Dataset Information

Novel secreted antigens of Mycobacterium paratuberculosis as serodiagnostic biomarkers for Johne's disease in cattle.

ABSTRACT: Johne's disease is a chronic gastroenteritis of cattle caused by Mycobacterium avium subsp. paratuberculosis that afflicts 40% of dairy herds worldwide. M. avium subsp. paratuberculosis-infected cattle can remain asymptomatic for years while transmitting the pathogen via fecal contamination and milk. Current serodiagnosis with enzyme-linked immunosorbent assays (ELISAs) fails to detect asymptomatic M. avium subsp. paratuberculosis-infected cattle due to the use of poorly defined antigens and knowledge gaps in our understanding of M. avium subsp. paratuberculosis components eliciting pathogen-specific immune responses. We set out to (i) define a subset of proteins that contain putative antigenic targets and (ii) screen these antigen pools for immunogens relevant in detecting infection. To accomplish our first objective, we captured and resolved M. avium subsp. paratuberculosis-secreted proteins using a 2-step fractionation method and reverse-phase liquid chromatography to identify 162 unique proteins, of which 66 had not been previously observed in M. avium subsp. paratuberculosis culture filtrates. Subsequent screening of M. avium subsp. paratuberculosis-secreted proteins showed four antigens, of which one or more reacted on immunoblotting with individual serum samples from 35 M. avium subsp. paratuberculosis-infected cows. Moreover, these novel antigens reacted with sera from 6 low M. avium subsp. paratuberculosis shedders and 3 fecal-culture-positive cows labeled as ELISA seronegative. The specificity of these antigens was demonstrated using negative-control sera from uninfected calves (n = 5) and uninfected cows (n = 5), which did not react to any of these antigens in immunoblotting. As three of the four antigens are novel, their characterization and incorporation into an ELISA-based format will aid in detecting asymptomatic cattle in early or subclinical stages of disease.

SUBMITTER: Facciuolo A

PROVIDER: S-EPMC3889510 | biostudies-literature | 2013 Dec

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Novel secreted antigens of Mycobacterium paratuberculosis as serodiagnostic biomarkers for Johne's disease in cattle.

Facciuolo Antonio A Kelton David F DF Mutharia Lucy M LM

Clinical and vaccine immunology : CVI 20131002 12

Johne's disease is a chronic gastroenteritis of cattle caused by Mycobacterium avium subsp. paratuberculosis that afflicts 40% of dairy herds worldwide. M. avium subsp. paratuberculosis-infected cattle can remain asymptomatic for years while transmitting the pathogen via fecal contamination and milk. Current serodiagnosis with enzyme-linked immunosorbent assays (ELISAs) fails to detect asymptomatic M. avium subsp. paratuberculosis-infected cattle due to the use of poorly defined antigens and kno ...[more]

PMID: 24089453

Similar Datasets

Project description:Johne's disease, a chronic gastrointestinal inflammatory disease caused by Mycobacterium avium subspecies paratuberculosis, is endemic in dairy cattle and other ruminants worldwide and remains a challenge to diagnose using traditional serological methods. Given the close phylogenetic relationship between M. avium subsp. paratuberculosis and the human pathogen Mycobacterium tuberculosis, here, we applied a whole-proteome M. tuberculosis protein array to identify seroreactive and diagnostic M. avium subsp. paratuberculosis antigens. A genome-scale pairwise analysis of amino acid identity levels between orthologous proteins in M. avium subsp. paratuberculosis and M. tuberculosis showed an average of 62% identity, with more than half the orthologous proteins sharing >75% identity. Analysis of the M. tuberculosis protein array probed with sera from M. avium subsp. paratuberculosis-infected cattle showed antibody binding to 729 M. tuberculosis proteins, with 58% of them having ?70% identity to M. avium subsp. paratuberculosis orthologs. The results showed that only 4 of the top 40 seroreactive M. tuberculosis antigens were orthologs of previously reported M. avium subsp. paratuberculosis antigens, revealing the existence of a large number of previously unrecognized candidate diagnostic antigens. Enzyme-linked immunosorbent assay (ELISA) testing of 20 M. avium subsp. paratuberculosis recombinant proteins, representing reactive and nonreactive M. tuberculosis orthologs, further confirmed that the M. tuberculosis array has utility as a screening tool for identifying candidate antigens for Johne's disease diagnostics. Additional ELISA testing of field serum samples collected from dairy herds around the United States revealed that MAP2942c had the strongest seroreactivity with Johne's disease-positive samples. Collectively, our studies have considerably expanded the number of candidate M. avium subsp. paratuberculosis proteins with potential utility in the next generation of rationally designed Johne's disease diagnostic assays.

Project description:Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of Johne's disease in ruminants, which has important health consequences for dairy cattle. The Regional Dairy Quality Management Alliance (RDQMA) project is a multistate research program involving MAP isolates taken from three intensively studied commercial dairy farms in the northeastern United States, which emphasized longitudinal data collection of both MAP isolates and animal health in three regional dairy herds for a period of about 7 years. This paper reports the results of a pan-GWAS analysis involving 318 MAP isolates and dairy cow Johne's disease phenotypes, taken from these three farms. Based on our highly curated accessory gene count the pan-GWAS analysis identified several MAP genes associated with bovine Johne's disease phenotypes scored from these three farms, with some of the genes having functions suggestive of possible cause/effect relationships to these phenotypes. This paper reports a pan-genomic comparative analysis between MAP and Mycobacterium tuberculosis, assessing functional Gene Ontology category enrichments between these taxa. Finally, we also provide a population genomic perspective on the effectiveness of herd isolation, involving closed dairy farms, in preventing MAP inter-farm cross infection on a micro-geographic scale.IMPORTANCE Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of Johne's disease in ruminants, which has important health consequences for dairy cattle, and enormous economic consequences for the dairy industry. Understanding which genes in this bacterium are correlated with key disease phenotypes can lead to functional experiments targeting these genes and ultimately lead to improved control strategies. This study represents a rare example of a prolonged longitudinal study of dairy cattle where the disease was measured and the bacteria were isolated from the same cows. The genome sequences of over 300 MAP isolates were analyzed for genes that were correlated with a wide range of Johne's disease phenotypes. A number of genes were identified that were significantly associated with several aspects of the disease and suggestive of further experimental follow-up.

Project description:Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of Johne's disease (JD), a chronic intestinal inflammatory disease of cattle and other ruminants. JD has a high herd prevalence and causes serious animal health problems and significant economic loss in domesticated ruminants throughout the world. Since serological detection of MAP infected animals during the early stages of infection remains challenging due to the low sensitivity of extant assays, we screened 180 well-characterized serum samples using a whole proteome microarray from Mycobacterium tuberculosis (MTB), a close relative of MAP. Based on extensive testing of serum and milk samples, fecal culture and qPCR for direct detection of MAP, the samples were previously assigned to one of 4 groups: negative low exposure (n = 30, NL); negative high exposure (n = 30, NH); fecal positive, ELISA negative (n = 60, F+E-); and fecal positive, ELISA positive (n = 60, F+E+). Of the 740 reactive proteins, several antigens were serologically recognized early but not late in infection, suggesting a complex and dynamic evolution of the MAP humoral immune response during disease progression. Ordinal logistic regression models identified a subset of 47 candidate proteins with significantly different normalized intensity values (p<0.05), including 12 in the NH and 23 in F+E- groups, suggesting potential utility for the early detection of MAP infected animals. Next, the diagnostic utility of four MAP orthologs (MAP1569, MAP2942c, MAP2609, and MAP1272c) was assessed and reveal moderate to high diagnostic sensitivities (range 48.3% to 76.7%) and specificity (range 96.7% to 100%), with a combined 88.3% sensitivity and 96.7% specificity. Taken together, the results of our analyses have identified several candidate MAP proteins of potential utility for the early detection of MAP infection, as well individual MAP proteins that may serve as the foundation for the next generation of well-defined serological diagnosis of JD in cattle.

Project description:Mycobacterium avium subsp. paratuberculosis (MAP) is the etiologic agent of Johne's disease in ruminants causing chronic diarrhea, malnutrition, and muscular wasting. Neonates and young animals are infected primarily by the fecal-oral route. MAP attaches to, translocates via the intestinal mucosa, and is phagocytosed by macrophages. The ensuing host cellular immune response leads to granulomatous enteritis characterized by a thick and corrugated intestinal wall. We review various tissue culture systems, ileal loops, and mice, goats, and cattle used to study MAP pathogenesis. MAP can be detected in clinical samples by microscopy, culturing, PCR, and an enzyme-linked immunosorbent assay. There are commercial vaccines that reduce clinical disease and shedding, unfortunately, their efficacies are limited and may not engender long-term protective immunity. Moreover, the potential linkage with Crohn's disease and other human diseases makes MAP a concern as a zoonotic pathogen. Potential therapies with anti-mycobacterial agents are also discussed. The completion of the MAP K-10 genome sequence has greatly improved our understanding of MAP pathogenesis. The analysis of this sequence has identified a wide range of gene functions involved in virulence, lipid metabolism, transcriptional regulation, and main metabolic pathways. We also review the transposons utilized to generate random transposon mutant libraries and the recent advances in the post-genomic era. This includes the generation and characterization of allelic exchange mutants, transcriptomic analysis, transposon mutant banks analysis, new efforts to generate comprehensive mutant libraries, and the application of transposon site hybridization mutagenesis and transposon sequencing for global analysis of the MAP genome. Further analysis of candidate vaccine strains development is also provided with critical discussions on their benefits and shortcomings, and strategies to develop a highly efficacious live-attenuated vaccine capable of differentiating infected from vaccinated animals.

Dataset Information

Novel secreted antigens of Mycobacterium paratuberculosis as serodiagnostic biomarkers for Johne's disease in cattle.

Publications

Novel secreted antigens of Mycobacterium paratuberculosis as serodiagnostic biomarkers for Johne's disease in cattle.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets