Genomic sequence analysis of a nucleopolyhedrovirus isolated from the diamondback moth, Plutella xylostella

Abstract

The CL3 plaque isolate of Plutella xylostella multiple nucleopolyhedrovirus (PlxyMNPV-CL3) exhibits a high degree of genetic similarity with the Autographa californica MNPV (AcMNPV) but is significantly more virulent against the diamondback moth, P. xylostella, than AcMNPV. To identify genetic differences between PlxyMNPV-CL3 and AcMNPV that may account for the difference in virulence against P. xylostella, the genome sequence of the CL3 plaque isolate of PlxyMNPV was determined and compared to the genome sequence of AcMNPV isolate C6. The PlxyMNPV genome is 134,417 bp, 523 bp larger than the AcMNPV-C6 genome, and the nucleotide sequence is almost completely co-linear with that of AcMNPV-C6. Of the 153 open reading frames (ORFs) identified in PlxyMNPV, 151 had homologues in AcMNPV-C6, with a mean amino acid sequence identity of 98.5%. The PlxyMNPV genome possessed two features previously reported for other variants of AcMNPV: (1) an extra baculovirus repeated orf (bro) sequence located between the plxy29/ac30 and sod ORFs, and (2) the deletion of the AcMNPV pnk/pnl polynucleotide kinase/ligase gene. In addition, an 817 bp insert of unknown origin located between the fp25K and lef-9 genes was discovered. This insert contained two small ORFs and was detected in both tissue culture- and larvae-derived PlxyMNPV DNA by PCR. Finally, the PlxyMNPV-CL3 ie-2 gene encodes a product with a low level (37.3%) of amino acid sequence identity with the AcMNPV-C6 ie-2 product. PlxyMNPV-CL3 apparently acquired this variant ie2 gene by recombination with an undescribed nucleopolyhedrovirus.

Genetic and biological variation among nucleopolyhedrovirus isolates from the fall armyworm, Spodoptera frugiperda (Lepidoptera: Noctuidae)

“…Prior to sequencing, amplimers were precipitated away from excess primers and deoxynucleotides as previously described [27]. In some cases, amplimers were purified by agarose gel electrophoresis or cloned using the TOPO TA Cloning Kit (Invitrogen, Carlsbad, CA).…”

Conserved baculoviral ORFs 10 and 14 from Bombyx mori multiple nucleopolyhedrovirus

“…The sequence that belongs to the ORF14 from BmMNPV had an identity of 99% with ORF23 of AcMNPV, which is related to viral folding protein (Gomi et al, 1999). It also had an identity of 88% with folding protein or F protein of PxMNPV (Harrison and Lynn, 2007), and with a folding protein of A. californica as well (Ayres et al, 1994). However, an identity of 84% was observed with a fusion protein of RoMNPV (Harrison and Bonning, 2003), whereas a 65% identity to MvMNPV was detected (Chen et al, 2008).…”

“…The analysis of ORF14 nucleotide sequence revealed that the sequence of MNPV subgroup of B. mori (BmMNPV) had identity equivalent to 99, 94, 93, 89, 79%, respectively, with: BmNPV genome, isolate T3 (Gomi et al, 1999); AcMNPV genome, clone C6 (Ayres et al, 1994); PxMNPV genome, isolate CL3 (Harrison and Lynn, 2007); RoMNPV genome (Harrison and Bonning, 2003), and MvMNPV genome (Chen et al, 2008). …”

Isolation of an Adoxophyes orana granulovirus (AdorGV) occlusion body morphology mutant: biological activity, genome sequence and relationship to other isolates of AdorGV

“…Two microlitres of a 100-fold dilution of the DNA templates was used in 50 ml PCRs as described by Rowley et al (2010) using primers designed to amplify the AdorGV ORFs p10, ORF 14, pep-p10, pep-2, ORF 30, p47, dbp, desmoplakin and AdorGV-E ORF 112 (Table S4). Amplimers were precipitated away from excess primers and nucleotides using 20 % PEG/2.5 M NaCl and subsequently sequenced with gene-specific primers as described by Harrison & Lynn (2007).…”