氟化物对变形链球菌耐氟菌株生物膜形成的影响

doi:10.13701/j.cnki.kqyxyj.2019.06.009

口腔医学研究 ›› 2019, Vol. 35 ›› Issue (6): 546-550.DOI: 10.13701/j.cnki.kqyxyj.2019.06.009

氟化物对变形链球菌耐氟菌株生物膜形成的影响

杨瑶瑶,张志民^*,赵洪岩^*,郭馨蔚,刘璐,凌晓旭,曲添

吉林大学口腔医院牙体牙髓病科吉林长春 130021

收稿日期:2019-01-14 出版日期:2019-06-28 发布日期:2019-06-27
通讯作者: 张志民,E-mail:zhangzm1964@sina.com
赵洪岩,E-mail:1973hero@163.com
作者简介:杨瑶瑶(1992~ ),女,硕士在读,从事龋病病因学研究。
基金资助:
吉林省科技厅自然科学基金 (编号:20160101003JC)

Effect of Fluoride on Biofilm Formation of Streptococcus Mutans Fluoride-resistant Strains

YANG Yao-yao, ZHANG Zhi-min^*, ZHAO Hong-yan^*, GUO Xin-wei, LIU Lu, LING Xiao-xu, QU Tian

Department of Endodontics, School of Stomatology, Jilin University. Changchun 130021, China.

Received:2019-01-14 Online:2019-06-28 Published:2019-06-27

摘要/Abstract

摘要： 目的:观察变形链球菌耐氟菌株体外成膜过程,并与亲代菌株进行比较;探讨氟化物对耐氟菌株生物膜形成的影响。方法:变形链球菌耐氟菌株及亲代菌株,分为亲代组(变形链球菌亲代菌株生长于无氟BHI培养基中)、耐氟组(耐氟菌株生长于无氟BHI培养基中)和含氟组(耐氟菌株生长于含氟量为1 g/L的BHI培养基中)。通过结晶紫染色、扫描电镜及激光共聚焦显微镜观察3组在6、12、18、24、36、48 h生物膜形成情况,采用SPSS 21.0单因素方差分析进行统计学比较。结果:(1)结晶紫染色法观察到生物膜生物量随时间增加,24~36 h达到最大值且相对稳定。亲代组及耐氟组生物膜生物量无统计学差异(P＞0.05);含氟组生物量在实验的6个时间点均明显少于耐氟组(P＜0.05)。(2)扫描电镜可见生物膜随时间的延长结构更加复杂,含氟组在实验时间内未见形成完整生物膜。(3)激光共聚焦实验通过细菌活死染观察到6~24 h生物膜呈绿色荧光,以活菌为主;36~48 h生物膜呈黄色荧光,以死菌为主。在不同时间段,实验组之间细菌密度及活菌比例均存在显著的统计学差异(P＜0.01)。结论:通过对变形链球菌耐氟菌株及亲代菌株体外成膜情况无明显差异;氟化物对耐氟菌株生物膜的形成有明显的抑制作用。本实验为后续药物对生物膜防治作用的研究及代谢组学的研究奠定基础。

关键词: 变形链球菌, 变形链球菌耐氟菌株, 细菌生物膜

Abstract: Objective: To observe the in vitro film formation process of Streptococcus mutans fluoride-resistant strains and compare them with parental strains; and to investigate the effect of fluoride on the biofilm formation of fluoride-resistant strains. Methods: Streptococcus mutans fluoride-resistant strain and parental strain were divided into parental group (Streptococcus mutans parental strain grown in fluorine-free BHI medium), fluoride-resistant group (fluorine-resistant strain grown in fluorine-free BHI medium), and fluorine-containing group (the fluoride-resistant strain was grown in BHI medium having a fluorine content of 1 g/L). The biofilm formation of three groups at 6h, 12h, 18h, 24h, 36h, and 48h was observed by crystal violet staining, scanning electron microscopy, and laser confocal microscopy. The statistical analysis was performed by SPSS 21.0 one-way analysis of variance. Results: (1) The biofilm biomass was observed to increase with time by crystal violet staining, which reached the maximum at 24-36 h and was relatively stable. There was no significant difference in biofilm biomass between the parental group and the fluoride-tolerant group (P>0.05). The biomass of the fluoride group was significantly less than the fluoride-resistant group at the six time points (P<0.05). (2) Scanning electron microscopy showed that the structure of biofilm became more complicated with the increase of time, and the fluorine-containing group did not form a complete biofilm during the experimental time. (3) The laser confocal experiment showed that the biofilm showed green fluorescence at 6h-24h through live bacterial death, and the biofilm was mainly composed of 36h-48h biofilm, which was mainly yellow. There were significant statistical differences in bacterial density and viable ratio between the experimental groups at different time points (P<0.01). Conclusion: There is no significant difference in the in vitro formation of fluoride-resistant strains and parental strains of Streptococcus mutans. Fluoride has a significant inhibitory effect on the formation of biofilms resistant to fluoride. This experiment lays the foundation for the research of the follow-up drugs on the prevention and treatment of biofilm and the study of metabolomics.

Key words: Streptococcus mutans, Streptococcus mutans fluoride-resistant strain, Bacterial biofilm

杨瑶瑶,张志民,赵洪岩,郭馨蔚,刘璐,凌晓旭,曲添. 氟化物对变形链球菌耐氟菌株生物膜形成的影响[J]. 口腔医学研究, 2019, 35(6): 546-550.

YANG Yao-yao, ZHANG Zhi-min, ZHAO Hong-yan, GUO Xin-wei, LIU Lu, LING Xiao-xu, QU Tian. Effect of Fluoride on Biofilm Formation of Streptococcus Mutans Fluoride-resistant Strains[J]. Journal of Oral Science Research, 2019, 35(6): 546-550.

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氟化物对变形链球菌耐氟菌株生物膜形成的影响

Effect of Fluoride on Biofilm Formation of Streptococcus Mutans Fluoride-resistant Strains

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