Tuesday, 16 November 2004

Mapping of the Diachasmimorpha longicaudata entomopoxvirus genome and analysis of selected ORFs

Yoshifumi Hashimoto, hashimot@ufl.edu and Pauline O. Lawrence, pol@ifas.ufl.edu. University of Florida, Entomology and Nematology, Building 970, Natural Area Drive, Gainesville, FL

The Diachasmimorpha longicaudata entomopoxvirus (DlEPV) is the first symbiotic EPV described from a parasitic wasp. In order to construct a genomic map and characterize selected gene components, MboI-partial digests of DlEPV genomic DNA were cloned into SuperCos1 cosmid to establish a genomic library. Using PCR, sequencing, and restriction enzyme (BamHI, SacI, and XhoI) digestion, approximately 200 cosmid clones were identified. Twenty-seven overlapping clones, selected to cover the entire genome, were arranged in contiguous sequence. The estimated DlEPV genome, excluding the redundant regions, is 244.75kb and contains 15 BamHI, 6 SacI, and 18 XhoI sites. Genetic variation in the number of BamHI sites and lengths of restriction fragments were found among some clones, a probable indication of a mixed (uncloned) DlEPV population. The predicted protein sequences of eight selected ORFs within the DlEPV genome were compared with those of poxviruses and other organisms. Five were homologous with the poxvirus putative metalloprotease, membrane protein, late transcription factor-3, virion surface protein, and poly (A) polymerase regulatory small subunit. Three were simiar to the gamma-glutamyltransferase, eukaryotic initiation factor 4A, and lambda phage integrase of Drosophila and bacteria. Phylogenetic analysis of multiple alignments of the five DlEPV ORFs and 20 poxvirus homologs suggested that DlEPV probably diverged from the ancestral node between the fowlpox virus and the genus B lepidopteran and orthopteran EPVs. These results confirm earlier estimates of DlEPV genome size, support our hypothesis of group C classification for DlEPV, and lay the foundation for in depth molecular characterization of this unique virus. Funded by NSF grant 9986076 to POL.

Species 1: Hymenoptera Braconidae Diachasmimorpha longicaudata
Species 2: Diptera Tephritidae Anastrepha suspensa (Caribbean fruit fly)
Keywords: entomopoxvirus, genomic map

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