Identification and characterization of soluble binding proteins associated with host foraging in the parasitoid wasp Diachasmimorpha longicaudata.

The communication and reproduction of insects are driven by chemical sensing. During this process, chemical compounds are transported across the sensillum lymph to the sensory neurons assisted by different types of soluble binding proteins: odorant-binding proteins (OBPs); chemosensory proteins (CSPs); some members of ML-family proteins (MD-2 (myeloid differentiation factor-2)-related Lipid-recognition), also known as NPC2-like proteins. Potential transcripts involved in chemosensing were identified by an in silico analysis of whole-body female and male transcriptomes of the parasitic wasp Diachasmimorpha longicaudata. This analysis facilitated the characterization of fourteen OBPs (all belonging to the Classic type), seven CSPs (and two possible isoforms), and four NPC2-like proteins. A differential expression analysis by qPCR showed that eleven of these proteins (CSPs 2 and 8, OBPs 2, 3, 4, 5, 6, 9, 10, and 11, and NPC2b) were over-expressed in female antenna and two (CSP 1 and OBP 12) in the body without antennae. Foraging behavior trials (linked to RNA interference) suggest that OBPs 9, 10, and 11 are potentially involved in the female orientation to chemical cues associated with the host. OBP 12 seems to be related to physiological processes of female longevity regulation. In addition, transcriptional silencing of CSP 3 showed that this protein is potentially associated with the regulation of foraging behavior. This study supports the hypothesis that soluble binding proteins are potentially linked to fundamental physiological processes and behaviors in D. longicaudata. The results obtained here contribute useful information to increase the parasitoid performance as a biological control agent of fruit fly pest species.

Transcriptome analysis of Anastrepha fraterculus sp. 1 males, females, and embryos: insights into development, courtship, and reproduction.

BACKGROUND: Anastrepha fraterculus sp. 1 is considered a quarantine pest in several American countries. Since chemical control applied in an integrated pest management program is the only strategy utilized against this pest, the development of pesticide-free methods, such as the Sterile Insect Technique, is being considered. The search for genes involved in sex-determination and differentiation, and in metabolic pathways associated with communication and mating behaviour, contributes with key information to the development of genetic control strategies. The aims of this work were to perform a comprehensive analysis of A. fraterculus sp. 1 transcriptome and to obtain an initial evaluation of genes associated with main metabolic pathways by the expression analysis of specific transcripts identified in embryos and adults. RESULTS: Sexually mature adults of both sexes and 72 h embryos were considered for transcriptome analysis. The de novo transcriptome assembly was fairly complete (62.9% complete BUSCO orthologs detected) with a total of 86,925 transcripts assembled and 28,756 GO annotated sequences. Paired-comparisons between libraries showed 319 transcripts differently expressed between embryos and females, 1242 between embryos and males, and 464 between sexes. Using this information and genes searches based on published studies from other tephritid species, we evaluated a set of transcripts involved in development, courtship and metabolic pathways. The qPCR analysis evidenced that the early genes serendipity alpha and transformer-2 displayed similar expression levels in the analyzed stages, while heat shock protein 27 is over-expressed in embryos and females in comparison to males. The expression of genes associated with courtship (takeout-like, odorant-binding protein 50a1) differed between males and females, independently of their reproductive status (virgin vs mated individuals). Genes associated with metabolic pathways (maltase 2-like, androgen-induced gene 1) showed differential expression between embryos and adults. Furthermore, 14,262 microsatellite motifs were identified, with 11,208 transcripts containing at least one simple sequence repeat, including 48% of di/trinucleotide motifs. CONCLUSION: Our results significantly expand the available gene space of A. fraterculus sp. 1, contributing with a fairly complete transcript database of embryos and adults. The expression analysis of the selected candidate genes, along with a set of microsatellite markers, provides a valuable resource for further genetic characterization of A. fraterculus sp. 1 and supports the development of specific genetic control strategies.

Genetic variation and heteroplasmy of Varroa destructor inferred from ND4 mtDNA sequences.

Varroa destructor, a parasitic mite of the western honey bee, Apis mellifera L., is a serious threat to colonies and beekeeping worldwide. Population genetics studies of the mite have provided information on two mitochondrial haplotypes infecting honey bee colonies, named K and J (after Korea and Japan, respectively, where they were originally identified). On the American continent, the K haplotype is much more prevalent, with the J haplotype only detected in some areas of Brazil. The aims of the present study were to assess the genetic diversity of V. destructor populations in the major beekeeping region of Argentina and to evaluate the presence of heteroplasmy at the nucleotide level. Phoretic mites were collected from managed A. mellifera colonies in ten localities, and four mitochondrial DNA (mtDNA) regions (COXI, ND4, ND4L, and ND5) were analyzed. Based on cytochrome oxidase subunit I (COXI) sequencing, exclusively the K haplotype of V. destructor was detected. Furthermore, two sub-haplotypes (KArg-N1 and KArg-N2) were identified from a variation in ND4 sequences and the frequency of these sub-haplotypes was found to significantly correlate with geographical latitude. The occurrence of site heteroplasmy was also evident for this gene. Therefore, ND4 appears to be a sensitive marker for detecting genetic variability in mite populations. Site heteroplasmy emerges as a phenomenon that could be relatively frequent in V. destructor.

Complementary sex determination in the parasitic wasp Diachasmimorpha longicaudata.

We studied the sex determination in Diachasmimorpha longicaudata, a parasitoid braconid wasp widely used as biological control agent of fruit pest tephritid flies. We tested the complementary sex determination hypothesis (CSD) known in at least 60 species of Hymenoptera. According to CSD, male or female development depends on the allelic composition of one sex locus (single-locus CSD) or multiple sex loci (multiple-locus CSD). Hemizygote individuals are normal haploid males, and heterozygotes for at least one sex locus are normal diploid females, but homozygotes for all the sex loci are diploid males. In order to force the occurrence of diploid males in D. longicaudata, we established highly inbred lines and examined their offspring using chromosome counting, flow cytometry, and sex ratio analysis. We found that when mother-son crosses were studied, this wasp produced about 20% of diploid males out of the total male progeny. Our results suggest that this parasitoid may represent the second genus with multiple-locus CSD in Hymenoptera. Knowledge about the sex determination system in D. longicaudata is relevant for the improvement of mass rearing protocols of this species. This information also provides the necessary background for further investigations on the underlying molecular mechanisms of sex determination in this species, and a better insight into the evolution of this pathway in Hymenoptera in particular and insects in general.