Calmodulin-specific Small Interfering RNA Induces Consistent Expression Suppression and Morphological Changes in Echinococcus Granulosus

Among parasitic helminths, biological features of Echinococcus granulosus have been a focus of particular interest in biology and medicine. The determinants and underlying molecular mechanisms of Echinococcus development in different host settings is largely unknown. The phenomenal bi-directional development of E. granulosus protoscoleces into multi-proglottid and/or microcysts, is a fascinating feature of the parasite cultivation. Calmodulin (CaM) is the major intracellular Ca2+ binding protein in plant and animal organisms. Many Ca-related processes in the physiology of eukaryotic organisms are CaM-dependent, however little is known on the role of CaM in platyhelminths growth and development. Small interfering (si) RNA-induced manipulations of the genes involving in the parasite development is an opportunity to explore novel approaches for cystic echinococcosis (CE) prevention and management. Regarding the fundamental role of CaM in cellular function of the parasites, in this study, we investigated the molecular and morphological changes induced by siRNA on CaM in different in vitro stages of E. granulosus. Three developmental stages of the tapeworm, protoscoleces, microcysts and strobilated worms, were cultivated in vitro in mono- and di-phasic media and three delivery methods, i.e. electroporation, soaking and electro-soaking, were used for RNA interference. The level of mRNA suppression as well as the phenotypic changes of the parasites were measured. Following RNA interference, EgCaM mRNA suppressions of 65-99% were recorded in different stages of the tapeworm as compared to untreated/unrelated siRNA controls. Lower viability, growth retardation, morphological abnormalities as well as EgCaM expression suppression were documented in the parasite implying potential of siRNA technology for the prevention and management of CE.