Particle film deters oviposition by Diaprepes abbreviatus (Coleoptera: Curculionidae)

A hydrophilic formulation of the inert silicate kaolin was tested in a screenhouse for its effect on the behavior of the root weevil Diaprepes abbreviatus (L.), a pest of citrus and ornamental plants in Florida and the Caribbean. Feeding by adults on treated foliage was reduced by 68-84% compared with adults fed untreated foliage. No insecticidal activity was detected after 14 d of exposure to kaolin-treated leaves. Oviposition was completely suppressed on treated foliage. Although females oviposited >19,000 eggs during two trials on untreated foliage, no egg masses were found on foliage treated with the kaolin formulation. These data indicate potential for kaolin as a barrier to oviposition in citrus groves and may prove to be an economically viable and environmentally sound component of an integrated approach for control of D. abbreviatus and related root weevils.     

Virulence of entomopathogenic nematodes to pecan weevil larvae, Curculio caryae (Coleoptera: Curculionidae), in the laboratory

The pecan weevil, Curculio caryae (Horn), is a key pest of pecans in the Southeast. Entomopathogenic nematodes have been shown to be pathogenic toward the larval stage of this pest. Before this research, only three species of nematodes had been tested against pecan weevil larvae. In this study, the virulence of the following nine species and 15 strains of nematodes toward fourth-instar pecan weevil was tested: Heterorhabditis bacteriophora Poinar (Baine, HP88, Oswego, NJ1, and Tf strains), H. indica Poinar, Karunakar & David (original and Homl strains), H. marelatus Liu & Berry (IN and Point Reyes strains), H. megidis Poinar, Jackson & Klein (UK211 strain), H. zealandica Poinar (NZH3 strain), Steinernema riobrave Cabanillas, Poinar & Raulston (355 strain), S. carpocapsae (Weiser) (All strain), S. feltiae (Filipjev) (SN strain), and S. glaseri (Steiner) (NJ43 strain). No significant difference in virulence was detected among nematode species or strains. Nematode-induced mortality was not significantly greater than control mortality (in any of the experiments conducted) for the following nematodes: H. bacteriophora (Baine), H. zealandica (NZH3), S. carpocapsae (All), S. feltiae (SN), S. glaseri (NJ43), and S. riobrave (355). All other nematodes caused greater mortality than the control in at least one experiment. Heterorhabditis megidis (UK211) but not H. indica (original) displayed a positive linear relationship between nematode concentration and larval mortality. Results suggested that, as pecan weevil larvae age, they may have become more resistant to infection with entomopathogenic nematodes.     

Cuticular hydrocarbons on elytra of the Diaprepes root weevil Diaprepes abbreviatus (L.) (Coleoptera: Curculionidae)

Abstract 1 External gland openings and associated structures on the elytra of teneral and mature Diaprepes root weevils, Diaprepes abbreviatus (L.), were elucidated by scanning electron microscopy (SEM). 2 There were clear differences between teneral, callow adults and fully mature adults. In the field, teneral adults remain in the pupal chamber in the soil until sclerotization of the cuticle is complete or nearly so. 3 Phenotypic variation of the elytra in this species consists of varying patterns and coloration of scaled intervals between a variable number of raised ridges devoid of scales. In addition to being thinner and lighter in colour than fully mature adults, the elytra of teneral adults were devoid of waxy hydrocarbon secretions. 4 External gland openings at the base of each scale were observed on teneral elytra and mature elytra washed with methylene chloride. 5 SEM evidence to document the production of waxy filaments by these glands and partial characterization of these by gas chromatography and mass spectrometry are presented.     

Effects of Culture Method and Formulation on the Virulence of Steinernema riobrave (Rhabditida: Steinernematidae) to Diaprepes abbreviatus (Coleoptera: Curculionidae)

The Diaprepes root weevil, Diaprepes abbreviatus, is a pest of vegetables, ornamental plants, sugarcane, and citrus in Florida and the Caribbean. The entomopathogenic nematode, Steinernema riobrave, can reduce larval populations of D. abbreviatus substantially. Efficacy of entomopathogenic nematodes, however, may be affected by culture method and formulation. Using D. abbreviatus as the host, we compared the efficacy of two commercial S. riobrave formulations, a liquid and a waterdispersible granule (WDG), with each other and with in vivo produced S. riobrave. Nematodes in the commercial formulations were produced in vitro through liquid fermentation; the in vivo nematodes were cultured in Galleria mellonella and applied in aqueous suspension. Laboratory experiments measured nematode virulence in plastic cups containing soil and seventh-eighth instar D. abbreviatus. One laboratory experiment was conducted using only fresh nematodes (less than 5 days old); another experiment included WDG nematodes that were stored for 25 days at 10 °C. Two field experiments were conducted in which nematodes were applied either to potted citrus (containing D. abbreviatus larvae) placed beneath mature citrus trees or to soil directly beneath the tree. In the latter experiment, efficacy was determined by measuring mortality of caged D. abbreviatus larvae that were buried beneath the soil surface prior to application. Mortality of D. abbreviatus treated with nematodes ranged from 80-98% and 50-75% in laboratory and field experiments, respectively. In all experiments, we did not detect any significant effects of culture method or formulation.     

Gut chitin synthase and sterols from larvae of Diaprepes abbreviatus (Coleoptera: Curculionidae)

Gut chitin synthase was characterized and the sterols and ecdysteroids in the sugarcane rootstalk borer weevil, Diaprepes abbreviatus, were identified. An in vitro cell-free chitin synthase assay was developed using larval gut tissues from D. abbreviatus. Subcellular fractionation experiments showed that the majority of chitin synthase activity was located in 10,000g pellets. The gut chitin synthase requires Mg²⁺ to be fully active: 7–8-fold increases in activity were obtained with 10 mM Mg²⁺ present in reaction mixture. Calcium also stimulated activity (4–5-fold with 10 mM Ca²⁺), while Cu⁺² completely inhibited at 1 mM. Other monovalent and divalent cations had little or no effect on activity. The pH and temperature optima were 7 and 25°C, respectively. Gut chitin synthesis was activated ca. 50% by trypsin treatments. GlcNAc stimulated chitin synthase activity, but Glc, GlcN and glycerin did not. Polyoxin D, UDP, and ADP inhibited the chitin synthase reaction with I₅₀'s of 75 μM, 2.3 mM, and 3.6 mM, respectively. Nikkomycin Z was a potent inhibitor of chitin synthase (91% inhibition at 10 μM). Tunicamycin and diflubenzuron had no effect on the enzyme. The apparent Km and V_max for the gut chitin synthase were, respectively, 122.5 ± 7.4 μM and 426 ± 19.7 pmol/h/mg protein utilizing UDP-GlcNAc as the substrate. Sterol analyses indicated that cholesterol was the major dietary and larval sterol. HPLC/RIA data indicated that 20-hydroxyecdysone was the major molting hormone.     

Sampling methods as abundance indices for adult Diaprepes abbreviatus (Coleoptera: Curculionidae) in citrus

Beat sampling and two type of traps, cup traps and Tedders traps, were evaluated as sampling methods to detect and estimate population densities of adult Diaprepes abbreviatus L. weevils newly colonizing young citrus trees. The study was conducted over a 65-wk period across a 0.25-ha area of 80 citrus trees [Citrus sinensis (L.) Osbeck] (1.2-1.5 m tall). Beat samples were taken weekly to determine the number of trees infested and number of new adult weevils per tree. Sixteen of the 80 trees studied were each monitored weekly using one of the following trapping methods: cup traps in trees, cup traps on a stake in the ground within the tree drip line, cup traps on a stake in the ground outside of the drip line, Tedders traps on the ground within the drip line, and Tedders traps on the ground outside of the drip line. Weevils collected each week from trees and traps were removed from the study site. Based on the coefficients associated with Taylor's power law, the optimum numbers of trees to sample for an SEM equal to 25% of the mean estimate decreased from 50 trees at a mean of 0.5 new weevils per tree to 30 trees at a mean of 0.8 new weevils per tree. A significant relationship was found between the weekly mean number of new weevils per tree and the proportion of trees infested, a binomial relationship that could be further explored in the search for a sampling program for adult D. abbreviatus. Regression analyses indicated that three of the trapping methods served at least as weak indices of the presence and abundance of new weevils: cup traps in trees, Tedders traps inside the dripline and Tedders traps outside the dripline. Cup traps in trees and Tedders traps inside the dripline captured the most weevils and most frequently detected weevils. Although relatively inefficient as abundance indices of populations of new weevils, these two trapping methods appeared to have some value with respect to signaling when weevils first appeared in trees during the spring.     

Virulence of entomopathogenic nematodes to larvae of the guava weevil, Conotrachelus psidii (Coleoptera: Curculionidae), in laboratory and greenhouse experiments

The guava weevil, Conotrachelus psidii, is a major pest of guava in Brazil and causes severe reduction in fruit quality. This weevil is difficult to control with insecticides because adults emerge over a long period, and larvae develop to the fourth-instar inside the fruit and move to the soil for pupation. We assessed the virulence of entomopathogenic nematodes to fourth-instar larvae in soil by comparing their susceptibility to nine species or strains: Heterorhabditis bacteriophora HP88, H. baujardi LPP7, and LPP1, H. indica Hom1, Steinernema carpocapsae All and Mexican, S. feltiae SN, S. glaseri NC, and S. riobrave 355. In petri dish assays with sterile sand at a concentration of 100 infective juveniles (IJs) of a given nematode species/strain, larval mortality ranged from 33.5 to 84.5%, with the heterorhabditids being the most virulent. In sand column assays with H. baujardi LPP7, H. indica Hom1, or S. riobrave 355 at concentrations of 100, 200, and 500 IJs, mortality was greater than the control only for H. baujardi (62.7%) and H. indica (68.3%) at the highest concentration. For H. baujardi LPP7 in a petri dish assay, the time required to kill 50 and 90% of the larvae (LT₅₀ and LT₉₀) for 100 IJs was 6.3 and 9.9 days, whereas the lethal concentration required to kill 50 and 90% of the larvae (LC₅₀ and LC₉₀) over 7 days was 52 and 122.2 IJs. In a greenhouse study with guava trees in 20-L pots, 10 weevil larvae per pot, and concentrations of 500, 1000 or 2000 IJs, H. baujardi LPP7 caused 30 and 58% mortality at the two highest concentrations. These results show that H. baujardi is virulent to fourth-instar larvae and has potential as a biological control agent in IPM programs.     

Reduced oviposition by Diaprepes abbreviatus (Coleoptera: Curculionidae) and growth enhancement of citrus by Surround particle film

Regularly applied sprays of a particle film, Surround WP, greatly enhanced the growth of citrus trees on a poorly drained Winder soil at Fort Pierce, FL. After 3 yr of applications every 3 or 4 wk, Surround-treated trees had at least 5 times the mass, 6 times the canopy volume, and approximately 4 times the cross-sectional area of the tree stems at the graft union compared with untreated trees. The larger Surround-treated trees attracted a higher number of adult weevil Diaprepes abbreviatus (L.) and to a lesser extent citrus root weevil, Pachnaeus litus (Germar), per tree, but there was an equivalent number of egg masses per tree compared with the control trees. The number of egg masses per female weevil oviposited on Surround-treated trees was significantly less than either the control trees or trees treated biannually with an entomopathogenic nematode, BioVector. The number of larvae per tree recovered from the roots of excavated trees was greater from trees treated with Surround once every 3 wk compared with control trees. The data suggest that Surround particle film greatly enhanced the growth of citrus trees grown in a poorly drained soil. The reduction in oviposition by D. abbreviatus was insufficient to significantly reduce the number of root weevil larvae per tree feeding on the roots. However, the more vigorous trees resulting from Surround applications may be more resistant or tolerant to root weevil feeding.     

Reproductive biology of Fidiobia dominica (Hymenoptera: Platygastridae), an egg parasitoid of Diaprepes abbreviatus (Coleoptera: Curculionidae)

The reproductive biology of Fidiobia dominica Evans (Hymenoptera: Platygastridae) was studied in the laboratory (25.6 +/- 1 degrees C) using host eggs of Diaprepes abbreviatus L. (Coleoptera: Curculionidae). F. dominica readily parasitized D. abbreviatus eggs on both host plant and wax paper substrates. The number of egg masses parasitized and the number of offspring produced were higher when females were offered more than one host egg mass but did not differ when either two or three egg masses were offered. Female parasitoids that were provided with host eggs and a honey food source lived significantly longer than those that were not provided a food source; however, they did not parasitize more D. abbreviatus eggs. Oviposition occurred in host eggs from 0 to 7 d old, and host mortality was relatively consistent for eggs 0-5 d old and lower for eggs 6-7 d old. Successful parasitoid emergence seldom occurred after host eggs were 4 d old, and by 7 d, no adults successfully emerged. Developmental time from egg to adult was 19.3 +/- 0.2 d for males, significantly more rapid than the females (20.4 +/- 0.1 d). The mean longevity of adult females was 8.0 +/- 0.4 d, with a mean oviposition period of 2.7 +/- 0.3 d; males survived 8.1 +/- 0.4 d. The demographic parameters including intrinsic rate of increase (r(m)), generation time (T), and net reproduction (R(o)) were 0.142/d, 22 d, and 22.4 female eggs/d, respectively.     

Abundance of Diaprepes abbreviatus (L.) (Coleoptera: Curculionidae) neonates falling to the soil under tree canopies in Florida citrus groves

The purpose of these experiments was to estimate the number and distribution of Diaprepes abbreviatus (L.) neonate larvae dropping from the canopy of infested citrus trees. The number of neonates was monitored in the field using passive funnel traps in two simultaneous experiments and a separate experiment for an additional year. In one experiment, traps were placed from trunk to dripline in the cardinal directions under each of five trees (132 traps total). In a second experiment, eight traps were placed under each tree in the cardinal directions, one trap 30 cm from the trunk and one trap 30 cm from the dripline/direction for 25 trees (200 traps total). Larvae were collected weekly for 50 wk in conical tubes containing ethylene glycol as a preservative. Traps closer to the tree trunk captured more larvae than traps nearer the dripline. The area under the tree canopy was positively correlated with the total estimated number of larvae captured per tree. The estimated number of total larvae/tree over the course of our experiments ranged from 955 to 7,290. The highest number of neonate larvae observed in 1 wk was 67 +/- 6/m2. There was an inverse relationship between the number of traps beneath a tree and the number of trees that needed to be sampled to estimate mean population density with a given precision. However, there was a direct relationship between number of traps/tree and the total number of traps needed for a given precision. This passive technique could be used to quantify the destructive larval stage and to assess D. abbreviatus management strategies.     

Storage proteins of the larval root weevil Diaprepes abbreviatus (Coleoptera: Curculionidae): riboflavin binding and subunit isolation.

Proteins present at high concentrations in hemolymph of the larval weevil Diaprepes abbreviatus were previously shown to bind a synthetic coumarin, 7-amino-3-phenyl coumarin (coumarin-10). One of the two native proteins previously identified (protein I) is now shown to separate into two distinct bands (proteins Ia and Ib) using native gradient pore-limiting electrophoresis. The high concentration of proteins Ia, Ib, and II in larval hemolymph, their disappearance from hemolymph upon pupation, and an apparent hexameric structure shown by chemical crosslinking identify them as hexameric storage proteins (hexamerins). At least one chromatographic form of Ib isolated by anion exchange HPLC is now shown to bind riboflavin (Rb). Binding was also demonstrated by quenching of Rb fluorescence by a partially isolated mixture of the storage proteins. Lipophorin did not quench Rb fluorescence. Rb was heat-extracted from whole hemolymph and identified by its fluorescence spectra and by reverse phase HPLC with fluorescence detection. The two subunits shared by the three holoproteins have been isolated by sequential density gradient ultracentrifugation, gel permeation HPLC, and reverse phase HPLC. All three holoproteins shared the alpha subunit (M(r) 75,000), while the beta subunit (M(r) 71,000) was lacking from one of the three. Repeated passage through an anion exchange column yielded two of the three proteins (Ib and II) in homogeneous form. Chemical crosslinking with dimethylsuberimidate indicated a hexameric structure for the holoproteins. All subunits and holoproteins stained as high mannose glycoproteins when probed with biotinylated concanavalin A on PVDF membranes. The alpha subunit was high in Met, His, and Thr, and the beta subunit was high in Lys. Both were high in Pro and had approximately 16% Phe+Tyr. Sequences of the 20 N-terminal amino acid residues of each subunit showed 45-60% homology between subunits. These coleopteran proteins also showed some sequential homology but no immunological cross-reactivity with storage proteins from the lepidopterans Galleria mellonella and Heliothis virescens.     

Larval growth of Diaprepes abbreviatus (Coleoptera: Curculionidae) and resulting root injury to three citrus varieties in two soil types

Larval growth and intraspecific competition of Diaprepes abbreviatus (L.) larvae and consequent root injury in container-grown citrus in the greenhouse were evaluated. Roots of Carrizo citrange, Citrus sinesis L. Osbeck x Poncirus trifoliata (L.) Raf.; Cleopatra mandarin, C. reticulata Blanco, and Swingle citrumelo, C. paradisi Macf. x P. trifoliata (L.) Raf. rootstock seedlings grown in Candler fine sand and potting soil were colonized with different populations of D. abbreviatus larvae. Larvae were exposed to the seedlings for 79 d. Larval growth and development increased steadily for approximately 70 d on all rootstock-soil combinations, at which time most larvae were instars 6-8. Most feeding injury occurred to roots when larvae were between instars 3 and 6. Larval weight reached a plateau at approximately 70 d, but often declined between 70 and 79 d. When larvae were small, injury to seedlings developed slowly, primarily on fibrous roots, then feeding increased rapidly, often resulting in total consumption of both fibrous root and bark tissue. Although not statistically significant, root injury developed slightly slower on Swingle citrumelo compared with Carrizo and Cleopatra rootstocks, but damage was comparable by 79 d. Little or no difference in consumptive benefit to the larvae was found between the rootstocks. Based on larval weight days, little feeding injury occurred during the first 21 d, but increased rapidly between 21 and 60 d. Soil type affected the rate of larval growth and development, with potting soil contributing to greater growth rates. Detritus in potting soil provided little or no nutritional resource, suggesting that the effect of potting soil on larval development was primarily physical. In addition, fewer inoculated larvae per seedling exhibited greater weight gains than higher infestation densities, suggesting that intraspecific competition for nutritional resources influenced larval development.     

Mitochondrial nucleotide variability in invasive populations of the root weevil Diaprepes abbreviatus (Coleoptera: Curculionidae) of Florida and preliminary assessment of Diaprepes sp. from Dominica

Diaprepes abbreviatus (L.) (Coleoptera: Curculionidae) is a root weevil introduced into the United States from the Caribbean in 1964. It is associated with >300 plants, including citrus, sugarcane, and potatoes. D. abbreviatus is widespread in Florida, and it has recently been detected in limited areas of California and Texas. The purpose of this research is to evaluate the utility of 16S ribosomal (16S rRNA) and cytochrome oxidase I (COI) mitochondrial markers for the delineation of genetic populations of D. abbreviatus in Florida and for the characterization of patterns of dispersion among these populations. We also assessed these markers as genetic tools for the clarification of taxonomic uncertainties in specimens from Dominica (Lesser Antilles). We analyzed 111 weevils from six Florida populations and six specimens from Dominica. In Florida, we found three haplotypes with only one haplotype in each population. Florida haplotypes differed by one to three nucleotide substitutions, possibly the result of a recent divergence from one source population or three different introductions from closely related populations from the Caribbean. In contrast, specimens from Dominica showed a high genetic variability with three 16S haplotypes and six unique COI haplotypes, delineating two mitochondrial clades. We show that these mitochondrial markers are useful for phylogeographic studies of D. abbreviatus.     

Status of biological control by egg parasitoids of Diaprepes abbreviatus (Coleoptera: Curculionidae) in citrus in Florida and Puerto Rico

Eggs of Diaprepes abbreviatus (L.) were routinely monitored in citrus groves at ten locations in Florida during 1997 and 1998 to study egg parasitism. One citrus location was studied in Puerto Rico. No native parasitoids were recovered from 1,337 D. abbreviatus egg masses studied in Florida citrus. In contrast, an average of 35.5% (range 12.5 to 68.8%) parasitism of egg masses was reported in Puerto Rico. The parasitoids Aprostocetus gala, Horismenus spp, and Quadrastichus haitiensis were recovered from the eggs of D. abbreviatusfstudied in Puerto Rico. The Horismenus parasitoids were suspected hyperparasitoids. Releases of the parasitoid Ceratogramma etiennei from Guadeloupe were initiated during 1998 at each of the Florida research sites. By the end of 1998, C. etiennei had been recovered from D. abbreviatus eggs at two of nine locations in Florida citrus. The parasitoid was recovered from 1 of 34 egg masses at one of these locations during the month of September and from 3 of 34 egg masses at the other location during the month of November. Whether or not C. etiennei establishes itself at one or more locations in Florida remains to be seen.     

Effect of neem seed extract on feeding,growth, survival,and reproduction of Diaprepes abbreviatus (Coleoptera: Curculionidae)

A commercially available neem seed extract, Neemix 4.5, containing 4.5% azadirachtin (AZA), was assessed for biological activity against the root weevil Diaprepes abbreviatus (L.), an important exotic insect pest of Florida citrus. Laboratory bioassays against neonatal and 3-wk-old larvae fed sliced carrot treated with Neemix produced dose-dependent larval mortality and reduced fresh weights among survivors of treatments. The weight response was greater than the mortality response for both larval age groups. Neonates treated with 45 mg/liter AZA weighed 60% less than those in the control after 4 wk. Three-week-old larvae treated with 45 mg/ liter AZA weighed 30% less than those in the control after 5 wk. When neonates were exposed to insect diet incorporated with Neemix, reductions in larval survival and weight were observed at concentrations as low as 4.8 mg/liter AZA after 6 wk. Larval growth was inhibited by >97% with 42.9 mg/liter AZA in the diet. A soil drench containing 30 mg/liter AZA reduced the survival and weight gain of neonates added to potted citrus and provided protection to the roots in a greenhouse experiment. A concentration of 90 mg/liter AZA was required to provide protection of citrus roots against 4-wk-old larvae. Reproductive effects were observed when adult weevils were fed foliage treated with Neemix. The numbers of larvae hatching per egg mass were reduced by 27% and 68% at 30 and 90 mg/liter AZA, respectively. These results suggest that Neemix should be further evaluated for use in integrated pest management (IPM) programs of citrus.     

Steinernema diaprepesi n. sp. (Rhabditida: Steinernematidae), a Parasite of the Citrus Root Weevil Diaprepes abbreviatus (L) (Coleoptera: Curculionidae)

A nematode collected from Diaprepes abbreviatus is identified and described as a new species, Steinernema diaprepesi n. sp. The new species is closely related to S. feltiae, S. glaseri, and S. oregonense and can be distinguished from these species by the following characteristics: Males: Spicule averaging 79 (71-90) µm and spicule shape; D% (distance from anterior end to excretory pore/ esophagus length × 100) about 80; the ratio SW (spicule length/anal body width) about 1.8. Females: Vulva with short, double- flapped epiptygma; tail terminus usually with 5 papillae-like structures. Infective juveniles: Body averaging 1,002 (880-1,133) µm, EP (distance from anterior end to excretory pore) = 74 (66-83) µm; tail length = 83 (65-91) µm, and E% (EP/tail length × 100) = 89.6 (78-114). Lateral field pattern variable, the formula for the arrangement of ridges from head to tail is: 2, 6, 7, 8, 4, 2. The portion with eight ridges is the longest. This new species can be differentiated further from three closest species (S. feltiae, S. glaseri, and S. oregonense) by characteristic sequences of their ITS regions, including sequence lengths, ratios of similarity, composition, and differences in base characters in sequence alignment.     

Cloning and characterization of a basic cysteine-like protease (cathepsin L1) expressed in the gut of larval Diaprepes abbreviatus L. (Coleoptera: Curculionidae)

Diaprepes abbreviatus is an important pest that causes extensive damage to citrus in the USA. Analysis of an expressed sequence tag (EST) library from the digestive tract of larvae and adult D. abbreviatus identified cathepsins as major putative digestive enzymes. One class, sharing amino acid sequence identity with cathepsin L’s, was the most abundant in the EST dataset representing 14.4% and 3.6% of the total sequences in feeding larvae and adults, respectively. The predominant cathepsin (Da-CTSL1) among this class was further studied. Three dimensional modeling of the protein sequence showed that the mature Da-CTSL1 protein folds into an expected cathepsin L structure producing a substrate binding pocket with appropriate positioning of conserved amino acid residues. A full-length cDNA was obtained and the proCTSL1 encoding sequence was expressed in Rosetta™ Escherichia coli cells engineered to express tRNAs specific for eukaryotic codon usage. The Da-CTSL1 was expressed as a fusion protein with GST and His₆ tags and purified in the presence of 1% Triton X-100 by Ni-NTA affinity and size exclusion chromatography. Recombinant mature Da-CTSL1 (23 KDa) exhibits optimal activity at pH 8, rather than at acidic pH that was shown of all previously characterized cathepsins L. Substrate specificity supports the hypothesis that Da-CTSL1 is a unique basic cathepsin L and protease inhibitor studies also suggest unique activity, unlike other characterized acidic cathepsin Ls. This paper describes for the first time a prokaryotic expression system for the production of a functional eukaryotic cathepsin L1 from larval gut of D. abbreviatus.     

Susceptibility of lady beetles (Coleoptera: Coccinellidae) to entomopathogenic nematodes

We investigated differential susceptibility of lady beetles to entomopathogenic nematodes, for two reasons: (1) to estimate potential nontarget effects on natural lady beetle populations, (2) to compare the susceptibility of exotic versus native lady beetle species. We hypothesize that successful establishment of some exotically introduced arthropods may be due, in part, to a lower susceptibility relative to competing native species. In laboratory studies, we compared the pathogenicity, virulence, and reproductive capacity of Heterorhabditis bacteriophora and Steinernema carpocapsae among two native (Coleomegilla maculata and Olla v-nigrum) and two successfully established exotic (Harmonia axyridis and Coccinella septempunctata) lady beetles, and a known susceptible lepidopteran host, Agrotis ipsilon. After 1 and 2 days of exposure to either nematode species, mortality of A. ipsilon was higher than in all lady beetles. Thus, we predict that nematode field applications would have significantly less impact on lady beetle populations than on a susceptible target pest. Additionally, the impact of soil-applied nematodes may be lower on lady beetles than on soil-dwelling hosts because the former spends relatively less time on the soil. Exotic lady beetles were less susceptible to nematode infection than native species. Reproductive capacity data also indicated lower host suitability in H. axyridis, but not in C. septempunctata. Overall, the hypothesis that low susceptibility to pathogens in certain exotic lady beetles may have contributed to competitive establishment was supported (especially for H. axyridis). Additional studies incorporating different hosts and pathogens from various geographic locations will be required to further address the hypothesis.