Individual transmembrane domains of SfABCC2 from Spodoptera frugiperda do not serve as functional Cry1F receptors.

The fall armyworm (Spodoptera frugiperda) is a major global pest causing severe damage to various crops, especially corn. Transgenic corn producing the Cry1F pesticidal protein from the bacterium Bacillus thuringiensis (Cry1F corn) showed effectiveness in controlling this pest until S. frugiperda populations at locations in North and South America evolved practical resistance. The mechanism for practical resistance involved disruptive mutations in an ATP binding cassette transporter subfamily C2 gene (SfABCC2), which serves as a functional Cry1F receptor in the midgut cells of susceptible S. frugiperda. The SfABCC2 protein contains two transmembrane domains (TMD1 and TMD2), each with a cytosolic nucleotide (ATP) binding domain (NBD1 and NBD2, respectively). Previous reports have demonstrated that disruptive mutations in TMD2 were linked with resistance to Cry1F, yet whether the complete SfABCC2 structure is needed for receptor functionality or if a single TMD-NBD protein can serve as functional Cry1F receptor remains unknown. In the present study, we separately expressed TMD1 and TMD2 with their corresponding NBDs in cultured insect cells and tested their Cry1F receptor functionality. Our results show that the complete SfABCC2 structure is required for Cry1F receptor functionality. Moreover, binding competition assays revealed that Cry1F specifically bound to SfABCC2, whereas neither SfTMD1-NBD1 nor SfTMD2-NBD2 exhibited any significant binding. These results provide insights into the molecular mechanism of Cry1F recognition by SfABCC2 in S. frugiperda, which could facilitate the development of more effective insecticidal proteins.

Development of a bioassay method to test activity of cry insecticidal proteins against Diatraea spp. (Lepidoptera: Crambidae) sugarcane stem borers.

The genus Diatraea (Lepidoptera: Crambidae) includes stem borers representing the most critical sugarcane pests in the Americas. Colombia's most widely distributed and damaging Diatraea species include Diatraea saccharalis, D. indigenella, D. busckella, and D. tabernella. The reduced efficacy of biological tools commonly used in controlling several species highlights the importance of evaluating alternative management strategies, such as transgenic plants expressing insecticidal proteins from the bacterium Bacillus thuringiensis (Bt). The selection of optimal Bt insecticidal proteins for Diatraea control depends on bioassays with purified Bt proteins. Because there is no described artificial diet for borer species other than D. saccharalis and availability of most purified Bt toxins is restricted, this study aimed at developing a bioassay method using fresh corn tissue and providing proof of concept by testing susceptibility to the Cry1Ac insecticidal protein from Bt. Toxicity was evaluated with a single Cry1Ac dose applied directly to corn discs. Stem borer mortality after seven days was higher than 90% for all four tested Diatraea species, while control mortality was below 8%. In addition, we observed that Cry1Ac caused more than 90% weight inhibition in all survivors and delayed development. These results validate the use of this method to determine mortality and growth inhibition due to the consumption of the Cry1Ac protein in each of the Diatraea species. Furthermore, this method could be used to assess other entomopathogenic substances to control these insect pests.

First documentation of major Vip3Aa resistance alleles in field populations of Helicoverpa zea (Boddie) (Lepidoptera: Noctuidae) in Texas, USA.

The corn earworm, Helicoverpa zea, is a major target pest of the insecticidal Vip3Aa protein used in pyramided transgenic Bt corn and cotton with Cry1 and Cry2 proteins in the U.S. The widespread resistance to Cry1 and Cry2 proteins in H. zea will challenge the long-term efficacy of Vip3Aa technology. Determining the frequency of resistant alleles to Vip3Aa in field populations of H. zea is critically important for resistance management. Here, we provided the first F2 screen study to estimate the resistance allele frequency for Vip3Aa in H. zea populations in Texas, U.S. In 2019, 128 H. zea neonates per isofamily for a total of 114 F2 families were screened with a diagnostic concentration of 3.0 µg/cm2 of Vip3Aa39 protein in diet-overlay bioassays. The F2 screen detected two families carrying a major Vip3Aa resistance allele. The estimated frequency of major resistance alleles against Vip3Aa39 in H. zea in Texas from this study was 0.0065 with a 95% CI of 0.0014-0.0157. A Vip3Aa-resistant strain (RR) derived from the F2 screen showed a high level of resistance to Vip3Aa39 protein, with a resistance ratio of >588.0-fold relative to a susceptible population (SS) based on diet-overlay bioassays. We provide the first documentation of a major resistance allele conferring high levels of Vip3Aa resistance in a field-derived strain of H. zea in the U.S. Data generated from this study contribute to development of management strategies for the sustainable use of the Vip3Aa technology to control H. zea in the U.S.

Productive performance of dairy cows fed with hydrolyzed sugarcane / Desempenho produtivo de vacas leiteiras alimentadas com cana-de-açúcar hidrolisada

This study aimed to evaluate the productive performance of dairy cows fed with sugarcane treated with 5g kg-1 of calcium oxide (CaO) or hydroxide [Ca(OH)2]. Eight Holstein cows with 638.01±12.52kg of body weight and milk yield of 20.32±1.5kg d-1 were randomly assigned into two 4x4 Latin squares, fed with the following diets composed of corn silage (CS), fresh sugarcane (FS), sugarcane treated with calcium oxide (STCO) or calcium hydroxide (STCH) as only forage. Data collection lasted five days, after 15 days of adaptation to diets and facilities. The dry matter intake (% of body weight) was higher in diets with CS (3.08) compared to those with FS (2.67), STCO (2.73) or STCH (2.73), which did not differ. Diets with CS determined milk production adjusted for 4% fat (20.05kg d-1) similar to diets containing STCO and STCH (18.01 and 17.89kg d-1, respectively) and higher than those with FS (17.33kg d-1). The experimental diets did not alter the composition of milk. The use of sugarcane treated with Ca(OH)2 is a viable option for feeding Holstein cows with average genetic potential for milk production because it allows production and composition similar to milk dairy cows fed with corn silage, besides benefiting the logistics of feeding in the rural properties.

Objetivou-se avaliar o desempenho produtivo de vacas leiteiras alimentadas com cana-de-açúcar tratada com 5g kg-1 de óxido (CaO) ou hidróxido de cálcio [Ca(OH)2]. Foram utilizadas oito vacas da raça Holandesas com 638,01±12,52kg de peso corporal e produção média de leite de 20,32±1,5kg dia-1, distribuídas aleatoriamente em dois quadrados latinos balanceados 4x4, alimentadas com dietas contendo silagem de milho (SM), cana-de açúcar in natura (CIN), cana-de-açúcar tratada com óxido de cálcio (CTOC) ou hidróxido de cálcio (CTHC) como único volumoso. O período de coleta de dados foi de cinco dias, após 15 dias de adaptação às instalações e dietas. O consumo de matéria seca (% do peso corporal) foi superior nas dietas com SM (3,08) em relação àquelas com CIN (2,67), CTOC (2,73) ou CTHC (2,73), as quais não diferiram entre si. Dietas com SM determinaram produção de leite ajustada para 4% de gordura (20,05kg dia-1), similar às dietas contendo CTOC e CTHC (18,01 e 17,89kg dia-1, respectivamente) e superior àquelas com CIN (17,33kg dia-1). As dietas experimentais não alteraram a composição do leite. A utilização da cana-de-açúcar tratada com Ca(OH)2 é uma alternativa alimentar viável para vacas Holandesas com potencial genético médio para produção leiteira, já que possibilita produção e composição do leite similar à vacas alimentadas com silagem de milho, além de beneficiar a logística de alimentação nas propriedades rurais.