Molecular Identification, Transcriptome Sequencing and Functional Annotation of Pulex irritans.

Abstract

Pulex irritans are vectors of various zoonotic pathogens. However, molecular studies on P. irritans and flea-borne diseases are limited due to the lack of molecular data. This study aimed to conduct transcriptome sequencing, functional annotation, and pathogen analysis of P. irritans. Fleas collected from a dog were identified morphologically and molecularly. RNA was extracted for transcriptome sequencing and functional annotation. Open reading frames (ORFs) of unigenes were confirmed by employing bioinformatics strategies, and maximum likelihood (ML) trees were reconstructed based on the highly expressed genes of ejaculation globulin-specific 3-like protein, salivary protein, and actin for phylogenetic relationship analysis. The obtained mitochondrial 16S rRNA gene sequences showed 99.71% of similarity with P. irritans obtained from GenBank database. Transcriptome sequencing generated 74,412 unigenes, of which 53,211 were functionally annotated. A total of 195 unigenes were assigned to fleas, of which 69 contained complete ORFs. Phylogenetic trees of both ejaculatory globulin and salivary protein genes demonstrated that P. irritans first clustered with Pulicidae sp., indicating the reliability of transcriptome data. It is noteworthy that 1070 unigenes were assigned to Hymenolepis microstoma and Dipylidium caninum, of which 62 contained complete ORFs. The phylogenetic tree of the actin gene showed that the unigenes had closer relationships with Echinococcus sp., suggesting the role of P. irritans as intermediate hosts of tapeworms. The results of this study provide the possibility for functional exploration of important genes and lay foundations for the prevention and control of P. irritans and flea-borne diseases.