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Rapid and accurate detection of<jats:italic>M<\/jats:italic>.<jats:italic>bovis<\/jats:italic>is fundamental for controlling the disease in animals and humans, and for the proper treatment of patients as one of the first-line anti-TB drug, pyrazinamide, is ineffective against<jats:italic>M<\/jats:italic>.<jats:italic>bovis<\/jats:italic>. Currently, there are no rapid, simplified and low-cost diagnostic methods that can be easily integrated for use in many developing countries. Here, we report the development of a loop-mediated isothermal amplification (LAMP) assay for specific identification of<jats:italic>M<\/jats:italic>.<jats:italic>bovis<\/jats:italic>by targeting the region of difference 4 (RD4), a 12.7 kb genomic region that is deleted solely in<jats:italic>M<\/jats:italic>.<jats:italic>bovis<\/jats:italic>. The assay's specificity was evaluated using 139 isolates comprising 65<jats:italic>M<\/jats:italic>.<jats:italic>bovis<\/jats:italic>isolates, 40<jats:italic>M<\/jats:italic>.<jats:italic>tuberculosis<\/jats:italic>isolates, seven<jats:italic>M<\/jats:italic>.<jats:italic>tuberculosis<\/jats:italic>complex reference strains, 22 non-tuberculous mycobacteria and five other bacteria. The established LAMP detected only<jats:italic>M<\/jats:italic>.<jats:italic>bovis<\/jats:italic>isolates as positive and no false positives were observed using the other mycobacteria and non-mycobacteria tested. Our LAMP assay detected as low as 10 copies of<jats:italic>M<\/jats:italic>.<jats:italic>bovis<\/jats:italic>genomic DNA within 40 minutes. The procedure of LAMP is simple with an incubation at a constant temperature. Results are observed with the naked eye by a color change, and there is no need for expensive equipment. The established LAMP can be used for the detection of<jats:italic>M<\/jats:italic>.<jats:italic>bovis<\/jats:italic>infections in cattle and humans in resource-limited areas.<\/jats:p>","DOI":"10.1371\/journal.pntd.0008996","type":"journal-article","created":{"date-parts":[[2021,1,25]],"date-time":"2021-01-25T18:23:16Z","timestamp":1611598996000},"page":"e0008996","update-policy":"https:\/\/doi.org\/10.1371\/journal.pntd.corrections_policy","source":"Crossref","is-referenced-by-count":29,"title":["Development of a loop-mediated isothermal amplification (LAMP) method for specific detection of Mycobacterium bovis"],"prefix":"10.1371","volume":"15","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-1127-3138","authenticated-orcid":true,"given":"Thoko Flav","family":"Kapalamula","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3447-6483","authenticated-orcid":true,"given":"Jeewan","family":"Thapa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8252-2798","authenticated-orcid":true,"given":"Mwangala Lonah","family":"Akapelwa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kyoko","family":"Hayashida","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4833-5542","authenticated-orcid":true,"given":"Stephen V.","family":"Gordon","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bernard Mudenda","family":"Hang' ombe","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7224-1071","authenticated-orcid":true,"given":"Musso","family":"Munyeme","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Eddie Samuneti","family":"Solo","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4212-4816","authenticated-orcid":true,"given":"Precious","family":"Bwalya","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9251-6482","authenticated-orcid":true,"given":"Mirriam Ethel","family":"Nyenje","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9078-7972","authenticated-orcid":true,"given":"Aki","family":"Tamaru","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1313-1494","authenticated-orcid":true,"given":"Yasuhiko","family":"Suzuki","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9194-1062","authenticated-orcid":true,"given":"Chie","family":"Nakajima","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"340","published-online":{"date-parts":[[2021,1,25]]},"reference":[{"key":"pntd.0008996.ref001","doi-asserted-by":"crossref","first-page":"670","DOI":"10.1038\/nrmicro1472","article-title":"de la Rua-Domenech R, Clifton-Hadley RS, Hewinson RG. 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