Triterpene acids from apple peel inhibit lepidopteran larval midgut lipases and larval growth.

Fruit extracts from apple, kiwifruit, feijoa, boysenberry, and blueberry were screened for the presence of lipase inhibitory compounds against lepidopteran larval midgut crude extracts. From 120 extracts, six showed significant inhibition with an extract from the peel of Malus × domestica cv. "Big Red" showing highest levels of inhibition. Because this sample was the only apple peel sample in the initial screen, a survey of peels from seven apple cultivars was undertaken and showed that, despite considerable variation, all had inhibitory activity. Successive solvent fractionation and LC-MS of cv. "Big Red" apple peel extract identified triterpene acids as the most important inhibitory compounds, of which ursolic acid and oleanolic acid were the major components and oxo- and hydroxyl-triterpene acids were minor components. When ursolic acid was incorporated into artificial diet and fed to Epiphyas postvittana Walker (Tortricidae: Lepidoptera) larvae at 0.16% w/v, a significant decrease in larval weight was observed after 21 days. This concentration of ursolic acid is less than half the concentration reported in the skin of some apple cultivars.

The effects of the broad-specificity lipase inhibitor, tetrahydrolipstatin, on the growth, development and survival of the larvae of Epiphyas postvittana (Walker) (Tortricidae, Lepidoptera).

The effects of the lipase inhibitor, tetrahydrolipstatin (THL), on neonate Epiphyas postvittana (Walker) (Lepidoptera, Tortricidae) larvae were investigated by feeding on control artificial diets (with and without 2% ethanol) and diets containing 2% ethanol and one of three concentrations of THL (0.011%, 0.037% and 0.11%). Small but significant reductions in growth rate, percent pupation and time to pupation were observed for larvae feeding on 2% ethanol control diet compared with standard control diet, but larger reductions in all parameters occurred with increasing THL concentration. Third instar larvae fed 0.011% THL in the diet had 40% of the midgut lipase activity in the relevant control larvae and showed up-regulation of gene expression of the gastric lipase-like family but not the pancreatic lipase-like family of midgut lipases.

Expression of various biotin-binding proteins in transgenic tobacco confers resistance to potato tuber moth, Phthorimaea operculella (Zeller) (fam. Gelechiidae).

The high affinity biotin-binding proteins (BBPs) avidin and streptavidin are established insecticidal agents, effective against a range of insect pests. Earlier work showed that, when expressed in planta, full length avidin and a truncated form of streptavidin are highly insecticidal. More recently, a wide range of BBPs, found in diverse organisms or engineered for various biotechnological applications have been reported. However, their effectiveness as plant-based insecticides has not been established. Here we report in planta expression of three different genes, designed to produce BBP variant proteins in the vacuole. The first was mature full length chicken avidin, the second a circularly permuted dual chain chicken avidin, and the third was an avidin homologue, a native bradavidin from Bradyrhyzobium japonicum. All three proteins were expressed in Nicotiana tabacum (tobacco). The transgenic tobacco lines were healthy, phenotypically normal and, when subjected to bioassay, resistant to the important cosmopolitan pest, potato tuber moth (Phthorimaea operculella) larvae at concentrations of ~50 ppm.

Expression of two heterologous proteins depends on the mode of expression: comparison of in vivo and in vitro methods.

The yield of two proteins, avidin and green fluorescent protein (GFP), expressed from a modified Autographa californica nucleopolyhedrovirus (AcMNPV), was compared in Sf9 cell culture monolayer, Sf21 cell suspension culture and intact Spodoptera litura larvae. GFP expressed from the p10 promoter yielded up to 1.5% of total soluble protein in larvae, 20-fold higher than that in monolayer suspension culture. Avidin, expressed from the polh promoter, yielded up to 2.3% of total soluble protein in larvae, 10-fold higher than that in suspension culture and 40-fold higher than that in monolayers. Avidin expression did not affect amounts of GFP in dual-expressing baculovirus compared with those detected from a GFP-only expressing AcMNPV. A biotin-binding assay showed that all avidin expressed in larvae was fully active. Glycosylation patterns of chicken-avidin and Spodoptera-avidin were very similar, though the latter showed a proportion of partially glycosylated material.

DNA diagnostics of three armored scale species on kiwifruit in New Zealand.

Three species of armored scale (Hemiptera: Diaspididae) are found on kiwifruit (Actinidia spp.) in New Zealand orchards: latania scale, Hemiberlesia lataniae (Signoret); greedy scale, Hemiberlesia rapax (Comstock); and oleander scale, Aspidiotus nerii (Bouché). Each of them is a quarantine pest in some of the markets to which New Zealand kiwifruit are exported. Adult females of the three species can be distinguished morphologically; however, the task is laborious when large numbers must be identified. Furthermore, it is not possible to distinguish among the immature stages. A DNA-based diagnostic using a multiplex polymerase chain reaction (PCR) method based on differences in the cytochrome oxidase I and II genes was developed to distinguish the three species. The test relies on the rapid isolation of amplifiable DNA by using a protease (prepGEM), followed by multiplex PCR using primers that distinguish the species at three or more nucleotide positions within cytochrome oxidase I and II, resulting in PCR products of characteristic size for each species. The test was validated in a double-blind experiment and then used to determine the relative distribution and abundance of the three species on leaves and fruit of 'Hayward' and 'Hortl6A' kiwifruit across the dominant growing regions throughout New Zealand during the 2007 season. In total, 3,418 scale insects were identified to species level: 1,904 (56%) were latania scale; 1,473 (43%) were greedy scale; and 41 (1%) were oleander scale. Since the last survey in 1988, latania scale has displaced greedy scale as the dominant species of armored scale on Hayward kiwifruit in the North Island and was found for the first time in the South Island. Only a single latania scale was found on Hortl6A fruit, consistent with previous reports of reduced rates of settlement on the fruit of this cultivar by latania scale compared with greedy scale.