title: Molecular identification of Trichinella spiralis nudix hydrolase and its induced protective immunity against trichinellosis in BALB/c mice.
abstract: BACKGROUND
  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.
  
  
  METHODS
  The full-length cDNA sequence of TsNd gene encoding a 46 kDa protein from T. spiralis intestinal infective larvae (IIL) was cloned and identified. The antigenicity of rTsNd was analyzed by Western blot. Transcription and expression of TsNd at T. spiralis different stages were observed by RT-PCR and IFT. The levels of the specific total IgG, IgG1 and IgG2a antibodies to rTsNd were determined by ELISA. The immune protection of rTsNd against T. spiralis infection was investigated.
  
  
  RESULTS
  Sequence and phylogenetic analysis revealed that TsNd had a nudix motif located at 226-244aa, which had high homology and the closest evolutionary status with T. pseudospiralis. The rTsNd was obtained after expression and purification. Western blot analysis showed that anti-rTsNd serum recognized the native TsNd protein in crude antigens of muscle larvae (ML), IIL, adult worms (AW) and newborn larvae (NBL), and ES antigens of ML. Transcription and expression of TsNd gene was observed in all developmental stages of T. spiralis (ML, IIL, AW and NBL), with high level expression in IIL. An immunolocalization analysis identified TsNd in the cuticle, stichocytes and reproductive organs of the parasite. Following immunization, anti-rTsNd IgG levels were increased, and the levels of IgG1 were more significantly higher than that of IgG2a. After a challenge infection with T. spiralis, mice immunized with the rTsNd displayed a 57.7% reduction in adult worms and a 56.9% reduction in muscle larval burden.
  
  
  CONCLUSIONS
  TsNd induced a partial protective immunity in mice and could be considered as a novel candidate vaccine antigen against trichinellosis.
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou, 450052, P. R. China. longshaorong@126.com.
authors.forename: Shao Rong
authors.initials: SR
authors.lastname: Long
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou, 450052, P. R. China. wangzq@zzu.edu.cn.
authors.forename: Zhong Quan
authors.initials: ZQ
authors.lastname: Wang
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou, 450052, P. R. China. liuruodan2006@126.com.
authors.forename: Ruo Dan
authors.initials: RD
authors.lastname: Liu
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou, 450052, P. R. China. liulina5965@163.com.
authors.forename: Li Na
authors.initials: LN
authors.lastname: Liu
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou, 450052, P. R. China. lilingge1314@163.com.
authors.forename: Ling Ge
authors.initials: LG
authors.lastname: Li
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou, 450052, P. R. China. jpdaisy@126.com.
authors.forename: Peng
authors.initials: P
authors.lastname: Jiang
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou, 450052, P. R. China. zhangxi_601@163.com.
authors.forename: Xi
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou, 450052, P. R. China. zifangzhang@zzu.edu.cn.
authors.forename: Zi Fang
authors.initials: ZF
authors.lastname: Zhang
authors.affiliation: Department of Immunology, Massachusetts General Hospital and Harvard Medical School, Charlestown, MA, 02129, USA. shiha@helix.mgh.harvard.edu.
authors.forename: Hai Ning
authors.initials: HN
authors.lastname: Shi
authors.affiliation: Department of Parasitology, Medical College of Zhengzhou University, Zhengzhou, 450052, P. R. China. cuij@zzu.edu.cn.
authors.forename: Jing
authors.initials: J
authors.lastname: Cui
mesh_terms: D000818:Animals; D000909:Antibodies, Helminth / Q000097:blood*; D000947:Antigens, Helminth / Q000276:immunology*; D018509:DNA, Helminth / Q000737:chemistry / Q000235:genetics; D005260:Female; D015801:Helminth Proteins / Q000235:genetics / Q000276:immunology; D056724:Immunity, Humoral; D007114:Immunization; D007074:Immunoglobulin G / Q000097:blood; D007814:Larva; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D010802:Phylogeny; D011755:Pyrophosphatases / Q000235:genetics / Q000276:immunology*; D011994:Recombinant Proteins; D017422:Sequence Analysis, DNA; D017160:Trichinella spiralis / Q000201:enzymology / Q000235:genetics / Q000276:immunology*; D014235:Trichinellosis / Q000276:immunology* / Q000469:parasitology; D000096964:Nudix Hydrolases
chemical_list: D000909:Antibodies, Helminth; D000947:Antigens, Helminth; D018509:DNA, Helminth; D015801:Helminth Proteins; D007074:Immunoglobulin G; D011755:Pyrophosphatases; D011994:Recombinant Proteins; D000096964:Nudix Hydrolases