Abstract
Although it has been known for many years that T. spiralis muscle larvae (ML) can not invade intestinal epithelial cells unless they are exposed to the intestinal milieu and activated into intestinal infective larvae (IIL), which genes in IIL are involved in the process of invasion is still unknown. In this study, suppression subtractive hybridization (SSH) was performed to identify differentially expressed genes between IIL and ML. SSH library was constructed using cDNA generated from IIL as the ‘tester’. About 110 positive clones were randomly selected from the library and sequenced, of which 33 T. spiralis genes were identified. Thirty encoded proteins were annotated according to Gene Ontology Annotation in terms of molecular function, biological process, and cellular localization. Out of 30 annotated proteins, 16 proteins (53.3%) had binding activity and 12 proteins (40.0%) had catalytic activity. The results of real-time PCR showed that the expression of nine genes (Ts7, Ndr family protein; Ts8, serine/threonine-protein kinase polo; Ts11, proteasome subunit beta type-7; Ts17, nudix hydrolase; Ts19, ovochymase-1; Ts22, fibronectin type III domain protein; Ts23, muscle cell intermediate filament protein OV71; Ts26, neutral and basic amino acid transport protein rBAT and Ts33, FACT complex subunit SPT16) from 33 T. spiralis genes in IIL were up-regulated compared with that of ML. The present study provide a group of the potential invasion-related candidate genes and will be helpful for further studies of mechanisms by which T. spiralis infective larvae recognize and invade the intestinal epithelial cells.
Highlights
Trichinella spiralis is a parasitic nematode that infects their vertebrate host by the consumption of raw or undercooked meat from infected animals [1,2]
The activation of the larvae by intestinal content or bile is one of the most pivotal requirements for the larval invasion of intestinal epithelial cells (IECs), but why is the procedure necessary before the invasion of IECs? What is the difference in gene expression between muscle larvae (ML) and infective larvae (IIL)? Which genes are differentially expressed significantly in T. spiralis larvae during the process of their activation and interaction with IECs? Among the proteins encoded by these up-regulated genes, which are related to the invasion by the parasite? These key questions related with the mechanism of the Trichinella invasion of host enterocytes are unknown
In subtracted cDNAs, 18S rRNA products were observed at 28 cycles, while the amplified products were seen at 18 cycles in the unsubtracted cDNAs
Summary
Trichinella spiralis is a parasitic nematode that infects their vertebrate host by the consumption of raw or undercooked meat from infected animals (e.g. pigs, wild animals) [1,2] Following their release in the stomach by digestion of meat, T. spiralis muscle larvae (ML) are activated by intestinal content or bile after 0.9 hour post-infection (hpi), and interacted with host intestinal epithelial cells (IECs). These activated larvae in intestine are named as ‘‘intestinal infective larvae (abbreviated as IIL)’’ [3]. The activation of the larvae by intestinal content or bile is one of the most pivotal requirements for the larval invasion of IECs, but why is the procedure necessary before the invasion of IECs? What is the difference in gene expression between ML and IIL? Which genes are differentially expressed significantly in T. spiralis larvae during the process of their activation and interaction with IECs? Among the proteins encoded by these up-regulated genes, which are related to the invasion by the parasite? These key questions related with the mechanism of the Trichinella invasion of host enterocytes are unknown
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