Silencing TUBB3 Expression Destroys the Tegument and Flame Cells of Echinococcus multilocularis Protoscoleces

SIMPLE SUMMARY: Echinococcus multilocularis is a fox and carnivore tapeworm with important zoonotic potential, as the larval stage can cause alveolar echinococcosis in both humans and livestock. An understanding of the molecular mechanisms of parasite growth and development is essential for disease diagnosis, management and control. This study investigates the potential role of E. multilocularis TUBB3 in protoscoleces through RNA interference in vitro with specific short interfering RNAs. Our results show that EmTUBB3 might contribute to tegument formation and protonephridial system flame cell integrity in E. multilocularis protoscoleces. ABSTRACT: Alveolar echinococcosis (AE), caused by infection with the larvae of Echinococcus multilocularis, is a neglected tropical disease and zoonosis that causes remarkable morbidity in humans and has economic importance in the livestock industry worldwide. The growth of this parasite resembles the invasion and proliferation of malignant tumours. Microtubules, especially the β-tubulin subunit in the exposed end, are the targets of many antitumour drugs. However, the role of TUBB3, which is the most studied isotype in solid tumours and is also a marker of biological aggressiveness associated with the modulation of tumour metastatic abilities in the growth and development of platyhelminths, is unknown. In this study, protoscoleces (PSCs) are cultivated in monophasic medium in vitro. Using electroporated short interfering RNA (siRNA), EmTUBB3 knockdown was performed with two EmTUBB3-specific siRNAs (siRNA-1 and siRNA-2). qRT–PCR was performed to detect the expression of TUBB3. PSCs viability and the evagination rate and number of body contractions were quantified under a light microscope. Scanning electron microscopy (SEM) and transmission electron microscopy (TEM) were used to observe the ultra-morphological changes of the parasites. After siRNA interference, the EmTUBB3 expression in E. multilocularis PSCs was significantly reduced. Reduced viability, a decreased evagination rate and a decreased number of body contractions were also documented. In particular, shrinkage and roughness of the tegument were observed. Ultrastructural changes included marked damage to flame cells, cracked cilia structures enclosed in the cell body and ruptured microtubule structures. EmTUBB3 possibly plays a crucial role in tegument and flame cell integrity in E. multilocularis PSCs. Novel drugs targeting this specific beta-tubulin isotype in E. multilocularis are potential methods for disease control and deserve further attention.