Resistance status of lepidopteran soybean pests following large-scale use of MON 87701 × MON 89788 soybean in Brazil

Widespread adoption of MON 87701 × MON 89788 soybean, expressing Cry1Ac Bt protein and glyphosate tolerance, has been observed in Brazil. A proactive program was implemented to phenotypically and genotypically monitor Cry1Ac resistance in Chrysodeixis includens (Walker). Recent cases of unexpected injury in MON 87701 × MON 89788 soybean were investigated and a large-scale sampling of larvae on commercial soybean fields was performed to assess the efficacy of this technology and the distribution of lepidopteran pests in Brazil. No significant shift in C. includens susceptibility to Cry1Ac was observed eight years after commercial introduction of this technology in Brazil. F2 screen results confirmed that the frequency of Cry1Ac resistance alleles remains low and stable in C. includens. Unexpected injury caused by Rachiplusia nu (Guenée) and Crocidosema aporema (Walsingham) in MON 87701 × MON 89788 soybean was detected during the 2020/21 season, and studies confirmed a genetically based alteration in their susceptibility to Cry1Ac. MON 87701 × MON 89788 soybean remains effective against Anticarsia gemmatalis (Hübner), C. includens, Chloridea virescents (Fabricius) and Helicoverpa armigera (Hübner) in Brazil. However, there is evidence of field-evolved resistance to MON 87701 × MON 89788 soybean by the secondary soybean pests R. nu and C. aporema.

Genetic diversity in Chrysodeixis includens nucleopolyhedrovirus and factors involved in its generation

In the present thesis, the genetic variability of field-collected isolates of Chrysodeixis includens nucleopolyhedrovirus (ChinNPV) was examined. These results have revealed the great variability present within natural ChinNPV isolates, confirm the differences between two different genetic variants from these isolates and, more important, appear to identify the ChinNPV-K variant as a generator of variability in these isolates. In conclusion, a great variability has been revealed within ChinNPV natural isolates, with a single variant showing a key role in the generation of this diversity in a concentration-dependent manner.

Generation of Variability in Chrysodeixis includens Nucleopolyhedrovirus (ChinNPV): The Role of a Single Variant

The mechanisms generating variability in viruses are diverse. Variability allows baculoviruses to evolve with their host and with changes in their environment. We examined the role of one genetic variant of Chrysodeixis includens nucleopolyhedrovirus (ChinNPV) and its contribution to the variability of the virus under laboratory conditions. A mixture of natural isolates (ChinNPV-Mex1) contained two genetic variants that dominated over other variants in individual larvae that consumed high (ChinNPV-K) and low (ChinNPV-E) concentrations of inoculum. Studies on the ChinNPV-K variant indicated that it was capable of generating novel variation in a concentration-dependent manner. In cell culture, cells inoculated with high concentrations of ChinNPV-K produced OBs with the ChinNPV-K REN profile, whereas a high diversity of ChinNPV variants was recovered following plaque purification of low concentrations of ChinNPV-K virion inoculum. Interestingly, the ChinNPV-K variant could not be recovered from plaques derived from low concentration inocula originating from budded virions or occlusion-derived virions of ChinNPV-K. Genome sequencing revealed marked differences between ChinNPV-K and ChinNPV-E, with high variation in the ChinNPV-K genome, mostly due to single nucleotide polymorphisms. We conclude that ChinNPV-K is an unstable genetic variant that is responsible for generating much of the detected variability in the natural ChinNPV isolates used in this study.

Large-scale assessment of lepidopteran soybean pests and efficacy of Cry1Ac soybean in Brazil

The soybean technology MON 87701 × MON 89788, expressing Cry1Ac and conferring tolerance to glyphosate, has been widely adopted in Brazil since 2013. However, pest shifts or resistance evolution could reduce the benefits of this technology. To assess Cry1Ac soybean performance and understand the composition of lepidopteran pest species attacking soybeans, we implemented large-scale sampling of larvae on commercial soybean fields during the 2019 and 2020 crop seasons to compare with data collected prior to the introduction of Cry1Ac soybeans. Chrysodeixis includens was the main lepidopteran pest in non-Bt fields. More than 98% of larvae found in Cry1Ac soybean were Spodoptera spp., although the numbers of Spodoptera were similar between Cry1Ac soybean and non-Bt fields. Cry1Ac soybean provided a high level of protection against Anticarsia gemmatalis, C. includens, Chloridea virescens and Helicoverpa spp. Significant reductions in insecticide sprays for lepidopteran control in soybean were observed from 2012 to 2019. Our study showed that C. includens and A. gemmatalis continue to be primary lepidopteran pests of soybean in Brazil and that Cry1Ac soybean continues to effectively manage the target lepidopteran pests. However, there was an increase in the relative abundance of non-target Spodoptera spp. larvae in both non-Bt and Cry1Ac soybeans.

Mixture compatibility of ChinNPV baculovirus with herbicides and fungicides used in soybean

Chrysodeixis includens Walker, 1858 (Lepidoptera: Noctuidae), commonly known as soybean looper, is one of the main pests that reduces soybean yield due to its defoliation capacity. Population outbreaks of this pest can occur in the vegetative stage of the crop, together with the occurrence of weeds or in the reproductive stage, at the same time as that of diseases. This often requires the use of pesticides against pests and weeds, or fungi at the same time. Thus, the objective of this study was to evaluate the compatibility of baculovirus ChinNPV with different synthetic chemical herbicides and fungicides used in soybean. Four bioassays were carried out, with or without the addition of ChinNPV to different herbicides and fungicides. The artificial diets were immersed in the solutions of the pesticides and their mixture and supplied to the caterpillars of C. includens, immediately and after one and two hours of mixing. The evaluation was performed by quantifying the number of dead caterpillars. The results showed that the ChinNPV baculovirus is compatible for mixing with all studied herbicides and fungicides, even after two hours of mixing. In all the studied scenarios, baculovirus mixed with chemical pesticides triggered the same mortality as sprayed with baculovirus alone, and values of over 80% mortality of C. includens have always been recorded.

Evaluation of Reference Genes and Expression Level of Genes Potentially Involved in the Mode of Action of Cry1Ac and Cry1F in a Susceptible Reference Strain of Chrysodeixis includens

Soybean looper (SBL), Chrysodeixis includens (Walker), is one of the major lepidopteran pests of soybean in the American continent. SBL control relies mostly on the use of insecticides and genetically modified crops expressing Bacillus thuringiensis (Bt) insecticidal Cry proteins. Due to the high selection pressure exerted by these control measures, resistance has developed to different insecticides and Bt proteins. Nevertheless, studies on the mechanistic background are still scarce. Here, the susceptibility of the laboratory SBL-Benzon strain to the Bt proteins Cry1Ac and Cry1F was determined in diet overlay assays and revealed a greater activity of Cry1Ac than Cry1F, thus confirming results obtained for other sensitive SBL strains. A reference gene study across larval stages with four candidate genes revealed that RPL10 and EF1 were the most stable genes for normalization of gene expression data obtained by RT-qPCR. Finally, the basal expression levels of eight potential Bt protein receptor genes in six larval instars were analyzed, including ATP-binding cassette (ABC) transporters, alkaline phosphatase, aminopeptidases, and cadherin. The results presented here provide fundamental knowledge to support future SBL resistance studies.

FORMULAÇÃO MULTICOMPONENTE PARA MONITORAMENTO DE LEPIDÓPTEROS PRAGA EM ÁREAS DE SOJA NO MATO GROSSO

Introdução: O complexo de lepidópteros-praga da soja, tem afetado diretamente a produção devido aos prejuízos pela destruição direta das vagens, além de reduzir a área fotossintética das plantas, pela desfolha. Com isso, faz-se necessário aplicações sucessivas de inseticidas, que impactam drasticamente o meio ambiente. Desta forma, uma alternativa sustentável para minimizar tais impactos, seria a utilização de feromônio sexual, que são sinais químicos emitidos pelos lepidópteros para o acasalamento. Estes feromônios podem conter compostos comuns atrativos a diferentes mariposas que conseguem ser usados para mais de uma espécie, ajudando assim o monitoramento nas lavouras de soja. Objetivo: Avaliar a eficiência de duas formulações multicomponentes no monitoramento de lepidópteros-praga da cultura da soja. Material e métodos: O experimento foi conduzido em delineamento inteiramente casualizado em uma área de soja no município de Tangará da Serra – MT, onde foram utilizadas armadilhas do tipo delta, de cor branca, para a captura dos adultos. As armadilhas ficaram dispostas a 100m da bordadura, distanciadas uma da outra a 50m. Foram avaliadas duas formulações multicomponentes e como controle utilizou-se hexano. Foram efetuadas cinco repetições e os tratamentos foram distribuídos em armadilhas tipo delta para a captura dos adultos e as avaliações eram realizadas semanalmente. Os dados foram submetidos ao Modelo Linear Generalizado (GLM), analisando a diferença entre os tratamentos pelo teste Tukey. Resultados: Foram coletadas 673 mariposas, com predominância para os noctuideos Spodoptera frugiperda e Chrysodeixis includens e o piralídeo Elasmopalpus lignosellus. A formulação 2 capturou 280 mariposas e diferiu estatisticamente da formulação 1 que coletou 241 e do hexano que teve uma coleta de 152 (p<0,05). Conclusão: As formulações avaliadas foram eficientes no monitoramento lepidópterospraga da soja, sendo uma alternativa promissora para o monitoramento deste complexo na cultura da soja.