title: Cloning, expression and characterization of a Trichinella spiralis serine protease gene encoding a 35.5 kDa protein.
abstract: Serine proteases are found in the excretory-secretory (ES) products from Trichinella spiralis muscle larvae, have collagenolytic and elastolytic activities, and may be related to the larval invasion of intestinal epithelial cells. In this study, the serine protease gene (TspSP-1.2, GenBank accession No. EU302800) encoding a 35.5 kDa protein from T. spiralis was cloned, and recombinant TspSP-1.2 protein was produced in an Escherichia coli expression system. An anti-TspSP-1.2 serum recognized the native protein migrating at 35.5 kDa by the Western blotting of the crude or ES antigens from muscle larvae at 42 days post infection. An immunolocalization analysis identified TspSP-1.2 in the cuticle and internal organs of the parasite. Transcription and expression of the TspSP-1.2 gene was observed at all developmental stages of T. spiralis (adult worms, newborn larvae, pre-encapsulated larvae and muscle larvae). An in vitro invasion assay showed that, when anti-TspSP-1.2 serum, serum of infected mice and normal mouse serum were added to the medium, the invasion rate of the infective larvae in an HCT-8 cell monolayer was 33.0%, 89.4%, and 96.2%, respectively (P<0.05), indicating that the anti-TspSP-1.2 serum partially prevented the larval invasion of intestinal epithelial cells. After a challenge infection with T. spiralis infective larvae, mice immunized with the recombinant TspSP-1.2 protein displayed a 34.92% reduction in adult worm burden and 52.24% reduction in muscle larval burden. The results showed that the recombinant TspSP-1.2 protein induced a partial protective immunity in mice and could be considered as a potential vaccine candidate against T. spiralis infection.
authors.affiliation: Department of Parasitology, Medical College, Zhengzhou University, 40 Daxue Road, Zhengzhou 450052, PR China.
authors.forename: Bin
authors.initials: B
authors.lastname: Wang
authors.forename: Zhong Quan
authors.initials: ZQ
authors.lastname: Wang
authors.forename: Jing
authors.initials: J
authors.lastname: Jin
authors.forename: Hui Jun
authors.initials: HJ
authors.lastname: Ren
authors.forename: Li Na
authors.initials: LN
authors.lastname: Liu
authors.forename: Jing
authors.initials: J
authors.lastname: Cui
mesh_terms: D000818:Animals; D000909:Antibodies, Helminth / Q000096:biosynthesis / Q000097:blood; D000947:Antigens, Helminth / Q000276:immunology; D015153:Blotting, Western; D045744:Cell Line, Tumor; D003001:Cloning, Molecular; D005260:Female; D005455:Fluorescent Antibody Technique; D015971:Gene Expression Regulation, Enzymologic; D015801:Helminth Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D006801:Humans; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D011897:Random Allocation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D057057:Serine Proteases / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D013047:Specific Pathogen-Free Organisms; D013552:Swine; D017160:Trichinella spiralis / Q000201:enzymology* / Q000235:genetics* / Q000276:immunology; D014235:Trichinellosis / Q000276:immunology / Q000517:prevention & control
chemical_list: D000909:Antibodies, Helminth; D000947:Antigens, Helminth; D015801:Helminth Proteins; D011994:Recombinant Proteins; D057057:Serine Proteases