Статті в журналах: "Mycobacterium avium subsp. paratublerculosis"

1

Thorel, M. F., M. Krichevsky, and V. Vincent Levy-Frebault. "Numerical Taxonomy of Mycobactin-Dependent Mycobacteria, Emended Description of Mycobacterium avium, and Description of Mycobacterium avium subsp. avium subsp. nov., Mycobacterium avium subsp. paratuberculosis subsp. nov., and Mycobacterium avium subsp. silvaticum subsp. nov." International Journal of Systematic Bacteriology 40, no. 3 (July 1, 1990): 254–60. http://dx.doi.org/10.1099/00207713-40-3-254.

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2

Weiss, Douglas J., Oral A. Evanson, Andreas Moritz, Ming Qi Deng, and Mitchell S. Abrahamsen. "Differential Responses of Bovine Macrophages to Mycobacterium avium subsp. paratuberculosis and Mycobacterium avium subsp. avium." Infection and Immunity 70, no. 10 (October 2002): 5556–61. http://dx.doi.org/10.1128/iai.70.10.5556-5561.2002.

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ABSTRACT Mycobacterium avium subsp. paratuberculosis and Mycobacterium avium subsp. avium are antigenically and genetically similar organisms; however, they differ in their virulence for cattle. M. avium subsp. paratuberculosis causes a chronic intestinal infection leading to a chronic wasting disease termed paratuberculosis or Johne's disease, whereas M. avium subsp. avium causes only a transient infection. We compared the response of bovine monocyte-derived macrophages to ingestion of M. avium subsp. paratuberculosis and M. avium subsp. avium organisms by determining organism survival, superoxide and nitric oxide production, and expression of the cytokines tumor necrosis factor alpha (TNF-α), gamma interferon (IFN-γ), interleukin-8 (IL-8), IL-10, IL-12, and granulocyte-monocyte colony-stimulating factor (GM-CSF). Unlike M. avium subsp. paratuberculosis, macrophages were able to kill approximately half of the M. avium subsp. avium organisms after 96 h of incubation. This difference in killing efficiency was not related to differences in nitric oxide or superoxide production. Compared to macrophages activated with IFN-γ and lipopolysaccharide, macrophages incubated with M. avium subsp. paratuberculosis showed greater expression of IL-10 and GM-CSF (all time points) and IL-8 (72 h) and less expression of IL-12 (72 h), IFN-γ (6 h), and TNF-α (6 h). When cytokine expression by macrophages incubated with M. avium subsp. paratuberculosis was compared to those of macrophages incubated with M. avium subsp. avium, M. avium subsp. paratuberculosis-infected cells showed greater expression of IL-10 (6 and 24 h) and less expression of TNF-α (6 h). Therefore, the combination of inherent resistance to intracellular degradation and suppression of macrophage activation through oversecretion of IL-10 may contribute to the virulence of M. avium subsp. paratuberculosis in cattle.

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Paustian, Michael L., Vivek Kapur, and John P. Bannantine. "Comparative Genomic Hybridizations Reveal Genetic Regions within the Mycobacterium avium Complex That Are Divergent from Mycobacterium avium subsp. paratuberculosis Isolates." Journal of Bacteriology 187, no. 7 (April 1, 2005): 2406–15. http://dx.doi.org/10.1128/jb.187.7.2406-2415.2005.

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ABSTRACT Mycobacterium avium subsp. paratuberculosis is genetically similar to other members of the Mycobacterium avium complex (MAC), some of which are nonpathogenic and widespread in the environment. We have utilized an M. avium subsp. paratuberculosis whole-genome microarray representing over 95% of the predicted coding sequences to examine the genetic conservation among 10 M. avium subsp. paratuberculosis isolates, two isolates each of Mycobacterium avium subsp. silvaticum and Mycobacterium avium subsp. avium, and a single isolate each of both Mycobacterium intracellulare and Mycobacterium smegmatis. Genomic DNA from each isolate was competitively hybridized with DNA from M. avium subsp. paratuberculosis K10, and open reading frames (ORFs) were classified as present, divergent, or intermediate. None of the M. avium subsp. paratuberculosis isolates had ORFs classified as divergent. The two M. avium subsp. avium isolates had 210 and 135 divergent ORFs, while the two M. avium subsp. silvaticum isolates examined had 77 and 103 divergent ORFs. Similarly, 130 divergent ORFs were identified in M. intracellulare. A set of 97 ORFs were classified as divergent or intermediate in all of the nonparatuberculosis MAC isolates tested. Many of these ORFs are clustered together on the genome in regions with relatively low average GC content compared with the entire genome and contain mobile genetic elements. One of these regions of sequence divergence contained genes homologous to a mammalian cell entry (mce) operon. Our results indicate that closely related MAC mycobacteria can be distinguished from M. avium subsp. paratuberculosis by multiple clusters of divergent ORFs.

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4

Stepanova, Hana, Barbora Pavlova, Nikola Stromerova, Petra Ondrackova, Karel Stejskal, Iva Slana, Zbynek Zdrahal, Ivo Pavlik, and Martin Faldyna. "Different immune response of pigs to Mycobacterium avium subsp. avium and Mycobacterium avium subsp. hominissuis infection." Veterinary Microbiology 159, no. 3-4 (October 2012): 343–50. http://dx.doi.org/10.1016/j.vetmic.2012.04.002.

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5

KLANICOVA, B., I. SLANA, H. VONDRUSKOVA, M. KAEVSKA, and I. PAVLIK. "Real-Time Quantitative PCR Detection of Mycobacterium avium Subspecies in Meat Products." Journal of Food Protection 74, no. 4 (April 1, 2011): 636–40. http://dx.doi.org/10.4315/0362-028x.jfp-10-332.

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The aim of this work was to examine various purchased meat products and to find out if any traces of Mycobacterium avium subsp. avium, M. avium subsp. hominissuis, and M. avium subsp. paratuberculosis could be detected in these samples. Analysis of the meat products (raw, cooked, and fermented) was performed using a real-time quantitative PCR (qPCR) method for the detection of specific insertion sequences: duplex qPCR for the detection of IS900 specific for M. avium subsp. paratuberculosis, and triplex qPCR for the detection of IS901 specific for Mycobacterium avium subsp. avium and IS1245 specific for M. avium subsp. hominissuis. Of the 77 analyzed meat samples, 17 (22%) were found to contain M. avium subsp. paratuberculosis DNA, 4 (5%) samples contained Mycobacterium avium subsp. avium DNA, and in 12 (16%) samples M. avium subsp. hominissuis DNA was detected. The concentration of M. avium subsp. paratuberculosis and M. avium subsp. hominissuis DNA in some meat products exceeded 104 genomes per g. Culture examination of these mycobacterial subspecies was negative. By analyzing a range of meat products, we have provided evidence to support the hypothesis that M. avium is present in everyday commodities sold to the general public

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Semret, Makeda, Gary Zhai, Serge Mostowy, Cynthia Cleto, David Alexander, Gerard Cangelosi, Debby Cousins, Desmond M. Collins, Dick van Soolingen, and Marcel A. Behr. "Extensive Genomic Polymorphism within Mycobacterium avium." Journal of Bacteriology 186, no. 18 (September 15, 2004): 6332–34. http://dx.doi.org/10.1128/jb.186.18.6332-6334.2004.

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ABSTRACT We have initiated comparative genomic analysis of Mycobacterium avium subspecies by DNA microarray, uncovering 14 large sequence polymorphisms (LSPs) comprising over 700 kb that distinguish M. avium subsp. avium from M. avium subsp. paratuberculosis. Genes predicted to encode metabolic pathways were overrepresented in the LSPs, and analysis revealed a polymorphism within the mycobactin biosynthesis operon that potentially explains the in vitro mycobactin dependence of M. avium subsp. paratuberculosis.

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LORENCOVA, ALENA, PETRA VASICKOVA, JITKA MAKOVCOVA, and IVA SLANA. "Presence of Mycobacterium avium Subspecies and Hepatitis E Virus in Raw Meat Products." Journal of Food Protection 77, no. 2 (February 1, 2014): 335–38. http://dx.doi.org/10.4315/0362-028x.jfp-13-252.

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Meat and meat products may be the source of various pathogenic and potentially pathogenic agents for humans. We ascertained the occurrence of Mycobacterium avium subsp. paratuberculosis, subsp. avium, and subsp. hominissuis, and hepatitis E virus in retail raw meat products. The DNA of at least one of the target M. avium subspecies was detected in 26 (29.2%) of 89 analyzed samples of meat products. Fourteen (15.7%), 1 (1.1%), and 17 (19.1%) samples contained the DNA of Mycobacterium avium subsp. paratuberculosis, subsp. avium, and subsp. hominissuis, respectively. The number of mycobacterial cells per gram of meat products determined by real-time quantitative PCR ranged from 1.15 × 102 to 6.97 × 103. Mycobacterium chitae and Mycobacterium nonchromogenicum were isolated from three (3.4%) samples. Culture examination was not positive for any M. avium subspecies. Hepatitis E virus RNA was not detected in any of the samples.

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8

Agdestein, Angelika, Tone B. Johansen, Øyvor Kolbjørnsen, Anne Jørgensen, Berit Djønne, and Ingrid Olsen. "A comparative study of Mycobacterium avium subsp. avium and Mycobacterium avium subsp. hominissuis in experimentally infected pigs." BMC Veterinary Research 8, no. 1 (2012): 11. http://dx.doi.org/10.1186/1746-6148-8-11.

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9

Bannantine, J. P., E. Baechler, Q. Zhang, L. Li, and V. Kapur. "Genome Scale Comparison of Mycobacterium avium subsp. paratuberculosis with Mycobacterium avium subsp. avium Reveals Potential Diagnostic Sequences." Journal of Clinical Microbiology 40, no. 4 (April 1, 2002): 1303–10. http://dx.doi.org/10.1128/jcm.40.4.1303-1310.2002.

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10

Svastova, P., I. Pavlik, and M. Bartos. "Rapid differentiation of Mycobacterium avium subsp. avium and Mycobacterium avium subsp. paratuberculosis by amplification of insertion element IS901." Veterinární Medicína 47, No. 5 (March 30, 2012): 117–21. http://dx.doi.org/10.17221/5814-vetmed.

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The aim of this study was to examine the specificity of primers designed to detect the insertion element IS901 commonly used in differentiation of Mycobacterium avium complex strains. This study shows that one of these primers non-specifically anneals to a sequence inside insertion element IS900, specific IS of M. avium subsp. paratuberculosis and to another sequence flanking this element. The resulting non-specific amplicon can be a product of amplification from some M. avium subsp. paratuberculosis strains and can simulate the presence of insertion element IS901 in these strains. However size difference between specific and non-specific amplicons allows such false-positive results to be distinguished. In addition the single PCR allows a rapid and simple differentiation between IS901+ M. avium subsp. avium and M. avium subsp. paratuberculosis strains.

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11

Lehtola, Markku J., Eila Torvinen, Ilkka T. Miettinen, and C. William Keevil. "Fluorescence In Situ Hybridization Using Peptide Nucleic Acid Probes for Rapid Detection of Mycobacterium avium subsp. avium and Mycobacterium avium subsp. paratuberculosis in Potable-Water Biofilms." Applied and Environmental Microbiology 72, no. 1 (January 2006): 848–53. http://dx.doi.org/10.1128/aem.72.1.848-853.2006.

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ABSTRACT Here, we present for the first time a high-affinity peptide nucleic acid (PNA) oligonucleotide sequence for detecting Mycobacterium avium bacteria, including the opportunistically pathogenic subspecies M. avium subsp. avium, M. avium subsp. paratuberculosis, and M. avium subsp. silvaticum, by the fluorescence in situ hybridization (FISH) method. There is evidence that M. avium subsp. avium especially is able to survive and grow in drinking-water biofilms and possibly transmit via drinking water. The designed PNA probe (MAV148) specificity was tested with several bacterial species, including other mycobacteria and mycolic acid-containing bacteria. From the range of bacterial strains tested, only M. avium subsp. avium and M. avium subsp. paratuberculosis strains were hybridized. The PNA FISH method was applied successfully to detect M. avium subsp. avium spiked in water samples and biofilm established within a Propella biofilm reactor fed with potable water from a distribution supply.

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12

Marri, Pradeep Reddy, John P. Bannantine, Michael L. Paustian, and G. Brian Golding. "Lateral gene transfer in Mycobacterium avium subspecies paratuberculosis." Canadian Journal of Microbiology 52, no. 6 (June 1, 2006): 560–69. http://dx.doi.org/10.1139/w06-001.

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Lateral gene transfer is an integral part of genome evolution in most bacteria. Bacteria can readily change the contents of their genomes to increase adaptability to ever-changing surroundings and to generate evolutionary novelty. Here, we report instances of lateral gene transfer in Mycobacterium avium subsp. paratuberculosis, a pathogenic bacteria that causes Johne's disease in cattle. A set of 275 genes are identified that are likely to have been recently acquired by lateral gene transfer. The analysis indicated that 53 of the 275 genes were acquired after the divergence of M. avium subsp. paratuberculosis from M. avium subsp. avium, whereas the remaining 222 genes were possibly acquired by a common ancestor of M. avium subsp. paratuberculosis and M. avium subsp. avium after its divergence from the ancestor of M. tuberculosis complex. Many of the acquired genes were from proteobacteria or soil dwelling actinobacteria. Prominent among the predicted laterally transferred genes is the gene rsbR, a possible regulator of sigma factor, and the genes designated MAP3614 and MAP3757, which are similar to genes in eukaryotes. The results of this study suggest that like most other bacteria, lateral gene transfers seem to be a common feature in M. avium subsp. paratuberculosis and that the proteobacteria contribute most of these genetic exchanges.Key words: mycobacteria, M. avium subsp. paratuberculosis, lateral gene transfer, unique genes, phylogeny.

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13

Beck, A., S. Špičić, M. Butorović-Dujmović, I. Račić, D. Huber, A. Gudan Kurilj, R. Beck, and Ž. Cvetnić. "Mucocutaneous Inflammatory Pseudotumours in Simultaneous Mycobacterium avium subsp. avium and Mycobacterium avium subsp. hominissuis Infection in a Cat." Journal of Comparative Pathology 153, no. 4 (November 2015): 227–30. http://dx.doi.org/10.1016/j.jcpa.2015.07.001.

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14

Kaevska, M., and K. Hruska. " Research on Mycobacterium avium during the period 1995 to 2009." Veterinární Medicína 55, No. 10 (November 10, 2010): 473–82. http://dx.doi.org/10.17221/2942-vetmed.

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Papers on Mycobacterium avium, published between 1995 and 2009 that are indexed in the databases Web of Science<sup>®</sup> (Thomson Reuters) and PubMed (U.S. National Library of Medicine) were analysed and 3377 papers, published by 11 197 authors from 2630 institution and 75 countries were compared. Mycobacterium avium is represented by four subspecies (M. avium subsp. avium, M. avium subsp. silvaticum, M. avium subsp. hominissuis, and M. avium subsp. paratuberculosis). Mycobacteria play an important role as human and animal pathogens and represent a potential risk to consumers as food and environmental pathogens and immunomodulators.

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Wu, Chia-wei, Jeremy Glasner, Michael Collins, Saleh Naser, and Adel M. Talaat. "Whole-Genome Plasticity among Mycobacterium avium Subspecies: Insights from Comparative Genomic Hybridizations." Journal of Bacteriology 188, no. 2 (January 15, 2006): 711–23. http://dx.doi.org/10.1128/jb.188.2.711-723.2006.

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ABSTRACT Infection with Mycobacterium avium subsp. paratuberculosis causes Johne's disease in cattle and is also implicated in cases of Crohn's disease in humans. Another closely related strain, M. avium subsp. avium, is a health problem for immunocompromised patients. To understand the molecular pathogenesis of M. avium subspecies, we analyzed the genome contents of isolates collected from humans and domesticated or wildlife animals. Comparative genomic hybridizations indicated distinct lineages for each subspecies where the closest genomic relatedness existed between M. avium subsp. paratuberculosis isolates collected from human and clinical cow samples. Genomic islands (n = 24) comprising 846 kb were present in the reference M. avium subsp. avium strain but absent from 95% of M. avium subsp. paratuberculosis isolates. Additional analysis identified a group of 18 M. avium subsp. paratuberculosis-associated islands comprising 240 kb that were absent from most of the M. avium subsp. avium isolates. Sequence analysis of DNA regions flanking the genomic islands identified three large inversions in addition to several small inversions that could play a role in regulation of gene expression. Analysis of genes encoded in the genomic islands reveals factors that are probably important for various mechanisms of virulence. Overall, M. avium subsp. avium isolates displayed a higher level of genomic diversity than M. avium subsp. paratuberculosis isolates. Among M. avium subsp. paratuberculosis isolates, those from wildlife animals displayed the highest level of genomic rearrangements that were not observed in other isolates. The presented findings will affect the future design of diagnostics and vaccines for Johne's and Crohn's diseases and provide a model for genomic analysis of closely related bacteria.

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Pate, M., D. Kušar, M. Žolnir-Dovč, and M. Ocepek. "MIRU–VNTR typing of Mycobacterium avium in animals and humans: Heterogeneity of Mycobacterium avium subsp. hominissuis versus homogeneity of Mycobacterium avium subsp. avium strains." Research in Veterinary Science 91, no. 3 (December 2011): 376–81. http://dx.doi.org/10.1016/j.rvsc.2010.10.001.

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Berger, Sven, Dominik Hinz, John P. Bannantine, and J. Frank T. Griffin. "Isolation of High-Affinity Single-Chain Antibodies against Mycobacterium avium subsp. paratuberculosis Surface Proteins from Sheep with Johne's Disease." Clinical and Vaccine Immunology 13, no. 9 (September 2006): 1022–29. http://dx.doi.org/10.1128/cvi.00163-06.

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ABSTRACT Johne's disease, caused by infection with Mycobacterium avium subsp. paratuberculosis, causes significant economic losses to the livestock farming industry. Improved investigative and diagnostic tools—necessary to understand disease processes and to identify subclinical infection—are much sought after. Here, we describe the production of single-chain antibodies with defined specificity for M. avium subsp. paratuberculosis surface proteins. Single-chain antibodies (scFv) were generated from sheep with Johne's disease by cloning heavy-chain and lambda light-chain variable regions and expressing these in fusion with gene III of filamentous phages. Two scFv clones (designated SurfS1.2 and SurfS2.2) were shown to be immunoreactive against M. avium subsp. paratuberculosis surface targets by flow cytometry, and immunoblotting identified specificity for a 34-kDa proteinase-susceptible determinant. Both antibodies were cross-reactive against Mycobacterium avium subsp. avium but nonreactive against Mycobacterium bovis or Mycobacterium phlei cells and were shown to be capable of enriching M. avium subsp. paratuberculosis cells by a factor of approximately 106-fold when employed in magnetic bead separation of mixed Mycobacterium sp. cultures. Further, magnetic bead separation using SurfS1.2 and SurfS2.2 was capable of isolating as few as 103 M. avium subsp. paratuberculosis cells from ovine fecal samples, indicating the diagnostic potential of these reagents. Finally, inclusion of SurfS1.2 or SurfS2.2 in in vitro broth culture with M. avium subsp. paratuberculosis indicated that surface binding activity did not impede bacterial growth, although colony clumping was prevented. These results are discussed in terms of the potential use of single-chain phage display monoclonal antibodies as novel diagnostic reagents.

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18

Harris, N. Beth, and Raúl G. Barletta. "Mycobacterium avium subsp. paratuberculosisin Veterinary Medicine." Clinical Microbiology Reviews 14, no. 3 (July 1, 2001): 489–512. http://dx.doi.org/10.1128/cmr.14.3.489-512.2001.

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SUMMARY Mycobacterium avium subsp. paratuberculosis (basonym M. paratuberculosis) is the etiologic agent of a severe gastroenteritis in ruminants known as Johne's disease. Economic losses to the cattle industry in the United States are staggering, reaching $1.5 billion annually. A potential pathogenic role in humans in the etiology of Crohn's disease is under investigation. In this article, we review the epidemiology, pathogenesis, diagnostics, and disease control measures of this important veterinary pathogen. We emphasize molecular genetic aspects including the description of markers used for strain identification, diagnostics, and phylogenetic analysis. Recent important advances in the development of animal models and genetic systems to study M. paratuberculosis virulence determinants are also discussed. We conclude with proposals for the applications of these models and recombinant technology to the development of diagnostic, control, and therapeutic measures.

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Pillai, Shreekumar R., Bhushan M. Jayarao, Jennifer D. Gummo, Eric C. Hue, Deepanker Tiwari, Judith R. Stabel, and Robert H. Whitlock. "Identification and sub-typing of Mycobacterium avium subsp. paratuberculosis and Mycobacterium avium subsp. avium by randomly amplified polymorphic DNA." Veterinary Microbiology 79, no. 3 (April 2001): 275–84. http://dx.doi.org/10.1016/s0378-1135(00)00358-8.

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Woo, Seng-Ryong, Raúl G. Barletta, and Charles J. Czuprynski. "Extracellular ATP Is Cytotoxic to Mononuclear Phagocytes but Does Not Induce Killing of Intracellular Mycobacterium avium subsp. paratuberculosis." Clinical and Vaccine Immunology 14, no. 9 (July 18, 2007): 1078–83. http://dx.doi.org/10.1128/cvi.00166-07.

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ABSTRACT Mycobacterium avium subsp. paratuberculosis is the etiologic agent of Johne's disease, a chronic granulomatous enteritis in ruminants. ATP has been reported to induce cell death of macrophages and killing of Mycobacterium species in human and murine macrophages. In this study we investigated the short-term effect of ATP on the viability of M. avium subsp. paratuberculosis-infected bovine mononuclear phagocytes and the bacilli within them. Addition of 5 mM ATP to M. avium subsp. paratuberculosis-infected bovine monocytes resulted in 50% cytotoxicity of bovine monocytes at 24 h. Addition of 2′(3′)-O-(4-benzoylbenzoyl) ATP triethylammonium salt (Bz-ATP), which is a longer-lived ATP homologue and purinergic receptor agonist, significantly increased the uptake of YO-PRO, which is a marker for membrane pore activation by P2X receptors. Addition of Bz-ATP also stimulated lactate dehydrogenase release and caspase-3 activity in infected bovine monocytes. Neither ATP nor Bz-ATP reduced the survival of M. avium subsp. paratuberculosis in bovine mononuclear phagocytes. Likewise, addition of ATP or Bz-ATP was cytotoxic to murine macrophage cell lines (RAW 264.7 and J774A.1 cells) but did not affect the intracellular survival of M. avium subsp. paratuberculosis, nor were the numbers of viable Mycobacterium avium subsp. avium or Mycobacterium bovis BCG cells altered in bovine mononuclear phagocytes or J774A.1 cells following ATP or Bz-ATP treatment. These data suggest that extracellular ATP does not induce the killing of intracellular M. avium subsp. paratuberculosis in bovine mononuclear phagocytes.

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Dhand, Navneet K., Jenny-Ann L. M. L. Toribio, and Richard J. Whittington. "Adsorption of Mycobacterium avium subsp. paratuberculosis to Soil Particles." Applied and Environmental Microbiology 75, no. 17 (June 26, 2009): 5581–85. http://dx.doi.org/10.1128/aem.00557-09.

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ABSTRACT Attachment of Mycobacterium avium subsp. paratuberculosis to soil particles could increase their availability to farm animals, as well as influence the transportation of M. avium subsp. paratuberculosis to water sources. To investigate the possibility of such attachment, we passed a known quantity of M. avium subsp. paratuberculosis through chromatography columns packed with clay soil, sandy soil, pure silica, clay-silica mixture, or clay-silica complexes and measured the organisms recovered in the eluent using culture or quantitative PCR. Experiments were repeated using buffer at a range of pH levels with pure silica to investigate the effect of pH on M. avium subsp. paratuberculosis attachment. Linear mixed-model analyses were conducted to compare the proportional recovery of M. avium subsp. paratuberculosis in the eluent between different substrates and pH levels. Of the organisms added to the columns, 83 to 100% were estimated to be retained in the columns after adjustment for those retained in empty control columns. The proportions recovered were significantly different across different substrates, with the retention being significantly greater (P < 0.05) in pure substrates (silica and clay-silica complexes) than in soil substrates (clay soil and sandy soil). However, there were no significant differences in the retention of M. avium subsp. paratuberculosis between silica and clay-silica complexes or between clay soil and sandy soil. The proportion retained decreased with increasing pH in one of the experiments, indicating greater adsorption of M. avium subsp. paratuberculosis to soil particles at an acidic pH (P < 0.05). The results suggest that under experimental conditions M. avium subsp. paratuberculosis adsorbs to a range of soil particles, and this attachment is influenced by soil pH.

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Bermudez, Luiz E., Mary Petrofsky, Sandra Sommer, and Raúl G. Barletta. "Peyer's Patch-Deficient Mice Demonstrate That Mycobacterium avium subsp. paratuberculosis Translocates across the Mucosal Barrier via both M Cells and Enterocytes but Has Inefficient Dissemination." Infection and Immunity 78, no. 8 (May 24, 2010): 3570–77. http://dx.doi.org/10.1128/iai.01411-09.

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ABSTRACT Mycobacterium avium subsp. paratuberculosis, the agent of Johne's disease, infects ruminant hosts by translocation through the intestinal mucosa. A number of studies have suggested that M. avium subsp. paratuberculosis interacts with M cells in the Peyer's patches of the small intestine. The invasion of the intestinal mucosa by M. avium subsp. paratuberculosis and Mycobacterium avium subsp. hominissuis, a pathogen known to interact with intestinal cells, was compared. M. avium subsp. paratuberculosis was capable of invading the mucosa, but it was significantly less efficient at dissemination than M. avium subsp. hominissuis. B-cell knockout (KO) mice, which lack Peyer's patches, were used to demonstrate that M. avium subsp. paratuberculosis enters the intestinal mucosa through enterocytes in the absence of M cells. In addition, the results indicated that M. avium subsp. paratuberculosis had equal abilities to cross the mucosa in both Peyer's patch and non-Peyer's patch segments of normal mice. M. avium subsp. paratuberculosis was also shown to interact with epithelial cells by an α5β1 integrin-independent pathway. Upon translocation, dendritic cells ingest M. avium subsp. paratuberculosis, but this process does not lead to efficient dissemination of the infection. In summary, M. avium subsp. paratuberculosis interacts with the intestinal mucosa by crossing both Peyer's patches and non-Peyer's patch areas but does not translocate or disseminate efficiently.

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Speer, C. A., M. Cathy Scott, John P. Bannantine, W. Ray Waters, Yasuyuki Mori, Robert H. Whitlock, and Shigetoshi Eda. "A Novel Enzyme-Linked Immunosorbent Assay for Diagnosis of Mycobacterium avium subsp. paratuberculosis Infections (Johne's Disease) in Cattle." Clinical and Vaccine Immunology 13, no. 5 (May 2006): 535–40. http://dx.doi.org/10.1128/cvi.13.5.535-540.2006.

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ABSTRACT Enzyme-linked immunosorbent assays (ELISAs) for the diagnosis of Johne's disease (JD), caused by Mycobacterium avium subsp. paratuberculosis, were developed using whole bacilli treated with formaldehyde (called WELISA) or surface antigens obtained by treatment of M. avium subsp. paratuberculosis bacilli with formaldehyde and then brief sonication (called SELISA). ELISA plates were coated with either whole bacilli or sonicated antigens and tested for reactivity against serum obtained from JD-positive and JD-negative cattle or from calves experimentally inoculated with M. avium subsp. paratuberculosis, Mycobacterium avium subsp. avium, or Mycobacterium bovis. Because the initial results obtained from the WELISA and SELISA were similar, most of the subsequent experiments reported herein were performed using the SELISA method. To optimize the SELISA test, various concentrations (3.7 to 37%) of formaldehyde and intervals of sonication (2 to 300 s) were tested. With an increase in formaldehyde concentration and a decreased interval of sonication, there was a concomitant decrease in nonspecific binding by the SELISA. SELISAs prepared by treating M. avium subsp. paratuberculosis with 37% formaldehyde and then a 2-s burst of sonication produced the greatest difference (7×) between M. avium subsp. paratuberculosis-negative and M. avium subsp. paratuberculosis-positive serum samples. The diagnostic sensitivity and specificity for JD by the SELISA were greater than 95%. The SELISA showed subspecies-specific detection of M. avium subsp. paratuberculosis infections in calves experimentally inoculated with M. avium subsp. paratuberculosis or other mycobacteria. Based on diagnostic sensitivity and specificity, the SELISA appears superior to the commercial ELISAs routinely used for the diagnosis of JD.

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KAEVSKA, M., J. STERBA, J. SVOBODOVA, and I. PAVLIK. "Mycobacterium avium subsp. avium and Mycobacterium neoaurum detection in an immunocompromised patient." Epidemiology and Infection 142, no. 4 (July 10, 2013): 882–85. http://dx.doi.org/10.1017/s0950268813001660.

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SUMMARYNon-tuberculous mycobacteria are increasingly described as infectious agents in immunocompromised patients. A 17-year-old male patient suffering from secondary non-Hodgkin's lymphoma and treated with chemotherapeutic agents was admitted to hospital due to pleuropneumonia. Mycobacterium neoaurum was cultured repeatedly from his sputum and, Mycobacterium avium subsp. avium (M. a. avium) was detected by IS901 qPCR from detached fragments of his intestinal mucosa. We attempted to determine the possible sources of infection by analysing environmental samples from the closed oncology unit and conventional unit in the hospital, and from the patient's home residence and places which he frequented. The environment of the patient harboured mycobacteria (41 isolates in total); however, M. neoaurum was not recovered. M. a. avium was detected by qPCR in the environmental samples from a small flock of hens kept by his neighbour. Although it was not confirmed by DNA fingerprinting methods, the M. a. avium infection could have been acquired through the eating of incompletely cooked eggs.

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Turenne, Christine Y., Desmond M. Collins, David C. Alexander, and Marcel A. Behr. "Mycobacterium avium subsp. paratuberculosis and M. avium subsp. avium Are Independently Evolved Pathogenic Clones of a Much Broader Group of M. avium Organisms." Journal of Bacteriology 190, no. 7 (February 1, 2008): 2479–87. http://dx.doi.org/10.1128/jb.01691-07.

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ABSTRACT Mycobacterium avium comprises organisms that share the same species designation despite considerable genomic and phenotypic variability. To determine the degree and nature of variability between subspecies and strains of M. avium, we used multilocus sequencing analysis, studying 56 genetically diverse strains of M. avium that included all described subspecies. In total, 8,064 bp of sequence from 10 gene loci were studied, with 205 (2.5%) representing variable positions. The majority (149/205) of these variations were found among M. avium subsp. hominissuis organisms. Recombination was also evident in this subspecies. In contrast, there was comparatively little variability and no evidence of recombination within the pathogenic subspecies, M. avium subsp. paratuberculosis, M. avium subsp. avium, and M. avium subsp. silvaticum. Phylogenetic analysis showed that M. avium subsp. avium and M. avium subsp. silvaticum strains clustered together on one branch, while a distinct branch defined M. avium subsp. paratuberculosis organisms. Despite the independent origin of these pathogenic subspecies, an analysis of their rates of nonsynonymous (dN) to synonymous (dS) substitutions showed increased dN/dS ratios for both: 0.67 for M. avium subsp. paratuberculosis and 0.50 for M. avium subsp. avium/M. avium subsp. silvaticum, while the value was 0.08 for M. avium subsp. hominissuis organisms. In conclusion, M. avium subsp. hominissuis represents a diverse group of organisms from which two pathogenic clones (M. avium subsp. paratuberculosis and M. avium subsp. avium/M. avium subsp. silvaticum) have evolved independently.

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Bannantine, John P., Michael L. Paustian, W. Ray Waters, Judith R. Stabel, Mitchell V. Palmer, Lingling Li, and Vivek Kapur. "Profiling Bovine Antibody Responses to Mycobacterium avium subsp. paratuberculosis Infection by Using Protein Arrays." Infection and Immunity 76, no. 2 (November 26, 2007): 739–49. http://dx.doi.org/10.1128/iai.00915-07.

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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.

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Weiss, Douglas J., Oral A. Evanson, David J. McClenahan, Mitchell S. Abrahamsen, and Bruce K. Walcheck. "Regulation of Expression of Major Histocompatibility Antigens by Bovine Macrophages Infected withMycobacterium avium subsp. paratuberculosis orMycobacterium avium subsp. avium." Infection and Immunity 69, no. 2 (February 1, 2001): 1002–8. http://dx.doi.org/10.1128/iai.69.2.1002-1008.2001.

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ABSTRACT Mycobacterium avium subsp. paratuberculosisand Mycobacterium avium subsp. avium are antigenically and genetically very similar organisms; however, they differ markedly in their virulence for cattle. We evaluated the capacity of bovine macrophages infected with M. aviumsubsp. paratuberculosis or M. avium subsp.avium to express major histocompatibility complex (MHC) class I and class II antigens on their surface and to interact with primed autologous lymphocytes. Our results indicate that infection of bovine macrophages with M. avium subsp.paratuberculosis promoted the downregulation of MHC class I and class II molecules on the macrophage surface within 24 and 12 h, respectively. Alternatively, MHC class II expression by M. avium subsp. avium-infected macrophages was not detected until 24 h after infection, and the magnitude of the decrease was smaller. Decreased MHC class I expression by M. avium subsp. avium-infected macrophages was not detected. Unlike M. avium subsp.paratuberculosis-infected macrophages, M. aviumsubsp. avium-infected macrophages upregulated MHC class I and class II expression after activation by gamma interferon or tumor necrosis factor alpha. Further, M. avium subsp.avium-infected macrophages were lysed by primed autologous lymphocytes, whereas M. avium subsp.paratuberculosis-infected macrophages were not. Overall, the results support the hypothesis that the difference in the virulence of M. avium subsp. paratuberculosis andM. avium subsp. avium for cattle is dependent on a difference in the capacity of the organisms to suppress mycobacterial antigen presentation to T lymphocytes.

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Secott, T. E., T. L. Lin, and C. C. Wu. "Fibronectin Attachment Protein Homologue Mediates Fibronectin Binding by Mycobacterium avium subsp.paratuberculosis." Infection and Immunity 69, no. 4 (April 1, 2001): 2075–82. http://dx.doi.org/10.1128/iai.69.4.2075-2082.2001.

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ABSTRACT Attachment of Mycobacterium avium subsp.paratuberculosis to host tissue and penetration of mucosal surfaces are pivotal events in the pathogenesis of Johne's disease. Fibronectin (FN) binding is required for attachment and internalization of several mycobacteria by epithelial cells in vitro. The objective of this study was to further characterize the FN binding activity ofM. avium subsp. paratuberculosis. Although the bacteria bound FN poorly at pH above 7, brief acid pretreatment greatly enhanced FN binding within the pH range (3 to 10) studied. A 4.6-kbp fragment from an M. avium subsp.paratuberculosis genomic library was found to contain a 1,107-bp open reading frame that shows very high nucleotide sequence identity with that of the FN attachment protein (FAP) gene of M. avium subsp. avium. Pretreatment of FN with an FN-binding peptide from M. avium subsp. aviumFAP abolished FN binding, indicating that M. avium subsp.paratuberculosis binds FN in a FAP-dependent manner. Pretreatment of M. avium subsp.paratuberculosis with anti-FAP immunoglobulin G did not abrogate FN binding; blocking occurred only when anti-FAP was added together with FN. FAP was detected by immunofluorescence only in lipid-extracted M. avium subsp.paratuberculosis. Western blotting and immunoelectron microscopy revealed that FAP is located near the interior of the cell envelope of M. avium subsp. paratuberculosis. The results indicate that a FAP homologue mediates the attachment of FN to M. avium subsp. paratuberculosis. Further, given the subcellular location of FAP, it is considered that this protein operates at the terminus of a coordinated FN binding system in the cell envelope of M. avium subsp.paratuberculosis.

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Waters, W. R., B. J. Nonnecke, M. V. Palmer, S. Robbe-Austermann, J. P. Bannantine, J. R. Stabel, D. L. Whipple, et al. "Use of Recombinant ESAT-6:CFP-10 Fusion Protein for Differentiation of Infections of Cattle by Mycobacterium bovis and by M. avium subsp. avium and M. avium subsp. paratuberculosis." Clinical Diagnostic Laboratory Immunology 11, no. 4 (July 2004): 729–35. http://dx.doi.org/10.1128/cdli.11.4.729-735.2004.

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ABSTRACT Immunological diagnosis of Mycobacterium bovis infection of cattle is often confounded by cross-reactive responses resulting from exposure to other mycobacterial species, especially Mycobacterium avium. Early secretory antigenic target 6 (ESAT-6) and culture filtrate protein 10 (CFP-10) are dominant gamma interferon (IFN-γ)-inducing antigens of tuberculous mycobacteria, and they are absent from many environmental nontuberculous mycobacteria. Because M. avium exposure is the primary confounding factor in the diagnosis of M. bovis-infected animals, in vitro responses to a recombinant ESAT-6:CFP-10 (rESAT-6:CFP-10) fusion protein by blood leukocytes from cattle naturally exposed to M. avium or experimentally challenged with Mycobacterium avium subsp. avium or Mycobacterium avium subsp. paratuberculosis were compared to responses by M. bovis-infected cattle. Responses to heterogeneous mycobacterial antigens (i.e., purified protein derivatives [PPDs] and whole-cell sonicates [WCSs]) were also evaluated. Tumor necrosis factor alpha (TNF-α), IFN-γ, and nitric oxide responses by M. bovis-infected cattle to rESAT-6:CFP-10 exceeded (P < 0.05) the corresponding responses by cattle naturally sensitized to M. avium. Experimental infection with M. bovis, M. avium, or M. avium subsp. paratuberculosis induced significant (P < 0.05) IFN-γ and nitric oxide production to WCS and PPD antigens, regardless of the mycobacterial species used for the preparation of the antigen. Responses to homologous crude antigens generally exceeded responses to heterologous antigens. Nitric oxide and IFN-γ responses to rESAT-6:CFP-10 by blood leukocytes from M. bovis-infected calves exceeded (P < 0.05) the corresponding responses of noninfected, M. avium-infected, and M. avium subsp. paratuberculosis-infected calves. Despite the reported potential for secretion of immunogenic ESAT-6 and CFP-10 proteins by M. avium and M. avium subsp. paratuberculosis, it appears that use of the rESAT-6:CFP-10 fusion protein will be useful for the detection of tuberculous cattle in herds with pre-existing sensitization to M. avium and/or M. avium subsp. paratuberculosis.

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Foddai, Antonio, Christopher T. Elliott, and Irene R. Grant. "Maximizing Capture Efficiency and Specificity of Magnetic Separation for Mycobacterium avium subsp. paratuberculosis Cells." Applied and Environmental Microbiology 76, no. 22 (September 17, 2010): 7550–58. http://dx.doi.org/10.1128/aem.01432-10.

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ABSTRACT In order to introduce specificity for Mycobacterium avium subsp. paratuberculosis prior to a phage amplification assay, various magnetic-separation approaches, involving either antibodies or peptides, were evaluated in terms of the efficiency of capture (expressed as a percentage) of M. avium subsp. paratuberculosis cells and the percentage of nonspecific binding by other Mycobacterium spp. A 50:50 mixture of MyOne Tosylactivated Dynabeads coated with the chemically synthesized M. avium subsp. paratuberculosis-specific peptides biotinylated aMp3 and biotinylated aMptD (i.e., peptide-mediated magnetic separation [PMS]) proved to be the best magnetic-separation approach for achieving 85 to 100% capture of M. avium subsp. paratuberculosis and minimal (<1%) nonspecific recovery of other Mycobacterium spp. (particularly if beads were blocked with 1% skim milk before use) from broth samples containing 103 to 104 CFU/ml. When PMS was coupled with a recently optimized phage amplification assay and used to detect M. avium subsp. paratuberculosis in 50-ml volumes of spiked milk, the mean 50% limit of detection (LOD50) was 14.4 PFU/50 ml of milk (equivalent to 0.3 PFU/ml). This PMS-phage assay represents a novel, rapid method for the detection and enumeration of viable M. avium subsp. paratuberculosis organisms in milk, and potentially other sample matrices, with results available within 48 h.

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Castellanos, Elena, Alicia Aranaz, Katherine A. Gould, Richard Linedale, Karen Stevenson, Julio Alvarez, Lucas Dominguez, Lucia de Juan, Jason Hinds, and Tim J. Bull. "Discovery of Stable and Variable Differences in the Mycobacterium avium subsp. paratuberculosis Type I, II, and III Genomes by Pan-Genome Microarray Analysis." Applied and Environmental Microbiology 75, no. 3 (December 1, 2008): 676–86. http://dx.doi.org/10.1128/aem.01683-08.

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ABSTRACT Mycobacterium avium subsp. paratuberculosis is an important animal pathogen widely disseminated in the environment that has also been associated with Crohn's disease in humans. Three M. avium subsp. paratuberculosis genomotypes are recognized, but genomic differences have not been fully described. To further investigate these potential differences, a 60-mer oligonucleotide microarray (designated the MAPAC array), based on the combined genomes of M. avium subsp. paratuberculosis (strain K-10) and Mycobacterium avium subsp. hominissuis (strain 104), was designed and validated. By use of a test panel of defined M. avium subsp. paratuberculosis strains, the MAPAC array was able to identify a set of large sequence polymorphisms (LSPs) diagnostic for each of the three major M. avium subsp. paratuberculosis types. M. avium subsp. paratuberculosis type II strains contained a smaller genomic complement than M. avium subsp. paratuberculosis type I and M. avium subsp. paratuberculosis type III genomotypes, which included a set of genomic regions also found in M. avium subsp. hominissuis 104. Specific PCRs for genes within LSPs that differentiated M. avium subsp. paratuberculosis types were devised and shown to accurately screen a panel (n = 78) of M. avium subsp. paratuberculosis strains. Analysis of insertion/deletion region INDEL12 showed deletion events causing a reduction in the complement of mycobacterial cell entry genes in M. avium subsp. paratuberculosis type II strains and significantly altering the coding of a major immunologic protein (MPT64) associated with persistence and granuloma formation. Analysis of MAPAC data also identified signal variations in several genomic regions, termed variable genomic islands (vGIs), suggestive of transient duplication/deletion events. vGIs contained significantly low GC% and were immediately flanked by insertion sequences, integrases, or short inverted repeat sequences. Quantitative PCR demonstrated that variation in vGI signals could be associated with colony growth rate and morphology.

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Whan, Lynne, Hywel J. Ball, Irene R. Grant, and Michael T. Rowe. "Occurrence of Mycobacterium avium subsp. paratuberculosis in Untreated Water in Northern Ireland." Applied and Environmental Microbiology 71, no. 11 (November 2005): 7107–12. http://dx.doi.org/10.1128/aem.71.11.7107-7112.2005.

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ABSTRACT Mycobacterium avium subsp. paratuberculosis is the known cause of Johne's disease of both domestic and wild ruminants and has been implicated as a possible cause of Crohn's disease in humans. The organism is shed in the feces of infected animals and can survive for protracted periods in the environment and hence could be present in catchment areas receiving agricultural runoff. A limited survey was undertaken in Northern Ireland to test for M. avium subsp. paratuberculosis in untreated water entering nine water treatment works (WTWs) over a 1-year period. Three detection methods were employed, viz., immunomagnetic separation-PCR and culture on Herrold's egg yolk medium (HEYM) and BACTEC 12B medium, the latter both supplemented with mycobactins. Of the 192 untreated water samples tested, 15 (8%) tested M. avium subsp. paratuberculosis positive by one or more of the three detection methods. M. avium subsp. paratuberculosis was successfully isolated from eight untreated water samples, three by BACTEC culture and five by culture on HEYM. Although the highest incidence of M. avium subsp. paratuberculosis was found in spring, overall, there was no statistically significant difference between the seasons. No significant correlation was found between numbers of coliforms or fecal coliforms and the presence of M. avium subsp. paratuberculosis. In general, a higher incidence of M. avium subsp. paratuberculosis was found in untreated water entering those WTWs that had a high mean water pH value over the sampling period. This work indicates the need to determine the efficacy of water treatment processes to either kill or remove M. avium subsp. paratuberculosis from untreated water and the possible risks posed by contact with recreational water sources.

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Stanley, Emma C., Richard J. Mole, Rebecca J. Smith, Sarah M. Glenn, Michael R. Barer, Michael McGowan, and Catherine E. D. Rees. "Development of a New, Combined Rapid Method Using Phage and PCR for Detection and Identification of Viable Mycobacterium paratuberculosis Bacteria within 48 Hours." Applied and Environmental Microbiology 73, no. 6 (January 26, 2007): 1851–57. http://dx.doi.org/10.1128/aem.01722-06.

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ABSTRACT The FASTPlaqueTB assay is an established diagnostic aid for the rapid detection of Mycobacterium tuberculosis from human sputum samples. Using the FASTPlaqueTB assay reagents, viable Mycobacterium avium subsp. paratuberculosis cells were detected as phage plaques in just 24 h. The bacteriophage used does not infect M. avium subsp. paratuberculosis alone, so to add specificity to this assay, a PCR-based identification method was introduced to amplify M. avium subsp. paratuberculosis-specific sequences from the DNA of the mycobacterial cell detected by the phage. To give further diagnostic information, a multiplex PCR method was developed to allow simultaneous amplification of either M. avium subsp. paratuberculosis or M. tuberculosis complex-specific sequences from plaque samples. Combining the plaque PCR technique with the phage-based detection assay allowed the rapid and specific detection of viable M. avium subsp. paratuberculosis in milk samples in just 48 h.

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Schrenzel, Mark, Melissa Nicolas, Carmel Witte, Rebecca Papendick, Tammy Tucker, Laura Keener, Meg Sutherland-Smith, et al. "Molecular epidemiology of Mycobacterium avium subsp. avium and Mycobacterium intracellulare in captive birds." Veterinary Microbiology 126, no. 1-3 (January 2008): 122–31. http://dx.doi.org/10.1016/j.vetmic.2007.06.016.

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Alexander, David C., Christine Y. Turenne, and Marcel A. Behr. "Insertion and Deletion Events That Define the Pathogen Mycobacterium avium subsp. paratuberculosis." Journal of Bacteriology 191, no. 3 (November 21, 2008): 1018–25. http://dx.doi.org/10.1128/jb.01340-08.

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ABSTRACT Mycobacterium avium comprises genetically related yet phenotypically distinct subspecies. Consistent with their common origin, whole-genome sequence comparisons have revealed extensive synteny among M. avium organisms. However, the sequenced strains also display numerous regions of heterogeneity that likely contribute to the diversity of the individual subspecies. Starting from a phylogenetic framework derived by multilocus sequence analysis, we examined the distribution of 25 large sequence polymorphisms across a panel of genetically defined M. avium strains. This distribution was most variable among M. avium subsp. hominissuis isolates. In contrast, M. avium subsp. paratuberculosis strains exhibited a characteristic profile, with all isolates containing a set of genomic insertions absent from other M. avium strains. The emergence of the pathogen from its putative M. avium subsp. hominissuis ancestor entailed the acquisition of approximately 125 kb of novel genetic material, followed by a second phase, characterized by reductive genomics. One genomic deletion is common to all isolates while additional deletions distinguish two major lineages of M. avium subsp. paratuberculosis. For the average strain, these losses total at least 38 kb (sheep lineage) to 90 kb (cattle lineage). This biphasic pattern of evolution, characterized by chromosomal gene acquisition with subsequent gene loss, describes the emergence of M. avium subsp. paratuberculosis and may serve as a general model for the origin of pathogenic mycobacteria.

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Möbius, P., P. Lentzsch, I. Moser, L. Naumann, G. Martin, and H. Köhler. "Comparative macrorestriction and RFLP analysis of Mycobacterium avium subsp. avium and Mycobacterium avium subsp. hominissuis isolates from man, pig, and cattle." Veterinary Microbiology 117, no. 2-4 (October 2006): 284–91. http://dx.doi.org/10.1016/j.vetmic.2006.05.005.

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Secott, T. E., T. L. Lin, and C. C. Wu. "Mycobacterium avium subsp. paratuberculosis Fibronectin Attachment Protein Facilitates M-Cell Targeting and Invasion through a Fibronectin Bridge with Host Integrins." Infection and Immunity 72, no. 7 (July 2004): 3724–32. http://dx.doi.org/10.1128/iai.72.7.3724-3732.2004.

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ABSTRACT Efficient attachment and ingestion of Mycobacterium avium subsp. paratuberculosis by cultured epithelial cells requires the expression of a fibronectin (FN) attachment protein homologue (FAP-P) which mediates FN binding by M. avium subsp. paratuberculosis. Invasion of Peyer's patches by M. avium subsp. paratuberculosis occurs through M cells, which, unlike other intestinal epithelial cells, express integrins on their luminal faces. We sought to determine if the interaction between FAP-P of M. avium subsp. paratuberculosis and soluble FN enabled targeting and invasion of M cells by M. avium subsp. paratuberculosis in vivo via these surface integrins. Wild-type and antisense FAP-P mutant M. avium subsp. paratuberculosis strains were injected alone or coinjected with blocking peptides or antibodies into murine gut loops, and immunofluorescence microscopy was performed to assess targeting and invasion of M cells by M. avium subsp. paratuberculosis. Nonopsonized M. avium subsp. paratuberculosis preferentially invaded M cells in murine gut loops. M-cell invasion was enhanced 2.6-fold when M. avium subsp. paratuberculosis was pretreated with FN. Invasion of M cells by the antisense FAP-P mutant of M. avium subsp. paratuberculosis was reduced by 77 to 90% relative to that observed for the control strains. Peptides corresponding to the RGD and synergy site integrin recognition regions of FN blocked M. avium subsp. paratuberculosis invasion of M cells by 75 and 45%, respectively, whereas the connecting segment 1 peptide was noninhibitory. Antibodies against the α5, αV, β1, and β3 integrin subunits inhibited M-cell invasion by 52 to 73%. The results indicate that targeting and invasion of M cells by M. avium subsp. paratuberculosis in vivo is mediated primarily by the formation of an FN bridge formed between FAP-P of M. avium subsp. paratuberculosis and integrins on M cells.

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Ssekitoleko, Judah, Lonzy Ojok, Ahmed Abd El Wahed, Joseph Erume, Ahmad Amanzada, ElSagad Eltayeb, Kamal H. Eltom, and Julius Boniface Okuni. "Mycobacterium avium subsp. paratuberculosis Virulence: A Review." Microorganisms 9, no. 12 (December 19, 2021): 2623. http://dx.doi.org/10.3390/microorganisms9122623.

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To propose a solution for control of Mycobacterium avium subsp. paratuberculosis (MAP) infections in animals as well as in humans, and develop effective prevention, diagnostic and treatment strategies, it is essential to understand the molecular mechanisms of MAP pathogenesis. In the present review, we discuss the mechanisms utilised by MAP to overcome the host defense system to achieve the virulence status. Putative MAP virulence genes are mentioned and their probable roles in view of other mycobacteria are discussed. This review provides information on MAP strain diversity, putative MAP virulence factors and highlights the knowledge gaps regarding MAP virulence mechanisms that may be important in control and prevention of paratuberculosis.

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Wang, Joyce, Jalal Moolji, Alex Dufort, Alfredo Staffa, Pilar Domenech, Michael B. Reed, and Marcel A. Behr. "Iron Acquisition in Mycobacterium avium subsp. paratuberculosis." Journal of Bacteriology 198, no. 5 (December 28, 2015): 857–66. http://dx.doi.org/10.1128/jb.00922-15.

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ABSTRACTMycobacterium aviumsubsp.paratuberculosisis a host-adapted pathogen that evolved from the environmental bacteriumM. aviumsubsp.hominissuisthrough gene loss and gene acquisition. Growth ofM. aviumsubsp.paratuberculosisin the laboratory is enhanced by supplementation of the media with the iron-binding siderophore mycobactin J. Here we examined the production of mycobactins by related organisms and searched for an alternative iron uptake system inM. aviumsubsp.paratuberculosis. Through thin-layer chromatography and radiolabeled iron-uptake studies, we showed thatM. aviumsubsp.paratuberculosisis impaired for both mycobactin synthesis and iron acquisition. Consistent with these observations, we identified several mutations, including deletions, inM. aviumsubsp.paratuberculosisgenes coding for mycobactin synthesis. Using a transposon-mediated mutagenesis screen conditional on growth without myobactin, we identified a potential mycobactin-independent iron uptake system on aM. aviumsubsp.paratuberculosis-specific genomic island, LSPP15. We obtained a transposon (Tn) mutant with a disruption in the LSPP15 geneMAP3776cfor targeted study. The mutant manifests increased iron uptake as well as intracellular iron content, with genes downstream of the transposon insertion (MAP3775ctoMAP3772c[MAP3775-2c]) upregulated as the result of a polar effect. As an independent confirmation, we observed the same iron uptake phenotypes by overexpressingMAP3775-2cin wild-typeM. aviumsubsp.paratuberculosis. These data indicate that the horizontally acquired LSPP15 genes contribute to iron acquisition byM. aviumsubsp.paratuberculosis, potentially allowing the subsequent loss of siderophore production by this pathogen.IMPORTANCEMany microbes are able to scavenge iron from their surroundings by producing iron-chelating siderophores. One exception isMycobacterium aviumsubsp.paratuberculosis, a fastidious, slow-growing animal pathogen whose growth needs to be supported by exogenous mycobacterial siderophore (mycobactin) in the laboratory. Data presented here demonstrate that, compared to other closely relatedM. aviumsubspecies, mycobactin production and iron uptake are different inM. aviumsubsp.paratuberculosis, and these phenotypes may be caused by numerous deletions in its mycobactin biosynthesis pathway. Using a genomic approach, supplemented by targeted genetic and biochemical studies, we identified that LSPP15, a horizontally acquired genomic island, may encode an alternative iron uptake system. These findings shed light on the potential physiological consequence of horizontal gene transfer inM. aviumsubsp.paratuberculosisevolution.

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40

Secott, T. E., T. L. Lin, and C. C. Wu. "Fibronectin Attachment Protein Is Necessary for Efficient Attachment and Invasion of Epithelial Cells by Mycobacterium avium subsp. paratuberculosis." Infection and Immunity 70, no. 5 (May 2002): 2670–75. http://dx.doi.org/10.1128/iai.70.5.2670-2675.2002.

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ABSTRACT Attachment and ingestion of Mycobacterium avium subsp. paratuberculosis by two epithelial cell lines were enhanced by soluble fibronectin (FN). Peptide blocking of the FN attachment protein (FAP-P) inhibited the internalization of M. avium subsp. paratuberculosis. Disruption of FAP-P expression significantly reduced attachment and ingestion of M. avium subsp. paratuberculosis by T-24 and Caco-2 cells. The results indicate that the interaction between FN and FAP-P facilitates attachment and internalization of M. avium subsp. paratuberculosis by epithelial cells.

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41

Dupont, Chris, Keith Thompson, Cord Heuer, Brigitte Gicquel, and Alan Murray. "Identification and characterization of an immunogenic 22 kDa exported protein of Mycobacterium avium subspecies paratuberculosis." Journal of Medical Microbiology 54, no. 11 (November 1, 2005): 1083–92. http://dx.doi.org/10.1099/jmm.0.46163-0.

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An exported 22 kDa putative lipoprotein was identified in an alkaline phosphatase gene fusion library of Mycobacterium avium subsp. paratuberculosis and expressed in Mycobacterium smegmatis. The full nucleic acid sequence of the gene encoding P22 was determined and the ORF was cloned into a mycobacterial expression vector, enabling full-length P22 to be produced as a C-terminal polyhistidine-tagged protein in M. smegmatis. N-terminal sequencing of the recombinant protein confirmed cleavage of a signal sequence. Native P22 was detected in culture supernatants and cell sonicates of M. avium subsp. paratuberculosis strain 316F using rabbit antibody raised to recombinant P22. Investigation of the presence of similar genes in other mycobacterial species revealed that the gene was present in Mycobacterium avium subsp. avium and similar genes existed in Mycobacterium intracellulare and Mycobacterium scrofulaceum. Database searches showed that P22 belonged to the LppX/LprAFG family of mycobacterial lipoproteins also found in Mycobacterium leprae and in members of the Mycobacterium tuberculosis complex. P22 shared less than 75 % identity to these proteins. Recombinant P22 was able to elicit interferon-gamma secretion in blood from eight of a group of nine sheep vaccinated with a live attenuated strain of M. avium subsp. paratuberculosis (strain 316F) compared to none from a group of five unvaccinated sheep. Antibody to P22 was detected by Western blot analysis in 10 out of 11 vaccinated sheep, in two out of two clinically affected cows and in 11 out of 13 subclinically infected cows.

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42

Uddin, Reaz, Bushra Siraj, Muhammad Rashid, Ajmal Khan, Sobia Ahsan Halim, and Ahmed Al-Harrasi. "Genome Subtraction and Comparison for the Identification of Novel Drug Targets against Mycobacterium avium subsp. hominissuis." Pathogens 9, no. 5 (May 12, 2020): 368. http://dx.doi.org/10.3390/pathogens9050368.

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Mycobacterium avium complex (MAC) is a major cause of non-tuberculous pulmonary and disseminated diseases worldwide, inducing bronchiectasis, and affects HIV and immunocompromised patients. In MAC, Mycobacterium avium subsp. hominissuis is a pathogen that infects humans and mammals, and that is why it is a focus of this study. It is crucial to find essential drug targets to eradicate the infections caused by these virulent microorganisms. The application of bioinformatics and proteomics has made a significant impact on discovering unique drug targets against the deadly pathogens. One successful bioinformatics methodology is the use of in silico subtractive genomics. In this study, the aim was to identify the unique, non-host and essential protein-based drug targets of Mycobacterium avium subsp. hominissuis via in silico a subtractive genomics approach. Therefore, an in silico subtractive genomics approach was applied in which complete proteome is subtracted systematically to shortlist potential drug targets. For this, the complete dataset of proteins of Mycobacterium avium subsp. hominissuis was retrieved. The applied subtractive genomics method, which involves the homology search between the host and the pathogen to subtract the non-druggable proteins, resulted in the identification of a few prioritized potential drug targets against the three strains of M. avium subsp. Hominissuis, i.e., MAH-TH135, OCU466 and A5. In conclusion, the current study resulted in the prioritization of vital drug targets, which opens future avenues to perform structural as well as biochemical studies on predicted drug targets against M. avium subsp. hominissuis.

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Huntley, J. F., J. R. Stabel, M. L. Paustian, T. A. Reinhardt, and J. P. Bannantine. "Expression Library Immunization Confers Protection against Mycobacterium avium subsp. paratuberculosis Infection." Infection and Immunity 73, no. 10 (October 2005): 6877–84. http://dx.doi.org/10.1128/iai.73.10.6877-6884.2005.

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ABSTRACT Currently, paratuberculosis vaccines are comprised of crude whole-cell preparations of Mycobacterium avium subsp. paratuberculosis. Although effective in reducing clinical disease and fecal shedding, these vaccines have severe disadvantages as well, including seroconversion of vaccinated animals and granulomatous lesions at the site of vaccination. DNA vaccines can offer an alternative approach that may be safer and elicit more protective responses. In an effort to identify protective M. avium subsp. paratuberculosis sequences, a genomic DNA expression library was generated and subdivided into pools of clones (∼1,500 clones/pool). The clone pools were evaluated to determine DNA vaccine efficacy by immunizing mice via gene gun delivery and challenging them with live, virulent M. avium subsp. paratuberculosis. Four clone pools resulted in a significant reduction in the amount of M. avium subsp. paratuberculosis recovered from mouse tissues compared to mice immunized with other clone pools and nonvaccinated, infected control mice. One of the protective clone pools was further partitioned into 10 clone arrays of 108 clones each, and four clone arrays provided significant protection from both spleen and mesenteric lymph node colonization by M. avium subsp. paratuberculosis. The nucleotide sequence of each clone present in the protective pools was determined, and coding region functions were predicted by computer analysis. Comparison of the protective clone array sequences implicated 26 antigens that may be responsible for protection in mice. This study is the first study to demonstrate protection against M. avium subsp. paratuberculosis infection with expression library immunization.

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Hostetter, J., R. Kagan, and E. Steadham. "Opsonization Effects on Mycobacterium avium subsp. paratuberculosis-Macrophage Interactions." Clinical Diagnostic Laboratory Immunology 12, no. 6 (June 2005): 793–96. http://dx.doi.org/10.1128/cdli.12.6.793-796.2005.

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ABSTRACT High antibody titers in ruminants infected with Mycobacterium avium subsp. paratuberculosis correlates with disease progression. Effects of humoral responses during mycobacterial infection are not completely understood. This study suggests that activation status may be an important factor in determining macrophage ability to limit proliferation of opsonized M. avium subsp. paratuberculosis.

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45

Olsen, Ingrid, Liv J. Reitan, and Harald G. Wiker. "Distinct Differences in Repertoires of Low-Molecular-Mass Secreted Antigens of Mycobacterium aviumComplex and Mycobacterium tuberculosis." Journal of Clinical Microbiology 38, no. 12 (2000): 4453–58. http://dx.doi.org/10.1128/jcm.38.12.4453-4458.2000.

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Antigens in a 4-week-old culture filtrate (CF) ofMycobacterium avium subsp. avium were separated in sodium dodecyl sulfate-polyacrylamide gel electrophoresis and identified by Western blotting. The culture had minimal lysis of bacilli, giving a CF preparation consisting mainly of secreted proteins. Comparison with a similar CF of Mycobacterium tuberculosis with almost no contamination with intracellular proteins showed the presence of cross-reactive antigens homologous to the four components of the antigen 85 complex, as well as MPT32. These were major constituents of the M. avium subsp.avium CF. In addition, there were several low-molecular-mass bands (<15 kDa) in both species that did not cross-react with polyclonal and polyvalent rabbit antibodies in Western blotting. Furthermore, these bands were not detected in corresponding sonicate preparations, indicating high localization indexes, which is typical of soluble secreted proteins. A 14-kDa protein was selected for purification and more detailed characterization. The N-terminal amino acid sequence was determined, and a matching gene was found within the genomic sequence of M. avium subsp. avium which was highly homologous to Rv0455c of M. tuberculosis. The gene encoded a signal peptide typical of secreted mycobacterial proteins. A rabbit antiserum was raised against the purified protein, and the antigen was demonstrated by Western blotting in CFs of M. avium subsp. avium, Mycobacterium aviumsubsp. paratuberculosis, Mycobacterium intracellulare, and Mycobacterium scrofulaceum but was not detected in M. tuberculosis. This is a new example of a highly homologous gene being differentially expressed by different mycobacterial species.

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46

Abukhalid, Norah, Sabrina Islam, Robert Ndzeidze, and Luiz E. Bermudez. "Mycobacterium avium Subsp. hominissuis Interactions with Macrophage Killing Mechanisms." Pathogens 10, no. 11 (October 22, 2021): 1365. http://dx.doi.org/10.3390/pathogens10111365.

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Non-tuberculosis mycobacteria (NTM) are ubiquitously found throughout the environment. NTM can cause respiratory infections in individuals with underlying lung conditions when inhaled, or systemic infections when ingested by patients with impaired immune systems. Current therapies can be ineffective at treating NTM respiratory infections, even after a long course or with multidrug treatment regimens. NTM, such as Mycobacterium avium subspecies hominissuis (M. avium), is an opportunistic pathogen that shares environments with ubiquitous free-living amoeba and other environmental hosts, possibly their evolutionary hosts. It is highly likely that interactions between M. avium and free-living amoeba have provided selective pressure on the bacteria to acquire survival mechanisms, which are also used against predation by macrophages. In macrophages, M. avium resides inside phagosomes and has been shown to exit it to infect other cells. M. avium’s adaptation to the hostile intra-phagosomal environment is due to many virulence mechanisms. M. avium is able to switch the phenotype of the macrophage to be anti-inflammatory (M2). Here, we have focused on and discussed the bacterial defense mechanisms associated with the intra-phagosome phase of infection. M. avium possesses a plethora of antioxidant enzymes, including the superoxide dismutases, catalase and alkyl hydroperoxide reductase. When these defenses fail or are overtaken by robust oxidative burst, many other enzymes exist to repair damage incurred on M. avium proteins, including thioredoxin/thioredoxin reductase. Finally, M. avium has several oxidant sensors that induce transcription of antioxidant enzymes, oxidation repair enzymes and biofilm- promoting genes. These expressions induce physiological changes that allow M. avium to survive in the face of leukocyte-generated oxidative stress. We will discuss the strategies used by M. avium to infect human macrophages that evolved during its evolution from free-living amoeba. The more insight we gain about M. avium’s mode of pathogenicity, the more targets we can have to direct new anti-virulence therapies toward.

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47

Marsh, Ian B., John P. Bannantine, Michael L. Paustian, Mark L. Tizard, Vivek Kapur, and Richard J. Whittington. "Genomic Comparison of Mycobacterium avium subsp. paratuberculosis Sheep and Cattle Strains by Microarray Hybridization." Journal of Bacteriology 188, no. 6 (March 15, 2006): 2290–93. http://dx.doi.org/10.1128/jb.188.6.2290-2293.2006.

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ABSTRACT Microarray-based comparisons of three Mycobacterium avium subsp. paratuberculosis isolates, including one sheep strain and two cattle strains, identified three large genomic deletions in the sheep strain, totaling 29,208 bp and involving 24 open reading frames. These deletions may help explain some of the differences in pathogenicity and host specificity observed between the cattle and sheep strains of Mycobacterium avium subsp. paratuberculosis.

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48

Dadvar, Ehsan. "Genomic and Phenotypic Variability of Mycobaterium avium Subspecies." McGill Science Undergraduate Research Journal 5, no. 1 (March 31, 2010): 11–17. http://dx.doi.org/10.26443/msurj.v5i1.67.

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Introduction: Mycobacterium avium complex consists of M. intracellulare and the subspecies of M. avium subsp. avium, M. avium subsp. paratuberculosis and M. avium subsp. hominissuis. Despite their taxonomic relationship, these subspecies are organisms with distinct phenotypes, ranging from environmental bacteria that cause infections in immunocompromised hosts to pathogens targeting birds and ruminants. The reasons for the variable pathogenicity and host range of M. avium subspecies are not known. We hypothesize that genotypic differences between M. avium subsp. avium and M. avium subsp. paraturberculosis can explain different pathogenic outcomes. Methods: We used tri-genomic comparisons to look for DNA fragments unique to each subspecies. We also used an acute model of mouse infection to determine different phenotypic outcomes in response to infection with different Mycobacterium subspecies. results: Through tri-genomic comparisons we identified genetic regions of interest that may contain genes to explain phenotypic or pathogenic differences among subspecies. In an 8 week course infection, mice infected with M. avium subspecies avium had the highest bacterial burden in their spleens and livers. at the same time, mice infected with M. avium subspecies paraturberculosis had the lowest bacterial burden. discussion: Differences in the genomic sequences of the M. avium subspecies suggests that these sequences encode pathogenic factors. Consequently, this study shows that the sequencing of M. avium subspecies genomes can be useful for predicting and explaining variation in pathogenesis.

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Tasara, T., and R. Stephan. "Development of an F57 Sequence-Based Real-Time PCR Assay for Detection of Mycobacterium avium subsp. paratuberculosis in Milk." Applied and Environmental Microbiology 71, no. 10 (October 2005): 5957–68. http://dx.doi.org/10.1128/aem.71.10.5957-5968.2005.

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ABSTRACT A light cycler-based real-time PCR (LC-PCR) assay that amplifies the F57 sequence of Mycobacterium avium subsp. paratuberculosis was developed. This assay also includes an internal amplification control template to monitor the amplification conditions in each reaction. The targeted F57 sequence element is unique for M.avium subsp. paratuberculosis and is not known to exist in any other bacterial species. The assay specificity was demonstrated by evaluation of 10 known M. avium subsp. paratuberculosis isolates and 33 other bacterial strains. The LC-PCR assay has a broad linear range (2 × 101 to 2 ×106 copies) for quantitative estimation of the number of M. avium subsp. paratuberculosis F57 target copies in positive samples. To maximize the assay's detection sensitivity, an efficient strategy for isolation of M. avium subsp. paratuberculosis DNA from spiked milk samples was also developed. The integrated procedure combining optimal M. avium subsp. paratuberculosis DNA isolation and real-time PCR detection had a reproducible detection limit of about 10 M. avium subsp. paratuberculosis cells per ml when a starting sample volume of 10 ml of M. avium subsp. paratuberculosis-spiked milk was analyzed. The entire process can be completed within a single working day and is suitable for routine monitoring of milk samples for M. avium subsp. paratuberculosis contamination. The applicability of this protocol for naturally contaminated milk was also demonstrated using milk samples from symptomatic M. avium subsp. paratuberculosis-infected cows, as well as pooled samples from a dairy herd with a confirmed history of paratuberculosis.

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Wu, Chia-wei, Shelly K. Schmoller, Sung Jae Shin, and Adel M. Talaat. "Defining the Stressome of Mycobacterium avium subsp. paratuberculosis In Vitro and in Naturally Infected Cows." Journal of Bacteriology 189, no. 21 (August 10, 2007): 7877–86. http://dx.doi.org/10.1128/jb.00780-07.

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ABSTRACT Mycobacterium avium subsp. paratuberculosis causes an enteric infection in cattle, with a great impact on the dairy industry in the United States and worldwide. Characterizing the gene expression profile of M. avium subsp. paratuberculosis exposed to different stress conditions, or shed in cow feces, could improve our understanding of the pathogenesis of M. avium subsp. paratuberculosis. In this report, the stress response of M. avium subsp. paratuberculosis on a genome-wide level (stressome) was defined for the first time using DNA microarrays. Expression data analysis revealed unique gene groups of M. avium subsp. paratuberculosis that were regulated under in vitro stressors while additional groups were regulated in the cow samples. Interestingly, acidic pH induced the regulation of a large number of genes (n = 597), suggesting the high sensitivity of M. avium subsp. paratuberculosis to acidic environments. Generally, responses to heat shock, acidity, and oxidative stress were similar in M. avium subsp. paratuberculosis and Mycobacterium tuberculosis, suggesting common pathways for mycobacterial defense against stressors. Several sigma factors (e.g., sigH and sigE) were differentially coregulated with a large number of genes depending on the type of each stressor. Subsequently, we analyzed the virulence of six M. avium subsp. paratuberculosis mutants with inactivation of differentially regulated genes using a murine model of paratuberculosis. Both bacterial and histopathological examinations indicated the attenuation of all gene mutants, especially those selected based on their expression in the cow samples (e.g., lipN). Overall, the employed approach profiled mycobacterial genetic networks triggered by variable stressors and identified a novel set of putative virulence genes. A similar approach could be applied to analyze other intracellular pathogens.

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