Parasitology

Biography

Biography: Jing Cui

Abstract

RNA interference (RNAi) technique was widely used in variety of organisms, including parasites, as a powerful tool for gene functional study. Th e aim of this study was to investigate the functions of T. spiralis Nudix hydrolase (TsNd) during the larval invasion of intestinal epithelial cells (IECs), development and survival in host by RNAi. Th e TsNd-specifi c doublestranded RNA (dsRNA) was designed to silence the expression of TsNd in T. spiralis larvae. DsRNA were delivered to the larvae by soaking incubation or electroporation. Silencing eff ect of TsNd transcription and expression was determined by real-time PCR and Western blotting, respectively. Th e infectivity of larvae treated with dsRNA was investigated by the in vitro larval invasion of IECs and experimental infection in mice. Aft er being soaked with 40 ng/μl of dsRNA-TsNd, the transcription and expression level of TsNd gene was inhibited 65.8% and 56.4%, respectively (P<0.05). Aft er being electroporated with 40 ng/μl of dsRNA-TsNd, the transcription and expression level of TsNd gene was inhibited 74.2% and 58.2%, respectively (P<0.05). Silencing TsNd expression by both soaking and electroporation inhibited signifi cantly the larval invasion of IECs in a dose-dependent manner (r1= -0.96798, r2=-0.98707). Th e invasion rate of the larvae soaked with 20, 30, 40, 50 or 60 ng/μl dsRNA-TsNd for 18 h was 54.4%, 44.7%, 39.2%, 35.3%, and 32.7%, respectively; while the invasion rate of the larvae treated with lip2000 or untreated was 61.7% and 63.1%, respectively. Th e invasion rate of the larvae eletroporated with the above-mentioned dose of dsRNA-TsNd for 18 h was 54.4%, 44.7%, 39.2%, 35.3%, and 32.7%, respectively; while the invasion rate of the larvae untreated was 58.3%. Mice inoculated with larvae soaked with TsNd dsRNA displayed a 49.9% reduction in intestinal adult worm burden and 39.9% reduction in muscle larval burden compared with the mice inoculated with untreated larvae (P<0.05). Mice infected with larvae electroporated with TsNd dsRNA displayed an 83.4% reduction in intestinal adult worm burden and 69.5% reduction in muscle larval burden compared with the mice inoculated with untreated larvae (P<0.05), indicating that electroporation had a higher effi ciency than soaking in inhibiting the larval development and survival in mice.Our results showed that silencing TsNd expression in T. spiralis inhibited signifi cantly the larval invasion and survival in host.