Identification and characterization of microRNAs in the screwworm flies Cochliomyia hominivorax and Cochliomyia macellaria (Diptera: Calliphoridae)

Abstract

MicroRNAs (miRNAs) are small noncoding RNAs that modulate gene expression through post-transcriptional regulation. Here, we report the identification and characterization of miRNAs in two closely related screwworm flies with different feeding habits: Cochliomyia hominivorax and Cochliomyia macellaria. The New World screwworm, C. hominivorax, is an obligatory parasite of warm-blooded vertebrates, whereas the secondary screwworm, C. macellaria, is a free-living organism that feeds on decaying organic matter. Here, the small RNA transcriptomes of adults and third-instar larvae of both species were sequenced. A total of 110 evolutionarily conserved miRNAs were identified, and 10 putative precursor miRNAs (pre-miRNAs) were predicted. The relative expression of six selected miRNAs was further investigated, including miRNAs that are related to reproduction and neural processes in other insects. Mature miRNAs were also characterized across an evolutionary time scale, suggesting that the majority of them have been conserved since the emergence of the Arthropoda [540 million years ago (Ma)], Hexapoda (488 Ma) and Brachycera (195 Ma) lineages. This study is the first report of miRNAs for screwworm flies. We also performed a comparative analysis with the hereby predicted miRNAs from the sheep blowfly, Lucilia cuprina. The results presented may advance our understanding of parasitic habits within Calliphoridae and assist further functional studies in blowflies.

Regulatory Pathways Controlling Female Insect Reproduction

“…Loss of function of miR-184 in adult females leads to abnormal mating receptivity and low levels of fecundity (55, 86). miR-184-3p has also been linked to mating and reproduction in the screwworm flies Cochliomyia hominivorax and C. macellaria (134). In D. melanogaster, knockout of miRNA let-7, which has been linked to ecdysteroid signaling, led to moderately severe defects in female fertility and oviposition (165).…”

microRNA Expression Dynamics in Culicoides sonorensis Biting Midges Following Blood-Feeding

“…While the miR-309 cluster is observed in other hematophagous dipterans, stable flies [67] and tsetse flies [55,56], these organisms lack miRNAs miR-2944a and miR-2944b. Interestingly, miR-1174 cluster is conserved across mosquito species [68] but not found in current miRNA libraries for other hematophagous Diptera such as Lutzomyia [58], Glossina [55,56], Cochliomyia [57], and Stomoxys [67]. Together, these data suggest that the expansion of the miR-309 cluster and the origin of the miR-1174 cluster are evolutionarily distinct to Culicomorpha, as mosquitoes (Culicidae) and midges (Ceratopogonidae) share an evolutionary origin for hematophagy that is separate from the independent evolutionary origins of hematophagy in sand flies (Psychodidae), tsetse flies (Glossinidae), screwworms (Calliphoridae), and stable flies (Muscidae) [69].…”

“…Our data revealed Culicoides cso-miR-956 was enriched in midgut tissues. In miRBase, miR-956 is only described in Drosophila but has been reported in multiple Dipteran taxa, including Culicomorpha (Culicidae) and Calyptratae muscoids (Sarcophagidae and Calliphoridae) [49,[55][56][57][58]. Work on Drosophila has alluded to multiple potential functions of miR-956, as this miRNA is thought to regulate the circadian rhythm [59] and has been recently shown to have a role in intestinal cell differentiation [60].…”

Identification and Characterization of Small RNA Markers of Age in the Blow Fly Cochliomyia macellaria (Fabricius) (Diptera: Calliphoridae)

“…To determine if there was age-dependent miRNA expression in the blow-fly Cochliomyia macellaria , we sequenced, identified, and analyzed the expression of known miRNAs across seven developmental time points in immature development (three third instars, four intrapuparial) of wild-type colony-raised flies. While miRNA have been identified in other flies of similar forensic, medical, and veterinary relevance [ 55 , 74 , 75 ], this is the first study outside of Drosophila , to our knowledge, which investigated patterns of miRNA expression using RNAseq in the whole insect in a developmental context. Our methods allowed us to identify 217 distinct, known animal miRNA sequences in C. macellaria ( Table S1 ).…”