Characterization of DNA from three nuclear polyhedrosis viruses pathogenic for Choristoneura sp

Three nuclear polyhedrosis viruses isolated from larvae of the insect genus Choristoneura showed polyhedrins of 28–30,000 daltons, genome sizes of 78–82 × 10⁶ daltons, and guanine plus cytosine contents of 47.9–49.4%. It was demonstrated by comparison of restriction endonuclease fragment patterns that two of the viruses are closely related genetically.     

Sublethal effects of Bacillus thuringiensis on the spruce budworm, Choristoneura fumiferana

Exposing larvae of the spruce budworm, Choristoneura fumiferana (Clemens), to sublethal ( 50% lethal dose) levels of Bacillus thuringiensis subsp. kurstaki at various stages of their development significantly increased development time to the pupal stage and reduced pupal size and number of eggs laid per female, but did not affect the proportion of embryonated eggs. The changes in larval development time, pupal weight and fecundity depended on the larval stage that was treated. Exposure of fourth instars delayed larval development and reduced only male pupal weights with no effects on fecundity. Exposure of sixth instars delayed larval development to a lesser extent than exposure of fourth instars but had a pronounced effect on weight of both male and female pupae. The effect on pupal weight was sex dependent, as males tended to be more affected than females. The reduction in male pupal weight did not appear to influence fecundity, because the effect of exposure was explained by the change in female pupal weight. Effects on larval growth and pupal weight were proportional to the dose that was ingested during exposure, and were observed at doses as low as one-tenth of the LD₅₀. Ingestion of an LD₅₀ caused a 29 or 45% delay in development of, respectively, female or male larvae when exposed as fourth instars and a 30% reduction in female pupal weight when larvae were exposed as sixth instars.     

Toxicity of Bacillus thuringiensis entomocidal protein toward cultured insect tissue

The entomocidal protein from crystalline inclusion bodies of Bacillus thuringiensis can be bioassayed in vitro using cultured insect tissue. Larval cells of the spruce budworm, Choristoneura fumiferana, are damaged by enzyme-digested (activated) protein isolated from B. thuringiensis crystals. Measurement of toxicity is accomplished by detection of adenosine triphosphate (ATP) in treated cultures using firefly bioluminescence. The ATP content of toxin-treated tissue is inversely proportional to the amount of toxin added. Tissue cells from the spruce budworm exhibited maximum susceptibility to activated δ-endotoxin after 120 hr incubation. Probit analysis of tissue ATP response to toxin dose indicated 50% of the cells were damaged by 14.6 μg or less of toxin protein per 2 × 10⁵ insect tissue cells. Activated δ-endotoxin was entomocidal to insects as well, as detemined by mortality studies with second-instar larvae of the European corn borer. Electron microscopic observations of insect tissue treated with activated δ-endotoxin protein for 60 min revealed massive outer membrane disruption and subsequent loss of cytoplasmic constituents, accompanied by swelling of the nuclear membrane.     

A bioassay technique for Entomophthora sphaerosperma on the spruce budworm,Choristoneura fumiferana

A technique for conducting bioassays of Entomophthora sphaerosperma on sixth-instar larvae of the spruce budworm, Choristoneura fumiferana, was developed. Four assays were conducted by showering conidia on 10 larvae for each of 10 to 20 doses per assay. Dose was estimated by averaging estimates of the concentration of spores falling on water agar dishes before and after insect exposure. Maximum-likelihood probit analysis indicated significant regressions between log dose and probit mortality for all four assays. LC₅₀ values ranged from 11.21 to 18.77 spores/mm² with a weighted mean of 16.13 spores/mm². Slope estimates ranged from 0.92 to 1.87 with a weighted mean of 1.13. These low slope values may have been indicative of a highly variable test insect population, but also suggested a nontoxic infection process by the pathogen.     

Genetically distinct island populations of the Egyptian vulture (Neophron percnopterus)

The Egyptian vulture (Neophron percnopterus) is a species in decline throughout Europe, with the largest remaining breeding populations found in northern Spain. Iberian Peninsula populations of this species (about 1000 pairs) migrate to Africa in winter, while small populations in both the Canary and Balearic Islands (less than 40 pairs in each case) are apparently sedentary. We found that Egyptian vultures from both of these island groups were significantly differentiated from Iberian Peninsula populations (R _ST = 0.065–0.129, p = 0.000–0.007), using nine microsatellite loci isolated in a related species, the bearded vulture. The greatest degree of genetic differentiation was observed between the two island groups (R _ST = 0.279, p = 0.000). These island populations were more distinct from mainland groups than was a small sample of a well-defined separate subspecies from India (N. p. ginginianus; R _ST = 0.083–0.091, p = 0.023–0.024). This implies that these two island populations have been isolated from peninsular populations for many generations, despite the long-distance migration capabilities of the species. In constrast, populations within the Peninsula were not differentiated from one another at these microsatellite loci (R _ST = $-$0.004–0.007, p = 0.442–0.675). Introductions of Egyptian vultures from the larger northern breeding groups might therefore be appropriate in southern Spain, if necessary, but mainland birds should not be introduced to the islands if the genetic distinctiveness of these groups is to be preserved. Independent conservation plans are urgently required to protect these two island populations from extinction. This revised version was published online in July 2006 with corrections to the Cover Date.     

Isolation and characterization of microsatellite loci in Eurycorymbus cavaleriei,a dioecious endemic tree species in China

Eleven dinucleotide microsatellites were developed and characterized for Eurycorymbus cavaleriei, a dioecious, rare and endemic tree species in China. A genomic DNA enrichment protocol was used to isolate microsatellite loci and polymorphism was explored using 30 trees from two natural populations. The observed heterozygosity (H_O) ranged from 0.379 to 0.931, with an average of 0.625. These markers provide powerful tools for the ongoing population and conservation genetics studies of E. cavaleriei.     

Contrasting microsatellite variation between subalpine and western larch, two closely related species with different distribution patterns

Subalpine larch (Larix lyallii Parl.) and western larch (Larix occidentalis Nutt.) represent two closely related species with contrasting abundance and distribution patterns in Western North America. Genetic diversity at seven informative microsatellite loci was determined for 19 populations of subalpine larch and nine populations of western larch. Contrasting genetic diversity and patterns of population differentiation were observed between the two species. The overall within-population genetic diversity parameters were lower in subalpine larch (A = 3.2; A(P) = 3.6; H(E) = 0.418) than in western larch (A(P) = 5.51; H(E) = 0.580), a pattern that is likely related to historical or demographic factors. No evidence of interspecific hybridization was observed. Significantly more population differentiation (theta = 0.15; R(ST) = 0.07), consistent with more restricted gene flow, was observed for subalpine larch as compared to western larch (theta = 0.05; R(ST) = 0.04). Under the assumption of an infinite allele mutation model, 12 of the 19 subalpine larch populations showed signs of deviation from the mutation-drift equilibrium, which suggests Holocene population bottlenecks and fluctuations in effective population size for this species. None of the western larch populations deviated significantly from the mutation-drift equilibrium. For both species, Mantel's test revealed a significant positive relationship between geographical and genetic distances indicative of isolation by distance. A similar geographical structure was detected in both species, suggesting at least two genetically distinct glacial populations in each species. The various implications for gene conservation are discussed.