Prevalence and Relative Virulence Properties of Circulatory Phylotypes of White Spot Syndrome Virus in Crustaceans

Abstract

White Spot Syndrome Virus (WSSV) is recognised as a highly prevalent and deadly virus worldwide which affects both shrimps and crabs in the coastal aquatic environment. This research work intended to observe the presence of deadly WSSV in selected ghers during the period of 2017 to 2019 and the virulence of WSSV’s phylotypes circulating in Bangladesh. We had collected 360 shrimp (Penaeus monodon) and 120 crab (Scylla sp.) samples from the South-East (Cox’s Bazar) and South-West (Satkhira) coastal regions of Bangladesh during monsoon, and the sampling was continued during post-monsoon also with the observation of same number of samples. PCR assays targeting VP28 gene uncovered statistically significant (p < 0.05, Kruskal Wallis test) differences in the WSSV’s prevalence in shrimps and crabs of the selected regions of this study (Cox’s Bazar and Satkhira), and also over the periods of this study (2017-2019). The mean Log copy numbers of WSSV varied from 8.40 (in Cox’s Bazar) to 10.48 (in Satkhira) per gram tissue indicating serious diseased condition. In the ghers where WSSV-positive crustacean samples were found, the mean values for salinity, dissolved oxygen, temperature and pH were 14.71±0.76 ppt, 3.7±0.1 ppm, 34.11±0.38 ˚C and 8.23±0.38, respectively. Phylogenetic analysis based on the envelope protein VP28 gene exposed a substitution in the amino acid (aa) sequences, (Glutamic acid→Glycine) at 167th position in the isolates of Cox’s Bazar (referred to as phylotype BD2) in comparison to the circulating one (BD1) worldwide. In the experimental infection assays, shrimp post larvae (PL) challenged with BD1 and BD2 phylotypes with filtrates of tissue containing 0.423 X 109 copies of WSSV per mL ensured a median LT50 value of 73 hrs and 75 hrs, respectively. The experimental infection assay demonstrated higher mean Log WSSV copies (6.47±2.07 per mg tissue) in shrimp PL challenged with BD1 compared to BD2 (4.75±0.35 per mg tissue). Crabs that were challenged with BD1 and BD2 through ingestion method demonstrated 100% mortality within 48 hrs and 62 hrs post-infection, respectively, having mean Log WSSV copies of 12.06±0.48 and 9.95±0.37 per gram tissue, respectively. Furthermore, expression of shrimp antimicrobial peptides (AMPs) penaeidin and lysozyme was at a low level in BD1 treated shrimp PL in comparison to BD2 treated shrimp PL. These results all together exhibited that relative virulence properties of WSSV based on the rate of mortality, viral load and expression of host immune genes in crustaceans could be affected by single aa substitution in VP28. Assessment of prevalence with a larger number of samples covering all the shrimp farming regions of Bangladesh in future along with gene expression profiling of all important immune genes of crustaceans infected with the circulating groups of WSSV can add values to get into the insights of the occurrence of WSSV infections.