IISSN 2410-7751 (Print)
ISSN 2410-776X (Online)
"Biotechnologia Acta" V. 11, No 1, 2018
Р. 58-63, Bibliography 17, English
Universal Decimal Classification: 576.858
https://doi.org/10.15407/biotech11.01.058
POLYMERASE CHAIN REACTION FOR IDENTIFICATION OF CYPRINID HERPESVIRUSES IN UKRAINE
Yu. P. Rud 1, M. I. Maistrenko 2, L. P. Buchatskiy 1, 2
1 Institute of Fisheries of the National Academy of Agriculture Sciences of Ukraine, Kyiv
2 ESC Institute of Biology and Medicine, Taras Shevchenko Kyiv National University, Ukraine
The aim of the research was to investigate diseased fish species Labeo (Labeo bicolor) and Sack-gill Catfish (Heteropneustes fossilis) by testing several diagnostic systems targeting fish herpesviruses in purpose to determine the etiology of the outbreak infection in aquarium fishes at the Kуiv Zoo during summer 2017. During a fish health inspection of aquariums in Kyiv Zoo, an Cyprinid herpesvirus 3 ? CyHV-3 was detected in Labeo and Sack-gill Catfish. Preliminary examination of infected fish revealed a range of lesions particularly in internal organs and tissues. The gills of diseased fish were characterized by hyperaemia and necrosis. The infringement of liver color and structure, kidney swelling and gallbladder necrosis were observed in both fish species. The virus did not grow in fish cell lines of RTG-2, FHM and EPC, that was evident by absence of any morphological changes in appropriate cell lines. Also initially fish were tested for parasitic and bacterial infections and they were determined to be non-infected. Our results demonstrated that specific oligonucleotide primers for thymidine kinase gene of CyHV-3 were successfully amplified the specific DNA fragments. The length of polymerase chain reaction product, as expected, was 264 nucleotide pairs. The amplified specific fragments were identical to the area of thymidine kinase gene CyHV-3, as was shown by sequence analysis. The identity of nucleotide sequences composed 97?99%. The same positive results were obtained using primers that recommended by the International Epizootic Bureau, fragments in size of 409 and 292 were also obtained. In our opinion, CyHV-3 was brought to Ukraine by import of aquarium fish avoiding sanitary control of transboundary transportation. Therefore, the uncontrolled import of aquarium and cultured fish species is a serious problem because imported fish could be as a source of highly pathogenic infections for industrial aquaculture.
Key words: polymerase chain reaction, CyHV-3 fish herpesvirus of Labeo bicolor and Sack-gill Catfish.
© Palladin Institute of Biochemistry of National Academy of Sciences of Ukraine, 2018
References
1. Waltzek T. B., Kelley G. O., Alfaro M. E., Kurobe T., Davison A. J., Hedrick R. P. Phylogenetic relationships in the family alloherpesviridae. Dis. Aquat. Org. 2009, V. 84, P. 179–194.
2. Jung S. J., Miyazaki T. . Herpesviral hematopoietic necrosis of goldfish, Carassius auratus (L.). J. Fish Dis. 1995, V. 18, P. 211–220.
3. Van Beurden S. J., Bossers A., Voorbergen-Laarman M. H. A., Haenen O. L. M., Peters S., Abma-Henkens M. H. C, Peeters B. P. H., Rottier P. J. M., Engelsma M. Y. Complete genome sequence and taxonomic position of anguillid herpesvirus 1. J. Gen. Virol. 2010, V. 91, P. 880–887.
4. Sano T., Fukuda H., Furukawa M., Hosoya H., Moriya Y. A herpesvirus isolated from carp papilloma in Japan. Fish Shell Pathol. 1995, V. 32, P. 307–311.
5. Waltzek T. B., Kelley G. O., Stone D. M., Way K., Hanson L., Fukuda H., Hirono I., Aoki T., Davison A. J., Hedrick R. P. Koi herpesvirus represents a third cyprinid herpesvirus (CyHV-3) in the family Herpesviridae. J. Gen. Virol. 2005, V. 86, P. 1659–1667.
6. Hedrick R. P., Gilad O., Yun S., Spangenberg J. V., Marty G. D., Nordhausen R. W., Kebus M. J., Bercovier H., Eldar A. A herpesvirus associated with mass mortality of juvenile and adult koi, a strain of common carp. J. Aquat. Anim. Health. 2000, V. 12, P. 44–57.
7. Bergmann S. M., Riechardt M., Fichtner D., Lee P., Kempter J. Investigation on the diagnostic sensitivity of molecular tools used for detection of koi herpesvirus. J. Virol. Meth. 2010, V. 163, P. 229–233.
8. Eide K. E., Miller-Morgan T., Heidel J. R., Kent M. L., Bildfell R. J., Lapatra S., Watson G., Jin L. Investigation of koi herpesvirus latency in koi. J. Virol. 2011, V. 85, P. 4954–4962.
9. Maistrenko M. I., Rud Y. P., Matvienko N. M., Holodna L. S., Buchatskiy L. P. Identification of СуНV-3 by the methods of electronic microscopy and polymerase chain reaction. Reports of Ukraine National Academy of Science. 2013, V. 4, P. 139–143. (In Ukrainian).
10. Aoki T., Hirono I., Kurokawa K., Fukuda H., Nahary R., Eldar A., Davidson A. J., Waltzek T. B., Bercuvier H., Hedrick R. P. Genome sequences of three koi herpesvirus isolates representing the expanding distribution of an emerging disease threatening koi and common carp worldwide. J. Virol. 2007, V. 81, P. 5058–5065.
11. Bercovier H., Fishman Y., Nahary R., Sinai S., Zlotkin A., Eyngor M., Gilad O., Eldar A., Hedrick R. Cloning of the koi herpesvirus (KHV) gene encoding thymidine kinase and its use for a highly sensitive PCR based diagnosis. BMC Microbiol. 2005, V. 5, P. 13.
12. Gray W. L., Mullis L., Lapatra S. E., Groff J. M., Goodwin A. Detection of koi herpesvirus DNA in tissues of infected fish. J. Fish Dis. 2002, V. 25, P. 171–178.
13. Ilouze M., Davidovich M., Diamant A., Kotler M. Dishon A. The outbreak of carp disease caused by CyHV-3 as a model for new emerging viral disease in aquaculture: a review. Ecol. Res. 2011, V. 26, P. 885–892.
14. Uchii K., Minamoto T., Honyo M., Kawabata Z. Seasonal reactivation enables Cyprinide Herpesvirus 3 to persist in wild host population. FEMS Microbiol. Ecol. 2014, V. 87, P. 536–542.
15. Davison A. J. Herpesvirus systematics. Vet. Microbiol. 2010, V. 143, P. 52–69.
16. Kempter J., Sadowski J., Schutze H., Fischer U., Dauber M., Fichtner D., Panicz R., Bergmann S. Koi herpes virus: do acipencerid restitution programs pose a threat to carp farms in the disease-free zones? Acta ichthyologica piscatorial. 2009, V. 2, P. 119–126.
17. Minamoto T., Honjo M., Yamanaka H., Tanaka N., Itajama T., Kawabata Z. Detection of cyprinid herpesvirus-3 DNA in lake plancton. Res.Veter. Sci. 2010, 90 (3), 530–532.