Persistence of Penaeus stylirostris densovirus delays mortality caused by white spot syndrome virus infection in black tiger shrimp (Penaeus monodon)

BMC Veterinary Research

Vanvimon Saksmerprome⁵  Timothy Flegel⁵  Pattira Pongtippatee³  Boonsirm Witchayachamnarnkul¹  Chuenchit Boonchird²  Yutthana Joyjinda⁴  Sarocha Jitrakorn⁵  Sudkhate Molthathong⁴ 

[1] Shrimp Genetics Improvement Center, Surat Thani, Thailand;Department of Biotechnology, Faculty of Science, Mahidol University, 10400, Bangkok, Thailand;Aquatic Animal Biotechnology Research Center, Surat Thani Campus, 84100, Surat Thani, Thailand;Centex Shrimp, Faculty of Science, Mahidol University, 10400, Bangkok, Thailand;National Center for Genetic Engineering and Biotechnology (BIOTEC), National Science and Technology Development Agency (NSTDA), Thailand Science Park, 12120, Pathum Thani, Thailand

关键词: Multiplex PCR;    Real-time PCR;    WSSV;    Shrimp;    IHHNV;    PstDNV;   

Others  :  1119605
DOI  :  10.1186/1746-6148-9-33

 received in 2012-10-08, accepted in 2013-02-06,  发布年份 2013

【 摘 要 】

Background

Persistent infection of Penaeus stylirostris densovirus (PstDNV) (also called IHHNV) and its non-infectious inserts in the black tiger shrimp, Penaeus monodon (P. monodon) genome are commonly found without apparent disease. Here, we introduced the method of multiplex PCR in order to differentiate shrimp with viral inserts from ones with the infectious virus. The method allowed us to study the effect of pre-infection of IHHNV, in comparison to IHHNV inserts, on WSSV resistance in P. monodon.

Results

A multiplex PCR system was developed to amplify the entire IHHNV genome, ensuring the accurate diagnosis. Field samples containing IHHNV DNA templates as low as 20 pg or equivalent 150 viral copies can be detected by this method. By challenging the two groups of diagnosed shrimp with WSSV, we found that shrimp with IHHNV infection and those with viral inserts responded to WSSV differently. Considering cumulative mortality, average time to death of shrimp in IHHNV-infected group (day 14) was significantly delayed relative to that (day 10) of IHHNV-inserted group. Real-time PCR analysis of WSSV copy number indicated the lower amount of WSSV in the IHHNV-infected group than the virus-inserted group. The ratio of IHHNV: WSSV copy number in all determined IHHNV-infected samples ranged from approximately 4 to 300-fold.

Conclusion

The multiplex PCR assay developed herein proved optimal for convenient differentiation of shrimp specimens with real IHHNV infection and those with insert types. Diagnosed shrimp were also found to exhibit different WSSV tolerance. After exposed to WSSV, the naturally pre-infected IHHNV P. monodon were less susceptible to WSSV and, consequently, survived longer than the IHHNV-inserted shrimp.

【 授权许可】

   
2013 Molthathong et al; licensee BioMed Central Ltd.

【 图 表 】

Figure 1.

【 参考文献 】

• [1]Tanticharoen M, Flegel TW, Meerod W, Grudloyma U, Pisamai N: Aquacultural biotechnology in Thailand: the case of the shrimp industry. Int J Biotechnol 2008, 10(6):588-603.
• [2]Lightner DV, Redman RM, Bell TA: Infectious hypodermal and hematopoietic necrosis, a newly recognized virus disease of penaeid shrimp. J Invertebr Pathol 1993, 41:62-70.
• [3]Fauquet CM, Mayo CMA, Maniloff J, Desselberger U, Ball LAE: Virus taxonomy classification and nomenclature of viruses eight report of the International Committee on Taxonomy of viruses. Massachusetts: Elsevier Academic Press; 2005:1259.
• [4]Bonami JR, Trumper B, Mari J, Brehelin M, Lightner DV: Purification and characterization of IHHN virus of penaeid shrimps. J Gen Virol 1990, 71:2657-2664.
• [5]Nunan LM, Poulos BT, Lightner DV: Use of polymerase chain reaction (PCR) for the detection of infectious hypodermal and hematopoietic necrosis virus (IHHNV) in penaeid shrimp. Marine Biotechnol 2000, 2:319-328.
• [6]Tang KFJ, Lightner DV: Low sequence variation among isolates of infectious hypodermal and hematopoietic necrosis virus (IHHNV) originating from Hawaii and the Americas. Dis Aquat Organ 2002, 49:93-97.
• [7]Tang KFJ, Navarro SA, Lightner DV: A PCR assay for discriminating between infectious hypodermal and hematopoietic necrosis virus (IHHNV) and the virus-related sequences in the genome of Penaeus monodon. Dis Aquat Organ 2007, 74:165-170.
• [8]Lightner DV: A Handbook of Shrimp Pathology and Diagnostic Procedures for Diseases of Cultured Penaeid Shrimp. Louisiana, USA: Baton Rouge; 1996.
• [9]Tang KFJ, Lightner DV: Infectious hypodermal and hematopoietic necrosis virus (IHHNV)-related sequences in the genome of the black tiger prawn Penaeus monodon from Africa and Australia. Virus Res 2006, 118:185-191.
• [10]Saksmerprome V, Jitrakorn S, Chayaburakul K, Laiphrom S, Boonsua K, Flegel TW: Additional random, single to multiple genome fragments of Penaeus stylirostris densovirus in the giant tiger shrimp genome have implications for viral disease diagnosis. Virus Res 2011, 160(1–2):180-190.
• [11]Flegel TW: Hypothesis for heritable, anti-viral immunity in crustaceans and insects. Biol Direct 2009, 4(32):1-8.
• [12]Bonnichon V, Lightner DV, Bonami JR: Viral interference between infectious hypodermal and hematopoietic necrosis virus and white spot syndrome virus in Litopenaeus vannamei. Dis Aquat Organ 2006, 72:179-184.
• [13]Tang KFJ, Durand SV, White BL, Redman RM, Mohney LL, Lightner DV: Induced resistance to white spot syndrome virus infection in Penaeus stylirostris through pre-infection with infectious hypodermal and hematopoietic necrosis virus - a preliminary study. Aquaculture 2003, 216:19-29.
• [14]Saksmerprome V, Puiprom O, Noonin C, Flegel TW: Detection of infectious hypodermal and haematopoietic necrosis virus (IHHNV) in farmed Australian Penaeus monodon by PCR analysis and DNA sequencing. Aquaculture 2010, 298(3–4):190-193.
• [15]Sambrook JFE, Maniatis T: Molecular cloning. A laboratory manual. Second edition. New York: Cold Spring Harbor Laboratory; 1989.
• [16]Molthathong S, Senapin S, Klinbunga S, Puanglarp N, Rojtinnakorn J, Flegel TW: Down-regulation of defender against apoptotic death (DAD1) after yellow head virus (YHV) challenge in black tiger shrimp Penaeus monodon. Fish Shellfish Immunol 2008, 24(2):173-179.
• [17]Rasmussen R: Rapid cycle real-time PCR, methods and applications. Heidenberg: Springer Press; 2001.
• [18]Johnson MR, Wang K, Smith JB, Heslin MJ, Diasio RB: Quantitation of dihydropyrimidine dehydrogenase expression by real-time reverse transcription polymerase chain reaction. Anal Biochem 2000, 278(2):175-184.
• [19]The PstDNV databasehttp://www.oie.int/fileadmin/Home/eng/Health_standards/aahm/2010/2.2.02_IHHN.pdf webcite
• [20]The WSSV databasehttp://www.oie.int/international-standard-setting/aquatic-manual/access-online/ webcite