Mutation in PgABCC2 confers low-level resistance to Cry1Ac in pink bollworm.

BACKGROUND: With the increasing incidence of pest resistance to transgenic crops producing Bacillus thuringiensis (Bt) proteins in the field, elucidating the molecular basis of resistance is important for monitoring, delaying and countering pest resistance. Previous work revealed that mutation or down-regulated expression of the cadherin gene (PgCad1) is associated with pink bollworm (Pectinophora gossypiella) resistance to Cry1Ac, and 20 mutant PgCad1 alleles (r1-r20) were characterized. Here, we tested the hypothesis that the ABC transporter PgABCC2 is a functional receptor for the Bt toxin Cry1Ac and that a mutation is associated with resistance. RESULTS: We identified and characterized the first resistance allele (rC2) of PgABCC2 in the laboratory-selected Cry1Ac-resistant strain AQ-C2 of pink bollworm. The rC2 allele had a one-base deletion in exon20, resulting in a frameshift and the introduction of a premature stop codon. This resulting PgABCC2 protein had a truncated C-terminus, including the loss of the NBD2 domain. AQ-C2 exhibited 20.2-fold greater resistance to Cry1Ac than the susceptible strain, and its inheritance of Cry1Ac resistance was recessive and genetically linked to PgABCC2. When produced in cultured insect cells, recombinant wild-type and rC2 mutant PgABCC2 proteins localized within the cell plasma membrane, although substantial cytoplasmic retention was also observed for the mutant protein, while the mutant PgABCC2 caused a 13.9-fold decrease in Cry1Ac toxicity versus the wild-type PgABCC2. CONCLUSIONS: PgABCC2 is a functional receptor of Cry1Ac and the loss of its carboxyl terminus (including its NBD2 domain) confers low-level resistance to Cry1Ac in both larvae and in cultured cells. © 2024 Society of Chemical Industry.

Efficacy and Adverse Reactions of Dahuang Mudan decoction Combined with Laparoscopy in the Treatment of Appendicitis: A Meta-Analysis.

Background: Appendicitis is a common acute abdominal disease. Traditional Chinese medicine believes that acute appendicitis is caused by the accumulation of heat and toxin, and the formation of carbuncle and pus in the colon due to stasis. Therefore, treatment should be carried out to clear heat and detoxify, clear the organs, and eliminate carbuncle. Dahuang Mudan Tang contains various traditional Chinese medicines for clearing heat and detoxifying, which can be used to treat appendicitis. This study observes the therapeutic effect of Dahuang Mudan Tang on patients undergoing laparoscopic surgery for acute appendicitis. Methods: Eight databases were searched by computer and inclusion criteria were pre determined before evaluation: (1) patients with appendicitis; (2) 18-70 years old; (3) Agree to this study and obtain randomized controlled trials at home and abroad on the combined treatment of appendicitis with caesarean section and rhubarb peony testing. Using RevMan 5.3 software, conduct a comprehensive evaluation of the cultivation quality and conduct data analysis. Results: The meta-analysis ultimately included 16 papers. They are all considered randomized controlled trials. The overall efficiency of the test unit and control unit was reported in 12 surveys. The total effective rate of the experimental group was significantly higher than that of the control group (Odds Ratio (OR): (1.16; 95% Cl: 1.11,1.20; P < .001), and the duration of bowel sounds was also significantly higher than that of the control group. Standardized mean deviation (SMD): (-7.39; 95% Cl: -8.48, -6.30; P < .01), defecation time SMD: (-1.60; 95% Cl: -2.07, -1.12; P < .01). Conclusion: Based on the total effective rate, defecation time, defecation time, CRP, IL-6, and adverse reactions of participants in this study, the combination of Dahuang Mudan Tang and laparoscopy in the treatment of appendicitis may be beneficial, which can improve clinical efficacy, inhibit inflammatory reactions, and promote postoperative recovery of patients. It is worth promoting and applying in clinical practice. However, these findings still require more high-quality research to confirm. Patients undergoing laparoscopic surgery for appendicitis were treated with Dahuang Mudan Tang combined with targeted intervention.

Cry1Ac Mixed with Gentamicin Influences the Intestinal Microbial Diversity and Community Composition of Pink Bollworms.

Pink bollworms severely affect the production of cotton. The method currently used for pink bollworm control is the planting of Bt (Bacillus thuringiensis) protein-expressing transgenic cotton. However, pink bollworms can develop strong resistance to Bt proteins in transgenic cotton because of the large planting area and long planting time of this crop, which severely affects the control of pink bollworms. Intestinal microorganisms play very important roles in insect growth, development and Bt resistance. However, the effect of intestinal microorganisms on pink bollworm Bt resistance is still unclear. The current study aimed to analyze the effect of intestinal microorganisms on the Bt resistance of pink bollworms. Intestinal microorganisms associated with Bt resistance were initially screened through Illumina MiSeq sequencing and analysis. The results showed that feeding with a mixture of gentamicin, Cry1Ac and an artificial diet could significantly increase the mortality of pink bollworm larvae compared with feeding with of a mixture of Cry1Ac and an artificial diet or an artificial diet alone. The microbial diversity, community structure and composition of the pink bollworm larval intestine were significantly influenced by feeding with a mixture of gentamicin, Cry1Ac and an artificial diet. Several intestinal bacteria with significantly altered abundances after treatment with gentamicin were preliminarily screened as potential resources for addressing Bt toxicity. This study provides useful strategies for addressing the Bt resistance of pink bollworms.

Mutation in the Cadherin Gene Is a Key Factor for Pink Bollworm Resistance to Bt Cotton in China.

Transgenic crops producing Bacillus thuringiensis (Bt) toxins are widely planted for insect control, but their efficacy may decrease as insects evolve resistance. Understanding the genetic basis of insect resistance is essential for developing an integrated strategy of resistance management. To understand the genetic basis of resistance in pink bollworm (Pectinophora gossypiella) to Bt cotton in the Yangtze River Valley of China, we conducted an F2 screening for alleles associated with resistance to the Bt (Cry1Ac) protein for the first time. A total of 145 valid single-paired lines were screened, among which seven lines were found to carry resistance alleles. All field parents in those seven lines carried recessive resistance alleles at the cadherin locus, including three known alleles, r1, r13 and r15, and two novel alleles, r19 and r20. The overall frequency of resistance alleles in 145 lines was 0.0241 (95% CI: 0.0106-0.0512). These results demonstrated that resistance was rare and that recessive mutation in the cadherin gene was the primary mechanism of pink bollworm resistance to Bt cotton in the Yangtze River Valley of China, which will provide a scientific basis for implementing targeted resistance management statics of pink bollworm in this region.

Cadherin repeat 5 mutation associated with Bt resistance in a field-derived strain of pink bollworm.

Evolution of resistance by pests reduces the benefits of transgenic crops that produce insecticidal proteins from Bacillus thuringiensis (Bt). Here we analyzed resistance to Bt toxin Cry1Ac in a field-derived strain of pink bollworm (Pectinophora gossypiella), a global pest of cotton. We discovered that the r14 allele of the pink bollworm cadherin gene (PgCad1) has a 234-bp insertion in exon 12 encoding a mutant PgCad1 protein that lacks 36 amino acids in cadherin repeat 5 (CR5). A strain homozygous for this allele had 237-fold resistance to Cry1Ac, 1.8-fold cross-resistance to Cry2Ab, and developed from neonate to adult on Bt cotton producing Cry1Ac. Inheritance of resistance to Cry1Ac was recessive and tightly linked with r14. PgCad1 transcript abundance in midgut tissues did not differ between resistant and susceptible larvae. Toxicity of Cry1Ac to transformed insect cells was lower for cells expressing r14 than for cells expressing wild-type PgCad1. Wild-type PgCad1 was transported to the cell membrane, whereas PgCad1 produced by r14 was not. In larval midgut tissue, PgCad1 protein occurred primarily on the brush border membrane only in susceptible larvae. The results imply r14 mediates pink bollworm resistance to Cry1Ac by reduced translation, increased degradation, and/or mislocalization of cadherin.

Bt resistance alleles in field populations of pink bollworm from China: Similarities with the United States and decreased frequency from 2012 to 2015.

BACKGROUND: Although most monitoring of pest resistance to widely cultivated transgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt) relies on bioassays, DNA screening for alleles associated with resistance has some advantages, particularly for rare, recessively inherited resistance. In China's Yangtze River Valley, where farmers first planted transgenic cotton producing Bt toxin Cry1Ac in 2000, bioassays have been used to monitor the recessive resistance of pink bollworm (Pectinophora gossypiella). Previous bioassay results show a small but significant increase in resistance to Cry1Ac during 2008-2010, followed by a significant decrease in resistance during 2011-2015 associated with extensive planting of second-generation hybrid cotton seeds that boosted the percentage of non-Bt cotton. Here we screened DNA from 19 748 pink bollworm collected during 2012-2015 from the Yangtze River Valley for seven alleles associated with resistance to Cry1Ac. These alleles were previously identified from lab-selected strains; three from the U.S. and four from China. RESULTS: The most common resistance allele was first identified from the U.S. and accounted for over 71% of all resistance alleles detected. Resistance was rare, with the total frequency of the seven resistance alleles showing a significant, 2.3-fold decrease from 0.0105 (95% CI: 0.0084-0.0132) in 2012 to 0.0046 (0.0031-0.0067) in 2015. CONCLUSIONS: The DNA screening data confirm results from bioassays showing pink bollworm resistance to Cry1Ac remained rare in the Yangtze River Valley from 2012-2015. The prevalence in China of the resistance allele identified from the U.S. implies a shared genetic basis of resistance that could facilitate molecular monitoring of resistance. © 2019 Society of Chemical Industry.

Host Range of Herpetogramma basalis (Lepidoptera: Crambidae), a Biological Control Agent for the Invasive Weed Alternanthera philoxeroides (Centrospermae: Amaranthaceae) in China.

Alternanthera philoxeroides (Mart.) Griseb. is an invasive herbaceous amphibious weed species in China. A pyralid moth Herpetogramma basalis (Walker) was discovered feeding on A. philoxeroides through field surveys and may be a potentially useful biocontrol agent. To determine the host range of H. basalis and evaluate its potential to control A. philoxeroides, no-choice and multiple-choice tests were conducted. Herpetogramma basalis fed on target weeds and 29 nontarget plant species. In addition to the target weed A. philoxeroides, H. basalis developed to adult on eight other nontarget species. Herpetogramma basalis survived to adulthood successfully on A. philoxeroides and less successfully on several other Amaranthaceae species. In multiple-choice studies, H. basalis showed a strong oviposition preference for A. philoxeroides over Amaranthus tricolor L. (Centrospermae: Amaranthaceae). Amaranthus tricolor was the only crop plant that supported the complete development of H. basalis. We cautiously recommend H. basalis for the biological control of A. philoxeroides in China.

Transposon insertion causes cadherin mis-splicing and confers resistance to Bt cotton in pink bollworm from China.

Transgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt) are cultivated extensively, but rapid evolution of resistance by pests reduces their efficacy. We report a 3,370-bp insertion in a cadherin gene associated with resistance to Bt toxin Cry1Ac in the pink bollworm (Pectinophora gossypiella), a devastating global cotton pest. We found the allele (r15) harboring this insertion in a field population from China. The insertion is a miniature inverted repeat transposable element (MITE) that contains two additional transposons and produces two mis-spliced transcript variants (r15A and r15B). A strain homozygous for r15 had 290-fold resistance to Cry1Ac, little or no cross-resistance to Cry2Ab, and completed its life cycle on Bt cotton producing Cry1Ac. Inheritance of resistance was recessive and tightly linked with r15. For transformed insect cells, susceptibility to Cry1Ac was greater for cells producing the wild-type cadherin than for cells producing the r15 mutant proteins. Recombinant cadherin protein occurred on the cell surface in cells transformed with the wild-type or r15A sequences, but not in cells transformed with the r15B sequence. The similar resistance of pink bollworm to Cry1Ac in laboratory- and field-selected insects from China, India and the U.S. provides a basis for developing international resistance management practices.

Pink Bollworm Resistance to Bt Toxin Cry1Ac Associated with an Insertion in Cadherin Exon 20.

Insecticidal proteins from Bacillus thuringiensis (Bt) are widely used to control insect pests, but their efficacy is reduced when pests evolve resistance. We report on a novel allele (r16) of the cadherin gene (PgCad1) in pink bollworm (Pectinophora gossypiella) associated with resistance to Bt toxin Cry1Ac, which is produced by transgenic cotton. The r16 allele isolated from a field population in China has 1545 base pairs of a degenerate transposon inserted in exon 20 of PgCad1, which generates a mis-spliced transcript containing a premature stop codon. A strain homozygous for r16 had 300-fold resistance to Cry1Ac, 2.6-fold cross-resistance to Cry2Ab, and completed its life cycle on transgenic Bt cotton producing Cry1Ac. Inheritance of Cry1Ac resistance was recessive and tightly linked with r16. Compared with transfected insect cells expressing wild-type PgCad1, cells expressing r16 were less susceptible to Cry1Ac. Recombinant cadherin protein was transported to the cell membrane in cells transfected with the wild-type PgCad1 allele, but not in cells transfected with r16. Cadherin occurred on brush border membrane vesicles (BBMVs) in the midgut of susceptible larvae, but not resistant larvae. These results imply that the r16 allele mediates Cry1Ac resistance in pink bollworm by interfering with the localization of cadherin.

Resistance to Bacillus thuringiensis linked with a cadherin transmembrane mutation affecting cellular trafficking in pink bollworm from China.

Evolution of pest resistance reduces the efficacy of insecticidal proteins from the gram-positive bacterium Bacillus thuringiensis (Bt) used widely in sprays and transgenic crops. In some previously studied strains of three major lepidopteran pests, resistance to Bt toxin Cry1Ac is associated with mutations disrupting the extracellular or cytoplasmic domains of cadherin proteins that bind Cry1Ac in the midgut of susceptible larvae. Here we report the first case of a cadherin transmembrane mutation associated with insect resistance to Bt. We discovered this mutation in a strain of the devastating global cotton pest, the pink bollworm (Pectinophora gossypiella), derived from a field population in the Yangtze River Valley of China. The mutant allele analyzed here has a 207 base pair deletion and encodes a cadherin protein lacking its transmembrane domain. Relative to a susceptible strain, a strain homozygous for this allele had 220-fold resistance to Cry1Ac and 2.1-fold cross-resistance to Cry2Ab. On transgenic cotton plants producing Cry1Ac, no susceptible larvae survived, but the resistant strain completed its life cycle. Inheritance of resistance to Cry1Ac was autosomal, recessive and tightly linked with the cadherin gene. Transportation of cadherin protein to the cell membrane and susceptibility to Cry1Ac occurred in transfected insect cells expressing the wild type cadherin allele, but not in transfected insect cells expressing the mutant cadherin allele. The results imply that the mutant allele analyzed here confers resistance to Cry1Ac by disrupting cellular trafficking of cadherin.

Hybridizing transgenic Bt cotton with non-Bt cotton counters resistance in pink bollworm.

Extensive cultivation of crops genetically engineered to produce insecticidal proteins from the bacterium Bacillus thuringiensis (Bt) has suppressed some major pests, reduced insecticide sprays, enhanced pest control by natural enemies, and increased grower profits. However, these benefits are being eroded by evolution of resistance in pests. We report a strategy for combating resistance by crossing transgenic Bt plants with conventional non-Bt plants and then crossing the resulting first-generation (F1) hybrid progeny and sowing the second-generation (F2) seeds. This strategy yields a random mixture within fields of three-quarters of plants that produce Bt toxin and one-quarter that does not. We hypothesized that the non-Bt plants in this mixture promote survival of susceptible insects, thereby delaying evolution of resistance. To test this hypothesis, we compared predictions from computer modeling with data monitoring pink bollworm (Pectinophora gossypiella) resistance to Bt toxin Cry1Ac produced by transgenic cotton in an 11-y study at 17 field sites in six provinces of China. The frequency of resistant individuals in the field increased before this strategy was widely deployed and then declined after its widespread adoption boosted the percentage of non-Bt cotton plants in the region. The correspondence between the predicted and observed outcomes implies that this strategy countered evolution of resistance. Despite the increased percentage of non-Bt cotton, suppression of pink bollworm was sustained. Unlike other resistance management tactics that require regulatory intervention, growers adopted this strategy voluntarily, apparently because of advantages that may include better performance as well as lower costs for seeds and insecticides.

Adult Exposure to Bt Toxin Cry1Ac Reduces Life Span and Reproduction of Resistant and Susceptible Pink Bollworm (Lepidoptera: Gelechiidae).

Insecticidal proteins from Bacillus thuringiensis (Bt) are used widely in sprays and transgenic plants to control insect pests. Although much research has elucidated the effects of Bt toxins on larvae, relatively little is known about their effects on adults. Here, we evaluated the effects of exposing adults to Bt toxin Cry1Ac on the life span and reproduction of two strains of pink bollworm ( Pectinophora gossypiella (Saunders)). In larval diet bioassays, the concentration of Cry1Ac killing 50% of larvae (LC 50 ) was 640 times higher for the laboratory-selected resistant strain (AZP-R) than the susceptible strain (APHIS-S). In experiments with adults, the highest concentrations of Cry1Ac tested (160 and 640 µg Cry1Ac per ml of 5% honey water) reduced life span for both strains. Treatments with 10, 40, and 160 µg Cry1Ac per ml reduced the duration of the oviposition period as well as the number of eggs laid by both strains, but did not affect the percentage of pairs producing eggs, the duration of the preoviposition period, or the percentage of eggs hatching for either strain. Adult life span did not differ between strains at low to moderate concentrations of Cry1Ac, but it was significantly greater for the resistant strain than the susceptible strain at the two highest concentrations of Cry1Ac tested. The reduced susceptibility to high concentrations of Cry1Ac in adults of the AZP-R strain relative to the APHIS-S strain provides the first evidence of expression of resistance to a Bt toxin in adult Lepidoptera.

Increased frequency of pink bollworm resistance to Bt toxin Cry1Ac in China.

Transgenic crops producing insecticidal proteins from Bacillus thuringiensis (Bt) kill some key insect pests, but evolution of resistance by pests can reduce their efficacy. The main approach for delaying pest adaptation to Bt crops uses non-Bt host plants as "refuges" to increase survival of susceptible pests. To delay evolution of pest resistance to transgenic cotton producing Bt toxin Cry1Ac, the United States and some other countries have required refuges of non-Bt cotton, while farmers in China have relied on "natural" refuges of non-Bt host plants other than cotton. The "natural" refuge strategy focuses on cotton bollworm (Helicoverpa armigera), the primary target of Bt cotton in China that attacks many crops, but it does not apply to another major pest, pink bollworm (Pectinophora gossypiella), which feeds almost entirely on cotton in China. Here we report data showing field-evolved resistance to Cry1Ac by pink bollworm in the Yangtze River Valley of China. Laboratory bioassay data from 51 field-derived strains show that the susceptibility to Cry1Ac was significantly lower during 2008 to 2010 than 2005 to 2007. The percentage of field populations yielding one or more survivors at a diagnostic concentration of Cry1Ac increased from 0% in 2005-2007 to 56% in 2008-2010. However, the median survival at the diagnostic concentration was only 1.6% from 2008 to 2010 and failure of Bt cotton to control pink bollworm has not been reported in China. The early detection of resistance reported here may promote proactive countermeasures, such as a switch to transgenic cotton producing toxins distinct from Cry1A toxins, increased planting of non-Bt cotton, and integration of other management tactics together with Bt cotton.