Bioinformatic Prediction and Production of Four Recombinant Proteins from Different Developmental Stages of and Testing of Their Diagnostic Sensitivity in Mice

Overview

Journal Iran J Parasitol

Publisher Tehran University of Medical Sciences

Specialty Parasitology

Date 2021 Mar 31

PMID 33786054

Authors

Cheng-Cheng Zhai

Xiao-Lei Liu

Xue Bai

Ze-Jun Jia

Shao-Hong Chen

Li-Guang Tian

Lin Ai

Bin Tang

Ming-Yuan Liu

Xiu-Ping Wu

Jia-Xu Chen

Affiliations

Soon will be listed here.

Abstract

Background: Trichinellosis is a serious food-borne parasitic zoonosis, thus finding high quality antigens is the key to serodiagnosis of trichinosis. This article reports the characterization and sensitivity of four recombinant proteins expressed by four genes (, , , and ) from different developmental stages of for the diagnosis of trichinellosis in mice.

Methods: This study was conducted in Jilin University and National Institute of Parasitic Diseases of Chinese Center for Disease Control and Prevention in 2017-2018. The structures and functions of the proteins encoded by four genes were predicted by bioinformatics analysis. The four genes were cloned and expressed, and the recombinant proteins were purified. Anti- IgM and IgG antibodies in the sera of mice infected with from 1-45 d post-infection (dpi) were evaluated by ELISA.

Results: The optimal antigen epitopes of four proteins (P1, P2, P3, and P4) encoded by the four genes from T- and B-cells were predicted, and four purified recombinant proteins (r-P1, r-P2, r-P3, and r-P4) were successfully produced. For IgM, the antibody levels detected by the four recombinant antigens were approximately equal to the cut-off value. Anti- IgG antibodies were first detected by r-P1 at 8 dpi, followed by r-P2, r-P3, and r-P4 at 10 dpi, 14 dpi, and 16 dpi, respectively, and the antibody levels remained high until 45 dpi.

Conclusion: The recombinant antigens r-P1, r-P2, r-P3, and r-P4 could be antigens that react with antibodies, they showed high sensitivity in the detection of anti- IgG antibodies in mice. Among these proteins, r-P1 may be a candidate antigen for the detection of anti- IgG antibodies in the early infection phase and exhibited the best sensitivity among the antigens.

References

Mayer-Scholl A, Pozio E, Gayda J, Thaben N, Bahn P, Nockler K . Magnetic Stirrer Method for the Detection of Trichinella Larvae in Muscle Samples. J Vis Exp. 2017; (121). PMC: 5409328. DOI: 10.3791/55354. View

Feidas H, Kouam M, Kantzoura V, Theodoropoulos G . Global geographic distribution of Trichinella species and genotypes. Infect Genet Evol. 2014; 26:255-66. DOI: 10.1016/j.meegid.2014.06.009. View

Wang Z, Shi Y, Liu R, Jiang P, Guan Y, Chen Y . New insights on serodiagnosis of trichinellosis during window period: early diagnostic antigens from Trichinella spiralis intestinal worms. Infect Dis Poverty. 2017; 6(1):41. PMC: 5319148. DOI: 10.1186/s40249-017-0252-z. View

Liu M, Wang X, Fu B, Li C, Wu X, Le Rhun D . Identification of stage-specifically expressed genes of Trichinella spiralis by suppression subtractive hybridization. Parasitology. 2007; 134(Pt 10):1443-55. DOI: 10.1017/S0031182007002855. View

Sun G, Liu R, Wang Z, Jiang P, Wang L, Liu X . New diagnostic antigens for early trichinellosis: the excretory-secretory antigens of Trichinella spiralis intestinal infective larvae. Parasitol Res. 2015; 114(12):4637-44. DOI: 10.1007/s00436-015-4709-3. View

Zocevic A, Lacour S, Mace P, Giovani B, Grasset-Chevillot A, Vallee I . Primary characterization and assessment of a T. spiralis antigen for the detection of Trichinella infection in pigs. Vet Parasitol. 2014; 205(3-4):558-67. DOI: 10.1016/j.vetpar.2014.09.004. View

Gottstein B, Pozio E, Nockler K . Epidemiology, diagnosis, treatment, and control of trichinellosis. Clin Microbiol Rev. 2009; 22(1):127-45, Table of Contents. PMC: 2620635. DOI: 10.1128/CMR.00026-08. View

Murrell K, Pozio E . Worldwide occurrence and impact of human trichinellosis, 1986-2009. Emerg Infect Dis. 2011; 17(12):2194-202. PMC: 3311199. DOI: 10.3201/eid1712.110896. View

Bien J, Cabaj W, Moskwa B . Proteomic analysis of potential immunoreactive proteins from muscle larvae and adult worms of Trichinella spiralis in experimentally infected pigs. Folia Parasitol (Praha). 2015; 62. DOI: 10.14411/fp.2015.022. View

10.

Wang L, Wang X, Bi K, Sun X, Yang J, Gu Y . Oral Vaccination with Attenuated Salmonella typhimurium-Delivered TsPmy DNA Vaccine Elicits Protective Immunity against Trichinella spiralis in BALB/c Mice. PLoS Negl Trop Dis. 2016; 10(9):e0004952. PMC: 5010209. DOI: 10.1371/journal.pntd.0004952. View

11.

Gomez-Morales M, Ludovisi A, Amati M, Blaga R, Zivojinovic M, Ribicich M . A distinctive Western blot pattern to recognize Trichinella infections in humans and pigs. Int J Parasitol. 2012; 42(11):1017-23. DOI: 10.1016/j.ijpara.2012.08.003. View

12.

Shimoni Z, Froom P . Uncertainties in diagnosis, treatment and prevention of trichinellosis. Expert Rev Anti Infect Ther. 2015; 13(10):1279-88. DOI: 10.1586/14787210.2015.1075394. View

13.

Wang L, Cui J, Hu D, Liu R, Wang Z . Identification of early diagnostic antigens from major excretory-secretory proteins of Trichinella spiralis muscle larvae using immunoproteomics. Parasit Vectors. 2014; 7:40. PMC: 3905669. DOI: 10.1186/1756-3305-7-40. View

14.

Mitreva M, Jasmer D, Zarlenga D, Wang Z, Abubucker S, Martin J . The draft genome of the parasitic nematode Trichinella spiralis. Nat Genet. 2011; 43(3):228-35. PMC: 3057868. DOI: 10.1038/ng.769. View

15.

Yang Y, Cai Y, Tong M, Sun N, Xuan Y, Kang Y . Serological tools for detection of infection in animals and humans. One Health. 2017; 2:25-30. PMC: 5462648. DOI: 10.1016/j.onehlt.2015.11.005. View

16.

Wu X, Fu B, Wang X, Yu L, Yu S, Deng H . Identification of antigenic genes in Trichinella spiralis by immunoscreening of cDNA libraries. Vet Parasitol. 2008; 159(3-4):272-5. DOI: 10.1016/j.vetpar.2008.10.035. View

17.

Xu J, Bai X, Wang L, Shi H, van der Giessen J, Boireau P . Immune responses in mice vaccinated with a DNA vaccine expressing serine protease-like protein from the new-born larval stage of Trichinella spiralis. Parasitology. 2017; 144(6):712-719. PMC: 5426336. DOI: 10.1017/S0031182016002493. View

18.

Rostami A, Gamble H, Dupouy-Camet J, Khazan H, Bruschi F . Meat sources of infection for outbreaks of human trichinellosis. Food Microbiol. 2017; 64:65-71. DOI: 10.1016/j.fm.2016.12.012. View

19.

Yera H, Andiva S, Perret C, Limonne D, Boireau P, Dupouy-Camet J . Development and evaluation of a Western blot kit for diagnosis of human trichinellosis. Clin Diagn Lab Immunol. 2003; 10(5):793-6. PMC: 193884. DOI: 10.1128/cdli.10.5.793-796.2003. View

20.

Yang Y, Wen Y, Cai Y, Vallee I, Boireau P, Liu M . Serine proteases of parasitic helminths. Korean J Parasitol. 2015; 53(1):1-11. PMC: 4384789. DOI: 10.3347/kjp.2015.53.1.1. View