MOE Key Laboratory of Industrial Fermentation Microbiology, College of Biotechnology, Tianjin University of Science & Technology, Tianjin, China.
Department of Pathogen Biology, College of Basic Medical Sciences, China Medical University, Shenyang, China.
Mol Oral Microbiol. 2020 Dec;35(6):260-270. doi: 10.1111/omi.12316. Epub 2020 Oct 15.
Fluoride has been used as an effective anticaries agent for more than 70 years, which might result in the emergence of fluoride-resistant strains. However, the fluoride resistance mechanism and the cariogenic properties of fluoride-resistant mutant for cariogenic bacterial species Streptococcus mutans remain largely unknown. We describe here the construction and characterization of a mariner-based transposon system designed to be used in S. mutans, which is also potentially applicable to other streptococci. To identify genetic determinants of fluoride resistance in S. mutans, we constructed a library of S. mutans transposon insertion mutants and screened this library to identify mutants exhibiting fluoride resistance phenotype. Two mutants were found to carry transposon insertion in two different genetic loci (smu.396 and smu.1291c), respectively. Our subsequent genetic study indicates the fluoride-resistant phenotype for the mutant with the insertion in smu.1291c is resulting from the constitutive overexpression of downstream operon smu.1290c-89c, which is consistent with the previous reports. We also demonstrate for the first time that the deletion of smu.396 is responsible for the fluoride-resistant phenotype and that the combining of smu1290c-89c overexpression and smu.396 deletion in one strain could attribute an additive effect on the fluoride resistance. In addition, our results suggest that the biological fitness of those fluoride-resistant mutants is reduced compared to that of wild-type strain. Overall, our identification and characterization of genetic determinants responsible for fluoride resistance in S. mutans expand our understanding of the fluoride resistance mechanism and the biological consequence of the fluoride resistance strains.
氟化物作为一种有效的抗龋剂已经使用了 70 多年，这可能导致氟化物耐药株的出现。然而，对于致龋菌变形链球菌的氟化物耐药机制和氟化物耐药突变体的致龋特性仍知之甚少。我们在这里描述了一种基于 mariner 的转座子系统的构建和特性，该系统设计用于变形链球菌，也可能适用于其他链球菌。为了鉴定变形链球菌中氟化物耐药的遗传决定因素，我们构建了一个变形链球菌转座子插入突变体文库，并筛选该文库以鉴定表现出氟化物耐药表型的突变体。发现两个突变体分别在两个不同的遗传位点（smu.396 和 smu.1291c）携带转座子插入。我们随后的遗传研究表明，插入 smu.1291c 的突变体的氟化物耐药表型是由于下游操纵子 smu.1290c-89c 的组成型过表达所致，这与之前的报道一致。我们还首次证明 smu.396 的缺失是氟化物耐药表型的原因，并且在一个菌株中 smu1290c-89c 的过表达和 smu.396 的缺失的组合对氟化物耐药具有加性效应。此外，我们的结果表明，与野生型菌株相比，这些氟化物耐药突变体的生物适应性降低。总的来说，我们鉴定和表征了导致变形链球菌氟化物耐药的遗传决定因素，这扩展了我们对氟化物耐药机制和氟化物耐药菌株的生物学后果的理解。
