变异链球菌hit基因功能及致龋力影响的初步分析

doi:10.13701/j.cnki.kqyxyj.2023.07.006

口腔医学研究 ›› 2023, Vol. 39 ›› Issue (7): 594-601.DOI: 10.13701/j.cnki.kqyxyj.2023.07.006

变异链球菌hit基因功能及致龋力影响的初步分析

樊紫萱¹, 乔里², 赖扬帆³, 王鹏³, 蒋代仁³, 梁燕³, 叶朝阳^2*, 罗鹏^1*

1.贵州医科大学公共卫生与健康学院贵州贵阳 550025;
2.贵州医科大学附属医院临床医学研究中心贵州贵阳 550025;
3.贵州医科大学附属口腔医院牙体牙髓科贵州贵阳 550025

收稿日期:2023-01-28 发布日期:2023-07-25
通讯作者: *叶朝阳,E-mail:yezy@gmc.edu.cn; 罗鹏,E-mail:luopeng@gmc.edu.cn
作者简介:樊紫萱(1996~ ),女,天津人,硕士在读,主要从事口腔医学研究和毒理学研究。
基金资助:
贵州省科技计划项目(黔教合基础-ZK[2021]一般438)

Preliminary Analysis on Functions of hit Gene in Streptococcus Mutans and Its Impact on Cariogenicity

FAN Zixuan¹, QIAO Li², LAI Yangfan³, WANG Peng³, JIANG Dairen³, LIANG Yan³, YE Zhaoyang^2*, LUO Peng^1*

1. School of Public Health, Guizhou Medical University, Guiyang 550025, China;
2. Research Center of Clinical Medicine, Affiliated Hospital of Guizhou Medical University, Guiyang 550025, China;
3. Department of Dentistry and Endodontics, School of Stomatology Affiliated to Guizhou Medical University, Guiyang 550025, China.

Received:2023-01-28 Published:2023-07-25

摘要/Abstract

摘要： 目的:初步探讨hit基因对变异链球菌生长和致龋力的影响。方法:通过同源重组构建变异链球菌hit基因缺陷株(Δhit)、hit基因回补株(ΔhitC、ΔhitC-eGFP);观察不同pH下Smu.Δhit、Smu.ΔhitC和ATCC25175模式菌株生长曲线;通过结晶紫染色、蒽酮染色、扫描电镜等实验,比较生长、生物膜形成、粘附、菌体形态差异及耐酸特性;通过人工龋实验,在体外模拟菌株对牙釉产生龋样损坏的差异。结果:成功构建Smu.Δhit、 Sum.ΔhitC和Smu.ΔhitC-eGFP菌株;观察发现Smu.Δhit较ATCC25175菌株生长迅速,随着pH值降低,Smu.Δhit生长受到部分抑制,ATCC25175则完全被抑制;Smu.Δhit菌体呈“椭圆球状平铺附着”分散在牙釉面上,胞外生物膜量较少;Smu.Δhit造成的龋样损坏明显强于ATCC25175。结论:hit 基因调控变异链球菌生长繁殖,Smu.Δhit缺陷株生长速率显著高于亲本株,更耐受酸性环境,对人牙釉面破坏性更强。

关键词: 变异链球菌, ATCC25175, hit基因, 致龋力, hit-样蛋白

Abstract: Objective: To investigate the function of hit gene and its effect on the cariogenicity of Streptococcus mutans. Methods: LFH-PCR was employed to construct the hit-deficient mutant strain (Smu.Δhit). The hit gene complemented strain (Smu.ΔhitC) and its added eGFP tagging strain (Smu.ΔhitC-eGFP) were also constructed. The growth rate and acid resistance with different pH values, the biofilm formation, as well as the morphology of the Smu.Δhit and its parental strains were compared. In vitro detection of artificial caries-like lesions of the Smu.Δhit on human dental enamel surfaces was performed. Results: The Smu.Δhit,Smu.ΔhitC, and Smu.ΔhitC-eGFP were successfully screened out. The growth rate of the Smu.Δhit versus their parental was increased greatly in BHI medium. When subjected to modified BHI medium (pH 5.6), the growth inhibition of the Smu.Δhit was much less than that of ATCC25175. Compared with ATCC25175, scanning electron microscopy (SEM), crystalline violet staining, and anthrone-sulfuric acid test analysis showed that deficiency of hit caused alterations in morphology and decreased in biofilm formation, being devoid of extracellular glucans. Much more serious artificial caries-like lesions of the Smu.Δhit on human dental enamel surfaces was also rapidly occurred. Conclusion: The hit gene is growth-phase regulated in the cell-cycle, and has significant impact on the growth rate, biofilm formation, and acid resistance, and rapidly causes caries-like lesions on human dental enamel surfaces.

Key words: Streptococcus mutans, ATCC25175, hit gene, cariogenicity, hit-like protein

樊紫萱, 乔里, 赖扬帆, 王鹏, 蒋代仁, 梁燕, 叶朝阳, 罗鹏. 变异链球菌hit基因功能及致龋力影响的初步分析[J]. 口腔医学研究, 2023, 39(7): 594-601.

FAN Zixuan, QIAO Li, LAI Yangfan, WANG Peng, JIANG Dairen, LIANG Yan, YE Zhaoyang, LUO Peng. Preliminary Analysis on Functions of hit Gene in Streptococcus Mutans and Its Impact on Cariogenicity[J]. Journal of Oral Science Research, 2023, 39(7): 594-601.

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变异链球菌hit基因功能及致龋力影响的初步分析

Preliminary Analysis on Functions of hit Gene in Streptococcus Mutans and Its Impact on Cariogenicity

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