Characterization of two variants of Panolis flammea multiple nucleocapsid nuclear polyhedrosis virus

Abstract

The baculovirus Panolisflammea multiple nucleocapsid nuclear polyhedrosis virus (PfMNPV) was originally isolated from a natural virus epizootic and shown to consist of a mixture of variants. Two subclasses of variants (PfMNPV A and B) were identified by Southern blot hybridization, their polyhedrin genes being located on different restriction fragments. The proportion of the A and B variants changed according to the larval host in which the virus was propagated; PfMNPV(B) predominated in P. flammea but PfMNPV(A) was predominant in Mamestra brassicae. Bioassays of the two pure virus variants in M. brassicae larvae have shown the LDso values to be 4610 polyhedron inclusion bodies (pibs) for PfMNPV(A) and 5937 pibs for PfMNPV(B). Genomic DNA from the two variants was compared using restriction endonuclease analysis, and dot blot and Southern blot hybridization. Reciprocal quantitative dot blot hybridization analysis in 50% formamide showed PfMNPV(A) and PfMNPV(B) to be only distantly related to Autographa californica MNPV (less than 1%) and more closely related to M. brassicae MNPV (21 to 26%). The two PfMNPV variants exhibited a very high degree of identity to each other (nearly 100%) and therefore are very closely related. This was confirmed by physical mapping of the virus genomes. The nucleotide sequence of the polyhedrin gene of PfMNPV(B) was determined and compared with the published DNA sequences of other polyhedrin genes.

Genotypic and phenotypic diversity of a baculovirus population within an individual insect host

“…Most of the alterations within the genome are concentrated in one region (HindIII X/R fragments), although there are polymorphisms in two other areas. An earlier analysis of one of our variants (Pf7) also demonstrated changes in these three areas (Weitzman et al, 1992). Preliminary DNA sequencing of an enlarged HindIII R fragment (from Pf9) has shown that sequences corresponding to a chitinase gene are found within this region, with the closest match being to chitinases found in other NPVs (Paul, 1997).…”

Host mediated selection of pathogen genotypes as a mechanism for the maintenance of baculovirus diversity in the field

“…1). HindIII-F fragment variation was conWrmed by the presence of a larger fragment (F) and two smaller fragments (F 1 and F 2 ), as described by Weitzman et al (1992). Fig.…”

“…Hybridization analysis was carried out under high stringency conditions using [ -32 P]dATP labeled PaXNPV HindIII-R and -X fragments, using the method described by Sambrook et al (1989). Due to its high degree of sequence similarity, the MbMNPV polh was used to probe the PaXNPV HindIII-F fragments as described by Weitzman et al (1992). The probed membrane was exposed against a storage phosphor screen (Molecular Dynamics) and scanned using a phosphorimager (Typhoon 8600 Variable Mode Imager, Molecular Dynamics).…”

“…They identiWed three 'hotspots' of variation on the virus genome with most variation occurring around the HindIII-X and -R fragments although preliminary sequence data of these regions only identiWed the presence of a putative chitinase (Cory et al, 2005). Weitzman et al (1992) found that passaging Weld-collected PaXNPV, which had been propagated in M. brassicae on semi-artiWcial diet, through M. brassicae resulted in one of the genotypes (A, as opposed to B) dominating the population, as determined by Southern hybridisation of the HindIII-F fragment. This was previously sequenced by Oakey et al (1989) and found to contain the polyhedrin gene (polh).…”

“…This was previously sequenced by Oakey et al (1989) and found to contain the polyhedrin gene (polh). Weitzman et al (1992) identiWed an extra HindIII restriction site within PaXNPV B genotypes at this site, which after digestion with HindIII and hybridisation with the AcMNPV HindIII-V fragment (containing polh) resulted in either a single 8.10 kbp fragment (F) or two smaller 5.78 and 2.2 kbp fragments (F 1 and F 2 , respectively).…”