Trichinella spiralis muscle larval (ML) excretion/secretion (ES) antigen is the most widely used diagnostic antigen of trichinellosis, but preparation of ES antigen requires collecting worms from infected animals, and detection of specific IgG against ML ES antigen may result in a false negative at the early stage of infection. The aim of the study was to characterize T. spiralis elastase-1 (TsEla) and to evaluate its potential as diagnostic antigen for trichinellosis.

We have previously reported that a 31 kDa protein was screened from the excretory-secretory (ES) proteins of Tichinella spiralis muscle larvae (ML) by immunoproteomics using early infection sera, and the gene encoding a 31 kDa protein from T. spiralis was cloned and expressed in an E. coli expression system. In this study, the recombinant 31 kDa antigens were used for detection of anti-Trichinella antibodies in serum of experimentally infected mice by ELISA.

The most commonly used serodiagnostic antigens for trichinellosis are the excretory-secretory (ES) antigens from T. spiralis muscle larvae (ML), but the specific antibodies against the ML ES antigens are usually negative during early stage of Trichinella infection. The recent studies demonstrated that T. spiralis adult worm (AW) antigens were recognized by mouse or swine infection sera on Western blot as early as 7-15 days post-infection (dpi), the AW antigens might contain the early diagnostic markers for trichinellosis. The purpose of this study was to screen early diagnostic antigens in T. spiralis AW ES proteins recognized by sera of early patients with trichinellosis. T. spiralis AW were collected at 72 h post-infection (hpi), and their ES antigens were analyzed by SDS-PAGE and Western blot. Our results showed that 5 protein bands (55, 48-50, 45, 44, and 36 kDa) were recognized by sera of early patients with trichinellosis collected at 19 dpi, and were subjected to shotgun LC-MS/MS and bioinformatics analyses. A total of 185 proteins were identified from T. spiralis protein database, of which 116 (67.2%) proteins had molecular weights of 30∼60 kDa, and 125 (67.6%) proteins with pI 4-7. Bioinformatic analyses showed that the identified proteins have a wide diversity of biological functions (binding of nucleotides, proteins, ions, carbohydrates, and lipids; hydrolase, transferase, and oxidoreductase, etc.). Several enzymes (e.g., adult-specific DNase II, serine protease and serine protease inhibitor) could be the invasion-related proteins and early diagnostic markers for trichinellosis. Moreover, recombinant T. spiralis serine protease (rTsSP-ZH68) was expressed in E. coli and its antigenicity was analyzed by Western blot with the early infection sera. The rTsSP-ZH68 was recognized by sera of infected mice at 8-10 dpi and sera of early patients with trichinellosis at 19 dpi. T. spiralis AW proteins identified in this study, especially serine protease, are the promising early diagnostic antigens and vaccine candidates for trichinellosis.

Nudix hydrolases (Nd) is a widespread superfamily, which is found in all classes of organism, hydrolyse a wide range of organic pyrophosphates and has a 'housecleaning' function. The previous study showed that Trichinella spiralis Nd (TsNd) bound to intestinal epithelial cells (IECs), and the vaccination of mice with T7 phage-displayed TsNd polypeptides produced protective immunity. The aim of this study was to clone, express and identify the full-length TsNd and to investigate its immune protection against T. spiralis infection.

Trichinellosis is a serious zoonositc parasitosis worldwide. Because its clinical manifestations aren't specific, the diagnosis of trichinellosis is not easy to be made. Trichinella spiralis muscle larva (ML) excretory-secretory (ES) antigens are the most widely applied diagnostic antigens for human trichinellosis, but the major drawback of the ES antigens for assaying anti-Trichinella antibodies is the false negative in the early Trichinella infection period. The aim of this study was to characterize the T. spiralis putative serine protease (TsSP) and to investigate its potential use for diagnosis of trichinellosis.

Trichinellosis is a serious food-borne zoonotic parasitic disease with global distribution, causing serious harm to public health and food safety. Molting is prerequisite for intestinal larval development in the life cycle of T. spiralis. Metalloproteinases play an important role in the molting process of T. spiralis intestinal infective larvae (IIL). In this study, the metalloproteinase Tsdpy31 was cloned, expressed and characterized. The results revealed that the Tsdpy31 was expressed at various T. spiralis stages and it was principally located in cuticle, hypodermis and embryos of the nematode. Recombinant Tsdpy31 (rTsdpy31) had the catalytic activity of natural metalloproteinase. Silencing of Tsdpy31 increased the permeability of larval new cuticle. When the mice were orally challenged with dsRNA treated- muscle larvae, the burden of intestinal adult and muscle larvae in Tsdpy31 dsRNA treatment group was significantly reduced, compared with the control green fluorescent protein (GFP) dsRNA and PBS groups (P < 0.05). Tsdpy31 may play a major role in the new cuticle synthesis and old cuticle shedding. Tsdpy31 also participates in T. spiralis embryonic development. We conclude that Tsdpy31 could be a candidate vaccine target molecule against intestinal T. spiralis ecdysis and development.

In China, the nematode Trichinella spiralis is the main aetiological agent of human trichinellosis. We performed multi-locus microsatellite typing of T. spiralis isolates to improve the current knowledge of the evolution and population diversity. First, seven polymorphic microsatellite loci were used to infer the genetic diversity of T. spiralis collected in 10 endemic regions. Then, a Bayesian model-based STRUCTURE analysis, a clustering based on the neighbor-joining method, and a principal coordinate analysis (PCA) were performed to identify the genetic structure. Finally, the phylogenetic position of Chinese isolates was explored based on six mitochondrial and nuclear genetic markers (cox1, cytb, 5S ISR, ESV, ITS1, and 18S rDNA) using the maximum likelihood and Bayesian methods. In addition, the divergence time was estimated with multiple genes using an uncorrelated log-normal relaxed molecular-clock model. A total of 16 alleles were detected in 2,310 individuals (1,650 muscle larvae and 660 adult worms) using seven loci. The STRUCTURE analysis indicated that the T. spiralis isolates could be organized and derived from the admixture of two ancestral clusters, which was also substantiated through the clustering analysis based on the allelic data. PCA separated most samples from Tiandong, Guangxi (GX-td), and Linzhi, Tibet (Tibet-lz), from the remaining isolates. However, both maximum likelihood and Bayesian inference supported the close relationship between Xiangfan, Hubei (HB-xf), and GX-td. The molecular dating analysis suggested that the Chinese isolates started to diverge during the Late Pleistocene (0.69 Mya). Generally, T. spiralis was observed to harbor low genetic variation, and further investigation with deeper sampling is needed to elucidate the population structure.