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Establishment of an on-site diagnostic procedure for detection of orf virus from oral lesions of Japanese serows (Capricornis crispus) by loop-mediated isothermal amplification
Orf virus infection has been prevalent continuously in the population of wild Japanese serows (Capricornis crispus), goat-like grazing cloven-hoofed mammal species that live mainly in mountainous areas of Japan. Currently, definitive diagnosis of infection requires time-consuming laboratory work. To...
Autores principales: | , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
The Japanese Society of Veterinary Science
2016
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5240763/ https://www.ncbi.nlm.nih.gov/pubmed/27628591 http://dx.doi.org/10.1292/jvms.16-0268 |
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author | INOSHIMA, Yasuo TAKASU, Masaki ISHIGURO, Naotaka |
author_facet | INOSHIMA, Yasuo TAKASU, Masaki ISHIGURO, Naotaka |
author_sort | INOSHIMA, Yasuo |
collection | PubMed |
description | Orf virus infection has been prevalent continuously in the population of wild Japanese serows (Capricornis crispus), goat-like grazing cloven-hoofed mammal species that live mainly in mountainous areas of Japan. Currently, definitive diagnosis of infection requires time-consuming laboratory work. To diagnose rapidly on-site, we developed a field-friendly procedure for the detection of orf virus from oral cavity lesions. DNA was extracted from goat saliva spiked with orf virus as a proxy for Japanese serows by a commercial kit without the use of electricity, and the quality of the extracted DNA was evaluated by conventional polymerase chain reaction (PCR). Extracted DNA was amenable to DNA amplification, the same as when extracted in a laboratory. Next, to find optimal conditions for DNA amplification by loop-mediated isothermal amplification (LAMP), Bst and Csa DNA polymerases and 3 colorimetric indicators for visual diagnosis, hydroxy naphthol blue (HNB), malachite green and D-QUICK, were compared using a portable cordless incubator. The combination of Bst or Csa DNA polymerase with HNB was found to be easiest for visual diagnosis by the naked eye, and viral DNA was successfully amplified from all orf virus strains used. These results suggest that the procedure established here can work completely on-site and can be useful for definitive diagnosis and differentiation of orf virus infection in Japanese serows in remote mountainous areas. |
format | Online Article Text |
id | pubmed-5240763 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2016 |
publisher | The Japanese Society of Veterinary Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-52407632017-01-30 Establishment of an on-site diagnostic procedure for detection of orf virus from oral lesions of Japanese serows (Capricornis crispus) by loop-mediated isothermal amplification INOSHIMA, Yasuo TAKASU, Masaki ISHIGURO, Naotaka J Vet Med Sci Wildlife Science Orf virus infection has been prevalent continuously in the population of wild Japanese serows (Capricornis crispus), goat-like grazing cloven-hoofed mammal species that live mainly in mountainous areas of Japan. Currently, definitive diagnosis of infection requires time-consuming laboratory work. To diagnose rapidly on-site, we developed a field-friendly procedure for the detection of orf virus from oral cavity lesions. DNA was extracted from goat saliva spiked with orf virus as a proxy for Japanese serows by a commercial kit without the use of electricity, and the quality of the extracted DNA was evaluated by conventional polymerase chain reaction (PCR). Extracted DNA was amenable to DNA amplification, the same as when extracted in a laboratory. Next, to find optimal conditions for DNA amplification by loop-mediated isothermal amplification (LAMP), Bst and Csa DNA polymerases and 3 colorimetric indicators for visual diagnosis, hydroxy naphthol blue (HNB), malachite green and D-QUICK, were compared using a portable cordless incubator. The combination of Bst or Csa DNA polymerase with HNB was found to be easiest for visual diagnosis by the naked eye, and viral DNA was successfully amplified from all orf virus strains used. These results suggest that the procedure established here can work completely on-site and can be useful for definitive diagnosis and differentiation of orf virus infection in Japanese serows in remote mountainous areas. The Japanese Society of Veterinary Science 2016-09-15 2016-12 /pmc/articles/PMC5240763/ /pubmed/27628591 http://dx.doi.org/10.1292/jvms.16-0268 Text en ©2016 The Japanese Society of Veterinary Science http://creativecommons.org/licenses/by-nc-nd/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution Non-Commercial No Derivatives (by-nc-nd) License. |
spellingShingle | Wildlife Science INOSHIMA, Yasuo TAKASU, Masaki ISHIGURO, Naotaka Establishment of an on-site diagnostic procedure for detection of orf virus from oral lesions of Japanese serows (Capricornis crispus) by loop-mediated isothermal amplification |
title | Establishment of an on-site diagnostic procedure for detection of orf virus
from oral lesions of Japanese serows (Capricornis crispus) by
loop-mediated isothermal amplification |
title_full | Establishment of an on-site diagnostic procedure for detection of orf virus
from oral lesions of Japanese serows (Capricornis crispus) by
loop-mediated isothermal amplification |
title_fullStr | Establishment of an on-site diagnostic procedure for detection of orf virus
from oral lesions of Japanese serows (Capricornis crispus) by
loop-mediated isothermal amplification |
title_full_unstemmed | Establishment of an on-site diagnostic procedure for detection of orf virus
from oral lesions of Japanese serows (Capricornis crispus) by
loop-mediated isothermal amplification |
title_short | Establishment of an on-site diagnostic procedure for detection of orf virus
from oral lesions of Japanese serows (Capricornis crispus) by
loop-mediated isothermal amplification |
title_sort | establishment of an on-site diagnostic procedure for detection of orf virus
from oral lesions of japanese serows (capricornis crispus) by
loop-mediated isothermal amplification |
topic | Wildlife Science |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5240763/ https://www.ncbi.nlm.nih.gov/pubmed/27628591 http://dx.doi.org/10.1292/jvms.16-0268 |
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