Experimental infection reveals mud crab, Scylla serrata is less susceptible than Scylla olivacea and shrimp, Penaeus vannamei to white spot syndrome virus (WSSV)

Abstract

White spot syndrome virus (WSSV) causes severe mortality in farmed shrimps, besides causing infection in a wide range of crustaceans. Among the hosts, mud crabs are reported to be relatively resistant to the virus. To validate this, the present study investigated the susceptibility of two species of mud crabs, Scylla olivacea and S. serrata compared with shrimp, Penaeus vannamei to WSSV through experimental infection. Initially, to find out the pathogenicity of the virus, a quantified, 10-fold serially diluted WSSV inoculum was injected intramuscularly into shrimp and mud crabs. Cumulative mortality of 100% was observed in shrimp injected with 10−1 and 10−2 WSSV dilution at 5 and 6 day post-injection (dpi), respectively. However, injection of the virus inoculum of 10−1 dilution resulted in 100% and 85% cumulative mortality at 7 dpi in S. olivacea and S. serrata, respectively. The LD50 (lethal dose 50% endpoint) value of WSSV in P. vannamei, S. olivacea and S. serrata was determined to be 1.74 × 105, 3.80 × 106 and 8.51 × 106 viral copies per animal, respectively. PCR amplification of DNA extracted from different tissues of the experimental animals at 24, 48 and 72 h post-injection (hpi) confirmed the infection. A SYBR green-based real-time PCR assay revealed comparatively higher viral load in shrimp tissues (4.1 × 103, 2.8 × 106 and 4.3 × 107 copies/ 100 ng DNA in stomach, 1.0 × 103, 2.4 × 106 and 3.5 × 107 copies/ 100 ng DNA in gill and 3.5 × 102, 1.1 × 104 and 1.1 × 105 copies/ 100 ng DNA in hepatopancreas at 24, 48 and 72 h, respectively). The viral load in S. olivacea was estimated to be 2.9 × 103, 2.2 × 106 and 3.0 × 107 copies/ 100 ng DNA in the stomach, 7.1 × 102, 1.6 × 106 and 2.4 × 107 copies/ 100 ng DNA in gill and 2.0 × 102, 8.6 × 103& 7.8 × 104 copies/ 100 ng DNA and in S. serrata the viral copy numbers detected were 1.1 × 103, 1.7 × 106 and 1.8 × 107 copies/ 100 ng DNA in stomach, 6.6 × 102, 1.3 × 106 and 1.4 × 107 copies/ 100 ng DNA in gill and 5.8 × 101, 7.7 × 102 & 1.1 × 104 copies/ 100 ng DNA at 24, 48 and 72 h, respectively. A higher viral load was detected in the stomach tissue compared to the gill and hepatopancreas of all the experimental animals. Histological examination of the stomach and gill of P. vannamei, S. olivacea and S. serrata at different time points post-infection revealed characteristic WSSV pathology. Hepatopancreatic tubular epithelium showed no viral inclusions, though pathological changes could be noticed in the intertubular connective tissue. The most pronounced changes were found in the stomach epithelium of P. vannamei and S. olivacea, which revealed significant nuclear hypertrophy and large-sized inclusions, compared to S. serrata which showed relatively smaller-sized inclusion bodies.