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Bacillus thuringiensis Cry and Vip proteins are highly effective at controlling agricultural pests and could be used in pyramided transgenic crops. However, the molecular mechanism underlying the Cry1Ah and Vip3Aa19 synergistic interaction has never been investigated at the molecular level in Yellow peach moth (YPM) Conogethes punctiferalis. Binding affinity and synergism of Cry1Ah and Vip3Aa19 proteins with ABC transporter subfamily C receptors ABCC1, ABCC2 and ABCC3 proteins from the midgut of YPM larva by using surface plasmon resonance (SPR) and pull-down assays. Both assays revealed that Cry1Ah could interact with ABCC1, ABCC2, and ABCC3, whereas Vip3Aa19 only interacts with ABCC1 and ABCC3, but not with ABCC2. Hence, when compared to the Vip3Aa19 protein, Cry1Ah had a higher binding affinity for ABCC1, ABCC2, and ABCC3. Furthermore, competitive binding assay between Cry1Ah and Vip3Aa19 protein with ABC transporter subfamily C receptors resulted in the final eluted protein samples displaying vibrant blue bands of Cry1Ah and very faint bands of Vip3Aa19. Suggesting that Cry and Vip proteins could deliver a synergistic effect after cleaving the midgut proteases. Therefore, this finding indicated that the Cry1Ah and Vip3Aa19 do not compete for interacting with midgut receptors and thus provide strong synergism against YPM. SPR and pull-down results revealed that Cry1Ah could interact with ABCC1, ABCC2, and ABCC3, whereas Vip3Aa19 only interacts with ABCC1 and ABCC3, but not with ABCC2. The competitive binding assay indicated that Cry1Ah and Vip3Aa19 do not compete for interacting with midgut receptors and thus provide strong synergism against YPM. [Display omitted] • Cry1Ah had a higher binding affinity for ABCC1, ABCC2, and ABCC3 than Vip3Aa19. • Cry1Ah and Vip3Aa19 does not compete for interacting with midgut receptors of Conogethes punctiferalis. • Cry1Ah and Vip3Aa19 combination provides strong synergism against C. punctiferalis. [ABSTRACT FROM AUTHOR]

• Bt protein levels in transgenic rice vary significantly across different growth stages and organs. • Bt protein levels higher in cry2A* than cry1C* transgenic early japonica rice. • Bt protein levels are highest in leaves, followed by stem sheaths and young panicles. • Expression levels of Bt proteins in brown rice are the lowest among all growth stages and organs. • Correlation between Bt gene transcription and protein levels is significant in various rice tissues. Transgenic insect-resistant rice offers an environmentally friendly approach to mitigate yield losses caused by lepidopteran pests, such as stem borers. Bt (Bacillus thuringiensis) genes encode insecticidal proteins and are widely used to confer insect resistance to genetically modified crops. This study investigated the integration, inheritance, and expression characteristics of codon-optimised synthetic Bt genes, cry1C* and cry2A*, in transgenic early japonica rice lines. The early japonica rice cultivar, Songgeng 9 (Oryza sativa), was transformed with cry1C* or cry2A*, which are driven by the ubi promoter via Agrobacterium tumefaciens-mediated transformation. Molecular analyses, including quantitative PCR (qPCR), enzyme-linked immunosorbent assay (ELISA), and Southern blot analysis were performed to confirm transgene integration, inheritance, transcriptional levels, and protein expression patterns across different tissues and developmental stages. Stable transgenic early japonica lines exhibiting single-copy transgene integration were established. Transcriptional analysis revealed variations in Bt gene expression among lines, tissues, and growth stages, with higher expression levels observed in leaves than in other organs. Notably, cry2A* exhibited consistently higher mRNA and protein levels than cry1C* across all examined tissues and developmental time points. Bt protein accumulation followed the trend of leaves > stem sheaths > young panicles > brown rice, with peak expression during the filling stage in the vegetative tissues. Synthetic cry2A* displayed markedly elevated transcription and translation compared to cry1C* in the transgenic early japonica rice lines examined. Distinct spatiotemporal patterns of Bt gene expression were elucidated, providing insights into the potential insect resistance conferred by these genes in rice. These findings will contribute to the development of insect-resistant japonica rice varieties and facilitate the rational deployment of Bt crops. [ABSTRACT FROM AUTHOR]

High expression of Bacillus thuringiensis ( Bt ) protein in transgenic Bt crops has dramatically improved the resistance of crops to some insect pests. However, some Bt crops were reported to have poorer adaptabilities to abiotic stress environments compared to their non- Bt counterparts. The biological reasons for poorer adaptabilities were still unclear. A two-year field experiment was conducted to investigate yield and physiological response of newly bred Bt rice lines with different Bt protein expression levels [MH63 ( Cry1C* ), MH63 ( Cry2A* ) and MH63 ( Cry1Ab/Ac )] and their non- Bt counterpart MH63 to drought stress. MH63 ( Cry1C* ), MH63 ( Cry2A* ) and MH63 ( Cry1Ab/Ac ) all showed lower grain yields and lower biomass under drought treatment compared to MH63. The lower grain yields and lower biomass of Bt -MH63 were mainly due to the earlier leaf senescence associated with disorder in protein metabolism under drought treatment. These variations in Bt -MH63 were found to be correlated to the BT/SP (ratio of Bt protein to soluble protein) in Bt -MH63 leaves. The impact of drought on traits of Bt -MH63 was aggravated with increase of the Bt protein expression level in the plant. These results suggested that Bt -MH63 had poorer adaptabilities to drought stress compared to their non- Bt counterpart MH63. “Bio-burden” caused by the high expression of Bt protein in the plant was proposed to be one possible reason for the poorer adaptability, and BT/SP was estimated to be an indicator for the assessment of the “bio-burden”. [ABSTRACT FROM AUTHOR]

Transgenic rice with Cry Bacillus thuringiensis ( Bt ) genes have been developed in the last few decades. The performance of transgenic rice, especially the yield and resistance, must be meticulously evaluated prior to commercial production. In this study, yield and resistance performance of newly bred transgenic Bt ( Cry1Ab/Ac ) rice lines with different genetic backgrounds were first investigated under pesticide-free field environment. Transgenic Bt ( Cry1Ab/Ac ) rice lines showed high insect resistances and superior agronomic traits, resulting in obvious yield advantages ranging from 9.1% to 38.0% over their respective non- Bt control lines. Bt protein content (BTC) was similar among the Bt ( Cry1Ab/Ac ) rice lines in different years, while it was much higher in leaves than in stems of the Bt ( Cry1Ab/Ac ) rice lines. Different genetic backgrounds of Bt ( Cry1Ab/Ac ) rice lines affected the yield advantage but had no influence on the resistance and BTC. The results suggested that Bt ( Cry1Ab/Ac ) rice lines have gained obvious yield advantages and high resistances regardless of genetic background under pesticide-free environment. [ABSTRACT FROM AUTHOR]

Using a transgenic Bacillus thuringiensis ( Bt ) crop is reportedly one of the most effective ways to improve insect resistance; however, the insertion of external Bt genes may affect the non-target growth characteristics of crop. Field experiments were conducted to investigate the yield performance and physiological traits in an elite restorer rice line, Minghui63 (MH63) with Cry1C * , Cry2A * or Cry1Ab/Ac genes under three fertilizer applications [zero nitrogen fertilizer (N0), zero potassium fertilizer (K0) and recommended nitrogen and potassium fertilizer (NK)]. In comparison with MH63, MH63 ( Cry1C * ) experienced declining grain yields of 15.3–31.0% under different fertilizer applications because of the decreased setting rates associated with the poorer matter transport-related traits. The grain yield of MH63 ( Cry2A * ) was averagely 18.4% lower than MH63 in only the N0 treatment due to the accelerated leaf senescence, which was correlated with the higher ratios of the Bt protein content to the soluble protein content (BTC/SPC) in the MH63 ( Cry2A * ) leaves at the filling stage (FS). The yield performance of MH63 ( Cry1Ab/Ac ) was identical to MH63. In addition, no significant differences were recorded between Bt- MH63 and MH63 regarding N and K assimilation. Hence, our results suggest that the Cry1C * and Cry2A * transgenes affect the yield performance and related traits of MH63 in response to nitrogen and/or potassium applications. [ABSTRACT FROM AUTHOR]

Abstract: DAS-59122-7 (59122) is a transgenic maize (Zea mays L.) that contains genes encoding Cry34Ab1 and Cry35Ab1 proteins from Bacillus thuringiensis Berliner strain 149B1 and phosphinothricin acetyltransferase (PAT) protein from Streptomyces viridochromogenes. Expression of these proteins in planta confers resistance to corn rootworms and other Coleopteran parasites and tolerance to herbicides containing glufosinate ammonium, respectively. In the current study, processed flours from 59122 maize grain or its near isogenic control line (091) were used at two concentrations (50% and 70% wt/wt) to produce diets that were fed to rats for 90 days in accordance with Chinese toxicology guidelines (GB15193.13-2003). A commercial AIN93G diet was used as an additional negative control. No significant differences in body weight and feed utilization were observed between rats consuming diets formulated with 59122 and 091 Control corn. Statistical differences (p <0.05) were observed in certain hematology and serum chemistry response variables between rats consuming diets formulated with 59122 or 091 Control flour compared to AIN93G diet. However, the mean value of these response variables in the 59122 groups were not statistically different from those observed in diets formulated with corresponding high and low concentrations of the flour from the 091 Control maize grain. Therefore, the statistical differences were considered to be related to consumption of diets containing high concentrations of maize flour (compared to AIN93G diets) regardless of source rather than to consumption of flour from 59122 maize grain. The results from this study demonstrated that 59122 maize grain is as safe as non-transgenic maize grain. [Copyright &y& Elsevier]

Abstract: Seedling responses of two transgenic Bacillus thuringiensis (Bt) cotton (Gossypium hirsutum L.) cultivars, 99B and SGK9708-41, to salinity stress were studied under 0, 50, 100 and 200mM NaCl treatments. Cotton seedling growth, indicated by height, leaf area, and fresh and dry weights, of both cultivars was reduced by NaCl in a concentration-dependent manner. Total N concentrations of seedling roots, stems and leaves of both cotton cultivars decreased with increasing NaCl concentrations. For both cultivars, soluble protein content increased in leaves and decreased in stems due to salt stress, and decreased in roots by 50 and 100mM NaCl. Bt protein concentrations in leaves of both cultivars were decreased by salt stress, but still were biologically effective to control the neonate bollworm larvae in seedling cotton. [Copyright &y& Elsevier]

Propylea japonica is a very important predator in agricultural ecosystems, which could be exposed to Bt protein. In this study, the bacterial community of P. japonica fed with normal food and food containing Cry2Ab protein was characterized for the first time using qPCR and high-throughput sequencing approaches. Results showed no effect of Cry2Ab on P. japonica development and reproduction. The most abundant bacterial phylum was Firmicutes , and the most abundant genus was Staphylococcus. The total bacteria copy number was not significantly different across four larval stages. Bacteria species composition was gathered more closely in feed on sucrose solution (sucrose-fed) than in larvae only fed on pea aphid (aphid-fed), the diversity indices of some operational taxonomic unit (OTU) were significantly different between sucrose-fed and aphid-fed samples. Different instar larval stages of P. japonica fed with sucrose solution containing Cry2Ab Bt protein and found no effect on microbial community composition and total bacteria copy numbers. However, effects on relative abundance of microbes, copy numbers of Corynebacterium 1 and Glutamicibacter arilaitensis were observed significantly lower in Bt -fed first and fourth larval stages. Low and high concentrations of Cry2Ab protein altered the microbial abundance relative to sucrose-fed P. japonica and copy numbers of G. arilaitensis and Staphylococcus xylosus were significantly lower in Bt -fed samples than control sucrose-fed. Our results are the first report showing that feeding on Cry2Ab protein does not alter microbial species composition in P. japonica , but effects gene copy number of some dominant bacteria. Further investigations are needed to assess the effect of copy number change on P. japonica. Image 1 • No effect of Cry2Ab on Propylea japonica development and reproduction. • Abundant bacterial was of Firmicutes & mostly was from genus Staphylococcus. • Feeding on Cry2Ab solution had effects on relative abundance of microbes in P. japonica. • Copy numbers of two bacteria species were significantly lower in Bt -fed P. japonica. [ABSTRACT FROM AUTHOR]