Genetic manipulation of probiotic bacterium Lactococcus lactis to produce OMP10 protein of Brucella abortus

authors:

avatar kazemi kazemi , avatar Abbas Doosti ORCID , * , avatar Mohammad Saeid Jami

Koomesh: Vol.25, issue 3; 279-288
published online: September 01, 2023
article type: Research Article
received: October 05, 2022
accepted: May 01, 2023

how to cite: kazemi K, Doosti A, Jami M S. Genetic manipulation of probiotic bacterium Lactococcus lactis to produce OMP10 protein of Brucella abortus. koomesh. 2023;25(3):e152826. 

Abstract

Introduction: In all species of Brucella bacteria, the Omp10 gene sequence is conserved. The high antigenic property of this gene product stimulates the host's immune system. Using the probiotic Lactococcus lactis bacterium as a live carrier, in addition to being safe, is suitable for the production of recombinant proteins and gene transfer to eukaryotic cells. Omp10 is one of the surface lipoproteins in brucella’s outer membrane with high immunogenicity which has a substantial role in up-taking nutrition, signaling, adherence, and antibiotic resistance. This protein has antigenic properties and can play an adjuvant role when attached to other antigens. Therefore, the purpose of this research is to create the probiotic bacteria Lactococcus lactis to produce the Omp10 protein of Brucella abortus. Materials and Methods: Brucella abortus Omp10 gene with Usp45 peptide signal was synthetically cloned in the pNZ8148 vector. The recombinant vector was first reproduction in Escherichia coli strain TOP10 and then purified by plasmid extraction kit. The concentration of extracted plasmid was measured by nanodrop. The recombinant pNZ8148-Usp45-Omp10 vector was then transformed into Lactococcus lactis bacteria through electroporation. Cloning and gene expression assessment and verification were done by SDS-PAGE, enzyme digestions, and RT-PCR techniques. Results: The results showed the successful expression of the target gene Omp10 in the Lactococcus lactis probiotic bacteria. The recombinant protein was well expressed in probiotic bacteria. Conclusion: The present study showed that the Omp10 protein is expressed in the probiotic bacterium Lactococcus lactis transformed by electroporation with the recombinant pNZ8148-Usp45-Omp10 vector. Lactococcus lactis transformed in the present study can be suitable as an oral subunit vaccine to prevent brucellosis in future studies.

References

  • 1.

    Mohammadi E, Golchin M. High protection of mice against Brucella abortus by oral immunization with recombinant probiotic Lactobacillus casei vector vaccine, expressing the outer membrane protein OMP19 of Brucella species. Comp Immunol Microbiol Infect Dis 2020; 70: 101470.

  • 2.

    Dadar M, Tiwari R, Sharun K, Dhama K. Importance of brucellosis control programs of livestock on the improvement of one health. Vet Q 2021; 41: 137-151.

  • 3.

    Seyedalizadeh N, Alesheikh AA, Ahmadkhani M. Spatio-statistical modeling of human brucellosis using environmental parameters: A case study of Northern Iran. Int Arch Photogram Remote Sens Spat Inf Sci 2019; 42: 969-973. (Persian).##https://doi.org/10.5194/isprs-archives-XLII-4-W18-969-2019.

  • 4.

    Rouzic N, Desmier L, Cariou ME, Gay E, Foster JT, Williamson CH, et al. First case of brucellosis caused by an amphibian-type brucella. Clin Infect Dis 2021; 72: e404-e407.

  • 5.

    Celli J. The changing nature of the Brucella-containing vacuole. Cell Microbiol 2015; 17: 951-958.

  • 6.

    Pakzad R, Pakzad I, Safiri S, Shirzadi MR, Mohammadpour M, Behroozi A, et al. Spatiotemporal analysis of brucellosis incidence in Iran from 2011 to 2014 using GIS. Int J Infect Dis 2018; 67: 129-136.

  • 7.

    Whatmore AM. Current understanding of the genetic diversity of Brucella, an expanding genus of zoonotic pathogens. Infect Genet Evol 2009; 9: 1168-1184.

  • 8.

    Al Jindan R. Scenario of pathogenesis and socioeconomic burden of human brucellosis in Saudi Arabia. Saudi J Biol Sci 2021; 28: 272-279.

  • 9.

    Whatmore AM, Davison N, Cloeckaert A, Al Dahouk S, Zygmunt MS, Brew SD, et al. Brucella papionis sp. nov., isolated from baboons (Papio spp.). Int J Syst Evol Microbiol 2014; 64: 4120-4128.

  • 10.

    Khurana SK, Sehrawat A, Tiwari R, Prasad M, Gulati B, Shabbir MZ, et al. Bovine brucellosis - a comprehensive review. Vet Q 2021; 41: 61-88.

  • 11.

    Doostdari S, Mahmoodi P, Mohammadzadeh A, Khafri A. Evaluation of humoral immune responses of sheep vaccinated with Razi institute Rev. 1 Brucellosis Vaccine in Comparison to a Spanish (CZV) Rev. 1 Vaccine. Avicenna J Clin Microbiol Infect 2019; 6: 95-99. (Persian).##https://doi.org/10.34172/ajcmi.2019.17.

  • 12.

    Jazayeri SD, Lim HX, Shameli K, Yeap SK, Poh CL. Nano and microparticles as potential oral vaccine carriers and adjuvants against infectious diseases. Front Pharmacol 2021; 12: 682286.

  • 13.

    El-Saadony MT, Alagawany M, Patra AK, Kar I, Tiwari R, Dawood MA, et al. The functionality of probiotics in aquaculture: An overview. Fish Shellfish Immunol 2021; 117: 36-52.

  • 14.

    Bajaj BK, Claes IJ, Lebeer S. Functional mechanisms of probiotics. J Microbiol Biotechnol Food Sci 2021; 2021: 321-327.

  • 15.

    Kumar M, Yadav AK, Verma V, Singh B, Mal G, Nagpal R, Hemalatha R. Bioengineered probiotics as a new hope for health and diseases: an overview of potential and prospects. Future Microbiol 2016; 11: 585-600.

  • 16.

    Song AA, In LLA, Lim SHE, Rahim RA. A review on Lactococcus lactis: from food to factory. Microb Cell Fact 2017; 16: 55.

  • 17.

    Shirdast H, Ebrahimzadeh F, Taromchi AH, Mortazavi Y, Esmaeilzadeh A, Sekhavati MH, et al. Recombinant lactococcus lactis displaying Omp31 antigen of brucella melitensis can induce an immunogenic response in BALB/c Mice. Probiotics Antimicrob Proteins 2021; 13: 80-89.

  • 18.

    Huy TX, Nguyen TT, Reyes AW, Vu SH, Min W, Lee HJ, et al. Immunization with a combination of four recombinant brucella abortus proteins Omp16, Omp19, Omp28, and L7/L12 Induces T Helper 1 immune response against Virulent B. abortus 544 Infection in BALB/c Mice. Front Vet Sci 2021; 7: 577026.

  • 19.

    Attar A, Afkhami H, Khaledi M, Sadati MS. Bactericidal activity of serum by brucella abortus RB51 outer membrane protein's combined by brucella abortus S99 lipopolysaccharide induction. PREPRINT 2021.##https://doi.org/10.21203/rs.3.rs-231416/v1.

  • 20.

    Tibor A, Decelle B, Letesson JJ. Outer membrane proteins Omp10, Omp16, and Omp19 of Brucella spp. are lipoproteins. Infect Immun 1999; 67: 4960-4962.

  • 21.

    Tibor A, Saman E, de Wergifosse P, Cloeckaert A, Limet JN, Letesson JJ. Molecular characterization, occurrence, and immunogenicity in infected sheep and cattle of two minor outer membrane proteins of Brucella abortus. Infect Immun 1996; 64: 100-107.

  • 22.

    Wallach JC, Ferrero MC, Victoria Delpino M, Fossati CA, Baldi PC. Occupational infection due to Brucella abortus S19 among workers involved in vaccine production in Argentina. Clin Microbiol Infect 2008; 14: 805-807.

  • 23.

    Miyoshi A, Bermdez-Humarn LG, Ribeiro LA, Le Loir Y, Oliveira SC, Langella P, Azevedo V. Heterologous expression of Brucella abortus GroEL heat-shock protein in Lactococcus lactis. Microb Cell Fact 2006; 5: 14.

  • 24.

    D'Souza R, Pandeya DR, Hong ST. Lactococcus lactis: an efficient Gram positive cell factory for the production and secretion of recombinant protein. Biomed Res 2012; 23: 1-7.

  • 25.

    Xin KQ, Hoshino Y, Toda Y, Igimi S, Kojima Y, Jounai N, et al. Immunogenicity and protective efficacy of orally administered recombinant Lactococcus lactis expressing surface-bound HIV Env. Blood 2003; 102: 223-228.

  • 26.

    Masjedian Jezi F, Razavi S, Mirnejad R, Zamani K. Immunogenic and protective antigens of Brucella as vaccine candidates. Comp Immunol Microbiol Infect Dis 2019; 65: 29-36.

  • 27.

    Bhattacharjee AK, Van de Verg L, Izadjoo MJ, Yuan L, Hadfield TL, Zollinger WD, Hoover DL. Protection of mice against brucellosis by intranasal immunization with Brucella melitensis lipopolysaccharide as a noncovalent complex with Neisseria meningitidis group B outer membrane protein. Infect Immun 2002; 70: 3324-3329.

  • 28.

    Liu X, Qi L, Lv J, Zhang Z, Zhou P, Ma Z, et al. The immune response to a recombinant Lactococcus lactis oral vaccine against foot-and-mouth disease virus in mice. Biotechnol Lett 2020; 42: 1907-1917.

  • 29.

    Letesson JJ, Tibor A, van Eynde G, Wansard V, Weynants V, Denoel P, Saman E. Humoral immune responses of Brucella-infected cattle, sheep, and goats to eight purified recombinant Brucella proteins in an indirect enzyme-linked immunosorbent assay. Clin Diagn Lab Immunol 1997; 4: 556-564.

  • 30.

    Tibor A, Wansard V, Bielartz V, Delrue RM, Danese I, Michel P, et al. Effect of omp10 or omp19 deletion on Brucella abortus outer membrane properties and virulence in mice. Infect Immun 2002; 70: 5540-5546.

  • 31.

    Li T, Huang M, Wang Z, Guo F, Zhang H, Chen C. Construction and characteristic analysis of Omp10 deletion mutant of Brucella abortus. Pak J Zool 2017; 49: 1809-1806.##https://doi.org/10.17582/journal.pjz/2017.49.5.1809.1816.

  • 32.

    Im YB, Park WB, Jung M, Kim S, Yoo HS. Comparative analysis of immune responses to outer membrane antigens OMP10, OMP19, and OMP28 of Brucella abortus. Jpn J Infect Dis 2018; 71: 197-204.

  • 33.

    Mahmoudi Vashian Z, Doosti A. Cloning and gene expression of ureG gene as a DNA vaccine candidate against Helicobacter pylori. J Guilan Univ Med Sci 2017; 26: 20-29. (Persian).

  • 34.

    Liu X, Qi L, Lv J, Zhang Z, Zhou P, Ma Z, et al. The immune response to a recombinant Lactococcus lactis oral vaccine against foot-and-mouth disease virus in mice. Biotechnol Lett 2020; 4: 1907-1917.

  • 35.

    Pan N, Liu B, Bao X, Zhang H, Sheng S, Liang Y, et al. Oral delivery of novel recombinant lactobacillus elicit high protection against staphylococcus aureus pulmonary and skin infections. Vaccines (Basel) 2021; 9: 984.

  • 36.

    Rezaei M, Rabbani-Khorasgani M, Zarkesh-Esfahani SH, Emamzadeh R, Abtahi H. Prediction of the Omp16 epitopes for the development of an epitope-based vaccine against brucellosis. Infect Disord Drug Targets 2019; 19: 36-45.

  • 37.

    Li R, Liu W, Yin X, Zheng F, Wang Z, Wu X, et al. Brucella spp. Omp25 promotes proteasome-mediated cGAS degradation to attenuate IFN- production. Front Microbiol 2021; 12: 702881.

  • 38.

    Hoseini Tavassol Z, Ejtahed H, Soroush A, Shahriari A, Siadat S, Hasani-Ranjbar S, et al. Clinical application of gut microbiota metabolites: A novel opportunity in personalized medicine. Koomesh 2022; 24: 197-209 (Persian).

  • 39.

    Rahmani N, Minuchehr Z, Bambai B, Sarhangi N, Allahyari Fard N. Evaluation of vitamins diversity produced by probiotics in the microbiome and introduction of selected species. Koomesh 2022; 24: 267-275. (Persian).