Binding analyses of Cry1Ab and Cry1Ac with membrane vesicles from Bacillus thuringiensis-resistant and -susceptible Ostrinia nubilalis

The binding properties of Bacillus thuringiensis toxins to brush border membrane vesicles of Dipel-resistant and -susceptible Ostrinia nubilalis larvae were compared using ligand-toxin immunoblot analysis, surface plasmon resonance (SPR), and radiolabeled toxin binding assays. In ligand-toxin immunoblot analysis, the number of Cry1Ab or Cry1Ac toxin binding proteins and the relative toxin binding intensity were similar in vesicles from resistant and susceptible larvae. Surface plasmon resonance with immobilized activated Cry1Ab toxin indicated that there were no significant differences in binding with fluid-phase vesicles from resistant and susceptible larvae. Homologous competition assays with radiolabeled Cry1Ab and Cry1Ac toxin and vesicles from resistant and susceptible larvae resulted in similar toxin dissociation constants and binding site concentrations. Heterologous competition binding assays indicated that Cry1Ab and Cry1Ac completely competed for binding, thus they share binding sites in the epithelium of the larval midguts of O. nubilalis. Overall, the binding analyses indicate that resistance to Cry1Ab and Cry1Ac in this Bt-resistant strain of O. nubilalis is not associated with a loss of toxin binding.     

Ten years of MON 810 resistance monitoring of field populations of Ostrinia nubilalis in Europe

From 2005 to 2015, Ostrinia nubilalis were collected in the most important maize‐growing areas in Europe where MON 810 was cultivated. The susceptibility of these O. nubilalis collections to the Cry1Ab protein was determined using overlay bioassays and compared to that of reference (control) strains. Larvae that died or did not moult after 7 days were used to calculate a moulting inhibition concentration (MIC). Two different batches of Cry1Ab protein were used over the course of this study. Between 2005 and 2015, 145 collections of O. nubilalis from 14 areas were analysed. The Cry1Ab susceptibility of populations from different geographic regions differed only slightly across years. The greatest variability in the MIC₅₀ for field samples collected from 2005 to 2011 and tested with batch 1 was 6.6‐fold in 2006. For field‐collected O. nubilalis, the difference between MIC₅₀ values of the most susceptible and most tolerant samples was 13.1‐fold for this period. For samples collected in 2012–2015 and tested with batch 2, the greatest variability was 4.1‐fold in 2014. A diagnostic concentration (MIC₉₉) was calculated for batch 1 (48 ng/cm²) using the results from all the collections in 2005–2012. Bridging experiments indicated that the diagnostic concentration for batch 2 was 28 ng/cm². From 2006 onwards, no O. nubilalis reached the 2nd larval stage when the diagnostic concentration of either batch of Cry1Ab was used. Only one insect collected from Romania in 2012 and two insects collected as reference strain from Spain in 2015 survived exposure to a dosage of 20 ng/cm², and none of these larvae survived on MON 810 maize. Our results indicate that there has been no significant change in susceptibility to Cry1Ab in European populations of O. nubilalis over the period 2005–2015.     

Larval survival and development of susceptible and resistant Ostrinia nubilalis (Lepidoptera: Pyralidae) on diet containing Bacillus thuringiensis

Abstract 1 Larval survival and development of Dipel‐susceptible and ‐resistant strains of European corn borer, Ostrinia nubilalis (Hübner), were assayed using diets incorporating low doses of a commercial formulation of Bacillus thuringiensis var. kurstaki Berliner (Dipel ES). 2 Larval mortality, growth and development, pupation rate and pupal weight were not significantly different between Dipel‐susceptible and ‐resistant strains when larvae were reared on a nontoxic control diet. 3 Larval mortality of Dipel‐resistant larvae did not significantly change as Dipel concentration increased at the tested concentrations, whereas mortality of Dipel‐susceptible larvae increased dramatically as Dipel concentration increased. 4 Larval development was significantly delayed when larvae were fed diets containing low doses of Dipel. 5 Pupation rate and pupal body weight declined as Dipel concentration increased but it decreased faster for the susceptible strain than for the resistant strain.     

Resistance monitoring of field populations of the corn borers Sesamia nonagrioides and Ostrinia nubilalis after 5 years of Bt maize cultivation in Spain

Approximately 22 000 hectares (5% of the total maize growing area) of transgenic maize expressing the Cry1Ab toxin from Bacillus thuringiensis (Bt maize) have been planted annually in Spain since 1998. Changes in the susceptibility to Cry1Ab of Spanish populations of the Mediterranean corn borer (MCB), Sesamia nonagrioides (Lefebvre) (Lepidoptera: Noctuidae), and the European corn borer (ECB), Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae), were assessed by annual monitoring on Bt maize fields. No increase in resistance was detected in the MCB populations from Ebro, Albacete, and Badajoz, nor in the ECB populations from Ebro and Badajoz during the period 1999–2002. The susceptibility of the MCB population from Madrid fluctuated from year to year, but a gradual trend towards higher levels of tolerance was not observed. Laboratory selection assays for eight generations yielded selected strains of MCB and ECB that were 21‐ and 10‐fold significantly more tolerant to Cry1Ab than the corresponding unselected strains, respectively. Nevertheless, none of the field‐collected or laboratory‐selected larvae were able to survive on Bt maize. Considering these data, no consistent shifts in susceptibility were found for Spanish populations of MCB nor ECB after 5 years of Bt maize cultivation, but systematic field monitoring needs to be continued.     

Larval feeding behavior of Dipel-resistant and susceptible Ostrinia nubilalis on diet containing Bacillus thuringiensis (Dipel ESTM)

Diet choice tests were conducted to examine the effects of Bacillus thuringiensis Berliner on larval feeding behavior of Dipel-resistant and susceptible strains of European corn borer, Ostrinia nubilalis (Hübner). Larvae (first through fourth instars) were presented an untreated standard diet and three diets incorporating different concentrations of a commercial formulation of B. thuringiensis, Dipel ES^TM. Significantly higher proportions of susceptible and resistant larvae were found on the control diet compared to the proportions recorded on any of the Dipel-treated diets. Soon after release (0.5 to 3 h), the occurrence of larvae on each of the diets was similar across different instars. The number of first and second instars on the control diet increased steadily over 72 h after release, when 43 to 75% of larvae were found on the control diet. The proportion of third and fourth instars on the control diet also increased initially, with an equilibrium occurring 12 to 24 h later when 30 to 40% of larvae were found on the control diet. Both strains appeared to avoid the Dipel-treated diets and their responses were similar over different Dipel concentrations. The Dipel-resistant strain showed higher ability to avoid the treated diets than the susceptible strain.     

Bt抗、感种群亚洲玉米螟幼虫取食Bt玉米后Cry1Ab杀虫蛋白在其体内的组织分布与含量

采用ELISA方法检测了实验室汰选的对Cry1Ab产生107倍抗性的亚洲玉米螟Ostrinia furnacalis (Guenée)种群与敏感种群3龄幼虫取食表达Cry1Ab杀虫蛋白的Bt玉米心叶后,杀虫蛋白在幼虫体内的分布情况。结果表明：Cry1Ab杀虫蛋白在抗性种群幼虫中的组织分布情况与敏感种群相近,主要存在于中肠组织和血淋巴中。抗、感种群中均以含有内含物的中肠组织中含量最高,分别为277.2 ng/g 和104.9 ng/g;其次为血淋巴,分别为93.7 ng/g 和69.5 ng/g;不含内含物的中肠组织中52.7 ng/g 和40.1 ng/g;在丝腺和马氏管组织的含量很低,丝腺中分别为8.5 ng/g和11.7ng/g,而马氏管中分别为6.7 ng/g和6.5 ng/g。脂肪体、生殖器官中未检测到杀虫蛋白。抗性种群中肠组织(含有内含物和不含内含物)中Cry1Ab的含量显著高于敏感种群。幼虫期取食过Bt玉米的亚洲玉米螟发育的蛹、成虫及其卵中均不含杀虫蛋白,说明Bt杀虫蛋白不会通过幼虫取食向蛹、成虫及卵传递。     

转Bt基因抗虫玉米对亚洲玉米螟幼虫取食行为的影响

室内研究了转cry1Ab杀虫蛋白基因的Bt抗虫玉米MON81 0和Bt1 1对亚洲玉米螟Ostriniafurnacalis初孵幼虫和3龄幼虫的取食行为、取食选择性和存活率的影响。在48h的非选择性试验中玉米螟初孵幼虫在MON81 0和Bt1 1玉米心叶组织上的幼虫取食率随时间的增加而下降,在对照玉米上的幼虫取食率随时间的增加而上升,两者间差异极显著。初孵幼虫接虫到MON81 0和Bt1 1玉米叶片48h的累计死亡率分别为67 .5 %和47 .5 % ,而在对照玉米上死亡率均为0. 3龄幼虫在Bt和非Bt玉米穗轴组织上的幼虫取食率随时间的增加呈上升趋势,第48h时在Bt和非Bt玉米上的幼虫取食率分别达到77 5 %和1 0 0 % ,差异极显著。选择性试验中,第4～48h内,初孵幼虫在Bt玉米上的幼虫取食率呈下降趋势,第48h时MON81 0和Bt1 1与各自非Bt对照的组合中初孵幼虫的累计死亡率分别为2 5 .0 %和1 7. 5 % ,二者差异不显著。3龄幼虫在Bt玉米和非Bt玉米上的幼虫取食率均随时间的延长而增加,但在非Bt玉米的幼虫取食率增加速度快,与Bt玉米差异极显著。Bt玉米对玉米螟幼虫取食有抑制和忌避作用。     

Linkage of an ABCC transporter to a single QTL that controls Ostrinia nubilalis larval resistance to the Bacillus thuringiensis Cry1Fa toxin

Field evolved resistance of insect populations to Bacillus thuringiensis (Bt) crystalline (Cry) toxins expressed by crop plants has resulted in reduced control of insect feeding damage to field crops, and threatens the sustainability of Bt transgenic technologies. A single quantitative trait locus (QTL) that determines resistance in Ostrinia nubilalis larvae capable of surviving on reproductive stage transgenic corn that express the Bt Cry1Fa toxin was previously mapped to linkage group 12 (LG12) in a backcross pedigree. Fine mapping with high-throughput single nucleotide polymorphism (SNP) anchor markers, a candidate ABC transporter (abcc2) marker, and de novo mutations predicted from a genotyping-by-sequencing (GBS) data redefined a 268.8 cM LG12. The single QTL on LG12 spanned an approximate 46.1 cM region, in which marker 02302.286 and abcc2 were ≤2.81 cM, and the GBS marker 697 was an estimated 1.89 cM distant from the causal genetic factor. This positional mapping data showed that an O. nubilalis genome region encoding an abcc2 transporter is in proximity to a single QTL involved in the inheritance of Cry1F resistance, and will assist in the future identification the mutation(s) involved with this phenotype.     

Effect of Cry1F maize on the behavior of susceptible and resistant Spodoptera frugiperda and Ostrinia nubilalis

Understanding the behavior of pests targeted with Bacillus thuringiensis Berliner (Bt) crops is important to define resistance management strategies. Particularly the study of larval movement between plants is important to determine the feasibility of refuge configurations. Exposure to Bt maize, Zea mays L. (Poaceae), has been suggested to increase larval movement in lepidopteran species but few studies have examined the potential for resistance to interact with behavioral responses to Bt toxins. Choice and no‐choice experiments were conducted with Spodoptera frugiperda (JE Smith) (Lepidoptera: Noctuidae) and Ostrinia nubilalis (Hübner) (Lepidoptera: Crambidae) to determine whether Cry1F resistance influences neonate movement. Leaf discs of Cry1F maize and the corresponding isoline were used to characterize behavioral responses. In both experiments, the location (on or off of plant tissues) and mortality of susceptible and Cry1F resistant neonates was recorded for 5 days, but the analysis of larvae location was performed until 7 h. Our results indicated no strong difference between resistant and susceptible phenotypes in S. frugiperda and O. nubilalis, although a small percentage of susceptible neonates in both species abandoned maize tissue expressing Cry1F. However, significant behavioral differences were observed between species. Ostrinia nubilalis exhibited increased movement between leaf discs, whereas S. frugiperda selected plant tissue within the first 30 min and remained on the chosen plant regardless of the presence of Cry1F. Spodoptera frugiperda reduced larval movement may have implications to refuge configuration. This study represents the first step toward understanding the effects of Cry1F resistance on Lepidoptera larval behavior. Information regarding behavioral differences between species could aid in developing better and more flexible resistance management strategies.     

Resistance of the fall armyworm,Spodoptera frugiperda,to transgenic Bacillus thuringiensis Cry1F corn in the Americas: lessons and implications for Bt corn IRM in China

The fall armyworm, Spodoptera frugiperda (J.E. Smith), is a major target pest of Bt crops (e.g., corn, cotton, and soybean) in North and South America. This pest has recently invaded Africa and Asia including China and the invasion has placed a great threat to the food security in many countries of these two continents. Due to the extensive use of Bt crops, practical resistance of S. frugiperda to Cry1F corn (TC 1507) with field control problems has widely occurred in Puerto Rico, Brazil, Argentina, and the mainland United States. Analyzing data generated from decade-long studies showed that several factors might have contributed to the wide development of the resistance. These factors include (1) limited modes of action of Bt proteins used in Bt crops; (2) cross-resistance among Cry1 proteins; (3) use of nonhigh dose Bt crop traits; (4) that the resistance is complete on Bt corn plants; (5) abundant in initial Cry1F resistance alleles; and (6) lack of fitness costs/recessive fitness costs of the resistance. The long-term use of Bt crop technology in the Americas suggests that Bt corn can be an effective tool for controlling S. frugiperda in China. IRM programs for Bt corn in China should be as simple as possible to be easily adopted by small-scale growers. The following aspects may be considered in its Bt corn IRM programs: (1) use of only “high dose” traits for both S. frugiperda and stalk borers; (2) developing and implementing a combined resistance monitoring program; (3) use “gene pyramiding” as a primary IRM strategy; and (4) if possible, Bt corn may not be planted in the areas where S. frugiperda overwinters. Lessons and experience gained from the global long-term use of Bt crops should have values in improving IRM programs in the Americas, as well as for a sustainable use of Bt corn technology in China.     

CRISPR/Cas9 mediated genome editing of Helicoverpa armigera with mutations of an ABC transporter gene HaABCA2 confers resistance to Bacillus thuringiensis Cry2A toxins

High levels of resistance to Bt toxin Cry2Ab have been identified to be genetically linked with loss of function mutations of an ABC transporter gene (ABCA2) in two lepidopteran insects, Helicoverpa armigera and Helicoverpa punctigera. To further confirm the causal relationship between the ABCA2 gene (HaABCA2) and Cry2Ab resistance in H. armigera, two HaABCA2 knockout strains were created from the susceptible SCD strain with the CRISPR/Cas9 genome editing system. One strain (SCD-A2KO1) is homozygous for a 2-bp deletion in exon 2 of HaABCA2 created by non-homologous end joining (NHEJ). The other strain (SCD-A2KO2) is homozygous for a 5-bp deletion in exon 18 of HaABCA2 made by homology-directed repair (HDR), which was produced to mimic the r2 resistance allele of a field-derived Cry2Ab-resistant strain from Australia. Both knockout strains obtained high levels of resistance to both Cry2Aa (>120-fold) and Cry2Ab (>100-fold) compared with the original SCD strain, but no or very limited resistance to Cry1Ac (<4-fold). Resistance to Cry2Ab in both knockouts is recessive, and genetic complementary tests confirmed Cry2Ab resistance alleles are at the same locus (i.e. HaABCA2) for the two strains. Brush border membrane vesicles (BBMVs) of midguts from both knockout strains lost binding with Cry2Ab, but maintained the same binding with Cry1Ac as the SCD strain. In vivo functional evidence from this study demonstrates knockout of HaABCA2 confers high levels of resistance to both Cry2Aa and Cry2Ab, confirming that HaABCA2 plays a key role in mediating toxicity of both Cry2Aa and Cry2Ab against H. armigera.     

转Bt基因玉米对甜菜夜蛾幼虫存活和发育的影响

在室内测定了2种转Cry1Ab基因的Bt玉米MON810和Bt11不同组织对甜菜夜蛾 Spodoptera exigua （Hübner）初孵幼虫以及心叶对4龄幼虫存活和发育的影响,在田间比较了甜菜夜蛾幼虫取食Bt 和非Bt玉米雌穗的存活和为害情况。结果表明,转Cry1Ab基因的Bt玉米的不同组织对甜菜夜蛾初孵幼虫都具有明显的杀虫活性,取食Bt玉米心叶、苞叶、籽粒时甜菜夜蛾均在幼虫期死亡; 取食MON810和Bt11雄穗的初孵幼虫化蛹率分别为5.2%和2.1%,羽化率为2.1%和1.0%;取食MON810和Bt11花丝的初孵幼虫化蛹率分别为1.0%和2.1%,但不能羽化。4龄幼虫取食MON810玉米心叶的化蛹率与对照差异不显著,而取食Bt11的化蛹率与对照差异显著; 取食两种Bt玉米心叶的４龄幼虫化蛹后的雌、雄蛹重和羽化率与对照组差异显著,但蛹期和平均单雌产卵量差异不显著,虽然对照组羽化的成虫平均产卵量高于Bt玉米组。田间接种初孵幼虫10 天后的调查结果表明,在MON810和Bt11玉米花丝上幼虫存活率分别为1.3%和0.3%, 而对照组分别为12.9%和16.2%;MON810和Bt11玉米雌穗被害率分别为18.3%和5.0%,而对照组分别为93.3%和95.0%,均显著低于对照组。     

Large scale production of Bacillus thuringiensis PS149B1 insecticidal proteins Cry34Ab1 and Cry35Ab1 from Pseudomonas fluorescens

The 14kDa (Cry34Ab1) and 44kDa (Cry35Ab1) binary insecticidal proteins are produced naturally by Bacillus thuringiensis PS149B1 as parasporal inclusion bodies. Here, we show production of these two insecticidal proteins in recombinant Pseudomonas fluorescens and their subsequent purification to near homogeneity to provide large quantities of protein for safety-assessment studies associated with the registration of transgenic corn plants. The gene sequence specific for each protein was expressed in P. fluorescens and fermented at the 75-L scale. For Cry34Ab1, the protein accumulated as insoluble inclusion bodies, and was purified by extraction directly from the cell pastes at pH 3.4 with a sodium acetate buffer, selective precipitation at pH 7.0, and differential centrifugation. For Cry35Ab1, the protein was extracted from the purified inclusion bodies with sodium acetate buffer (pH 3.5) containing 0.5M urea, followed by diafiltration. No chromatography steps were required to produce over 30g of lyophilized protein powder with purity greater than 98%, while retaining full insecticidal activity against Western corn rootworm larvae. The proteins were further characterized to assure identity and suitability for use in safety-assessment studies.     

A hybrid Bacillus thuringiensis delta-endotoxin gives resistance against a coleopteran and a lepidopteran pest in transgenic potato

Expression of Bacillus thuringiensis delta-endotoxins has proven to be a successful strategy for obtaining insect resistance in transgenic plants. Drawbacks of expression of a single resistance gene are the limited target spectrum and the potential for rapid adaptation of the pest. Hybrid toxins with a wider target spectrum in combination with existing toxins may be used as tool to mitigate these problems. In this study, Desiree potato plants were genetically modified to resist attack by insect species belonging to the orders Coleoptera and Lepidoptera, through the insertion of such a hybrid gene, SN19. Transgenic plants were shown to be resistant against Colorado potato beetle larvae and adults, potato tuber moth larvae, and European corn borer larvae. These are the first transgenic plants resistant to pests belonging to two different insect orders. In addition, the target receptor recognition of this hybrid protein is expected to be different from Cry proteins currently in use for these pests. This makes it a useful tool for resistance management strategies.     

Midgut aminopeptidase N isoforms from Ostrinia nubilalis: Activity characterization and differential binding to Cry1Ab and Cry1Fa proteins from Bacillus thuringiensis

Aminopeptidase N (APN) isoforms from Lepidoptera are known for their involvement in the mode of action of insecticidal Cry proteins from Bacillus thuringiensis. These enzymes belong to a protein family with at least eight different members that are expressed simultaneously in the midgut of lepidopteran larvae. Here, we focus on the characterization of the APNs from Ostrinia nubilalis (OnAPNs) to identify potential Cry receptors. We expressed OnAPNs in insect cells using a baculovirus system and analyzed their enzymatic activity by probing substrate specificity and inhibitor susceptibility. The interaction with Cry1Ab and Cry1Fa proteins (both found in transgenic insect-resistant maize) was evaluated by ligand blot assays and immunocytochemistry. Ligand blots of brush border membrane proteins showed that both Cry proteins bound mainly to a 150 kDa-band, in which OnAPNs were greatly represented. Binding analysis of Cry proteins to the cell-expressed OnAPNs showed that OnAPN1 interacted with both Cry1Ab and Cry1Fa, whereas OnAPN3a and OnAPN8 only bound to Cry1Fa. Two isoforms, OnAPN2 and OnAPN3b, did not interact with any of these two proteins. This work provides the first evidence of a differential role of OnAPN isoforms in the mode of action of Cry proteins in O. nubilalis.     

取食转Bt基因抗虫玉米后亚洲玉米螟幼虫中肠的组织病理变化

利用透射显微镜（TEM）观察亚洲玉米螟Ostrinia furnacalis (Guenée)幼虫取食了表达Cry1Ab杀虫蛋白的转Bt基因玉米心叶组织后中肠的组织病理变化, 以探讨转Bt基因玉米对亚洲玉米螟的致病机理, 为其合理、安全和持续利用提供理论依据。结果表明：亚洲玉米螟取食Bt玉米后中肠细胞及其细胞器发生了明显的病变。取食Bt玉米12 h后中肠细胞开始病变, 首先微绒毛脱落、内质网开始肿胀, 24 h后内质网肿胀、增多, 杯状细胞杯腔增大, 48 h后微绒毛大量脱落, 细胞开始空泡化, 随着取食时间的增加, 细胞空泡化程度加剧, 在感染前期细胞间的病变程度差异较大。微绒毛脱落、内质网肿胀断裂是在多数取食Bt玉米的亚洲玉米螟中肠细胞发生的普遍病变。由此表明, 人工修饰的Cry1Ab基因导入到玉米染色体组中所表达的杀虫蛋白可使玉米螟幼虫中肠细胞发生病变, 最终导致其死亡。     

Relative fitness of susceptible and Cry1A.105/Cry2Ab2-single-/dual-protein-resistant Helicoverpa zea (Boddie) (Lepidoptera: Noctuidae) on non-Bt diet and a diet containing a low concentration of two proteins

Helicoverpa zea (Boddie) is a destructive agricultural pest species that is targeted by both Bacillus thuringiensis (Bt) maize and cotton in the United States. Cry1A.105 and Cry2Ab2 are two Bt proteins expressed in a widely planted maize event MON 89034. In this study, two tests (Test-I and Test-II) were conducted to evaluate the relative fitness of Bt-susceptible and -resistant H. zea on non-Bt diet (Test-I and Test-II) and a diet containing a mix of Cry1A.105 and Cry2Ab2 at a low concentration (Test-II only). Insect populations evaluated in Test-I were two Bt-susceptible strains and three Bt-resistant strains (a single-protein Cry1A.105-, a single-protein Cry2Ab2-, and a dual-protein Cry1A.105/Cry2Ab2-resistant strains). Test-II analyzed the same two susceptible strains, three backcrossed-and-reselected Cry1A.105/Cry2Ab2-single-/dual-protein-resistant strains, and three F₁ heterozygous strains. Measurements of life table parameters showed that neither the single- nor dual-protein Cry1A.105/Cry2Ab2 resistance in H. zea was associated with fitness costs under the test conditions. The single Cry protein resistances at a concentration of a mix of Cry1A.105 and Cry2Ab2 that resulted in a zero net reproductive rate for the two susceptible strains were functionally incomplete recessive or codominant, and the dual-protein resistance was completely dominant. The lack of fitness costs could be a factor contributing to the rapid revolution of resistance to the Cry proteins in this species. Data generated from this study should aid our understanding of Cry protein resistance evolution and help in refining IRM programs for H. zea.     

Genetics of resistance to Cry1C toxin from Bacillus thuringiensis in Spodoptera litura (Fab.)

The present study was undertaken to determine the genetics of Cry1C resistance in Spodoptera litura. Selection of S. litura (Fab.) with Cry1C was done for eight generations to develop resistance. Reciprocal crosses between resistant and susceptible populations were made to understand the population genetics of Cry1C resistance in S. litura. Generation wise selection with Cry1C was evaluated for resistance development in S. litura. The LC₅₀ of Cry1C was 0.14 µg/cm² for the first selected generation and it increased to 23.98 µg/cm² after eight selected generations, which is a 285.47-fold increase in resistance compared with the susceptible strain. The estimated realized heritability (h²) after eight generations of selection with Cry1C insecticidal protein was 0.44. The number of generations required for the tenfold increase in LC₅₀ (1/R) was estimated to be 3.33. Response to Cry1C selection in S. litura was 0.30, the estimated selection differential was 0.69 and the pheonotypic standard deviation (dP) was 0.24. Reciprocal crosses between Cry1C resistant and susceptible strain of S. litura showed autosomal resistance.     

Characterization of six antibacterial response genes from the European corn borer (Ostrinia nubilalis) larval gut and their expression in response to bacterial challenge

Six cDNAs encoding putative antibacterial response proteins were identified and characterized from the larval gut of the European corn borer (Ostrinia nubilalis). These antibacterial response proteins include four peptidoglycan recognition proteins (PGRPs), one β-1,3-glucanase-1 (βglu-1), and one lysozyme. Tissue-specific expression analysis showed that these genes were highly expressed in the midgut, except for lysozyme. Analysis of expression of these genes in different developmental stage showed that they were expressed in larval stages, but little or no detectable expression was found in egg, pupa and adult. When larvae were challenged with Gram-negative bacteria (Enterobacter aerogenes), the expression of all six genes was up-regulated in the fatbodies. However, when larvae were challenged with Gram-positive bacteria (Micrococcus luteus), only PGRP-C and lysozyme genes were up-regulated. This study provides additional insights into the expression of antibacterial response genes in O. nubilalis larvae and helps us better understand the immune defense response in O. nubilalis.