Regulating Effect Of MiR-36f On Larvae Development Of Ascaris Suum And Immunological Evaluation Of DNA Vaccines Of Its Targets

Ascaris suum(A. suum) belongs to Ascaridida, ascarididae, and Ascaris, and is the largest parasitic nematode in pigs causing huge economic losses worldwide. A. suum consumes the host nutrients and causes intestinal blockage in pigs. A. suum larvae shift between host organs results in mechanical injury and further infection. Although chemical drugs have certain effect, drug resistance and residues have occurred due to long-term and unscientificlly usage. Therefore, development of new drugs for parasite control is becoming imperative to scientific researchers.MicroRNA(miRNA) is a class of non-coding, small RNA, which regulates gene expression at the post transcriptional level. It is known now that as an important post transcriptional regulator, miRNA plays important roles in parasite development, which may suggest new ways for the parasite control. DNA vaccine, also known as naked DNA vaccine, not only can stimulate the body humoral immune response, but also induce cellular immune responses, with fine security.We herein carried out two experiments to study miR-36 f and its targets:1. Preliminary evaluation of the effect of asu-miR-36 f on the development of A. suum larvae.In our previous research, asu-miR-36 f is only found to be expressed in eggs and larvae stage, indicating an important role of the miRNA to early development of the parasite. We therefore designed and synthesized the miRNA mimics and inhibitor to evaluate miRNA function via soaking method. Results showed that, not only the recovered number of larvae, but also the length and width of the recovered worms in the inhibitor group were significantly smaller to the control group, suggesting a regulating effect of the asu-miR-36 f on larvae development. However, significant difference was not found between the mimics group and control groups, which suggested that the function of miRNA might depend on a certain threshold.2. DNA vaccine preparation and immune effect evaluation.By the bioinformatics analysis, from the 153 miR-36 f targets, we selected 2 key target genes(NCS and AG) existed only in A. suum for DNA vaccine preparation. Results showed that the pVAX-NCS effectively induced experimental mice to produce humoral and cellular immune responses, with the worm reduction rate of 35.96%(P < 0.05); pVAX-AG also induced the mice to produce humoral and cellular immune responses, but the responses were weaker than that induced by pVAX-NCS(P > 0.05). Therefore, PVAX-NCS was proved to be of protective immunity on mice against A. suum infection.In summary, the present study demonstrated the regulating effect of mir-36 f on A. suum larvae development. The DNA vaccines based on its targets were successfully constructed and evaluated, which provided foundation for the development of DNA vaccines against A. suum. Furthermore, because miR-36 f is also existed in other worms, the present study also provides foundation for the control of other parasites by DNA vaccination.