Pheromone-gland-specific fatty-acyl reductase in the adzuki bean borer,Ostrinia scapulalis (Lepidoptera: Crambidae)

The adzuki bean borer moth, Ostrinia scapulalis, uses a mixture of (E)-11- and (Z)-11-tetradecenyl acetates as a sex pheromone. At a step in the pheromone biosynthetic pathway, fatty-acyl precursors are converted to corresponding alcohols by an enzyme, fatty-acyl reductase (FAR). Here we report the cloning of FAR-like genes expressed in the pheromone gland of female O. scapulalis, and the characterization of a single pheromone-gland-specific FAR (pgFAR) and its functional assay using an insect cell expression system. As many as thirteen FAR-like genes (FAR-I–FAR-XIII) were expressed in the pheromone gland of O. scapulalis; however, only one (FAR-XIII) was pheromone-gland-specific. The deduced amino acid sequence of FAR-XIII predicted a 462-aa protein with a conserved NAD(P)H-binding motif in the N-terminal region, showing overall identity of 34% with the pgFAR of Bombyx mori. A functional assay using Sf9 cells transfected with an expression vector containing the open reading frame of the FAR-XIII gene has proven that FAR-XIII protein has the ability to convert a natural substrate, (Z)-11-tetradecenoic acid, to a corresponding alcohol, (Z)-11-tetradecenol.     

Transfection of Wolbachia in Lepidoptera: the feminizer of the adzuki bean borer Ostrinia scapulalis causes male killing in the Mediterranean flour moth Ephestia kuehniella

Two species of Lepidoptera, Ostrinia scapulalis and Ephestia kuehniella, harbour Wolbachia, which are maternally transmitted intracellular bacteria that often cause reproductive abnormalities in arthropods. While the infection in O. scapulalis causes conversion of genetic males into functional females (feminization), that in E. kuehniella induces cytoplasmic incompatibility. In the present study, we investigated the relative importance of host and Wolbachia factors in the differential expression of reproductive alterations in these insects. We transferred the Wolbachia harboured by O. scapulalis to E. kuehniella in which the original infection had been cured by tetracycline treatment. The transfected strain of E. kuehniella expressed a maternally inherited, female-biased sex ratio. Unexpectedly, two lines of evidence suggested that the sex ratio distortion was due to male killing. First, higher mortality of young larvae was observed. Second, the removal of the transferred Wolbachia resulted in the recovery of a 1:1 sex ratio, whereas the removal of a feminizer should result in a male-biased sex ratio among offspring. To the authors' knowledge, this is the first report that a single Wolbachia strain can cause two distinct sexual abnormalities in different hosts. Our observations highlighted the importance of host-Wolbachia interactions in determining the phenotype of reproductive alterations.     

Sexual mosaics induced by tetracycline treatment in the Wolbachia-infected adzuki bean borer, Ostrinia scapulalis.

Wolbachia-infected Ostrinia scapulalis (Lepidoptera: Crambidae) females generate all-female or nearly all-female broods. Curing the infection by tetracycline treatment during larval stages results in the generation of all-male broods in the next generation. Here we show that sexually mosaic offspring are produced by Wolbachia-infected females treated with tetracycline at the adult stage. The sexual mosaics had wings that were composed of distinctive female and male sectors. Besides wings, the sexually dimorphic mid tibiae displayed an intermediate morphology in some of the mosaics. Many of the mosaic individuals had an abnormal structure of the external genitalia as well, a combination of the male uncus and the female ovipositor. We assume that Wolbachia has a feminizing effect on O. scapulalis genetic males and, hence, incomplete curing of the Wolbachia infection results in the generation of sexually mosaic individuals.     

Feminizing Wolbachia in an insect,Ostrinia furnacalis (Lepidoptera: Crambidae)

Wolbachia, which forms a group of maternally inherited bacteria in arthropods, often cause reproduction alterations in their hosts, such as cytoplasmic incompatibility, parthenogenesis, male-killing, hybrid breakdown and feminization. To date, Wolbachia-induced feminization has been reported only in isopods. Here we report that a Wolbachia strain feminizes an insect host, Ostrinia furnacalis. Among 79 wild females of O. furnacalis examined, Wolbachia infection was detected in 13 females. Twelve of the 13 infected females produced all-female progenies, and this trait was maternally inherited. Tetracycline treatment of thelygenic matrilines resulted in the production of all-male progenies. The present findings indicate that the Wolbachia infection induces feminization of genetic males in O. furnacalis. Differences in the Wolbachia-induced feminization in O. furnacalis and that in isopods are discussed along with the differences in sex determination mechanisms between insects and isopods. Phylogenetic analysis of the wsp gene sequence of Wolbachia suggests independent evolutionary origins for the Wolbachia-induced feminizations in O. furnacalis and in isopods. Our findings over 5 years suggest that the infection has been maintained at a low prevalence in the O. furnacalis population.     

Conventional resistance of experimental maize lines to corn earworm (Lepidoptera: Noctuidae), fall armyworm (Lepidoptera: Noctuidae), southwestern corn borer (Lepidoptera: Crambidae), and sugarcane borer (Lepidoptera: Crambidae)

Plant resistance is a useful component of integrated pest management for several insects that are economically damaging to maize, Zea mays L. In this study, 15 experimental lines of maize derived from a backcross breeding program were evaluated for resistance to corn earworm, Helicoverpa zea (Boddie); fall armyworm, Spodoptera frugiperda (J. E. Smith); southwestern corn borer, Diatraea grandiosella Dyar; and sugarcane borer, Diatraea saccharalis (F.). Experimental line 100-R-3 was resistant in the field to leaf feeding by fall armyworm and line 116-B-10 was resistant in the field to leaf feeding by fall armyworm and leaf and stalk feeding by southwestern corn borer. When corn earworm larvae were fed field harvested silks from experimental line 81-9-B in the laboratory, their pupal weights were significantly lower than the pupal weights of larvae that were fed silks from the resistant control, Zapalote Chico. Maysin levels lower than those commonly associated with corn earworm resistance were present in the resistant experimental line, 107-8-7, indicating a new basis confers resistance to corn earworm in this line. These resistant experimental lines will provide plant breeders with new sources of resistance to lepidopterous insects for the development of improved maize breeding populations.     

cDNA cloning and in situ hybridization of Delta11-desaturase, a key enzyme of pheromone biosynthesis in Ostrinia scapulalis (Lepidoptera: Crambidae)

Female sex pheromones are considered to be produced in a "pheromone gland" located in the terminal abdominal segments (8th-10th, TAS) of a moth; however, in many moth species, the cells that produce pheromones have not actually been specified. We investigated cells in the TAS that synthesize pheromones in the adzuki bean borer Ostrinia scapulalis, by locating pheromones and their precursors, and mRNA for Delta11-desaturase, a key enzyme in pheromone biosynthesis. We demonstrated that the pheromone components, (E)-11- and (Z)-11-tetradecenyl acetates, and their fatty acyl precursors were specifically contained in the dorsal part of the TAS. A cDNA (OscaZ/E11) that encodes a Delta11-desaturase was cloned from the TAS. RT-PCR and in situ hybridization unequivocally showed that OscaZ/E11 is specifically expressed in the modified epidermal cells located at the dorsal end of the 8th-9th intersegmental membrane.     

Two mitochondrial lineages occur in the Asian corn borer, Ostrinia furnacalis (Lepidoptera: Crambidae), in Japan

The genealogy and diversity of the mitochondrial cytochrome oxidase subunit II (COII) gene were investigated for Ostrinia furnacalis in Japan. A preliminary examination of mitochondrial lineages in China and the Philippines was also made. Two lineages (A and B) were found in the COII gene. Lineage A was frequent throughout the Japanese main islands (Hokkaido, Honshu, Shikoku and Kyushu), while the frequency of lineage B varied among these islands. No clear patterns of geographical population structure were found. Population genetic features suggested that the O. furnacalis population harboring the lineage A mitochondria expanded in the recent past, while lineage B showed weak signals of a population expansion. It is not clear whether the two lineages of mtDNA evolved in separate or identical geographical regions. We discuss two hypotheses regarding the two lineages of mtDNA: a cryptic race/species hypothesis and a selective sweep hypothesis.     

Identification, cloning and expression of a Cry1Ab cadherin receptor from European corn borer, Ostrinia nubilalis (Hubner) (Lepidoptera: Crambidae)

Transgenic corn expressing the Cry1Ab toxin from Bacillus thuringiensis is highly toxic to European corn borer, Ostrinia nubilalis, larvae. A putative Cry1Ab receptor (OnBt-R(1)) molecule was cloned and sequenced from a cDNA library prepared from midgut tissue of O. nubilalis larvae. The 5.6 Kb gene is homologous with a number of cadherin genes identified as Cry1 binding proteins in other lepidopterans. Brush border membrane vesicles were prepared using dissected midguts from late instars. A 220-kDa protein was identified as a cadherin-like molecule, which bound to Cry1Ab toxin and cross-reacted with an anti-cadherin serum developed from recombinant expression of a partial O. nubilalis cadherin peptide. Two additional proteins of smaller size cross-reacted with the anti-cadherin serum indicating that Cry1Ab binds to multiple receptors or to different forms of the same protein. Spodoptera frugiperda (SF9) cells transfected with the OnBt-R(1) gene were shown to express the receptor molecule which caused functional susceptibility to Cry1Ab at concentrations as low as 0.1 microg/ml. These results in combination suggest strongly that a cadherin-like protein acts as receptor and is involved with Cry1Ab toxicity in O. nubilalis.     

Monitoring of the European corn borer (Lepidoptera: Crambidae) in central Maine

Pheromone trap types and within-field trap locations were compared for their effectiveness in monitoring the flight activity of European corn borer, Ostrinia nubilalis (Hübner), and its relationship to egg mass density and crop damage in sweet corn in central Maine from 1995 to 1996. The use of both 3:97 Z:E-11-tetradecenyl acetate and 97:3 Z:E-11 tetradecenyl acetate pheromone blends confirmed that European corn borer in central Maine is attracted to both pheromone lure types. European corn borer moths were captured predominantly with the E-lure type than with the Z-lure type in both years. The Scentry Heliothis trap was more effective than the Multi-Pher trap, but similar to the pheromone-baited water pan trap for monitoring European corn borer flights. With the Scentry Heliothis trap, the grassy border and 1st corn rows were the best locations for moth capture during the early flight period, but during the peak flight period, traps located in the middle of the field caught the most moths. Corn damage was recorded before moth captures in some sites and before egg mass counts in others, indicating poor efficacy of traps for early flights. Significant and positive correlations were found between moth captures in the midfield location and egg mass counts, and corn leaf damage, and between egg mass counts and corn leaf damage. However, low coefficients of variation suggest that pheromone trap captures were not good predictors of European corn borer leaf damage in sweet corn.     

Predicting economic losses from the continued spread of the Mexican rice borer (Lepidoptera: Crambidae)

The Mexican rice borer, Eoreuma loftini (Dyar) (Lepidoptera: Crambidae), is an invasive species that originated from Mexico, and it is threatening to cause major economic losses to sugarcane, Saccharum spp., and rice, Oryza sativa L., industries in Louisiana. The insect is expected to reach sugarcane and rice production areas in Louisiana by 2008, and infest all of Louisiana sugarcane and rice industries by 2035. When all sugarcane in Louisiana becomes infested, annual yield losses of $220 million would be expected for a cultivar of comparable susceptibility to LCP 85-384 (assuming this cultivar is planted on 100% of the production area). This also assumes the use of the current practice of rainfed production and one application of insecticide, which is presently used by farmers in Louisiana. Irrigation with 30 cm of water is predicted to reduce estimated losses by 29%, whereas four applications of a biorational insecticide such as tebufenozide are expected to reduce the loss in revenue by 53%. The use of the resistant 'HoCP 85-845' would reduce the projected loss in revenue by 24%. Combining all three management tactics on sugarcane, anticipated net loss in revenue would decrease by 66%. The rice industry in Louisiana is projected to suffer from a loss in revenue of $45 million when the entire state is infested. A 77% reduction in loss in revenue is expected with one application of lambda-cyhalothrin. A quarantine on east Texas sugarcane is estimated to save the Louisiana industry between $1.1 billion and $3.2 billion (depending on management) during the time needed for the insect to fully invade the state's sugarcane and rice producing area by natural migration rather than by accidental introduction. The rapid deployment of appropriate management tactics will have a key role in reducing the anticipated economic impact of E. loftini once it becomes a pest in Louisiana sugarcane and rice.     

Evaluating popcorn as a potential refuge of Ostrinia nubilalis (Lepidoptera: Crambidae)

Popcorn was evaluated in a series of experiments conducted over four growing seasons for its potential as a refuge for European corn borer, Ostrinia nubilalis (Hübner). Objectives of these studies were to determine whether more larvae were produced in popcorn than in field corn and to determine how popcorn influenced female oviposition and larval distribution in neighboring field corn. Two varieties of popcorn (M140, 105d and M3374Y, 118d), one mixture of popcorn (50% 105d and 50% 118d), and field corn (DK580, 108d) were evaluated. Number of egg masses, eggs per egg mass, and larvae were significantly higher in popcorn compared with field corn. Moth oviposition and larval distribution were evaluated using 105d popcorn embedded in several cornfields across Iowa. The row of field corn adjacent to popcorn had significantly more larvae compared with background field corn. In larger field experiments, O. nubilalis larval survival after overwintering was significantly different, with 2.2-18.7 times more O. nubilalis larvae surviving in popcorn than field corn. The potential use of popcorn as an O. nubilalis refuge for genetically engineered corn is considered.     

Sugarcane borer (Lepidoptera: Crambidae) resistance to transgenic Bacillus thuringiensis maize

Transgenic maize, Zea mays L., expressing the Bacillus thuringiensis (Bt) CrylAb toxin has been planted to extensive areas across the United States and several other countries, but no resistance has been documented in field populations oflepidopteran target pests. This article describes the first report of resistance alleles to commercially available Cry1Ab Bt maize in a Louisiana population of sugarcane borer, Diatraea saccharalis (F.) (Lepidoptera: Crambidae). Two hundred thirteen two-parent isolines of D. saccharalis were screened for Cry1Ab resistance on Bt maize leaf tissue using an F2 screening technique. Larvae representing three isolines survived >15 d on Bt tissue in the F2 generation. The second generation backcross progeny (B1F2) derived from isoline 52 completed larval development on Bt maize in the greenhouse. Segregation and resistance frequency analysis associated with isoline 52 suggested that Bt resistance is probably determined by a nearly completely recessive allele at a single locus. With this assumption, the estimated resistance allele frequency in this population is 0.0023, within a 95% confidence interval of 0.0003-0.0064.     

Analyses of Cry1Ab binding in resistant and susceptible strains of the European corn borer, Ostrinia nubilalis (Hubner) (Lepidoptera: Crambidae)

Cry1Ab toxin binding analysis was performed to determine whether resistance in laboratory-selected Ostrinia nubilalis strains is associated with target site alteration. Brush border membrane vesicles were prepared using dissected midguts from late instars of susceptible and resistant strains (Europe-R and RSTT) of O. nubilalis. Immunoblot analysis indicated that three different proteins bound to Cry1Ab toxin and were recognized by an anticadherin serum. In a comparison of resistant and susceptible strains, reduced Cry1Ab binding was apparent for all three bands corresponding to cadherin-like proteins in the Europe-R strain, while reduced binding was apparent in only one band for the RSTT strain. Real-time analysis of Cry1Ab binding to gut receptors using surface plasmon resonance suggested slight differences in affinity in both resistant strains. Additional binding analysis was conducted using 125I-labeled Cry1Ab, Cry1Ac, and Cry1Aa. Slight differences were again observed between the resistant and susceptible strains for Cry1Ab binding. However, when binding of 125I-labeled Cry1Aa was tested, a 10-fold reduction in the concentration of binding sites was observed in the Europe-R strain. Expression of the O. nubilalis cadherin gene was similar in both the resistant and susceptible strains and did not account for differences in binding. In combination, the results of the present work suggest that differences in susceptibility to Cry1A toxins in the Europe-R strain of O. nubilalis are associated with altered receptor binding, although the precise nature of this mechanism is still uncertain.     

Southwestern corn borer (Lepidoptera: Crambidae) damage and aflatoxin accumulation in maize

Aflatoxin, a potent carcinogen, is produced by the fungus Aspergillus flavus Link: Fr. Drought, high temperatures, and insect damage contribute to increased levels of aflatoxin contamination in corn, Zea mays L. Plant resistance is widely considered a desirable method of reducing aflatoxin contamination. Germplasm lines with aflatoxin resistance have been developed. This investigation was undertaken to determine whether crosses among these lines exhibited resistance to southwestern corn borer, Diatraea grandiosella Dyar, and to assess the effects of southwestern corn borer feeding on aflatoxin accumulation. Differences in ear damage among southwestern corn borer infested hybrids were significant. Estimates of general combining ability effects indicated that the lines Mp80:04, Mp420, and Mp488 contributed to reduced ear damage, and SC213 and T165 contributed to greater damage when used in hybrids. Mean aflatoxin levels were 254 ng/g for hybrids infested with southwestern corn borer larvae and 164 ng/g for noninfested hybrids in 2000 when environmental conditions were conducive to aflatoxin production. In contrast, the overall mean aflatoxin level for southwestern corn borer infested hybrids was only 5 ng/g in 1999 when environmental conditions did not favor aflatoxin accumulation. Crosses that included lines selected for aflatoxin resistance as parents (Mp80:04 and Mp313E) exhibited lower levels of aflatoxin contamination both with and without southwestern corn borer infestation in 2000. Only the experimental line Mp80:04 contributed significantly to both reduced southwestern corn borer damage and reduced aflatoxin contamination.     

Movement of Mexican rice borer (Lepidoptera: Crambidae) through the Texas rice belt

Pheromone-baited traps were used to monitor the movement of the Mexican rice borer, Eoreuma loftini (Dyar) (Lepidoptera: Crambidae), through the Texas rice belt from 2000 to 2005. Based on location of discovery in each county and year, the average rate of spread from 1980 to 2005 was 23 km/yr. From 2000 to 2005, the leading edge of the infestation has moved 16.5 km/yr toward Louisiana. The 1.8-fold increase (99% confidence interval) of the area occupied from 2000 to 2005 in the Texas rice belt indicates an expansion of the distribution of E. loftini. If movement continues to occur at similar rates, E. loftini will reach Louisiana by 2008.     

Olfactory neuron responsiveness and pheromone blend preference in hybrids between Ostrinia furnacalis and Ostrinia nubilalis (Lepidoptera: Crambidae)

The olfactory receptor neuron (ORN) and behavioral responses of hybrids between the Asian corn borer (ACB), Ostrinia furnacalis, and the E-strain European corn borer (ECB(E)), Ostrinia nubilalis were examined and compared to the parental populations. In hybrids and both parents, the large-spike-size ORN was capable of responding to all four pheromone components of ACB and ECB, despite differences in which compounds elicited the greatest spike frequency in each population. There was a small-spiking ORN more narrowly tuned to the minor pheromone components in both ACB and ECB(E). In hybrids the homologous small-spiking ORN was tuned primarily to the ECB(E) minor pheromone component, with some responsiveness to the ACB minor component. Both species and all the hybrids had an intermediate spike-size ORN tuned primarily to their common behavioral antagonist. Dominance of responsiveness to the ECB(E) versus the ACB minor pheromone component on the small-spiking ORN may explain the greater tendency of hybrids to fly upwind to the ECB(E) pheromone blend than the ACB blend. This finding points toward a distinct evolutionary role for this ORN in allowing a pheromone shift.     

Two multiplex sets of eight and five microsatellite markers for the European corn borer,Ostrinia nubilalis Hübner (Lepidoptera: Crambidae)

Primer sequence and polymorphism data are presented for 13 microsatellite loci isolated from the European corn borer moth, Ostrinia nubilalis, as part of a project to construct a linkage map for the two pheromone strains. Experimental conditions are described for polymerase chain reaction (PCR) multiplexing, which allows genotyping in two electrophoresis runs of eight and five markers each. In a sample of 27 individuals coming from one European locality, the number of alleles per locus ranged from one to 12, and gene diversity from 0 to 0.859. Seven loci showed a deficit of heterozygotes. Eleven loci cross‐amplify in the related Ostrinia furnacalis.     

Tolerance and compensatory response of rice to sugarcane borer (Lepidoptera: Crambidae) injury

A 3-yr field experiment was conducted to evaluate the tolerance and compensatory response of rice (Oryza sativa L.) to injury caused by sugarcane borer, Diatraea saccharalis (F.), as affected by cultivar (Cocodrie, Francis, and Jefferson), stage of crop growth during which the injury occurred (third tiller stage, panicle differentiation stage, and heading stage), and sugarcane borer density. The proportion of rice tillers with sugarcane borer injury (leaf and leaf sheath injury and/or stem injury) was lower when injury occurred at the third tiller stage (0.05) than at panicle differentiation (0.19) and heading (0.18). When injury occurred at the two latter stages, both the proportion of tillers with injury and the proportion of tillers with stem injury were negatively correlated with rainfall. Rainfall resulted in dislodgement and mortality of sugarcane borer eggs and larvae before the larvae entered the stems. Rice plant density in this study (111.1 plants/m2) was higher than recorded for previous research on rice compensation using potted rice or conducted in low-density hill production systems (26.7-51.3 plants/m2). Two mechanisms of within-plant tolerance/compensation were observed. Stem injured plants produced approximately 0.69 more tillers than uninjured plants, whereas tillers with leaf and leaf sheath injury produced larger panicles, up to 39.5 and 21.0% heavier than uninjured tillers, when injury occurred at third tiller stage and at panicle differentiation, respectively. Rice yield was not reduced with up to 23% injured tiller and up to 10% injured stems at the third tiller stage, 42% injured tillers and 17% injured stems at panicle differentiation, and 28% injured tillers and 14% injured stems at heading. Significant between-plant compensation was not detected, suggesting competition between adjacent plants is not significantly reduced by injury. Our results suggest that rice can tolerate and/or compensate for a level of stem borer injury previously considered to be economically damaging.     

Corn borer (Lepidoptera: Noctuidae and Crambidae) resistance of main races of maize from North America

Resistance to corn borers, Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae) and Sesamia nonagrioides (Lefebvre) (Lepidoptera: Noctuidae) in maize, Zea mays L., populations is partial, and more resistant populations are needed. The objective of this research was to compare resistance to corn borers of the main maize races from North America. Twenty open-pollinated maize populations belonging to the races Southern Dent, Corn Belt Dent, and Northern Flint, and three check populations, were evaluated under artificial infestation with S. nonagrioides and O. nubilalis. None of the populations had complete resistance. Northern Flint had the lowest yield under corn borer infestation, whereas Southern Dent had the highest yield but also the largest damage. Corn Belt Dent had a shorter growing cycle and similar yield of infected plants than Southern Dent. The checks had intermediate yield and resistance and were not significantly different from Corn Belt Dent for any trait. The Southern Dent populations Tennessee Red Cob and White Dent (PI221885 and PI311232) could be used as sources of tolerance to corn borers, although they are not expected to provide great gains compared with the levels of tolerance already present in some Corn Belt Dent and European Flint populations and would require adaptation to short growing cycle. The Corn Belt Dent synthetic BS17 had the highest yield and general agronomic performance under corn borer infestation, along with Rustler and Silver King, and the European Flint composite EPS13.     

Sampling for the sugarcane borer (Lepidoptera: Crambidae) on sugarcane in Louisiana.

A 3-yr study was conducted in 0.6- to 2.0-ha sugarcane fields throughout south Louisiana under varying sugarcane borer, Diatraea saccharalis (F.), density levels to determine the spatial dispersion of infestations and to develop a sequential sampling plan. Infestations of D. saccharalis were randomly dispersed. Infestation levels (percentage of stalks infested) ranged from 0.6 to 33.3%. Frequency distributions of the number of infested stalks indicated that the Poisson distribution best fit the data Tests of other distributions (negative binomial [aggregated], binomial [uniform], geometric, and hypergeometric) resulted in poorer fits. The sequential sampling plan devised, with lower and upper D. saccharalis infestation limits of 2 and 5% and 5 and 10%, required maximum average sample numbers of 7.1 and 5.5 (20-stalk samples), respectively, to make terminating management decisions. It is our assessment that implementation of these plans would decrease sampling effort by 50-60% when compared with sampling programs currently in use for D. saccharalis management decisions in Louisiana.