Abstract
BackgroundThe biting midge, Forcipomyia taiwana, is one of the most annoying blood-sucking pests in Taiwan. Current chemical control methods only target the adult, not the immature stages (egg to pupa), of F. taiwana. Discovering new or alternative tactics to enhance or replace existing methods are urgently needed to improve the effectiveness of F. taiwana control. The egg is the least understood life stage in this pest species but may offer a novel point of control as addition of NaCl to the egg environment inhibits development. Thus, the objective of this study was to use RNA profiling to better understand the developmental differences between wild-type melanized (black) and NaCl-induced un-melanized (pink), infertile F. taiwana eggs.ResultsAfter de novo assembly with Trinity, 87,415 non-redundant transcripts (Ft-nr) with an N50 of 1099 were obtained. Of these, 26,247 (30%) transcripts were predicted to have long open reading frames (ORFs, defined here as ≥300 nt) and 15,270 (17.5%) transcripts have at least one predicted functional domain. A comparison between two biological replicates each of black and pink egg samples, although limited in sample size, revealed 5898 differentially expressed genes (DEGs; 40.9% of the transcripts with long ORFs) with ≥2-fold difference. Of these, 2030 were annotated to a Gene Ontology biological process and along with gene expression patterns can be separated into 5 clusters. KEGG pathway analysis revealed that 1589 transcripts could be assigned to 18 significantly enriched pathways in 2 main categories (metabolism and environmental information processing). As expected, most (88.32%) of these DEGs were down-regulated in the pink eggs. Surprisingly, the majority of genes associated with the pigmentation GO term were up-regulated in the pink egg samples. However, the two key terminal genes of the melanin synthesis pathway, laccase2 and DCE/yellow, were significantly down-regulated, and further verified by qRT-PCR.ConclusionWe have assembled and annotated the first egg transcriptome for F. taiwana, a biting midge. Our results suggest that down-regulation of the laccase2 and DCE/yellow genes might be the mechanism responsible for the NaCl-induced inhibition of melanization of F. taiwana eggs.
Highlights
The biting midge, Forcipomyia taiwana, is one of the most annoying blood-sucking pests in Taiwan
In an attempt to explore treatments that are effective for F. taiwana control with minimal adverse effects for humans or the environment, we fortuitously found that the eggs of F. taiwana often did not undergo normal darkening of color and were inviable when laid in salt-containing milieu (Fig. 1)
F. taiwana transcriptome assembly and annotation To study F. taiwana gene expression differences between black and pink (NaCl treated; Fig. 1B) embryos, we sequenced RNA from two biological replicates of each sample type, generating 12 to 41 million raw paired-end reads per sample
Summary
The biting midge, Forcipomyia taiwana, is one of the most annoying blood-sucking pests in Taiwan. Current chemical control methods only target the adult, not the immature stages (egg to pupa), of F. taiwana. Over 6200 species of biting midges (Ceratopogonidae, Diptera) in 112 genera have been described worldwide [1] and some of them can be severe biting pests of humans, pets, livestock, and wildlife. They are minute to tiny, their bite can inflict a burning sensation which results in different reactions in humans, ranging from a small reddish welt at the bite site to strong local allergic reactions accompanied by significant itching [2]. Discovering new or alternative tactics to enhance or replace existing methods are urgently needed to improve the effectiveness of F. taiwana control
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