Egg Albumin as a Protein Marker to Study Dispersal of Noctuidae in the Agroecosystem.

Knowledge of dispersal and spatial dynamics of pest populations is fundamental for implementation of integrated pest management and integrated resistance management. This study evaluated 1) the effectiveness of egg white albumin protein to mark larvae and adults of two polyphagous and highly mobile pests, Spodoptera frugiperda (J.E. Smith) (fall armyworm) and Helicoverpa zea (Boddie) (corn earworm) (Lepidoptera: Noctuidae), and 2) the sensitivity of polyvinylidene difluoride membrane (dot blot) in detecting albumin on marked insects. Laboratory and field experiments tested egg albumin as a protein marker, which was detected using two enzyme-linked immunosorbent assay (ELISA), microplate, and dot blot. In the laboratory, 100% of the moths sprayed with 20% egg white solution acquired the albumin marker, which was detected through the last time point tested (5 d) after application. Egg albumin was not effective at long-term marking of larvae, detected only prior the molting to the next instar. Albumin application in field cages resulted in a high percentage of moths detected as marked at 24 h and 5 d for both species. Egg albumin applied in the open field resulted in 15% of the recaptured corn earworm moths marked with most of them collected 150 m from the application area, although some were captured as far as 1,600 m within approximately 6 d after adult emergence. The results indicated egg albumin is a suitable marker to study the dispersion of fall armyworm and corn earworm in the agroecosystem and dot blot was as effective to detect egg albumin as was indirect ELISA.

Assessing resistance of sugarcane varieties to sugarcane borer Diatraea saccharalis Fab. (Lepidoptera: Crambidae).

In this study, we investigated resistance traits to the sugarcane borer Diatraea saccharalis Fab. (Lepidoptera: Crambidae) in the leaves and stalks of six sugarcane cultivars in a series of greenhouse and laboratory assays. Investigation of plant factors and infestation rates to better discriminate stalk damage by the sugarcane borer indicated that infestation of 7-month-old, single plants with 20 larvae at the third or fourth instar per plant was suitable to assess tunneling length. Three cultivars (i.e. SP803280, RB928064, and RB835486) had lower stalk damage (i.e. tunnel length) than cultivar SP891115, which exhibited relatively greater susceptibility to tunneling by the borer. The time required for the larvae to enter the sugarcane stalk was longer for cultivar SP803280, indicating resistance traits on the stalk surface, which correlated with lower stalk damage. Larvae feeding on SP813250 stalks had the lowest weight gain, indicating that this cultivar has resistance traits to larval development within its stalks. Cultivars RB867515 and SP891115 resulted in the highest mortality of early-stage larvae feeding on leaves, indicating the presence of resistance factors in their leaves. Multi-trait cluster and principal component analyses placed the cultivars into three and four clusters, respectively. The cultivars placed in different groups that exhibited resistance to leaf feeding, stalk entrance, and tunneling by the sugarcane borer could be used for crossings in sugarcane breeding programs with the goal of obtaining higher levels of resistance to D. saccharalis.

Magnitude and Allele Frequency of Cry1F Resistance in Field Populations of the Fall Armyworm (Lepidoptera: Noctuidae) in Brazil.

Bacillus thuringiensis (Bt) corn producing the Cry1F protein was the first highly efficacious Bt corn deployed against the fall armyworm Spodoptera frugiperda (J.E. Smith) (Lepidoptera: Noctuidae) in Brazil, but reduced efficacy of this technology against the fall armyworm has been reported in some regions of the country. Here, we surveyed Cry1F resistance allele frequency and susceptibility of eight S. frugiperda populations collected in 2013 from non-Bt fields in different regions of Brazil. In F1 screen experiments, the overall frequency of the Cry1F resistance alleles in Brazilian populations was estimated at 0.24, with 95% credibility interval between 0.18 and 0.25. In concentration-response bioassays, five of the eight populations surveyed exhibited significant resistance levels, which were over 32 times higher than that of the standard susceptible laboratory strain. The estimates of Cry1F resistance allele frequency were positively correlated with those of median effective or lethal concentrations (i.e., EC50 or LC50). These results show that the allelic frequency and the magnitude of Cry1F resistance are high in field populations of S. frugiperda in Brazil, indicating a challenging situation for resistance management.

Altered cysteine proteinase activity in insecticide-resistant strains of the maize weevil: purification and characterization.

Insecticide resistance is usually associated with fitness costs, but such costs may be mitigated by increased energy and amino acid accumulation and mobilization as has been suggested in the maize weevil Sitophilus zeamais (Coleoptera: Curculionidae). To address this adaptation, cysteine proteinases (E.C. 3.4.22), one of the main proteinases in weevils, was purified from an insecticide-susceptible and two insecticide-resistant strains of the maize weevil (one with fitness costs, referred as resistant-cost, and the other without it, referred to as resistant no-cost) using thiol-sepharose affinity chromatography. Purification of the cysteine proteinases revealed a single 74,000 Da molecular mass band in the susceptible strain, two bands of 72,000 and 83,000 Da in the resistant cost strain, and two bands of 68,000 and 74,000 Da in the resistant no-cost strain. Purified cysteine proteinases of the three strains behaved differently regarding casein degradation and inhibition; the proteinases least sensitive to inhibition by the specific cysteine proteinase inhibitor E-64 were those from the resistant no-cost strain as indicated by their highest I(50) value. The pH and temperature profile of cysteine proteinase activity differed among strains and although substrate affinity (i.e. K(M)) of the cysteine proteinases was similar, the V(max) value for cysteine-proteinases from the resistant cost strain was 3-fold and 5-fold higher than V(max) values for the resistant no-cost and susceptible strains respectively. Cysteine proteinase activity was highest for the resistant cost strain rather than the resistant no-cost. Therefore enhanced cysteine proteinase activity is unlikely to be playing significant role in mitigating the costs usually associated with insecticide resistance.

Influence of crop management practices on bean foliage arthropods.

Crop management practices can affect the population of phytophagous pest species and beneficial arthropods with consequences for integrated pest management. In this study, we determined the effect of no-tillage and crop residue management on the arthropod community associated with the canopy of common beans (Phaseolus vulgaris L.). Abundance and species composition of herbivorous, detritivorous, predaceous and parasitoid arthropods were recorded during the growing seasons of 2003 and 2004 in Coimbra County, Minas Gerais State, Brazil. Arthropod diversity and guild composition were similar among crop management systems, but their abundance was higher under no-tillage relative to conventional cultivation and where residues from the preceding crop were maintained in the field. Thirty-four arthropod species were recorded, and those most representative of the impact of the crop management practices were Hypogastrura springtails, Empoasca kraemeri and Circulifer leafhoppers, and Solenopsis ants. The infestation levels of major insect-pests, especially leafhoppers (Hemiptera: Cicadellidae), was on average seven-fold lower under no-tillage with retention of crop residues relative to the conventional system with removal of residues, whereas the abundance of predatory ants (Hymenoptera: Formicidae) and springtails (Collembola: Hypogastruridae) were, respectively, about seven- and 15-fold higher in that treatment. Importantly, a significant trophic interaction among crop residues, detritivores, predators and herbivores was observed. Plots managed with no-tillage and retention of crop residues had the highest bean yield, while those with conventional cultivation and removal of the crop residues yielded significantly less beans. This research shows that cropping systems that include zero tillage and crop residue retention can reduce infestation by foliar insect-pests and increase abundance of predators and detritivores, thus having direct consequences for insect pest management.

Partial purification and characterization of trypsin-like proteinases from insecticide-resistant and -susceptible strains of the maize weevil, Sitophilus zeamais.

Serine proteinases from three strains of Sitophilus zeamais (Coleoptera: Curculionidae), one susceptible and two resistant to insecticides--one exhibiting fitness cost (resistant cost strain) and the other lacking it (resistant no-cost strain), were partially purified using an aprotinin-agarose affinity column providing purification factors ranging from 36.5 to 51.2%, with yields between 10 and 15% and activity between 529 and 875 microM/min/mg protein with the substrate N-alpha-benzoyl-L-Arg-p-nitroanilide (L-BApNA). SDS-PAGE of the purified fraction revealed a 56,000 Da molecular mass band in all strains and a 70,000 Da band more visible in the resistant no-cost strain. The purified proteinases from all strains were inhibited by phenylmethyl sulphonyl fluoride (PMSF), N-alpha-tosyl-L-lysine chloromethyl ketone (TLCK), aprotinin, benzamidine and soybean trypsin inhibitor (SBTI) characterizing them as trypsin-like serine proteinases. Trypsin-like proteinases from the resistant strains exhibited higher affinity for L-BApNA. The resistant no-cost strain exhibited V(max)-values 1.5- and 1.7-fold higher than the susceptible and resistance cost strains, respectively. A similar trend was also observed when using N-alpha-p-tosyl-L-Arg methyl ester (L-TAME) as substrate. These results provide support to the hypothesis that the enhanced serine proteinase activity may be playing a role in mitigating physiological costs associated with the maintenance of insecticide resistance mechanisms in some maize weevil strains.

Inheritance of Cry1F resistance in laboratory-selected European corn borer and its survival on transgenic corn expressing the Cry1F toxin.

A major assumption of the high-dose/refuge strategy proposed for insect resistance management strategies for transgenic crop plants that express toxins from Bacillus thuringiensis is that resistance traits that evolve in pest species will be recessive. The inheritance of Cry1F resistance and larval survival on commercially available Cry1F corn hybrids were determined in a laboratory-selected strain of European corn borer, Ostrinia nubilalis (Hübner), displaying more than 3000-fold resistance to Cry1F. Concentration-response bioassays of reciprocal parental crosses indicated that the resistance is autosomal and recessive. Bioassays of the backcross of the F1 generation with the selected strain were consistent with the hypothesis that a single locus, or a set of tightly linked loci, is responsible for the resistance. Greenhouse experiments with Cry1F-expressing corn hybrids indicated that some resistant larvae survived the high dose of toxin delivered by Cry1F-expressing plants although F1 progeny of susceptible by resistant crosses had fitness close to zero. These results provide the first direct evidence that the high dose/refuge strategy currently in place to manage resistance in Cry1F-expressing corn is appropriate.

Seasonal mortality factors of the coffee leafminer, Leucoptera coffeella.

Seasonal population fluctuation of the coffee leafminer, Leucoptera coffeella (Guérin-Méneville & Perrottet) (Lepidoptera: Lyonetiidae), led to an investigation of its natural mortality factors during the rainy season when the population level is low and during the dry season when population peaks occur. Life-table data were collected from insecticide-free plots within a 3 ha coffee plantation on the upper, medium and lower canopy. Leafminer mortality was similar among the canopy parts but varied in the two seasons studied. During the rainy season, the generational mortality averaged 94.3%, with 50.2, 33.7 and 10.4% occurring during egg, larval and pupal stages, respectively. During the dry season, total mortality was 89%, with 13.2, 61.0 and 14.8% occurring during egg, larval and pupal stages, respectively. Marginal mortality rates during the rainy season were highest for physiological disturbances, rainfall and egg inviability; but, in the dry season, they were highest for predaceous wasps, physiological disturbances and parasitoids. Egg and larval stages accounted for most of the mortality variation in the rainy season, while the combination of larval and pupal mortality better described the generational mortality in the dry season. Variation in mortality during the rainy season was primarily associated with egg inviability, rainfall and parasitoids. In contrast, predatory wasps and physiological disturbances were the main factors associated with mortality variation during the dry season. These results suggest that weather conditions, natural enemies and plant quality attributes are the main determinants of the population dynamics of L. coffeella.