Application of gene probes as diagnostic tools for White Spot Baculovirus (WSBV) of penaeid shrimp

Abstract

Since 1993, similar baculoviruses, which cause high mortalities in penaeid shrimp, have been reported from China, Japan, Korea, Thailand and Taiwan. All these baculovirus-caused diseases are characterized by the presence of white spots in the cuticle. To isolate the agent of the disease referred to as White Spot Syndrome (WSS) and White Spot Baculovirus (WSBV), in this paper, Penaeus vannamei and P. stylirostris were experimentally inoculated with homogenates of infected Penaeus monodon from Thailand. In transmission electron microscopy (TEM), the enveloped WSBV virions have a size of about 350 nm long and 130 nm wide. The nucleocapsids range from 300 to 420 nm in length and 70 to 95 nm in diameter and show a superficially segmented appearance. Also present are unique particles, which have not been observed before, measuring 400 nm in length and 120 nm in width. These particles are larger than most of the usual nucleocapsids and have a cross- batched superficial appearance. After nucleic acid extraction, EcoRI-digested fragments of the WSBV genome were cloned. Four of these fragments were characterized and used as non-radioactive probes lableled with DIG-11-dUTP. By in situ hybridization, the probes hybridized with material located in nuclei of all WSBV-infected tissues. Previously reported target tissues, such as connective tissue, epithelial tissue and also hemocytes, clearly showed positive hybridization with the probes. In addition, some light infection was revealed in the muscle and nervous tissue.