Project description:This study reports the identification of splice variants for the calcium/calmodulin-dependent protein kinase II (CaMKII) gene from Nilaparvata lugens, Laodelphax striatellus, and Sogatella furcifera. CaMKII is a multifunctional serine/threonine protein kinase that transduces Ca2+ signals in cells to control a range of cellular processes in the nervous system and muscular tissue. Sequence analysis showed that CaMKII was 99.0% identical at the amino acid level among three rice planthoppers, with the exception of a variable region located in the association domain. Four kinds of 20-81 amino acid "inserts" were found in the variable region. The phylogenetic tree of the deduced amino acid sequences showed that the NlCaMKII isoforms were more closely related to the LsCaMKII isoforms and were slightly distinct from SfCaMKII. CaMKII-E was the dominant type among the five main isoforms. CaMKII genes were constitutively expressed in various nymphal and adult stages and in tested tissues with the predominant transcription occurring in the head. There was no major tissue specificity of isoform expression, but the expression pattern and relative abundance of isoforms varied when compared with the RT-PCR between tissues. In addition, RNAi in N. lugens with dsRNA at a concentration of 200 ng nymph-1 induced a mortality of 77.7% on the 10th day and a reduction in the mRNA expression level of 67.2%. Unlike the holometabolous insect Helicoverpa armigera, the knockdown of NlCaMKII did not suppress the expression of 20E response genes, such as ECR, USP1, and HR3, in N. lugens. These results indicate that the role of CaMKII in hemimetabolous insects may be different from that in holometabolous insects.
Project description:Sogatella furcifera and Laodelphax striatellus are economically important rice pests in China by acting as vectors of several rice viruses, sucking the phloem sap and blocking the phloem vessels. Ecdysteroid hormone 20-hydroxyecdysone regulates insect development and reproduction. A cytochrome P450 monooxygenase CYP302A1 (22-hydroxylase), encoded by the Halloween gene disembodied (dib), plays a critical role in ecdysteroidogenesis. The objective of this study is to test whether dib genes are potential targets for RNA interference-based management of S. furcifera and L. striatellus. We cloned and characterized Sfdib and Lsdib. The open reading frame regions of dib genes were generated and used for designing and constructing dsRNA fragments. Experiments were conducted using oral delivery of dsdib to investigate the effectiveness of RNAi in S. furcifera and L. striatellus nymphs. Real-time quantitative reverse transcriptase-PCR analysis demonstrated that continuous ingestion of dsdib at the concentration of 0.01, 0.05 and 0.50 mg/ml diminished Sfdib expression levels by 35.9%, 45.1% and 66.2%, and ecdysone receptor (SfEcR) gene mRNA levels by 34.0%, 36.2% and 58.5% respectively in S. furcifera, and decreased Lsdib expression level by 18.8%, 35.8% and 56.7%, and LsEcR mRNA levels by 25.2%, 46.8% and 68.8% respectively in L. striatellus. The reduction in dib and EcR transcript abundance resulted in observable phenotypes. The development of nymphs was impaired and the survival was negatively affected. Our data will enable the development of new insect control strategies and functional analysis of vital genes in S. furcifera and L. striatellus nymphs.
Project description:Brown planthopper (BPH), Nilaparvata lugens (Stål) and white-backed planthopper (WBPH), Sogatella furcifera (Horváth) are the most destructive sucking insect pests of rice in all rice growing parts of the world. For their accurate identification at early stages, we have developed two species-specific markers (SNL4F and SNL4R for BPH; SNF2F and SNF2R for WBPH) based on mitochondrial cytochrome oxidase I (COI) for their easy detection using Polymerase Chain Reaction (PCR). The markers were developed based on nucleotide differences in COI gene and were subjected to various tests based on PCR-based gel images. The designed primers were cross-checked with five other species, which confirmed their specificity. The primers were also found to be efficient in identification of their respective species (BPH and WBPH) in all the individuals sampled from different regions of India. The lowest detection sensitivity of both the primers was up to 1 ng/µl DNA after testing them through a series of varied DNA concentrations. The species-specific primers developed in this study will help in easy and rapid identification of BPH and WBPH in all the stages of their development and in turn facilitate their timely management.Supplementary informationThe online version contains supplementary material available at 10.1007/s13205-023-03693-x.
Project description:Interspecific interactions are complex in agro-ecosystems and could be affected by agricultural technologies including transgenic crop planting. Few studies focused on the effects of Bt crops on the interspecific interactions of non-target organisms. Here we assessed the effects of transgenic cry2A rice (Cry2A rice) on the interspecific interaction between two rice planthoppers, namely, Nilaparvata lugens (the brown planthopper, BPH) and Sogatella furcifera (the white-backed planthopper, WBPH). Cry2A rice showed no significant effects on most biological parameters of these two rice planthoppers, except for wet weight of BPH female adults and development duration of WBPH female nymphs. In contrast, interspecific interactions between BPH and WBPH showed significant impacts on their biological parameters, no matter on Cry2A rice or non-transgenic control. In two-factor analysis combing rice line and interspecific interaction together, the interaction between these two factors did not affect most biological parameters of neither planthopper species, except for development duration of BPH female nymphs and WBPH nymphs (both male and female). Additionally, the egg distributions of BPH and WBPH had no significant differences between Cry2A and non-Cry2A treatments. Results of field experiments showed that Cry2A rice did not affect their population densities at most sampling dates in a five-year survey, and the interaction between BPH and WBPH showed no significant differences in both Cry2A and non-Cry2A rice paddies. In conclusion, our tested Cry2A rice would not affect the interspecific interactions between BPH and WBPH based both laboratory and field results.
Project description:BackgroundGlutathione S-transferase (GST) genes control crucial traits for the metabolism of various toxins encountered by insects in host plants and the wider environment, including insecticides. The planthoppers Nilaparvata lugens and Sogatella furcifera are serious specialist pests of rice throughout eastern Asia. Their capacity to rapidly adapt to resistant rice varieties and to develop resistance to various insecticides has led to severe outbreaks over the last decade.Methodology/principal findingsUsing the genome sequence of N. lugens, we identified for the first time the complete GST gene family of a delphacid insect whilst nine GST gene orthologs were identified from the closely related species S. furcifera. Nilaparvata lugens has 11 GST genes belonging to six cytosolic subclasses and a microsomal class, many fewer than seen in other insects with known genomes. Sigma is the largest GST subclass, and the intron-exon pattern deviates significantly from that of other species. Higher GST gene expression in the N. lugens adult migratory form reflects the higher risk of this life stage in encountering the toxins of non-host plants. After exposure to a sub-lethal dose of four insecticides, chlorpyrifos, imidacloprid, buprofezin or beta-cypermethrin, more GST genes were upregulated in S. furcifera than in N. lugens. RNA interference targeting two N. lugens GST genes, NlGSTe1 and NlGSTm2, significantly increased the sensitivity of fourth instar nymphs to chlorpyrifos but not to beta-cypermethrin.Conclusions/significanceThis study provides the first elucidation of the nature of the GST gene family in a delphacid species, offering new insights into the evolution of metabolic enzyme genes in insects. Further, the use of RNA interference to identify the GST genes induced by insecticides illustrates likely mechanisms for the tolerance of these insects.
Project description:BACKGROUND: Nilaparvata lugens (the brown planthopper, BPH) and Laodelphax striatellus (the small brown planthopper, SBPH) are two of the most important pests of rice. Up to now, there was only one mitochondrial genome of rice planthopper has been sequenced and very few dependable information of mitochondria could be used for research on population genetics, phylogeographics and phylogenetic evolution of these pests. To get more valuable information from the mitochondria, we sequenced the complete mitochondrial genomes of BPH and SBPH. These two planthoppers were infected with two different functional Wolbachia (intracellular endosymbiont) strains (wLug and wStri). Since both mitochondria and Wolbachia are transmitted by cytoplasmic inheritance and it was difficult to separate them when purified the Wolbachia particles, concomitantly sequencing the genome of Wolbachia using next generation sequencing method, we also got nearly complete mitochondrial genome sequences of these two rice planthoppers. After gap closing, we present high quality and reliable complete mitochondrial genomes of these two planthoppers. RESULTS: The mitogenomes of N. lugens (BPH) and L. striatellus (SBPH) are 17, 619 bp and 16, 431 bp long with A + T contents of 76.95% and 77.17%, respectively. Both species have typical circular mitochondrial genomes that encode the complete set of 37 genes which are usually found in metazoans. However, the BPH mitogenome also possesses two additional copies of the trnC gene. In both mitochondrial genomes, the lengths of the atp8 gene were conspicuously shorter than that of all other known insect mitochondrial genomes (99 bp for BPH, 102 bp for SBPH). That two rearrangement regions (trnC-trnW and nad6-trnP-trnT) of mitochondrial genomes differing from other known insect were found in these two distantly related planthoppers revealed that the gene order of mitochondria might be conservative in Delphacidae. The large non-coding fragment (the A+T-rich region) putatively corresponding responsible for the control of replication and transcription of mitochondria contained a variable number of tandem repeats (VNTRs) block in different natural individuals of these two planthoppers. Comparison with a previously sequenced individual of SBPH revealed that the mitochondrial genetic variation within a species exists not only in the sequence and secondary structure of genes, but also in the gene order (the different location of trnH gene). CONCLUSION: The mitochondrial genome arrangement pattern found in planthoppers was involved in rearrangements of both tRNA genes and protein-coding genes (PCGs). Different species from different genera of Delphacidae possessing the same mitochondrial gene rearrangement suggests that gene rearrangements of mitochondrial genome probably occurred before the differentiation of this family. After comparatively analyzing the gene order of different species of Hemiptera, we propose that except for some specific taxonomical group (e.g. the whiteflies) the gene order might have diversified in family level of this order. The VNTRs detected in the control region might provide additional genetic markers for studying population genetics, individual difference and phylogeographics of planthoppers.
Project description:The brown planthopper, Nilaparvata lugens, and the white-backed planthopper, Sogatella furcifera, are significant pest insects for rice. Both species are able to survive year-round in northern Vietnam's Red River Delta, which includes a large rice-producing region. This study aimed to evaluate insecticide use by farmers and its effect on the density of planthoppers in the region. Through interviews conducted with farmers in study sites in Nam Dinh (ND) and Vinh Phuc (VP) Provinces, we learned that farmers frequently used imidacloprid, fipronil, and emamectin-benzoate in ND and pymetrozine and thiamethoxam in VP. Farmers applied insecticides when the local government announcements regarding plant protection were broadcast. Generalized linear model analysis indicated that the selective insecticides applied did not contribute to reducing the densities of planthoppers in the farmers' fields. Our results indicate the possible development of insecticide resistance by planthoppers or improper insecticide application by farmers.