ScienceAsia 32 (2006): 395-401 |doi: 10.2306/scienceasia1513-1874.2006.32.395

Selection and Testing Phage-displayed YHV-binding Peptides for Possible Detection of Yellow Head Virus (YHV) in Shrimp

Warachin Gangnonngiw¹, Pongrama Ramasootra², Chumporn Soowannayan^1,3, Paisarn Sithigorngul⁴, Siripron Sriurairatana¹, Kallaya Sritunyalucksana^1,3 and Timothy W. Flegel^1*

 
ABSTRACT:Yellow head virus (YHV) causes a serious disease that can result in high mortality of penaeid shrimp within 2-3 days after the first appearance of gross signs of disease in rearing ponds. Current detection systems for YHV are based on molecular and immunological techniques. Immunological detection methods require continual production of relatively large, complex proteins using living animals or hybridoma cells. An alternative possibility is to use the phage display technique to find short peptides that can bind strongly with YHV particles and replace antibodies in such tests. Once identified, these peptides could also be tested for efficacy in blocking YHV infection. YHV was purified and then immobilized on microtiter plates for 4 rounds of biopanning against a pool of phage displayed peptide variants. From 89 sequentially generated phage pools, 13 variants were selected for strong binding with YHV by ELISA assay. DNA sequencing led to the deduced amino acid sequences LNAKSRN, KSKKSSS, GPQRKRS, KLKRLSS, RTNKKNA, SNISNAS, SNSKKRN, RTKKMRT, NTKRPAR, GPQRKRS, VSNKKRS, RKKSNAS and GPKKNRS. Preliminary tests using two representative phages (SNSKKRN and GPQRKRS) with high YHV binding activity showed that they were unable to block YHV infection in shrimp. However, when immobilized, these clones could bind YHV from probe solutions, indicating that they have potential for use as YHV detection reagents.

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¹ Centex Shrimp, Faculty of Science, Mahidol University, Rama VI rd., Bangkok, Thailand.
² Department of Social and Environmental Medicine, Faculty of Tropical Medicine, Ratchawithi Rd., Bangkok, Thailand.
³ National Center for Genetic Engineering and Biotechnology (BIOTEC), National Science and Technology Development Agency (NSTDA), Klong Luang, Patumthani, Thailand.
⁴ Department of Biology, Faculty of Science, Srinakarinwirot University, Sukhumvit Soi 23, Bangkok, Thailand.

* Corresponding author, E-mail: sctwf@mahidol.ac.th

Received 18 Jan 2006, Accepted 10 May 2006