ◽  
Mycobacterium Avium Subsp Paratuberculosis

Abstract Background Paratuberculosis, is a widespread chronic infection of Mycobacterium avium subsp. paratuberculosis (Map), causes significant economic losses to the sheep industry. The current study investigated this disease, which causes diarrhea in sheep, particularly, in Bayannaoer, Inner Mongolia. Diagnosis was based on clinical symptoms, pathological autopsy, histopathological inspection, and serological and molecular methods. Results Paratuberculosis was confirmed via a polymerase chain reaction using DNA extracted from tissue and fecal samples. Serum samples from 472 individual sheep were obtained to detect antibodies against MAP by ELISA test and MAP antibodies were separately detected in 17.86% (35/196) and 18.48% (51/276) of sheep herds at 6 months and 1 years of age respectively. The results of tissue lesion and pathological section were consistent with paratuberculosis infection. Conclusions This is the first report of Mycobacterium avium subspecies paratuberculosis seroprevalence in Bayannaoer sheep in Inner Mongolia. Our findings show that MAP was prevalent and potentially threaten in this region and more further investigations on long-term epidemiological

scholarly journals The Effect of Mycobacterium avium subsp. Paratuberculosis Infection on the Productivity of Cows in Two Dairy Herds with a Low Seroprevalence of Paratuberculosis

Animals ◽  
2020 ◽  
Vol 10 (3) ◽  
pp. 490 ◽  
Author(s):  
Agnieszka Wiszniewska-Łaszczych ◽  
Katarzyna G. Liedtke ◽  
Joanna M. Szteyn ◽  
Tomasz Lachowicz
Keyword(s):  
Dairy Cows ◽  
Milk Yield ◽  
Mycobacterium Avium ◽  
Mycobacterium Avium Subspecies Paratuberculosis ◽  
Dairy Herds ◽  
Lactation Period ◽  
Serum Samples ◽  
Milk Components ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
Daily Milk Yield

Paratuberculosis is a chronic, progressive enteritis of ruminants, caused by Mycobacterium avium subspecies paratuberculosis. It affects the productivity of infected dairy cows, causing a reduction in the daily milk yield and basic milk components. The aim of the study was to determine the effect of Mycobacterium avium subspecies paratuberculosis on the productivity of dairy cows in two herds. The research materials were serum and milk samples taken from cows from two naturally infected dairy herds. All serum samples were serologically tested using the Mycobacterium paratuberculosis Antibody ELISA Kit by IDEXX—Screening and Verification. Seroprevalence differed between the herds (5.7% and 11.3%). Seroprevalence varied also between the groups of lactation. The highest seroprevalence was found in the first lactation group in both herds. The milk yield evaluation and analysis of the basic milk components’ content (protein and fat total solids) were tested once a month during one lactation period. The content of the basic milk components varied depending on the lactation group, as well as the serological status of the cows. A decrease in milk yield was observed in cows in herds with a higher seroprevalence (>11%). The largest decrease in milk yield and basic milk components was observed in older animals (>three lactations).

Detection of Mycobacterium avium subsp. paratuberculosis (MAP) from Subclinical Caprine Paratuberculosis Cases of Odisha

2020 ◽  
Author(s):  
Sangram Biswal ◽  
Adya Prakash Rath ◽  
Shoor Vir Singh ◽  
Niranjana Sahoo ◽  
Saurabh Gupta ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Animal Health ◽  
Restriction Endonuclease Analysis ◽  
Mycobacterium Avium Subsp ◽  
Serum Samples ◽  
Blood Samples ◽  
Blind Review ◽  
Faecal Samples ◽  
Goat Population ◽  
Mycobacterium Avium Subsp Paratuberculosis

Paratuberculosis is caused by Mycobacterium avium subsp. Paratuberculosis (MAP) and is a chronic, intestinal tract infection in the ruminant sector globally. A total of 122 EDTA mixed blood samples, 121 serum and 16 pooled faecal samples were collected from farms of 4 different districts i.e. Nayagarh, Cuttack, Khordha and Angul and a blind review was conducted at the Animal Health Division, CIRG, Mathura. Microscopic examination of 16 pooled faecal samples revealed +2 reactivity to Acid-Fast Bacilli. All the serum samples were subjected to indirect ELISA. Out of them, 23 (19.01%), 85 (70.25%), shows strongly positive, positive, antibody titre respectively. EDTA blood samples of 23 ELISA-strongly positive were subjected to 413 bp IS900 PCR and 11 (9%) of them were found positive for Mycobacterium avium subsp. Paratuberculosis (MAP). MAP isolates were further subjected to genotyping using 608 bpIS1311 PCR and restriction endonuclease analysis (IS1311 PCR-REA) and 2 (1.64%) of them matched with “Indian Bison Type”. Genotyping of the isolates using IS1311 PCR-REA revealed that goat population of Odisha are primarily infected with “Indian Bison Type” strains.

scholarly journals Mycobacterium avium subsp. paratuberculosis Proteome Changes Profoundly in Milk

Metabolites ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 549
Author(s):  
Kristina J. H. Kleinwort ◽  
Bernhard F. Hobmaier ◽  
Ricarda Mayer ◽  
Christina Hölzel ◽  
Roxane L. Degroote ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Stress Responses ◽  
Dairy Products ◽  
Molecular Mechanisms ◽  
Acid Metabolism ◽  
Mycobacterium Avium Subspecies Paratuberculosis ◽  
Specific Lipids ◽  
Stressful Environment ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
Proteome Changes

Mycobacterium avium subspecies paratuberculosis (MAP) are detectable viable in milk and other dairy products. The molecular mechanisms allowing the adaptation of MAP in these products are still poorly understood. To obtain information about respective adaptation of MAP in milk, we differentially analyzed the proteomes of MAP cultivated for 48 h in either milk at 37 °C or 4 °C or Middlebrook 7H9 broth as a control. From a total of 2197 MAP proteins identified, 242 proteins were at least fivefold higher in abundance in milk. MAP responded to the nutritional shortage in milk with upregulation of 32% of proteins with function in metabolism and 17% in fatty acid metabolism/synthesis. Additionally, MAP upregulated clusters of 19% proteins with roles in stress responses and immune evasion, 19% in transcription/translation, and 13% in bacterial cell wall synthesis. Dut, MmpL4_1, and RecA were only detected in MAP incubated in milk, pointing to very important roles of these proteins for MAP coping with a stressful environment. Dut is essential and plays an exclusive role for growth, MmpL4_1 for virulence through secretion of specific lipids, and RecA for SOS response of mycobacteria. Further, 35 candidates with stable expression in all conditions were detected, which could serve as targets for detection. Data are available via ProteomeXchange with identifier PXD027444.

scholarly journals Molecular characterization of Mycobacterium avium subsp. paratuberculosis (MAP) in dairy cattle of Nepal

2020 ◽  
Vol 5 (6) ◽  
pp. 212-217
Author(s):  
S. Singh ◽  
◽  
I. P. Dhakal ◽  
U. M. Singh ◽  
B. Devkota ◽  
...  
Keyword(s):  
Dairy Cattle ◽  
Molecular Characterization ◽  
Mycobacterium Avium ◽  
Mycobacterium Avium Subspecies Paratuberculosis ◽  
Restriction Endonuclease Analysis ◽  
Cattle Population ◽  
Control And Prevention ◽  
Mycobacterium Avium Subsp Paratuberculosis ◽  
Specific Sequences

Paratuberculosis caused by Mycobacterium avium subspecies paratuberculosis (MAP), is an economically important, endemic in many parts of the globe, and regarded as high prevalent disease of domestic and wild animals, especially ruminants, which is manifest as chronic granulomatous enteritis with decreased milk production, with serious cases resulting in progressive emaciation and death. Understanding the genetic variability of MAP, strains are important in diagnosis, epidemiological investigation, and therefore the formation of strategies for prevention and control of the disease. Thus, this study was designed to grasp the molecular characterization of MAP isolates of Nepal, as pioneer research of this area. Total of 46 MAP isolates obtained from cattle population of three different locations of dairy pocket areas of Chitwan, Nepal were typed using IS1311 polymerase chain reaction-restriction endonuclease analysis (PCR-REA) to research the MAP genotype of Nepal. The extracted DNA samples (n=46) were analyzed for the presence of MAP specific sequences (IS900) using PCR and DNA samples were further subjected to genotype differentiation using IS1311 PCR-REA and IS1311 L2 PCR-REA methods. All the DNA samples were positive for the entire three MAP specific sequences based PCRs. This study revealed that ‘Bison type’ strain is the single most prevalent MAP genotype circulating within the domestic cattle population of Nepal. IS1311 PCR-REA showed that MAP DNA samples of Nepal origin belonged to ‘Bison type’, whereas, IS1311 L2 PCR-REA method showed similarity with "Indian Bison type" and different restriction profiles of ‘Bison type’ genotype as compared to non-Indian strains. The study concludes that in Nepal, "Bison type" MAP stains was prevalent in all the MAP samples obtained from dairy cattle. These results have important epidemiological implications regarding control and prevention of paratuberculosis in Nepal.

SEROPREVALENCE AND MOLECULAR IDENTIFICATION OF MYCOBACTERIUM AVIUM SUBSPECIES PARATUBERCULOSIS IN FARMED GOATS IN CENTRAL-SOUTHERN TAIWAN

2018 ◽  
Vol 44 (04) ◽  
pp. 189-196
Author(s):  
Heng-Ching Lin ◽  
Wai How Chong ◽  
Han Hsiang Huang ◽  
Chi-Chung Chou ◽  
Yi-Lun Tsai ◽  
...  
Keyword(s):  
Molecular Identification ◽  
Mycobacterium Avium ◽  
Mycobacterium Avium Subspecies Paratuberculosis ◽  
Seroprevalence Rate ◽  
Serum Samples ◽  
Fecal Samples ◽  
Identification Rate ◽  
Individual Level ◽  
Seroprevalence Data ◽  
Southern Taiwan

Mycobacterium avium subspecies paratuberculosis (MAP) are Gram positive, aerobic, acid-fast, catalase positive bacteria. The Johne’s disease, caused by MAP, is a wasting disease found in all ruminants including cattle, sheep, goats, deer, camelids and wildlife ruminants. MAP has attracted hygienic attention due to the link between Crohn’s disease (CD) in humans and MAP presence in the gut of patients. The aims of this study are to investigate and monitor the serological prevalence and molecular identification rate of MAP in caprine feces and verify the MAP-negative goat farms in central-southern Taiwan. A total of 8486 blood samples were randomly collected between the years 2011 and 2015 from 321 caprine farms. The serum samples were assessed by commercial ELISA while 3015 fecal samples from 201 anti-MAP antibodies (MAP-Ab) negative herds were further molecularly examined by polymerase chain reaction (PCR) from year 2014 to 2015. The individual seroprevalence rate of caprine MAP in 2011, 2012, 2013, 2014 and 2015 was 0% (0/1032), 0% (0/429), 0% (0/1402), 0.14% (4/2917) and 0.07% (2/2706), respectively. Molecular identification rate of MAP in caprine fecal samples at MAP-Ab negative farms accounted for 0.92% (14/1515) and 0.93 (14/1500), respectively, in the years 2014 and 2015. Meanwhile, there was no association between the MAP seroprevalence and the sampling regions or years at farm or individual level. The seroprevalence data revealed in this study highlighted the rising prevalence of caprine MAP and the link of MAP to farmed ruminant species and its possible implications in hygienic aspects.

scholarly journals 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

2014 ◽  
Vol 21 (5) ◽  
pp. 698-703 ◽  
Author(s):  
Deepanker Tewari ◽  
Ernest Hovingh ◽  
Rick Linscott ◽  
Edmond Martel ◽  
John Lawrence ◽  
...  
Keyword(s):  
Mycobacterium Bovis ◽  
Mycobacterium Avium ◽  
Cross Reactivity ◽  
Johne's Disease ◽  
Antibody Responses ◽  
Johne’S Disease ◽  
Serum Samples ◽  
Content Type ◽  
Fecal Shedding ◽  
Cattle Herds

ABSTRACTVaccination 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 ofMycobacterium aviumsubsp.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 serumM. aviumsubsp.paratuberculosisantibody 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 toM. aviumsubsp.paratuberculosis.M. aviumsubsp.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 againstM. aviumsubsp.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 toMycobacterium bovis, no cross-reactivity was observed.

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