Field‐evolved resistance to Bt maize in sugarcane borer (Diatraea saccharalis) in Argentina

BACKGROUND

Maize technologies expressing Bacillus thuringiensis (Bt) insecticidal proteins are widely used in Argentina to control sugarcane borer (Diatraea saccharalis Fabricius). Unexpected D. saccharalis damage was observed to Bt maize events TC1507 (expressing Cry1F) and MON 89034 × MON 88017 (expressing Cry1A.105 and Cry2Ab2) in an isolated area of San Luis Province. Diatraea saccharalis larvae were sampled from MON 89034 × MON 88017 fields in the area to generate a resistant strain (RR), which was subsequently characterized in plant and diet bioassays.

RESULTS

Survivorship of the RR strain was high on TC1507 leaf tissue, intermediate on MON 89034 × MON 88017, and low on MON 810 (expressing Cry1Ab). The RR strain had high resistance to Cry1A.105 (186.74‐fold) and no resistance to Cry2Ab2 in diet bioassays. These results indicate resistance to Cry1F and Cry1A.105 (and likely cross‐resistance between them) but not to Cry1Ab or Cry2Ab2. Resistance to MON 89034 × MON 88017 was functionally recessive. Reviews of grower records suggest that resistance initially evolved to Cry1F, conferring cross‐resistance to Cry1A.105, with low refuge compliance as the primary cause. A mitigation plan was implemented in San Luis that included technology rotation, field monitoring, and grower education on best management practices (BMPs) including refuges.

CONCLUSION

In the affected area, the resistance to Cry1F and Cry1A.105 is being managed effectively through use of MON 89034 × MON 88017 and MON 810 in combination with BMPs, and no spread of resistance to other regions has been observed. © 2017 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.     

Development and characterisation of transgenic rice expressing two Bacillus thuringiensis genes

BACKGROUND: Transgenic crops that produce insecticidal toxins from the bacterium Bacillus thuringiensis Berliner (Bt) were first commercialised in 1996. The risk that pests have the potential to evolve resistance to Bt toxins is one of the most serious challenges to this technology. Gene stacking, pyramiding two Bt genes into one variety, is considered to be an effective insect resistance management (IRM) strategy. In this study, insect‐resistant rice expressing two Bt genes was developed by sexual crossing, and then characterised. RESULTS: Homozygous rice lines of two pyramided Bt genes were obtained in the F₃ generation. Quantification of Bt toxin showed that protein concentrations of Cry1Ab, Cry1Ac and Cry2A in the two‐gene lines were comparable with their single‐gene parents, while the expression of cry1C gene decreased after gene stacking. Four two‐gene lines showed higher activity to striped stem borer (Chilo suppressalis Walker) than parental lines in the laboratory bioassay. All pyramided lines and their hybrids exhibited excellent efficacy against stemborers and leaffolders in field evaluation, while most pyramided lines had no significant differences from original variety in yield under spraying of insecticide. CONCLUSION: These results demonstrate that the two‐gene lines have commercial potential and could serve as a valuable IRM strategy. Copyright © 2011 Society of Chemical Industry     

Bt maize and integrated pest management ‐ a European perspective

The European corn borer (Ostrinia nubilalis), the Mediterranean corn borer (Sesamia nonagrioides) and the western corn rootworm (Diabrotica virgifera virgifera) are the main arthropod pests in European maize production. Practised pest control includes chemical control, biological control and cultural control such as ploughing and crop rotation. A pest control option that is available since 1996 is maize varieties that are genetically engineered (GE) to produce insecticidal compounds. GE maize varieties available today express one or several genes from Bacillus thuringiensis (Bt) that target corn borers or corn rootworms. Incentives to growing Bt maize are simplified farm operations, high pest control efficiency, improved grain quality and ecological benefits. Limitations include the risk of resistance evolution in target pest populations, risk of secondary pest outbreaks and increased administration to comply with licence agreements. Growers willing to plant Bt maize in the European Union (EU) often face the problem that authorisation is denied. Only one Bt maize transformation event (MON810) is currently authorised for commercial cultivation, and some national authorities have banned cultivation. Spain is the only EU member state where Bt maize adoption levels are currently delivering farm income gains near full potential levels. In an integrated pest management (IPM) context, Bt maize can be regarded as a preventive (host plant resistance) or a responsive pest control measure. In any case, Bt maize is a highly specific tool that efficiently controls the main pests and allows combination with other preventive or responsive measures to solve other agricultural problems including those with secondary pests. Copyright © 2011 Society of Chemical Industry     

转cry1Ab/cry2Aj、cry1Ab/vip3DA玉米对棉铃虫、甜菜夜蛾和斜纹夜蛾的抗虫性评价

为评估转cry1Ab/cry2Aj、cry1Ab/vip3DA玉米对棉铃虫Helicoverpa armigera(Hübner)、甜菜夜蛾Spodoptera exigua(Hübner)和斜纹夜蛾Prodenia litura(Fabricius)的抗虫性,在室内测定了3个转cry1Ab/cry2Aj玉米品系和1个转cry1Ab/vip3DA玉米品系对3种害虫幼虫存活和生长发育的影响,研究了该系列Bt玉米不同组织器官对害虫的杀虫活性和控制效果。结果显示,棉铃虫初孵幼虫取食各品系Bt玉米叶片96h后死亡率为87.50%~90.00%,取食花丝和雌穗的幼虫96h后几乎全部死亡;甜菜夜蛾初孵幼虫取食各品系Bt玉米叶片、花丝和雌穗168h后死亡率为22.50%~68.33%,幼虫的生长发育受到明显抑制,体重抑制率达85.00%~95.00%;斜纹夜蛾初孵幼虫取食各品系Bt玉米叶片、花丝和雌穗96 h后死亡率显著高于非转基因亲本对照,168h后幼虫死亡率达90.00%以上。研究表明,转cry1Ab/cry2Aj和cry1Ab/vip3DA玉米品系对棉铃虫和斜纹夜蛾的初孵幼虫表现出较好的抗性,可以作为转多基因抗虫玉米育种的备选材料。     

Impact of Cry3Bb1-expressing Bt maize on adults of the western corn rootworm, Diabrotica virgifera virgifera (Coleoptera: Chrysomelidae)

BACKGROUND: Genetically engineered maize producing insecticidal Cry3Bb1 protein from Bacillus thuringiensis (Bt) is protected from root damage by corn rootworm larvae. An examination was made to establish whether western corn rootworm (Diabrotica virgifera virgifera) adults are affected by Cry3Bb1‐expressing maize (MON88017) when feeding on above‐ground tissue. RESULTS: In laboratory bioassays, adult D. v. virgifera were fed for 7 weeks with silk, leaves or pollen from Bt maize or the corresponding near‐isoline. Male, but not female, survival was reduced in the Bt‐leaf treatment compared with the control. Female weight was lower when fed Bt maize, and egg production was reduced in the Bt‐silk treatment. ELISA measurements demonstrated that beetles feeding on silk were exposed to higher Cry3Bb1 concentrations than beetles collected from Bt‐maize fields in the United States. In contrast to silk and pollen, feeding on leaves resulted in high mortality and low fecundity. Females feeding on pollen produced more eggs than on silk. C:N ratios indicated that silk does not provide enough nitrogen for optimal egg production. CONCLUSIONS: Direct effects of Cry3Bb1 on adult beetles could explain the observed effects, but varietal differences between Bt and control maize are also possible. The impact of Bt maize on adult populations, however, is likely to be limited. Copyright © 2011 Society of Chemical Industry     

Bt Transgenic Crops: Risks and Benefits

Bt crops, predominantly maize and cotton hybrids, transgenically expressing cry genes derived from Bacillus thuringiensis, were planted on approximately 14 million hectares (worldwide) in 1999. Preliminary reports suggest that in most cases pesticide use was reduced, and in some situations there were significant increases in yields and profits. However, assemblages of secondary pests – such as aphids, plant bugs and thrips – also exist in Bt crops, and although the overall need for scouting and chemical control is reduced in Bt crops, there may be a requirement for additional, conventionally applied chemicals to control such non-target pests.Naturally-occurring Bt toxins with activity against a wide variety of pest species have been discovered and are thus potentially available for engineering into Bt crops to control a broader spectrum of pests than are currently targeted. New Bt crops and second-generation products incorporating an expanding range of Cry toxins and other arthropod targeted genes are in development and could become available for introduction to the market within the next few years.Insecticide resistance management (IRM) strategies for Bt crops are reviewed in the context of studies on selection pressures and the potential for resistance development in target populations. The so-called, high dose strategy, combined with the use of refuges, is widely agreed to be the best technical approach for managing resistance, and evidence is accumulating that separate refuges are more effective at conserving pest susceptibility than mixed refuges. A widespread consensus on the necessity for such measures, and an appreciation of the importance of multi-tactical approaches, has developed. Monitoring programmes, protocols and studies relevant to detecting the early development of resistance to Bt Cry toxins are described.Field monitoring of non-target entomofauna has not revealed significant differences in the abundance or biodiversity of beneficial insects associated with Bt maize. Indeed, laboratory studies of effects on parasitoids suggest that Bt plants may even have an environmental advantage over broad spectrum pesticides. However, more complex, multi-trophic, long-term experiments are needed to thoroughly assess the compatibility of Bt crops with non-target invertebrates and to define the complex relationship between IRM, target species and their natural enemy assemblages. Studies on the effects of transgenically-expressed Cry toxins on non-target insects, and their persistence in soil and on leaves, is reviewed. It is suggested that there is currently no generally agreed framework, or methodology, within which ad hoc experimental results can be accommodated, and each crop-transgene combination has to be assessed on a case-by-case basis. Studies proposing a conceptual approach to evaluating risks associated with Bt crops are highlighted and potential benefits and hazards are reviewed.     

Microarray analysis of global gene regulation in the Cry1Ab-resistant and Cry1Ab-susceptible strains of Diatraea saccharalis

BACKGROUND: Extensive adoption of transgenic Bt corn in recent years for stalk borer control has increased risk of resistance evolution in the target pest populations. A Bt‐resistant strain of the sugarcane borer, Diatraea saccharalis, was approximately 100‐fold more tolerant to Cry1Ab toxin than the susceptible counterpart. To gain a better understanding of the molecular mechanisms of Bt resistance, the Cry1Ab‐susceptible (Cry1Ab‐SS) and Cry1Ab‐resistant (Cry1Ab‐RR) strains of D. saccharalis were subjected to a microarray analysis. RESULTS: Results showed that the expression levels of many genes were significantly different between the Cry1Ab‐RR and Cry1Ab‐SS strains. Microarray analysis of 7145 cDNAs revealed 384 differentially expressed genes. A total of 273 genes were significantly upregulated 2–51.6‐fold, and 111 genes were significantly downregulated 2–22.6‐fold in the Cry1Ab‐RR strain. The upregulation of three potential resistance‐related genes, coding for a glutathione S‐transferase (GST), a chymotrypsin‐like protease (CHY) and a lipase (LP), was confirmed using real‐time PCR, indicating a reproducibility of the microarray data. Ontology analysis revealed that more than twice the number of metabolic‐related genes were upregulated compared with downregulated genes with the same biological function. Up to 35.2% of the upregulated genes in the resistant strain were associated with catalytic activity, while only 9.5% of the downregulated genes were related to the same catalytic molecular function. CONCLUSION: The large portion of metabolic‐ or catalytic‐related genes with significant upregulations indicated a potential large increase in metabolic or catalytic activities in the Cry1Ab‐RR strain. This cDNA microarray gene expression data could be used to characterize and identify new genes that may be associated with Bt resistance in D. saccharalis. Copyright © 2012 Society of Chemical Industry     

Developmental studies of transgenic maize expressing Cry1Ab on the African stem borer, Busseola fusca; effects on midgut cellular structure

BACKGROUND: Busseola fusca is a major pest of maize in Africa but unfortunately is difficult to control using chemical insecticides. Insect‐resistant transgenic crops may provide an alternative viable strategy to control this pest. RESULTS: Recombinant Cry1Ab (1%) reduced larval weight by 60% over the trial period, while larval weight in the control group increased by 25%; no effects on mortality were observed. Insect survival, developmental rate and pupal and adult weight were significantly reduced (P < 0.05) on maize expressing Cry1Ab (MON810) compared with the non‐transformed parental line. These differences were more pronounced with second‐instar larvae than with third‐instar larvae. Leaf area consumed by Bacillus thuringiensis (Bt)‐fed larvae was significantly lower (0.5 cm² larva^?1 day^?1) compared with the area consumed by control‐fed insects (3.3 cm² larva^?1 day^?1). EM studies revealed that consumption of Bt maize deleteriously affected gut integrity. Effects were observed in columnar cells of the midgut epithelium, with the cytoplasm becoming highly vacuolated; the microvilli were disorganised, the mitochondria were abnormal and there was an increase in the number of lysosomal bodies. The rough endoplasmic reticulum had also become dilated. CONCLUSION: This study confirms the potential for Bt maize, when used as part of an IPM programme, for control of B. fusca. Copyright © 2011 Society of Chemical Industry     

Assessment of the high-dose concept and level of control provided by MON 87701 × MON 89788 soybean against Anticarsia gemmatalis and Pseudoplusia includens (Lepidoptera: Noctuidae) in Brazil

BACKGROUND: Genetically modified MON 87701 × MON 89788 soybean (Glycine max), which expresses the Cry1Ac and EPSP‐synthase proteins, has been registered for commercial use in Brazil. To develop an Insect Resistance Management (IRM) program for this event, laboratory and field studies were conducted to assess the high‐dose concept and level of control it provides against Anticarsia gemmatalis and Pseudoplusia includens. RESULTS: The purified Cry1Ac protein was more active against A. gemmatalis [LC₅₀ (FL 95%) = 0.23 (0.15–0.34) µg Cry1Ac mL^?1 diet] than P. includens [LC₅₀ (FL 95%) = 3.72 (2.65–4.86) µg Cry1Ac mL^?1 diet]. In bioassays with freeze‐dried MON 87701 × MON 89788 soybean tissue diluted 25 times in an artificial diet, there was 100% mortality of A. gemmatalis and up to 95.79% mortality for P. includens. In leaf‐disc bioassays and under conditions of high artificial infestation in the greenhouse and natural infestation in the field, MON 87701 × MON 89788 soybean showed a high level of efficacy against both target pests. CONCLUSIONS: The MON 87701 × MON 89788 soybean provides a high level of control against A. gemmatalis and P. includes, but a high‐dose event only to A. gemmatalis. Copyright © 2012 Society of Chemical Industry     

The western corn rootworm,a new threat to European agriculture: opportunities for biotechnology?

BACKGROUND: During the early 1990s, the western corn rootworm, Diabrotica virgifera virgifera Le Conte (WCR), a maize pest, invaded the European continent. The continuous spread of the pest has introduced a new constraint into European maize production. As the damage caused by the invasive species is highly variable and different crop protection (CP) strategies are available, farmers' optimal strategies are not obvious. This study uses a simulation model to assess the competitiveness of different CP strategies in seven Central European countries. RESULTS: Results indicate a high degree of heterogeneity in the profitability of different CP strategies, depending on the production parameters in each country. In general, crop rotation and Bt maize offer the best solutions to farmers, but, in continuous (non‐rotated) maize cultivation, chemical CP options may capture part of the market. For Austrian continuous maize production it is found that not deregulating Bt maize implies that farmers forego revenues of up to €59 ha^?1. CONCLUSIONS: In the presence of WCR, producing maize by an economically sound method requires incorporating country‐ and farm‐specific characteristics into the decision framework. Also, not deregulating Bt maize has direct monetary consequences for many farmers that could influence total maize output and resistance management. Copyright © 2010 Society of Chemical Industry     

Inheritance of resistance to Bacillus thuringiensis Cry1Ac toxin in Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) from India

BACKGROUND: The cotton bollworm, Helicoverpa armigera (Hübner), is one of the most serious insect pests of cotton. It has developed resistance to almost all groups of chemical insecticides because of their intensive use. The failure of insecticides to control H. armigera has been a strong incentive for the adoption of transgenic cotton (Bt cotton). However, the value of Bt could be diminished by widespread resistance development to Bt toxins in insect populations. Therefore, understanding the genetic basis of resistance is essential for developing and implementing strategies to delay and monitor resistance. RESULTS: A resistant strain designated as BM‐R was obtained from the cross of adults from Bathinda () and Muktsar (), Punjab, India, which showed the highest survival (60.68%) and LC₅₀ value (1.396 µg mL^?1 diet). Similarly, a laboratory‐maintained strain from Hoshiarpur, Punjab, showed maximum susceptibility to Cry1Ac toxin with the lowest LC₅₀ value (0.087 µg mL^?1), and was designated as HP‐S. The genetic purity of both strains was confirmed by RAPD profile analysis at each generation, and genetic similarity reached more than 90% after the third generation. Continuous maintenance of the resistant BM‐R strain on Cry1Ac resulted in an increase in LC₅₀ from 0.531 µg mL^?1 in F₀ to 4.28 µg mL^?1 in F₁₄ and 7.493 µg mL^?1 in F₁₉, while the LC₅₀ values for HP‐S larvae on diet without Cry1Ac increased to 0.106 and 0.104 µg mL^?1, which lay within the fiducial limits of the baseline LC₅₀ value. The mode of inheritance of resistance was elucidated through bioassay response of resistant, susceptible heterozygotes and backcross progeny to Cry1Ac incorporated in semi‐synthetic diet. CONCLUSION: Based on dominance, degree of dominance and backcross values, resistance was inferred to be polygenic, autosomal and inherited as a recessive trait. Copyright © 2011 Society of Chemical Industry     

Testing insecticide resistance management strategies: mosaic versus rotations

BACKGROUND: Developing scientifically valid, economically acceptable insecticide resistance management (IRM) programs is critical for sustainable insect management. The diamondback moth, Plutella xylostella (L.), has demonstrated an ability to develop resistance to many different classes of insecticides, including proteins produced by the bacterium Bacillus thuringiensis Berliner (Bt). Recently it has developed resistance to the novel compounds spinosad and indoxacarb. In greenhouse cage experiments, a laboratory‐selected population of P. xylostella resistant to spinosad, indoxacarb and Bt was used to compare population growth and resistance evolution if these three insecticides were rotated or used in a mosaic fashion. RESULTS: The average population density through nine generations was lowest in the treatment in which the insecticide was rotated every generation (R‐1) (x? = 20.7 ± 3.20) compared with the treatment in which the insecticide was rotated every third generation (R‐3) (x? = 41.4 ± 17.6) or where the insecticides were applied as a mosaic (M) (x? = 41.8 ± 6.53). After nine generations, the survival of resistant individuals increased for each insecticide (7.2–73.5%) compared with the population without selection (CK) (0.73–3.1%). Survival on spinosad was significantly lower (23.7%) in the single‐generation rotation than for the other two treatments, both of which exceeded 72%. The calculated survival on all three insecticides treated simultaneously, according to the survival on each insecticide, was 0.26, 0.81 and 1.6% for R‐1, R‐3 and M treatments respectively. CONCLUSION: Results of both population density and resistance development indicated that insecticide rotation every generation was better for IRM than if the insecticide was rotated every third generation or if the three insecticides were applied as a mosaic. Copyright © 2010 Society of Chemical Industry     

Elucidating resistance in cotton genotypes to whitefly, <Emphasis Type="Italic">Bemisia tabaci,</Emphasis> by population buildup studies

Plant resistance has become an important component of integrated pest management (IPM) for management of whitefly, Bemisia tabaci (Gennadius) (Homoptera: Aleyrodidae), an important pest of cotton in India. The present studies were undertaken to standardize the plant stage and identify resistant cotton genotypes against whitefly. Nine plant stages of F846, a susceptible cotton genotype, were exposed to whitefly for 25 days under no-choice conditions. The population buildup (eggs, nymphs, pupae and adults) was recorded. The 12-, 14- and 16-leaf stages were suitable for plant resistance studies against whitefly, and the 14-leaf stage was taken for further studies. Ten cotton genotypes of Gossypium hirsutum and two of G. arboreum were covered with split cages in which five pairs of B. tabaci (F₁) were released. The population buildup was recorded to categorize genotypes as resistant, moderately resistant, moderately susceptible or susceptible. The experiment was repeated with F₂ and F₃ generation whiteflies. Based on overall average score of three experiments, LD694 was rated as resistant; LK861, Supriya, RS2013, CNH911 and PA183 as moderately resistant; IS-376/4/1/20/72, NHH44, TxMaroon2-78, Bt 6304 and RS2098 as moderately susceptible; and F846 as susceptible. LD694 was found to be resistant in three consecutive generations of whitefly.     

Pyramiding Ty‐2 and Ty‐3 genes for resistance to monopartite and bipartite tomato leaf curl viruses of India

Breeding resistance to whitefly‐transmitted begomoviruses is an important goal of tomato breeding programmes worldwide. So far, resistance to begomoviruses in tomato has been achieved using wild species, and at least five resistance genes (Ty genes) have been studied. The present study was undertaken to combine Ty‐2 and Ty‐3 and to determine the effect of pyramiding on infection of tomato by three diverse begomovirus species. The diagnostic ability of the markers linked to Ty genes was assessed and marker‐assisted selection was used to develop pyramided tomato lines from the crosses between Ty stocks. Five stable pyramided tomato lines that differ in fruit morphology and yield potential were developed. The horticultural performance of pyramided lines in field trials showed that the yield and horticultural traits are well maintained in the plants. The response of these lines was assessed using agroinoculation and field tests in a disease hotspot. The pyramided lines and Ty‐3‐carrying lines exhibited a high level of resistance to the monopartite and two bipartite begomoviruses tested. The pyramided tomato lines developed in this study could be important genetic resources for sustainable tomato production in areas affected by tomato leaf curl virus disease. The combined results of disease resistance tests also showed that Ty‐3 is critical for achieving broad‐spectrum resistance. The limitations of relying on a single gene and the importance of pyramiding are discussed in the light of available evidence on frequent recombination in begomoviruses.     

Impact of seed blend and structured maize refuge on Helicoverpa zea (Lepidoptera: Noctuidae) potential phenological resistance development parameters in pupae and adults

BACKGROUND

Helicoverpa zea, an economic pest in the south-eastern United States, has evolved practical resistance to Bacillus thuringiensis (Bt) Cry toxins in maize and cotton. Insect resistance management (IRM) programs have historically required planting of structured non-Bt maize, but because of its low adoption, the use of seed blends has been considered. To generate knowledge on target pest biology and ecology to help improve IRM strategies, nine field trials were conducted in 2019 and 2020 in Florida, Georgia, North Carolina, and South Carolina to evaluate the impact of Bt (Cry1Ab + Cry1F or Cry1Ab + Cry1F + Vip3A) and non-Bt maize plants in blended and structured refuge treatments on H. zea pupal survival, weight, soil pupation depth, adult flight parameters, and adult time to eclosion.

RESULTS

From a very large sample size and geography, we found a significant difference in pupal mortality and weight among treatments in seed blends with Vip3A, implying that cross-pollination occurred between Bt and non-Bt maize ears. There was no treatment effect for pupation depth, adult flight distance, and eclosion time.

CONCLUSION

Results of this study demonstrate the potential impact of different refuge strategies on phenological development and survival of an important pest species of regulatory concern. © 2023 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.     

Interaction of proteinase inhibitors with Cry1Ac toxicity and the presence of 15 chymotrypsin cDNAs in the midgut of the tobacco budworm, Heliothis virescens (F.) (Lepidoptera: Noctuidae)

BACKGROUND: The potential development of resistance to Bacillus thuringiensis (Bt) cotton and surging of non‐targeted insects is a major risk in the durability of Bt plant technology. Midgut proteinases are involved in Bt activation and degradation. Proteinase inhibitors may be used to control a wide range of insects and delay Bt resistance development. Proactive action to examine proteinase inhibitors for synergistic interaction with Bt toxin and cloning of proteinase cDNAs for RNAi is necessary to make transgenic cotton more versatile and durable. RESULTS: A sublethal dose (15 ppb) of Cry1Ac, 0.5% benzamidine and 0.02% phenylmethylsulfonyl fluoride significantly suppressed midgut azocaseinase, tryptic and chymotryptic activities, and resulted in reductions in larval and pupal length and mass of Heliothis virescens. The combination of proteinase inhibitor and Bt suppressed 20–37% more larval body mass and 26–80% more enzymatic activities than the inhibitor only or Bt only. To facilitate knockdown‐resistance‐related proteinase genes, 15 midgut chymotrypsin cDNAs were sequenced. Most predicted chymotrypsins contained the conserved N‐termini IVGG, three catalytic center residues (His, Asp and Ser), substrate specificity determinant (Ser or Gly) and cysteines for disulfide bridges. These putative chymotrypsins were separated into three distinct groups, indicating the diverse proteinases evolved in this polyphagous insect. CONCLUSION: H. virescens has evolved diverse midgut proteinase genes. Proteinase inhibitors have potential insecticidal activity, and the interaction of Bt with proteinase inhibitors is desirable for enhancing Bt toxicity and delaying resistance development. Intensive sequencing of chymotrypsin cDNAs will facilitate future functional examinations of individual roles in Bt toxicity and resistance development and facilitate targeted control using RNAi and/or proteinase inhibitors. Copyright © 2012 Society of Chemical Industry     

Evidence for Resistance to Bacillus thuringiensis (Bt) subsp. kurstaki HD-1, Bt subsp. aizawai and Abamectin in Field Populations of Plutella xylostella from Malaysia

The efficacy of Bacillus thuringiensis (Bt) subsp. kurstaki HD-1 (‘Dipel’™; Btk; CryIA & CryII) and Bt. subsp. aizawai (‘Florbac’™; Bta; CryIA & CryIC) was assessed against larvae from various field populations of Plutella xylostella (F2 generation) collected in the Cameron Highlands, Malaysia in April 1994 and a lowland population (SERD 2; F10 generation) collected in December 1993. Evidence of resistance to Btk and to a lesser extent Bta is reported in these populations (LC₅₀ Toxicity Ratios [TR]=3–14 and 2–8 respectively), most notably in SERD 2. The first recorded evidence of resistance to abamectin (TR=17–195-fold) in field populations of P. xylostella is also reported. In an unselected sub-population of SERD 2, the TR values for Btk, Bta and abamectin declined 2- to 3-fold (P<0·01) over six generations in the laboratory (F10–F16) while in sub-populations of SERD 2 selected with these products (F11–F15) there was a significant (P<0·01) increase in the TR (15-, 3- and 2·5-fold respectively) when compared with the F10 generation. This suggests the presence of marked resistance to Btk and some resistance to Bta and abamectin. There is also evidence of slight cross-resistance to Btk in the Bta-selected sub-population but no evidence for the reverse selection of resistance or for cross-resistance between Btk and abamectin. Concurrent selection studies (F11–F15) with another sub-population of SERD 2 demonstrated resistance to the acylurea insect growth regulator, teflubenzuron (‘Nomolt’™) (29-fold increase in TR). Based on the selection experiments with SERD 2, estimates of realised heritability (h²) of resistance gave very high values for teflubenzuron and Btk (c.0·7) and moderate values for abamectin and Bta (c.0·3). The results are discussed in relation to integrated pest management (IPM) and insecticide resistance management (IRM) strategies for P. xylostella.     

Molecular basis of resistance to acetolactate synthase-inhibiting herbicides in Sisymbrium orientale and Brassica tournefortii

Three Australian Sisymbrium orientale and one Brassica tournefortii biotypes are resistant to acetolactate synthase (ALS)-inhibiting herbicides due to their possession of an ALS enzyme with decreased sensitivity to these herbicides. Enzyme kinetic studies revealed no interbiotypic differences within species in K_m (pyruvate) (the substrate concentration at which the reaction rate is half maximal) but a greater V_max (the rate when the enzyme is fully saturated with substrate) for two of the resistant S orientale biotypes over susceptible levels. F₁ hybrids from reciprocal crosses between resistant and susceptible biotypes of S orientale showed an intermediate response to chlorsulfuron compared to the parental plants. ALS herbicide resistance in S orientale segregated in a 3:1 (resistant:susceptible) ratio in F₂ plants with a single rate of chlorsulfuron, indicating that resistance is inherited as a single, incompletely dominant nuclear gene. Two regions of the ALS structural gene known to vary in ALS-resistant biotypes were amplified and sequenced. Resistant S orientale biotypes NS01 and SS03 contained a single nucleotide substitution in Domain B, predicting a Trp (in susceptible) to Leu (in resistant) amino acid change. Two adjacent nucleotide substitutions (CC T to AT T) predicting a Pro (in susceptible) to Ile (in resistant) change in the primary amino acid sequence were identified in Domain A of resistant S orientale biotype SS01. Likewise, a single nucleotide substitution at the same site in the resistant B tournefortii biotype predicts a Pro (in susceptible) to Ala (in resistant) substitution. No other interbiotypic nucleotide differences predicted amino acid changes in the sequenced regions, suggesting that the amino acid substitutions reported above are responsible for resistance to ALS-inhibiting herbicides in the respective biotypes. © 1999 Society of Chemical Industry     

Field-evolved resistance to Bt toxin Cry1Ac in the pink bollworm, Pectinophora gossypiella (Saunders) (Lepidoptera: Gelechiidae), from India

BACKGROUND: The pink bollworm is one of the most destructive pests of cotton. Transgenic cotton producing Bt toxin Cry1Ac or a combination of Cry1Ac and Cry2Ab2 has been used effectively against this pest. However, some other insects have evolved resistance to Bt toxins in the field. During the 2007–2008 and 2008–2009 seasons, pink bollworm populations in India were surveyed to evaluate their responses to Cry1Ac and seed powder containing Cry1Ac and Cry2Ab2. RESULTS: The results provide evidence that resistance to Cry1Ac had evolved by 2008 in a population sampled from non‐Bt cotton in the Amreli district of Gujarat in western India. The median lethal concentration of Cry1Ac for five‐day‐old larvae (LC₅₀) was significantly higher for insects derived in 2008 from Amreli than for any of the other field populations tested from four locations in India. For Cry1Ac, the mean LC₅₀ for the strain derived from Amreli in 2008 was 44 times higher than for the most susceptible population. However, for seed powder of Bollgard II containing primarily Cry2Ab2, the 2008 Amreli population was only slightly less susceptible than the most susceptible population. CONCLUSIONS: The data reported here constitute the first evidence of field‐evolved resistance of pink bollworm to Cry1Ac. This initial evidence spurred more extensive evaluations during the 2009–2010 growing season, which confirmed field‐evolved resistance to Cry1Ac in Amreli. The lack of cross‐resistance to Cry2Ab2 suggests that plants producing this toxin are likely to be more effective against resistant populations than plants producing only Cry1Ac. Copyright © 2011 Society of Chemical Industry     

Leaf chlorophyll fluorescence discriminates herbicide resistance in Echinochloa species

Timely detection of herbicide resistance at an early stage of crop cultivation is essential to help farmers find alternative solutions to manage herbicide resistance in their fields. In this study, maximum quantum yield of PS II [F_v/F_m = (F_m – F_o)/F_m] was measured at the 4–5 leaf stage to discriminate between herbicide‐resistant and susceptible biotypes of Echinochloa species. The differences in F_v/F_m between herbicide‐resistant and susceptible Echinochloa spp. were consistent with the whole‐plant assay based on I₅₀ (herbicide doses causing a 50% inhibition of F_v/F_m) and GR₅₀ (herbicide doses causing a 50% reduction in plant fresh weight) values and R/S ratios (herbicide resistance index), regardless of the mode of action of the tested herbicides. A PS II inhibitor caused the fastest inhibition of F_v/F_m, compared with ACCase and ALS inhibitors, after herbicide treatment. The required time for discrimination between herbicide‐resistant and susceptible Echinochloa spp. was 64 h after PS II inhibitor treatment, much shorter than those of ACCase and ALS inhibitor‐treated plants, which required 168 and 192 h respectively. The leaf chlorophyll fluorescence assay provided reliable diagnostics of herbicide resistance in Echinochloa spp. with significant time savings and convenient measurement in field conditions compared with the conventional whole‐plant assay.