Campoletis sonorensis Virus in Heliothis virescens : and preliminary studies on the organization of the viral genome

Abstract

The expression of Campoletis sonorensis Virus (CsV) in the parasitized host, Heliothis Virescens, was investigated. At least 12 major CsV mRNAs were detected in parasitized H. virescens larvae between 2 hours and 9 days after parasitization. Cloned CsV DNA restriction fragments which contained expressed sequences were used as hybridization probes for mapping individual CsV transcripts to superhelical regions of the CsV genome. Transcriptional mapping of several CsV mRNAs showed that the CsV genes studied are segregated on different superhelical DNAs or different sets of superhelical DNAs, thus demonstrating that the CsV genome is transcriptionally multipartite. Significant cross homology detected between CsV superhelical DNAs suggests that either (a) some CsV genes are shared on several superhelical DNAs or (b) separate superhelical DNAs may contain genes which are partially homologous. Also, the cross homology observed between different CsV superhelical DNAs was not limited to the coding regions of the genes examined in this study. Putative CsV proteins were identified by hybridization selection and in vitro translation of specific CsV mRNAs. cDNA clones homologous to a gene encoding an abundant CsV mRNA were identified. Based on Southern hybridizations of these cDNA clones to viral DNA, the cDNA clones were divided into two classes. DNA sequencing and northern blot analysis of two representative cDNA clones indicated that the two cDNA clones were synthesized from two CsV mRNAs (1.6 and 1.0 kb) which share several regions of homology. Analysis of the long open reading frame from each cDNA revealed several regions of conserved amino acid sequences although the two putative proteins have otherwise diverged in amino acid sequence. Comparison of the DNA sequence of a CsV genomic DNA restriction fragment to the sequence from a cDNA clone revealed that the CsV gene is a spliced gene which contains at least two introns. A 15.8 kbp superhelical DNA (WSp-16kA) was cloned from CsV superhelical DNA region W. The two cDNA classes were mapped to different regions of the circular map of WSp-16kA. These data show that two related CsV genes are located on a single CsV superhelical DNA molecule.