Neuropeptides predicted from the transcriptome analysis of the gray garden slug Deroceras reticulatum.

Abstract

The gray garden slug, Deroceras reticulatum (Gastropoda: Pulmonata), is one of the most common terrestrial molluscs. Research for this slug has focused mainly on its ecology, biology, and management due to the severe damage it causes on a wide range of vegetables and field crops. However, little is known about neuropeptides and hormonal signalings. This study, therefore, aimed to establish the transcriptome of D. reticulatum and to identify a comprehensive repertoire of neuropeptides in this slug. Illumina high-throughput sequencing of the whole body transcriptome of D. reticulatum generated a total of 5.9 billion raw paired-end reads. De novo assembly by Trinity resulted in 143,575 transcripts and further filtration selected 120,553 unigenes. Gene Ontology (GO) terms were assigned to 30,588 unigenes, composed of biological processes (36.9%), cellular components (30.2%) and molecular functions (32.9%). Functional annotation by BLASTx revealed 39,987 unigenes with hits, which were further categorized into important functional groups based on sequence abundance. Neuropeptides, ion channels, ribosomal proteins, G protein-coupled receptors, detoxification, immunity and cytoskeleton-related sequences were dominant among the transcripts. BLAST searches and PCR amplification were used to identify 65 putative neuropeptide precursor genes from the D. reticulatum transcriptome, which include achatin, AKH, allatostatin A, B and C, allatotropin, APGWamide, CCAP, cerebrin, conopressin, cysteine-knot protein hormones (bursicon alpha/beta and GPA2/GPB5), elevenin, FCAP, FFamide, FVamide (enterin, fulicin, MIP and PRQFVamide), GGNG, GnRH, insulin, NdWFamide, NKY, PKYMDT, PRXamide (myomodulin, pleurin and sCAP), RFamide (CCK/SK, FMRFamide, FxRIamide, LFRFamide, luqin and NPF), and tachykinin. Over 330 putative peptides were encoded by these precursors. Comparative analysis among different molluscan species clearly revealed that, while D. reticulatum neuropeptide sequences are conserved in Mollusca, there are also some unique features distinct from other members of this species. This is the first transcriptome-wide report of neuropeptides in terrestrial slugs. Our results provide comprehensive transcriptome data of the gray garden slug, with a more detailed focus on the rich repertoire of putative neuropeptide sequences, laying the foundation for molecular studies in this terrestrial slug pest.