Development of an In Situ Hybridization Method for Detection of Akabane Virus

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Nihat Toplu
Tuba Çiğdem Oğuzoğlu
Ayşe Nur Akkoç


Akabane virus, in situ hybridization, method, pathology


Akabane virus (AKAV) is an arbovirus belonging to the family Bunyaviridae, genus Orthobunyavirus. AKAV consists of three-segment (L, M, and S RNA segments), negative single-stranded RNA. The aim of this study was to investigate an in situ hybridization method (ISH) in a Vero E6 cell line infected with Akabane virus. The 320 base pair amplicon was obtained by RT-PCR with a primer pair and labeled with digoxigenin. Akabane virus RNAs were seen as a granular pattern in the cytoplasm of infected cells. As a result, the expression of the particular Akabane virus gene area was successfully disclosed in the current investigation using the ISH method with a digoxigenin-labeled probe.


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1. Kurogi H, Akiba K, Inaba Y, Matumoto M. Isolation of Akabane virus from the biting midge Culicoides oxystoma in Japan. Vet Microbiol 1987: Jan; 27(1): 243-248. 1987-11-01 PMID: 3124329
2. Oya A, Okuno T, Ogata T, Kobayashi I, Matsuyama T. Akabane, a new arbovirus isolated in Japan. Jap J Med Sci Biol 1961: Jun; 14:101-108. https:/10.7883/yoken1952.14.101 PMID: 14482934
3. Purnomo Edi S, Ibrahim A, Sukoco R, Bunali L, Taguchi M et al. Molecular characterization of an Akabane virus isolate from West Java, Indonesia. J Vet Med Sci 2017: Apr; 8:79(4):774-779. PMID: 28302930
4. Konno S, Moriwaki M, Nakagawa M. Akabane disease in cattle: congenital abnormalities caused by viral infection. Spontaneous disease. Veterinary Pathology 1982: May; 19(3):246-66. PMID: 7200278
5. Mohamed ME, Mellor PS, Taylor WP. Akabane virus: serological survey of antibodies in livestock in the Sudan. Rev Elev Med Vet Pays Trop 1996: 49(4):285-8. PMID: 9239934.
6. Kono R, Hirata M, Kaji M, Goto Y, Ikeda S, Yanase T, Kato T, Shogo T, Tsutsui T, Imada T, Yamakawa M. Bovine epizootic encephalomyelitis caused by Akabane virus in southern Japan. BMC Vet Res 2008: Jun; 13(4):1-10. PMID:18554406
7. Hayama Y, Moriguchi S, Yanase T, Ishikura Y, Abe S, Higashi T , Ishikawa H 5, Yamamoto T, Kobayashi S, Murai K, Tsutsui T. Spatial epidemiological analysis of bovine encephalomyelitis outbreaks caused by Akabane virus infection in western Japan in 2011. Trop Anim Health and Prod 2016: Apr; 48(4):843-847. PMID: 26898692
8. Miyazato S, Miura Y, Hase M, Kubo M, Goto Y, Kono, Y. Encephalitis of cattle caused by Iriki isolate, a new strain belonging to Akabane virus. Nihon Juigaku Zasshi 1989: Feb; 51(1): 128-136. PMID: 2494374
9. Ahi MR, Pourmahdi Borujeni M, Haji Hajikolaei MR, Seifi Abad Shapouri MR. A serological survey on antibodies against Akabane virus in sheep in southwest of Iran. Iran J Virol 2017: 9(2):20-25. Corpus ID: 55907548
10. Oem JK, Lee KH, Kim HR, Bae YC, Chung JY et al. Bovine epizootic encephalomyelitis caused by Akabane virus infection in Korea. J Comp Pathol 2012: Aug-Oct; 147(2-3):101-105. PMID: 22520820
11. Oğuzoğlu TÇ, Toplu N, Koç BT, Doğan F, Epikmen ET et al. First molecular detection and characterization of Akabane virus in small ruminants in Turkey. Arch Virol 2015: Oct;160(10):2623-2627. PMID: 26215442
12. Toplu N, Alçığır G. Pathologic findings and immunohistochemical distribution of viral antigen in experimental avian encephalomyelitis of the chick and quail. Kafkas Univ Vet Fak Derg 2009: 15 (5): 725-732.
13. Toplu N, Alçığır G, Tunca R. Pathologic changes and use of immunohistochemical methods in naturally occuring avian encephalomyelitis of the chick. Kafkas Univ Vet Fak Derg 2009: 15 (5): 717-724. 10.9775/kvfd.2009.094-A Corpus ID: 26138346
14. Resende TP, Marthaler D, Vannucci FA. In situ hybridization detection and subtyping of rotaviruses in swine samples. J Vet Diag Invest 2019: Jan;31(1):113-117. PMID: 30541408
15. Toplu N, Oğuzoğlu TÇ, Ural K, Albayrak H, Ozan E, Erturk A, Epikmen ET. West Nile Virus infection in horses: Detection by immunohistochemistry, in Situ Hybridization, and ELISA. Vet Pathol 2015: Nov;52(6):1073-10766. PMID: 25677341
16. Toplu N, Oğuzoğlu TÇ, Epikmen ET, Aydoğan A. neuropathologic study of Border Disease virus in naturally infected fetal and neonatal small ruminants and its association with apoptosis. Vet Pathol 2011: May;48(3):576-583. PMID: 20460448
17. H Akashi , S Onuma, H Nagano, M Ohta, T Fukutomi. Detection and differentiation of Aino and Akabane Simbu serogroup bunyaviruses by nested polymerase chain reaction. Arch Virol 1999:144(11);2101-2109. https:// doi: 10.1007/s007050050625 PMID: 10603165
18. Jensen E. Technical review: In situ hybridization. Anat Rec (Hoboken) 2014: Aug;297(8):1349-1353. PMID: 24810158
19. Gall JG, Pardue ML. Formation and detection of RNA-DNA hybrid molecules in cytological preparations. Proc Natl Acad Sci USA 1969: Jun; 63(2):378-383. PMID: 4895535
20. Warford A, Lauder I. In situ hybridisation in perspective. J Clin Pathol 1991: Mar; 44(3):177-181. PMID: 2013617
21. Cassidy A, Jones J. Developments in in situ hybridisation. Methods 2014: Nov; 70(1):39-45. PMID: 24747923
22. McNicol AM, Farquharson MA. In situ hybridization and its diagnostic applications in pathology. J Pathol 1997: Jul;182(3):250-261.<250:AID-PATH837>3.0.CO;2-S PMID: 9349226
23. Mabruk MJ. In situ hybridization: detecting viral nucleic acid in formalin-fixed, paraffin-embedded tissue samples. Expert Rev Mol Diag 2004: Sep;4(5):653-661. PMID: 15347259
24. Kimura K, Yanase T, Kato T. Histopathological, immunohistochemical and in-situ hybridization findings in suckling rats experimentally infected with Akabane genogroups Ⅰ and Ⅱ, Aino and Peaton Viruses. J Comp Pathol 2021: Aug;187:27-39. PMID: 34503652