The identification of alternative oxidase in intermediate host snails of Schistosoma and its potential role in protecting Oncomelania hupensis against niclosamide-induced stress
Authors: Jiang Ni , Li Shi-Zhu , Zhang Yang-Wen-Qing , Habib Mohamed R. , Xiong Tao , Xu Sha , Dong Huifen , Zhao Qin-Ping 
Published: March 2022

Background

Snail intermediate hosts are mandatory for the transmission of schistosomiasis, which has to date infected more than 200 million people worldwide. Our previous studies showed that niclosamide treatment caused the inhibition of aerobic respiration and oxidative phosphorylation, and the disruption of energy supply, in one of the intermediate hosts of schistosomiasis,Oncomelania hupensis, which eventually led to the death of the snails. Meanwhile, the terminal oxidase in the mitochondrial respiratory chain, alternative oxidase (AOX), was significantly up-regulated, which was thought to counterbalance the oxidative stress and maintain metabolic homeostasis in the snails. The aims of the present study are to identify the AOXs in several species of snails and investigate the potential activation ofO. hupensisAOX(OhAOX) under niclosamide-induced stress, leading to enhanced survival of the snail when exposed to this molluscicide.

Methods

The complete complementary DNA was amplified from the AOXs ofO. hupensisand three species ofBiomphalaria; the sequence characteristics were analysed and the phylogenetics investigated. The dynamic expression and localisation of theAOXgene and protein inO. hupensisunder niclosamide-induced stress were examined. In addition, the expression pattern of genes in the mitochondrial respiratory complex was determined and the production of reactive oxygen species (ROS) calculated. Finally, the molluscicidal effect of niclosamide was compared between snails with and without inhibition of AOX activity.

Results

AOXs containing the invertebrate AOX-specific motif NP-[YF]-XPG-[KQE] were identified from four species of snail, which phylogenetically clustered together into Gastropoda AOXs and further into Mollusca AOXs. After niclosamide treatment, the levels ofOhAOXmessenger RNA (mRNA) and OhAOX protein in the whole snail were 14.8 and 2.6 times those in untreated snails, respectively, but varied widely among tissues. Meanwhile, the level of cytochrome C reductase mRNA showed a significant decrease in the whole snail, and ROS production showed a significant decrease in the liver plus gonad (liver-gonad) of the snails. At 24 h post-treatment, the mortality of snails treated with 0.06–0.1 mg/L niclosamide and AOX inhibitor was 56.31–76.12% higher than that of snails treated with 0.1 mg/L niclosamide alone.

Conclusions

AOX was found in all the snail intermediate hosts ofSchistosomaexamined here. AOX was significantly activated inO. hupensisunder niclosamide-induced stress, which led to a reduction in oxidative stress in the snail. The inhibition of AOX activity in snails can dramatically enhance the molluscicidal effect of niclosamide. A potential target for the development of an environmentally safe snail control method, which acts by inhibiting the activity of AOX, was identified in this study.

Graphical abstract