Whole genome molecular characterization of Infectious Pancreatic Necrosis Viruses isolated in Turkey

Abstract

Infectious pancreatic necrosis virus (IPNV; Birnaviridae, Aquabirnavirus) causes infectious pancreatic necrosis (IPN) in fish. IPN disease was first found in 2002 in Turkey. In this study, 10 IPNV isolates were isolated between 2005 and 2013 and propagated in RTG-2 cell cultures. RNAs obtained from cell lysates were used as template and VP1, VP2, VP3, VP4 and VP5 genes of the virus were amplified in full length. Sequence analyses of the genes were made. The obtained sequences were compared with international reference strains from GenBank and phylogenetic analyses and genogrouping of the viruses were conducted. Turkish isolates were found to show a genetic similarity of between 93.5% and 99.8% in terms of molecules. When segment A sequences were compared with each other, a similarity between 97.8% and 99.8% was found; when they were compared with international reference strains, they were found to have the highest similarity (99.3%) with France AJ622822 isolate and the lowest (65.7%) similarity with Canada NC001915 isolate. When segment B sequences of the isolates were compared with each other, a similarity between 93.5% and 99.5% was found; when they were compared with international reference strains, they were found to have the highest similarity (98.9%) with Canada M58757 isolate and the lowest (52.7%) similarity with Finland KY548519 strain. As a result of phylogenetic analyses conducted, Turkish isolates were found to be closely related with France, Spain and Iran strains in serotype A2 and genogroup 5 they were placed in. 

Keywords

• molecular characterization,IPNV,RT-PCR,Turkey,whole genome

References

1. 1. Ahmadivand S, Soltani M, Behdani M, et al (2018): VP2 (PTA motif) encoding DNA vaccine confers protection against lethal challenge with infectious pancreatic necrosis virus (IPNV) in trout. Mol Immunol, 94, 61-67.
2. 2. Albayrak H (2017): The determination of molecular characterization and genetic analysis of IPNV field strains isolated from Turkey: The classic and recombinant vaccine trials and the development of diagnostic kit. Ondokuz Mayıs University Faculty of Veterinary Medicine, Samsun, Project of TUBITAK, Project Number: 213O156, 1-114.
3. 3. Blake S, Ma JY, Caporale DA, et al (2001): Phylogenetic relationships of aquatic birnaviruses based on deduced amino acid sequences of genome segment A cDNA. Dis Aquat Org, 45, 89-102.
4. 4. Candan A (2002): First report on the diagnosis of infectious pancreatic necrosis (IPN) based on reverse transcription polymerase chain reaction (RT-PCR) in Turkey. Bull Eur Ass Fish Pathol, 22, 45-48.
5. 5. Cuesta A, Chaves-Pozo E, De Las Heras AI, et al (2010): An active DNA vaccine against infectious pancreatic necrosis virus (IPNV) with a different mode of action than fish rhabdovirus DNA vaccines. Vaccine, 28, 3291-3300.
6. 6. Dadar M, Peyghan R, Rajabi-Memari H, et al (2014): Infectious pancreatic necrosis virus (IPNV) based on deduced amina acid sequences of genom segment A and B cDNA. Iran J Fish Sci, 13, 560-575.
7. 7. Davies KR, McColl KA, Wang LF, et al (2010): Molecular characterisation of Australasian isolates of aquatic birnaviruses. Dis Aquat Org, 93, 1- 15.
8. 8. Dobos P (1976): Size and structure of the genome of infectious pancreatic necrosis virus. Nucleic Acids Res, 3, 1903-1924.

9. 9. Dobos P (1995): The molecular biology of infectious pancreatic necrosis virus (IPNV). Ann Rev Fish Dis, 5, 25-54.
10. 10. Duncan R, Mason CL, Nagy E, et al (1991): Sequence analysis of infectious pancreatic necrosis virus genome segment B and its encoded VP1 protein: a putative RNA-dependent RNA polymerase lacking the Gly-Asp-Asp motif. Virology, 181, 541-552.
11. 11. Estévez RA, Mostazo MGC, Rodriguez E, et al (2018): Inducers of salmon innate immunity: An in vitro and in vivo approach. Fish Shellfish Immunol, 72, 247-258.
12. 12. Heppell J, Tarrab E, Berthiaume L, et al (1995): Characterization of the small open reading frame on genome segment A of infectious pancreatic necrosis virus. J Gen Virol, 76, 2091-2096.
13. 13. Ji F, Zhao JZ, Liu M, et al (2017): Complete genomic sequence of an infectious pancreatic necrosisvirus isolated from rainbow trout (Oncorhynchus mykiss) in China. Virus Genes, 53, 215-225.
14. 14. Julin K, Mennen S, Sommer A-I (2013): Study of virulence in field isolates of infectious pancreatic necrosis virus obtained from the northern part of Norway. J Fish Dis, 36, 89-102.
15. 15. Manríquez RA, Vera T, Villalba VM, et al (2017): Molecular characterization of infectious pancreatic necrosis virus strains isolated from the three types of salmonids farmed in Chile. Virol J, 14, 1-16.
16. 16. Mutoloki S, Evensen Ø (2011): Sequence similarities of the capsid gene of Chilean and European isolates of infectious pancreatic necrosis virus point towards a common origin. J Gen Virol, 92, 1721-1726.
17. 17. Nishizawa T, Kinoshita S, Yoshimizu M (2005): An approach for genogrouping of Japanese isolates of aquabirnaviruses in a new genogroup VII based on the VP2/NS junction region. J Gen Virol, 86, 1973-1978.
18. 18. Ørpetveit I. Mikalsen AB, Sindre H, et al (2010): Detection of infectious pancreatic necrosis virus in subclinically infected atlantic salmon by virus isolation in cell culture or real-time reverse transcription polymerase chain reaction: influence of sample preservation and storage. J Vet Diagn Invest, 22, 886-895.
19. 19. Salgado-Miranda C, Rojas-Anaya E, Garcia-Espinosa G, et al (2014): Molecular characterization of the VP2 gene of infectious pancreatic necrosis virus (IPNV) isolates from Mexico. J Aquat Anim Health, 26, 43-51.
20. 20. Santi N, Vakharia VN, Evensen Ø (2004): Identification of putative motifs involved in the virulence of infectious pancreatic necrosis virus. Virology, 322, 31-40.
21. 21. Santi N, Sandtro A, Sindre H, et al (2005): Infectious pancreatic necrosis virus induces apoptosis in vitro and in vivo independent of VP5 expression. Virology, 342, 13-25.
22. 22. Santi N, Song H, Vakharia VN, et al (2005): Infectious pancreatic necrosis virus VP5 is dispensable for virulence and persistence. J Virol, 79, 9206-9216.
23. 23. Shivappa RB, Song H, Yao K, et al (2004): Molecular characterization of Sp serotype strains of infectious pancreatic necrosis virus exhibiting differences in virulence. Dis Aquat Org, 61, 23-32.
24. 24. Skjesol A, Skjæveland I, Elnæs M, et al (2011): IPNV with high and low virulence: host immune responses and viral mutations during infection. Virol J, 8, 396-410.
25. 25. Song H, Santi N, Evensen Ø, et al (2005): Molecular determinants of infectious pancreatic necrosis virus virulence and cell culture adaptation. J Virol 2005, 79, 10289-10299.
26. 26. Weber S, Fichtner D, Mettenleiter TC, et al (2001): Expression of VP5 of infectious pancreatic necrosis virus strain VR299 is initiated at the second in-frame start codon. J Gen Virol, 82, 805-812.
27. 27. Wen CM (2017): Complete genome sequence and phylogenetic analyses of an aquabirnavirus isolated from a diseased marbled eel culture in Taiwan. Arch Virol, 162, 2467-2471.
28. 28. Zhu L, Wang X, Wang K, et al (2017): Outbreak of infectious pancreatic necrosis virus (IPNV) in farmed rainbow trout in China. Acta Trop, 170, 63-69.