A footprint in the genome of freshwater crayfish whispovirus (WSSV-Cc)

A white spot syndrome virus strain (Cambarus clarkii whispovirus, WSSV-Cc or Cc strain) was isolated from naturally infected and moribund freshwater crayfish (C. clarkii or Procambarus clarkii, P. clarkii), which is a new strain with a smaller genome. In order to find the footprints in the genome generated during virus evolution, microscopic and ultramicroscopic investigations, genome architecture and phylogenetic analysis, and specific gene amplification were performed. The products of six genes (74L, 86L, 87R, 88R, 92R and 95R) of Cc strain were selected to construct a phylogenetic tree with their homologs from eight white spot virus strains. Results showed that the strains can be divided into two clusters: one included P. clarkii nimavirus (Cc, CN02, and Pc) and the other contained marine shrimp nimaviruses (CN, CN01, CN03, CN04, TW and KR). Multiple sequence alignments of homologous proteins from different viral strains showed that Cc-87R is significantly different from its homologous membrane proteins of marine shrimp nimavirus. It lacks the transmembrane domain (TMD) and its adjacent 287 aa sequence, but still has a complete PI3K_rbd region. Further nucleic acid amplification was performed with two pairs of primers, 87R-F/87R-R and 238-F/87R-R, using the genomes of P. clarkii nimavirus Cc strain and marine shrimp nimavirus CN strain as templates respectively. A fragment with size of 709 bp containing the entire sequence of Cc-87R was amplified from the Cc genomic template. However, the fragments with size of 1 600 and 4 810 bp, only containing partial sequence of Cc-87R, were amplified from the template of the CN strain. The results provided experimental evidence that Cc-87R is a footprint with unique sequence structure of the Cc strain. This finding will benefit the detection of the P. clarkii nimavirus pathogen and the warning of its epidemic trend.