Additional random, single to multiple genome fragments of Penaeus stylirostris densovirus in the giant tiger shrimp genome have implications for viral disease diagnosis

Scattered reports of viral inserts in shrimp and insect genomes led to the hypothesis that random, autonomous insertion of such sequences occurs in these organisms and leads to specific, heritable immunity. To test the prediction regarding random insertion of viral sequences into the shrimp genome, we examined the giant tiger shrimp for random genomic insertions of Penaeus stylirostris densovirus (also called IHHNV). By PCR analysis using a set of 7 overlapping primer pairs to cover the whole IHHNV genome (4 kb), PCR failure with some pairs indicated sequence gaps that revealed a random pattern of putative viral inserts in the genomes of individual shrimp. Targeting a putative insert from one arbitrarily selected specimen, we used genome walking to reveal a viral insert linked to a host microsattelite-like fragment. This differed from 2 previously reported inserted fragments of IHHNV in P. monodon. In one specimen, 2 slightly different inserts were revealed, probably on paired chromosomes. By design and use of chimeric shrimp/virus primer pairs we proved that similar insertions occurred in several shrimp specimens, including those infected with IHHNV but showing no signs of disease. For the infected specimens, the inserts gave false positive PCR test results using 309F/R primers and a new IQ2000 test protocol currently recommended for detection of infectious IHHNV. This is the first experimental support for the hypothesis-based prediction that a random number and length of sequence fragments from a single virus genome may occur in the shrimp genome. Since some inserts can give false positive results for infectious IHHNV with the recommended methods above, they may have a negative effect on international seafood trade. In addition, discard of domesticated shrimp breeding stocks based on such false positive results might have negative consequences, if such inserts are related to shrimp viral disease tolerance, as also hypothesized. (C) 2011 Elsevier B.V. All rights reserved.