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Identification of the pXO1 plasmid in attenuated Bacillus anthracis vaccine strains.
Virulence 2016; 7(5):578-86V

Abstract

Anthrax toxins and capsule are the major virulence factors of Bacillus anthracis. They are encoded by genes located on the plasmids pXO1 and pXO2, respectively. The vaccine strain Pasteur II was produced from high temperature subcultures of B. anthracis, which resulted in virulence attenuation through the loss of the plasmid pXO1. However, it is unclear whether the high temperature culture completely abolishes the plasmid DNA or affects the replication of the plasmid pXO1. In this study, we tested 3 B. anthracis vaccine strains, including Pasteur II from France, Qiankefusiji II from Russia, and Rentian II from Japan, which were all generated from subcultures at high temperatures. Surprisingly, we detected the presence of pXO1 plasmid DNA using overlap PCR in all these vaccine strains. DNA sequencing analysis of overlap PCR products further confirmed the presence of pXO1. Moreover, the expression of the protective antigen (PA) encoded on pXO1 was determined by using SDS-PAGE and western blotting. In addition, we mimicked Pasteur's method and exposed the A16R vaccine strain, which lacks the pXO2 plasmid, to high temperature, and identified the pXO1 plasmid in the subcultures at high temperatures. This indicated that the high temperature treatment at 42.5°C was unable to eliminate pXO1 plasmid DNA from B. anthracis. Our results suggest that the attenuation of the Pasteur II vaccine strain is likely due to the impact of high temperature stress on plasmid replication, which in turn limits the copy number of pXO1. Our data provide new insights into the mechanisms of the remaining immunogenicity and toxicity of the vaccine strains.

Authors+Show Affiliations

a National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory for Infectious Disease Prevention and Control , Beijing , China.a National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory for Infectious Disease Prevention and Control , Beijing , China.a National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory for Infectious Disease Prevention and Control , Beijing , China.a National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory for Infectious Disease Prevention and Control , Beijing , China.a National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, State Key Laboratory for Infectious Disease Prevention and Control , Beijing , China.b Lanzhou Institute of Biological Products Co., Ltd , Lanzhou , China.b Lanzhou Institute of Biological Products Co., Ltd , Lanzhou , China.c Huadong Medical Institute of Biotechniques , Nanjing , China.

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

27029580

Citation

Liang, Xudong, et al. "Identification of the pXO1 Plasmid in Attenuated Bacillus Anthracis Vaccine Strains." Virulence, vol. 7, no. 5, 2016, pp. 578-86.
Liang X, Zhang H, Zhang E, et al. Identification of the pXO1 plasmid in attenuated Bacillus anthracis vaccine strains. Virulence. 2016;7(5):578-86.
Liang, X., Zhang, H., Zhang, E., Wei, J., Li, W., Wang, B., ... Zhu, J. (2016). Identification of the pXO1 plasmid in attenuated Bacillus anthracis vaccine strains. Virulence, 7(5), pp. 578-86. doi:10.1080/21505594.2016.1164366.
Liang X, et al. Identification of the pXO1 Plasmid in Attenuated Bacillus Anthracis Vaccine Strains. Virulence. 2016 07 3;7(5):578-86. PubMed PMID: 27029580.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Identification of the pXO1 plasmid in attenuated Bacillus anthracis vaccine strains. AU - Liang,Xudong, AU - Zhang,Huijuan, AU - Zhang,Enmin, AU - Wei,Jianchun, AU - Li,Wei, AU - Wang,Bingxiang, AU - Dong,Shulin, AU - Zhu,Jin, Y1 - 2016/03/30/ PY - 2016/4/1/entrez PY - 2016/4/1/pubmed PY - 2017/7/27/medline KW - attenuated KW - bacillus anthracis KW - pXO1 plasmid KW - vaccine strains SP - 578 EP - 86 JF - Virulence JO - Virulence VL - 7 IS - 5 N2 - Anthrax toxins and capsule are the major virulence factors of Bacillus anthracis. They are encoded by genes located on the plasmids pXO1 and pXO2, respectively. The vaccine strain Pasteur II was produced from high temperature subcultures of B. anthracis, which resulted in virulence attenuation through the loss of the plasmid pXO1. However, it is unclear whether the high temperature culture completely abolishes the plasmid DNA or affects the replication of the plasmid pXO1. In this study, we tested 3 B. anthracis vaccine strains, including Pasteur II from France, Qiankefusiji II from Russia, and Rentian II from Japan, which were all generated from subcultures at high temperatures. Surprisingly, we detected the presence of pXO1 plasmid DNA using overlap PCR in all these vaccine strains. DNA sequencing analysis of overlap PCR products further confirmed the presence of pXO1. Moreover, the expression of the protective antigen (PA) encoded on pXO1 was determined by using SDS-PAGE and western blotting. In addition, we mimicked Pasteur's method and exposed the A16R vaccine strain, which lacks the pXO2 plasmid, to high temperature, and identified the pXO1 plasmid in the subcultures at high temperatures. This indicated that the high temperature treatment at 42.5°C was unable to eliminate pXO1 plasmid DNA from B. anthracis. Our results suggest that the attenuation of the Pasteur II vaccine strain is likely due to the impact of high temperature stress on plasmid replication, which in turn limits the copy number of pXO1. Our data provide new insights into the mechanisms of the remaining immunogenicity and toxicity of the vaccine strains. SN - 2150-5608 UR - https://www.unboundmedicine.com/medline/citation/27029580/Identification_of_the_pXO1_plasmid_in_attenuated_Bacillus_anthracis_vaccine_strains_ L2 - http://www.tandfonline.com/doi/full/10.1080/21505594.2016.1164366 DB - PRIME DP - Unbound Medicine ER -