A transcriptomic survey of Migdolus fryanus (sugarcane rhizome borer) larvae.
ABSTRACT: Sugarcane, a major crop grown in the tropical and subtropical areas of the world, is produced mainly for sucrose, which is used as a sweetener or for the production of bioethanol. Among the numerous pests that significantly affect the yield of sugarcane, the sugarcane rhizome borer (Migdolus fryanus, a cerambycidae beetle) is known to cause severe damage to the crops in Brazil. The absence of molecular information about this insect reinforces the need for studies and an effective method to control this pest. In this study, RNA-Seq technology was employed to study different parts of M. fryanus larvae. The generated data will help in further investigations about the taxonomy, development, and adaptation of this insect. RNA was extracted from six different parts (head, fat body, integument, hindgut, midgut, and foregut) using Trizol methodology. Using Illumina paired-end sequencing technology and the Trinity platform, trimming and de novo assembly was performed, resulting in 44,567 contigs longer than 200 nt for a reunion of data from all transcriptomes, with a mean length of 1,095.27 nt. Transcripts were annotated using BLAST against different protein databanks (Uniprot/Swissprot, PFAM, KEEG, SignalP 4.1, Gene Ontology, and CAZY) and were compared for similarity using a Venn diagram. Differential expression patterns were studied for select genes through qPCR and FPKM comprising important protein families (digestive peptidases, glucosyl hydrolases, serine protease inhibitors and otopetrin), which allowed a better understanding of the insect's digestion, immunity and gravity sensorial mechanisms.
Project description:Cacosceles newmannii (Coleoptera: Cerambycidae) is an emerging pest of sugarcane in South Africa. The larvae of this cerambycid beetle live within the sugarcane stalk and drill galleries that considerably reduce sugar production. To provide an alternative to chemical control, entomopathogenic nematodes and fungus were investigated as potential biological control agents to be used in an integrated pest management system. The nematodes Steinernema yirgalemense, S. jeffreyense, Heterorhabditis indica, and different concentrations of the fungus Metarhizium pinghaense were screened for efficacy (i.e., mortality rate) against larvae of C. newmannii. The different biocontrol agents used, revealed a low level of pathogenicity to C. newmannii larvae, when compared to control treatments.
Project description:MicroRNA-regulation of gene expression plays a key role in the development and response to biotic and abiotic stresses. Deep sequencing analyses accelerate the process of small RNA discovery in many plants and expand our understanding of miRNA-regulated processes. We therefore undertook small RNA sequencing of sugarcane miRNAs in order to understand their complexity and to explore their role in sugarcane biology.A bioinformatics search was carried out to discover novel miRNAs that can be regulated in sugarcane plants submitted to drought and salt stresses, and under pathogen infection. By means of the presence of miRNA precursors in the related sorghum genome, we identified 623 candidates of new mature miRNAs in sugarcane. Of these, 44 were classified as high confidence miRNAs. The biological function of the new miRNAs candidates was assessed by analyzing their putative targets. The set of bona fide sugarcane miRNA includes those likely targeting serine/threonine kinases, Myb and zinc finger proteins. Additionally, a MADS-box transcription factor and an RPP2B protein, which act in development and disease resistant processes, could be regulated by cleavage (21-nt-species) and DNA methylation (24-nt-species), respectively.A large scale investigation of sRNA in sugarcane using a computational approach has identified a substantial number of new miRNAs and provides detailed genotype-tissue-culture miRNA expression profiles. Comparative analysis between monocots was valuable to clarify aspects about conservation of miRNA and their targets in a plant whose genome has not yet been sequenced. Our findings contribute to knowledge of miRNA roles in regulatory pathways in the complex, polyploidy sugarcane genome.
Project description:Plants respond to pathogens and insect attacks by inducing and accumulating a large set of defense-related proteins. Two homologues of a barley wound-inducible protein (BARWIN) have been characterized in sugarcane, SUGARWIN1 and SUGARWIN2 (sugarcane wound-inducible proteins). Induction of SUGARWINs occurs in response to Diatraea saccharalis damage but not to pathogen infection. In addition, the protein itself does not show any effect on insect development; instead, it has antimicrobial activities toward Fusarium verticillioides, an opportunistic fungus that usually occurs after D. saccharalis borer attacks on sugarcane. In this study, we sought to evaluate the specificity of SUGARWIN2 to better understand its mechanism of action against phytopathogens and the associations between fungi and insects that affect plants. We used Colletotrichum falcatum, a fungus that causes red rot disease in sugarcane fields infested by D. saccharalis, and Ceratocystis paradoxa, which causes pineapple disease in sugarcane. We also tested whether SUGARWIN2 is able to cause cell death in Aspergillus nidulans, a fungus that does not infect sugarcane, and in the model yeast Saccharomyces cerevisiae, which is used for bioethanol production. Recombinant SUGARWIN2 altered C. falcatum morphology by increasing vacuolization, points of fractures and a leak of intracellular material, leading to germling apoptosis. In C. paradoxa, SUGARWIN2 showed increased vacuolization in hyphae but did not kill the fungi. Neither the non-pathogenic fungus A. nidulans nor the yeast S. cerevisiae was affected by recombinant SUGARWIN2, suggesting that the protein is specific to sugarcane opportunistic fungal pathogens.
Project description:Otopetrin 1 (Otop1) encodes a multi-transmembrane domain protein with no homology to known transporters, channels, exchangers, or receptors. Otop1 is necessary for the formation of otoconia and otoliths, calcium carbonate biominerals within the inner ear of mammals and teleost fish that are required for the detection of linear acceleration and gravity. Vertebrate Otop1 and its paralogues Otop2 and Otop3 define a new gene family with homology to the invertebrate Domain of Unknown Function 270 genes (DUF270; pfam03189).Multi-species comparison of the predicted primary sequences and predicted secondary structures of 62 vertebrate otopetrin, and arthropod and nematode DUF270 proteins, has established that the genes encoding these proteins constitute a single family that we renamed the Otopetrin Domain Protein (ODP) gene family. Signature features of ODP proteins are three "Otopetrin Domains" that are highly conserved between vertebrates, arthropods and nematodes, and a highly constrained predicted loop structure.Our studies suggest a refined topologic model for ODP insertion into the lipid bilayer of 12 transmembrane domains, and highlight conserved amino-acid residues that will aid in the biochemical examination of ODP family function. The high degree of sequence and structural similarity of the ODP proteins may suggest a conserved role in the intracellular trafficking of calcium and the formation of biominerals.
Project description:The longhorn beetle Dorysthenes paradoxus (Faldermann, 1833) (Coleoptera: Cerambycidae) is not only a serious agricultural pest but also a traditionally edible insect in China. However, no genetic information on this species has been acquired. In the present study, we report the mitochondrial genome (mitogenome) of Do. paradoxus, as the first complete mitogenome of Prioninae. The circular mitogenome of 15,922 bp encodes 13 protein-coding genes (PCGs), 22 transfer RNAs (tRNAs), and two ribosomal RNAs (rRNAs), and it contains an A+T-rich region. This mitogenome exhibits the lowest A+T content (71.13%) but harbors the largest AT skew (0.116) among the completely sequenced Cerambycidae species. Eleven of the 13 PCGs have a typical ATN start codon, whereas COI and ND1 are tentatively designated by AAT and TTG, respectively. Only 4 of the 13 PCGs harbor a complete termination codon, and the remaining 9 possess incomplete termination codons (T or TA). Apart from tRNASer(AGN), the other 21 tRNAs can fold into a typical clover-leaf secondary structures. The Do. paradoxus A+T-rich region contains two poly-T stretches and a tandem repeat that comprises two 47-bp-long copies. Both Bayesian inference and Maximum likelihood analyses confirmed the subfamily ranks of Cerambycidae ([Prioninae + Cerambycinae] + Lamiinae) and the close relationship between Philinae and Prioninae/Cerambycinae. However, the data did not support the monophyly of Prioninae and Cerambycinae. The mitogenome presented here provides basic genetic information for this economically important species.
Project description:Global climate change is expected to affect temperature and precipitation patterns worldwide, which in turn is likely to affect insect phenology, distribution and diversity. To improve our understanding of such processes, it is important to understand how insects may respond to changes in seasonality, and how these affect their activity, patterns of distribution and species richness. The tropical dry forest (TDF) is a highly seasonal ecosystem, for which two seasons are commonly described (rainy and dry) and there is a lack of information on the combined effect of both precipitation and temperature on the insect communities. In order to evaluate the seasonal patterns in the community of Cerambycidae in a TDF, historical climatic variables were obtained, and an annual sampling of the family was carried out, using three collection techniques. We found that the Cerambycidae family showed a more complex response to climate, than simply the rainy and dry season of the year. The relationship between diversity and composition of cerambycids with changes in temperature and precipitation showed four seasonal communities which were synchronized with phenological processes of the TDF. Climate change could reduce biodiversity, causing seasonal patterns to lose complexity, either because the climatic characteristics of a season disappear and/or because the duration of a season expands, these changes will modify the ecological processes of the TDF, since they would generate changes in the flora and fauna associated with the different seasons.
Project description:Human-mediated changes in landscapes can facilitate niche expansion and accelerate the adaptation of insect species. The interaction between the evolutionary history of the sugarcane borer, Diatraea saccharalis Fabricius, and historical and modern agricultural activity in Brazil shaped its spatial genetic structure, facilitating ecological divergence and incipient host shifting. Based on microsatellite data, STRUCTURE analyses identified two (K = 2) and three (K = 3) significant genetic clusters that corresponded to: (a) a strong signal of spatial genetic structure and, (b) a cryptic signal of host differentiation. We inferred that K = 2 reflects the footprint of agricultural activity, such as expansion of crop production (sugarcane and maize), unintentional dispersion of pests, and management practices. In contrast, K = 3 indicated incipient host differentiation between larvae collected from sugarcane or maize. Our estimates of population size changes indicated that a historical bottleneck was associated with a reduction of sugarcane production ≈200 years ago. However, a more recent population expansion was detected (>1950s), associated with agricultural expansion of large crop production into previously unfarmed land. Partial Mantel tests supported our hypothesis of incipient host adaptation, and identified isolation-by-environment (e.g., host plant) in São Paulo and Minas Gerais states, where sugarcane has been traditionally produced in Brazil. The impact of agricultural production on D. saccharalis may continue, as the current population structure may hinder the efficacy of refuge plants in delaying insect resistance evolution to Bt toxin.
Project description:The sugarcane borer or corn stalk borer, Diatraea Guilding is polyphagous insect pest of many important crops such as corn, sorghum and sugarcane. Losses arising from the attack of Diatraea species have been a serious problem, which may cause loss in sugarcane production around 0.25% in sugar, 0.20% in alcohol and 0.77% of body weight for every 1% infestation and up to 21% in corn production fields. In Brazil, the most commonly reported species are Diatraea saccharalis (Fabricius, 1794) and Diatraea impersonatella (Walker, 1863) (= D. flavipennella). However, multiple other species of Diatraea have been identified in Brazil according to the literature. Currently, little information exists on the presence of the other species causing injury to sugarcane and corn. The objectives of this study were to improve the accuracy of species assignment, evaluate the population genetic structure, and address many of the outstanding questions of systematics and evolution of Brazilian populations of D. saccharalis. To address these main questions, classical taxonomic methods were used, focused on morphological characterization of the reproductive organs, especially the male genitalia. In addition, genetic studies were performed using simple sequence repeats (SSR) and a fragment of cytochrome C oxidase subunit I (COI) gene. The data and findings from this research will contribute to the understanding of evolutionary aspects of insect pests in order to develop more effective and sustainable population management practices.
Project description:Plants have developed multiple regulatory mechanisms to respond and adapt to stress. Drought stress is one of the major constraints to agricultural productivity worldwide and recent reports have highlighted the importance of plant sRNA in the response and adaptation to water availability. In order to increase our understanding of the roles of sRNA in response to water depletion, cultivars of sugarcane were submitted to treatment of ceasing drip irrigation for 24 hours. Deep sequencing analysis was carried out to identify the sRNA regulated in leaves and roots of sugarcane cultivars with different drought sensitivities. The pool of sRNA selected allowed the analysis of different sRNA classes (miRNA and siRNA). Twenty-eight and 36 families of conserved miRNA were identified in leaf and root libraries, respectively. Dynamic regulation of miRNA was observed and the expression profiles of eight miRNA were verified in leaf samples from three biological replicates by stem-loop qRT-PCR assay using the cultivars: SP90-1638--sensitive cultivar--and SP83-2847 and SP83-5073--tolerant cultivars. Altered miRNA regulation was correlated with changes in mRNA levels of specific targets. Two leaf libraries from individual sugarcane cultivars with contrasting drought-tolerance properties were also analyzed. An enrichment of 22-nt sRNA species was observed in leaf libraries. 22-nt miRNA triggered siRNA production by cleavage of their targets in response to water depletion. A number of genes of the sRNA biogenesis pathway were down-regulated in tolerant genotypes and up-regulated in sensitive in response to water depletion treatment. Our analysis contributes to increase the knowledge on the roles of sRNA in sugarcane submitted to water depletion.
Project description:Lepidoptera (butterflies and moths) is a major insect order including important pollinators and agricultural pests, however their microbiomes are little studied. Here, using next-generation sequencing (NGS)-based shotgun metagenomics, we characterize both the biodiversity and functional potential of gut microbiota of a lepidopteran model insect, the silkworm Bombyx mori. Two metagenomes, including the standard inbred strain Dazao (P50) and an improved hybrid strain Qiufeng × Baiyu (QB) widely used in commercial silk production, were generated, containing 45,505,084 and 69,127,002 raw reads, respectively. Taxonomic analysis revealed that a total of 663 bacterial species were identified in P50 silkworms, while 322 unique species in QB silkworms. Notably, Enterobacter, Acinetobacter and Enterococcus were dominated in both strains. The further functional annotation was performed by both BlastP and MG-RAST against various databases including Nr, COG, KEGG, CAZy and SignalP, which revealed >5 × 106 protein-coding genes. These datasets not only provide first insights into all bacterial genes in silkworm guts, but also help to generate hypotheses for subsequently testing functional traits of gut microbiota in an important insect group.