Identification and characterization of two novel peritrophic membrane (PM) genes in the Chinese mitten crab Eriocheir sinensis that exhibit activity against high-pH stress and Aeromonas hydrophila challenge

The peritrophic membrane (PM) is located in the gut of most insects and protects them from invasion by pathogens or toxins ingested during feeding. Peritrophins are a principal component of the PM and contribute to the structure and function of the PM. The PM has also been observed in the gastrointestinal tract of the Chinese mitten crab (Eriocheir sinensis). In this study, full-length cDNA sequences of two PM genes from E. sinensis were obtained, namely, Es-peritrophin1 and Es-peritrophin2, which were 855 bp and 1,611 bp respectively. The deduced proteins were composed of 267 and 468 amino acids and contained 3 and 1 chitin binding domains respectively. Both Es-peritrophin1 and Es-peritrophin2 contained a signal peptide sequence at the N terminus, indicating that both proteins were secretory proteins. Phylogenetic analysis revealed that Es-peritrophin1 and Es-peritrophin2 were divided into the CPAP3-B and CPAP1-H branches respectively. Nevertheless, sequence alignment demonstrated that they had lower homology to other known PMs, indicating they were two novel peritrophins. Quantitative real-time PCR (qRT-PCR) analysis showed that both the Es-peritrophin1 and Es-peritrophin2 transcript levels were high in the stomach and hindgut and lower in the midgut, indicating that the PM might be assembled locally in E. sinensis. The expression of Es-peritrophin1 and Es-peritrophin2 was downregulated by high-pH stress and Aeromonas hydrophila challenge in the examined tissues except in the hindgut, where the expression of both genes was upregulated by A. hydrophila challenge. These results indicated that Es-peritrophin1 and Es-peritrophin2 are two novel peritrophin genes related to immune defence in E. sinensis.