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Background: Mycobacterium avium subspecies paratuberculosis (MAP) is an infectious factor recently found in association with multiple sclerosis (MS) in Sardinia., Objectives: The objectives of this study were to confirm this association and evaluate its role in clinical features., Methods: A total of 436 patients and 264 healthy controls (HCs) were included. We examined the blood of each individual for MAPDNA and MAP2694 antibodies using IS900-specific PCR and ELISA, respectively. Differences in MAP presence between the MS group and HCs were evaluated. In MS patients, we considered: gender, age, age at onset, duration of disease, course, EDSS, therapy, relapse/steroids at study time, and oligoclonal bands (OBs)., Results: MAPDNA and MAP2694 antibodies were detected in 68 MS and six HCs (p = 1.14 × 10(-11)), and 123 MS and 10 HCs (p = 2.59 × 10(-23)), respectively. OBs were found with reduced frequency in MAP-positive patients (OR = 0.52; p = 0.02). MAP2694 antibodies were detected more in patients receiving MS treatments (OR = 2.26; p = 0.01), and MAPDNA in subjects on steroids (OR = 2.65; p = 0.02)., Conclusion: Our study confirmed the association of MAP and MS in Sardinia. The low OB frequency in MAP patients suggests a peripheral role as a trigger in autoimmunity. MAP positivity might be influenced by steroids and MS therapy. Studies in other populations are needed to confirm the role of MAP in MS.

Introduction: Recent findings propose that Mycobacterium avium subsp. paratuberculosis (MAP) infection could act as risk factor in favoring multiple sclerosis (MS) progression. SLC11A1 is a gene associated with mycobacterial survival in the host and it may be involved in the induction and maintenance of autoimmune disease., Methodology: In this preliminary study, 100 MS patients and 100 healthy controls (HCs) from Sardinia were enrolled. Eight single nucleotide polymorphisms (SNPs) in the SLC11A gene were searched by PCR RFLP-genotyping. IS900 specie specific PCR was undertaken to search for MAP presence. Indirect ELISA was performed to asses if MS patients displayed a stronger humoral response against MAP2694 protein compared to the HCs., Results: Only rs2276631 SNP was associated with MS. MAP DNA was detected in 23 out of 100 MS patients (23%) and in 7 out of 100 HCs (7%). A strong humoral response against MAP2694 protein was detected in 36% of MS patients and only in 3% of HCs. A correlation between ELISA sero-positivity and the rs2276631 SNP was also found., Conclusion: Our preliminary results suggest that the Sardinian population might be prone to develop autoimmune disease due to polymorphisms in immunomodulating the SLC11A1 gene, which is important in the immune response against intracellular bacteria such as MAP.

Sardinia acts as an ideal setting for multiple sclerosis (MS) studies because its prevalence of MS is one of the highest worldwide. Several pathogens have been investigated amongst 119 Sardinian MS patients and 117 healthy controls to determine whether they might have a role in triggering MS in genetically predisposed individuals. Mycobacterium avium subsp. paratuberculosis (MAP) and Epstein Barr virus DNA were detected in 27.5% and 17.3%, respectively, of the MS patients. Moreover an extremely high humoral immune response against MAP recombinant protein MAP FprB (homologous to human myelin P0) was observed, whereas no significant results were found against Mycobacterium tuberculosis FprA and Helicobacter pylori HP986 protein.

Mycobacterium avium subsp. paratuberculosis (Map) is the causative agent of Johne's disease, a chronic inflammation of ruminants' intestine. Recent studies have linked Map to type I Diabetes mellitus (T1DM). We searched the presence of antibodies against two specific proteins of Map (MptD and MAP3738c) in sera of patients affected by T1DM and type II Diabetes mellitus (T2DM). MptD protein (MAP3733c) has been recognized as a Map virulent factor whereas MAP3738c has not yet been studied. Both proteins are encoded by genes belonging to a Map specific pathogenicity island. Forty three T1DM patients' sera, 56 T2DM patients' sera and 48 healthy subjects' sera were screened by ELISA to evaluate the immunoresponse against MptD or MAP3738c recombinant proteins. Results showed a positive response to both proteins in T1DM patients whereas no difference with controls was found for T2DM patients. Results suggest a potential relation between T1DM and the bacterial infection., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Mycobacterium avium subsp. paratuberculosis (MAP) infection is highly spread in the ruminant herds of Sardinia, in the Western Mediterranean. The objective of this study was to investigate prevalence of MAP infection in association with Multiple Sclerosis (MS) using clinical specimen from patients and controls. We analyzed samples for the presence of MAP specific DNA and to demonstrate humoral response to a MAP protein (MAP2694), a predicted homologue of the T-cell receptor gamma-chain/complement component 1 of the host. We found presence of MAP DNA in 42% of the MS patients and an extremely significant humoral immune response revealed by the MS patients against the MAP protein. In our opinion, this is the first report that significantly associates MAP infection with MS. Further studies will be required to confirm if MAP could be one of the triggers of MS, according to the molecular mimicry theory, in susceptible (and genetically at risk) individuals.

Here we present the development of a specific DNA detection method using fluorescent semiconductor quantum dots (QDs) and magnetic beads (MBs) for fast detection of Mycobacterium spp., dispensing with the need for DNA amplification. Two biotinylated oligonucleotide probes were used to recognize and detect specific complementary mycobacterial target DNA through a sandwich hybridization reaction. Cadmium selenite QDs conjugated with streptavidin and species-specific probes were used to produce a fluorescent signal. MBs conjugated with streptavidin and a genus-specific probe were used to isolate and concentrate the DNA targets. The application of the proposed method to isolated bacteria produced the expected result in all cases. The minimum detection limit of the assay was defined as 12.5 ng of DNA diluted in a sample volume of 20 microl. In order to obtain an indication of the method's performance with clinical samples, we applied the optimized assay to the detection of Mycobacterium tuberculosis in DNA isolated from bronchoalveolar lavage specimens from patients with tuberculosis and Mycobacterium avium subsp. paratuberculosis in DNA isolated from feces and paraffin-embedded tissues in comparison with culture, Ziehl-Neelsen staining, and real-time PCR. The concordance of these methods compared to the proposed method with regard to positive and negative samples varied between 53.84% and 87.23% and between 84.61% and 100%, respectively. The overall accuracy of the QD assay compared to real-time PCR was 70 to 90% depending on the type of clinical material. The proposed diagnostic assay offers a simple, rapid, specific, and cost-effective method for direct detection and identification of mycobacterial DNA in clinical samples.