Chitin Synthases Are Critical for Reproduction, Molting, and Digestion in the Salmon Louse (Lepeophtheirus salmonis).

Chitin synthase (CHS) is a large transmembrane enzyme that polymerizes Uridine diphosphate N-acetylglucosamine into chitin. The genomes of insects often encode two chitin synthases, CHS1 and CHS2. Their functional roles have been investigated in several insects: CHS1 is mainly responsible for synthesizing chitin in the cuticle and CHS2 in the midgut. Lepeophtheirus salmonis is an ectoparasitic copepod on salmonid fish, which causes significant economic losses in aquaculture. In the present study, the tissue-specific localization, expression, and functional role of L. salmonis chitin synthases, LsCHS1 and LsCHS2, were investigated. The expressions of LsCHS1 and LsCHS2 were found in oocytes, ovaries, intestine, and integument. Wheat germ agglutinin (WGA) chitin staining signals were detected in ovaries, oocytes, intestine, cuticle, and intestine in adult female L. salmonis. The functional roles of the LsCHSs were investigated using RNA interference (RNAi) to silence the expression of LsCHS1 and LsCHS2. Knockdown of LsCHS1 in pre-adult I lice resulted in lethal phenotypes with cuticle deformation and deformation of ovaries and oocytes in adult lice. RNAi knockdown of LsCHS2 in adult female L. salmonis affected digestion, damaged the gut microvilli, reduced muscular tissues around the gut, and affected offspring. The results demonstrate that both LsCHS1 and LsCHS2 are important for the survival and reproduction in L. salmonis.

Effects of chitin synthesis inhibitor treatment on Lepeophtheirus salmonis (Copepoda, Caligidae) larvae.

The salmon louse (Lepeophtheirus salmonis) is an ectoparasite infecting Atlantic salmon (Salmo salar), which causes substantial problems to the salmon aquaculture and threatens wild salmon. Chitin synthesis inhibitors (CSIs) are used to control L. salmonis in aquaculture. CSIs act by interfering with chitin formation and molting. In the present study, we investigated the action of four CSIs: diflubenzuron (DFB), hexaflumuron (HX), lufenuron (LF), and teflubenzuron (TFB) on larval molt. As the mode of action of CSIs remains unknown, we selected key enzymes in chitin metabolism and investigated if CSI treatment influenced the transcriptional level of these genes. All four CSIs interfered with the nauplius II molt to copepodids in a dose-dependent manner. The EC50 values were 93.2 nM for diflubenzuron, 1.2 nM for hexaflumuron, 22.4 nM for lufenuron, and 11.7 nM for teflubenzuron. Of the investigated genes, only the transcriptional level of L. salmonis chitin synthase 1 decreased significantly in hexaflumuron and diflubenzuron-treated larvae. All the tested CSIs affected the molt of nauplius II L. salmonis larvae but at different concentrations. The larvae were most sensitive to hexaflumuron and less sensitive to diflubenzuron. None of the CSIs applied had a strong impact on the transcriptional level of chitin synthesis or chitinases genes in L. salmonis. Further research is necessary to get more knowledge of the nature of the inhibition of CSI and may require methods such as studies of protein structure and enzymological studies.

Chitin synthesis and degradation in Lepeophtheirus salmonis: Molecular characterization and gene expression profile during synthesis of a new exoskeleton.

Animals with exoskeleton need to molt to grow and develop. Molting is well described in some arthropods especially insects. Chitin is a polymer of N-acetylglucosamine, and one of the major components of the exoskeleton of arthropods. Chitin is synthesized and degraded by a series of enzymes during the molting cycle. However, the presence and function of these enzymes are largely unknown in copepods such as the ectoparasite salmon louse (Lepeophtheirus salmonis) a major pest in salmonid aquaculture. Here we describe six genes found in the L. salmonis genome (LsCHS1, LsCHS2, LsGFAT, LsGNA1, LsAGM, and LsUAP) with high homology to enzymes in the chitin synthesis pathway. The transcription profiles of these enzymes together with three chitinases enzymes (LsChi1, LsChi2, and LsChi4), which have been characterized before, were examined during the synthesis of a new exoskeleton and revealed a dynamical expression concurrent with the morphological changes during the molt cycle. Further understanding of chitin metabolism and its regulation may prove useful tool to develop new pesticides.