htrA、clpP基因对变异链球菌生长和耐酸性影响
发布时间：2018-08-13 18:39
【摘要】：目的研究htrA. clpP基因缺失对变异链球菌生长及耐酸性影响。 方法利用变异链球菌标准株及前期实验成功构建的htrA单基因缺陷株、clpP单基因缺陷株及htrA-clpP双基因缺陷株,测定4种菌株在正常培养条件下不同时间段的吸光度,采用MTT试验测定4种菌株在不同时间段的甲躜吸光度变化情况并探究其与平板活菌计数法所得的活菌量之间关系；将4种菌株培养至对数中期,分成两组,一组作为实验组进行5小时预酸化处理(pH=5.5),一组作为对照组在普通TPY培养液中进行5小时处理,然后将两组菌株在致死性酸环境(pH=3.0)处理3小时,每隔1小时取菌液采用平板活菌计数法测定活菌数,计算得到生存率,统计学分析。 结果1.在正常营养条件下,标准株的生长快于htrA、htrA-clpP缺陷株(P0.05),除3-4小时外标准株与clpP缺陷株生长无明显差异(P0.05),clpP缺陷株生长快于其他两种缺陷株(P0.05),但htrA缺陷株与htrA-clpP缺陷株生长无明显差异(P0.05)；2.未经预酸化理4种菌株的生存率中,3小时内,标准株、htrA单基因缺陷株和clpP单基因缺陷株的生存率并无明显差异(P0.05),而htrA-clpP双基因缺陷株变化明显,差异有统计学意义(P0.05),预酸化处理后,与标准株相比,htrA单基因缺陷株、clpP单基因缺陷株及htrA-clpP双基因缺陷株的生存率都有下降(P0.05)；3种缺陷株相比生存率差异没有明显差异(P0.05),预酸化处理的标准株和htrA单基因缺陷株、htrA-clpP双基因缺陷株生存率均明显高于未经预酸化处理(P0.05),预酸化处理对生存率的影响大于两种基因缺失(P0.05)。 结论1.本实验中htrA、clpP缺失均可抑制变异链球菌的生长,并且双基因缺陷对细菌生长抑制作用与htrA单独缺失相似,MTT试验可在一定的菌数范围内反映标准株与三种缺陷株的活菌数；2.与变异链球菌标准株相比,htrA、clpP基因单独或同时缺失都使菌株耐酸能力有不同程度的下降。
[Abstract]:Objective to study the effect of htrA. clpP gene deletion on the growth and acid tolerance of Streptococcus mutans. Methods using the standard strain of Streptococcus mutans and the successful construction of htrA single gene deficiency strain and htrA-clpP double gene deficiency strain, the absorbance of four strains were measured under normal culture conditions. MTT test was used to determine the absorbance changes of the four strains in different time periods and to explore the relationship between them and the live bacteria quantity obtained by the method of counting live bacteria on a flat plate, and the four strains were cultured to the middle of logarithmic period and divided into two groups. One group was treated with 5-hour preacidification (pH=5.5) as experimental group, the other group was treated in normal TPY culture medium for 5 hours as control group, then the two groups were treated in lethal acid environment (pH=3.0) for 3 hours. The number of viable bacteria was determined by plate counting method every 1 hour. The survival rate was calculated and analyzed statistically. Result 1. Under normal nutrition condition, the growth of the standard strain was faster than that of the htrAgna htrA-clpP deficiency strain (P0.05). There was no significant difference between the standard strain and the clpP deficient strain except 3-4 hours (P0.05), but there was no significant difference between the htrA deficient strain and the htrA-clpP deficient strain (P0.05). 2. The survival rate of the four strains without preacidification was not significantly different from that of the standard strain within 3 hours (P0.05), but that of the htrA-clpP double gene deficient strain was significantly different from that of the standard strain (P0.05), while the survival rate of the standard strain was not significantly different from that of the htrA-clpP strain (P0.05). The difference was statistically significant (P0.05). Compared with the standard strain, the survival rate of clpP single gene deficient strain and htrA-clpP double gene deficient strain decreased after preacidification (P0.05). There was no significant difference in survival rate among the three defective strains (P0.05). The survival rates of standard strains and htrA single-gene deficient strains were significantly higher than those of non-preacidified strains (P0.05). The effect of preacidification on survival rate was significant. The noise was greater than that of two gene deletions (P0.05). Conclusion 1. In this experiment, the growth of Streptococcus mutans could be inhibited by the deletion of htrAfion clpP, and the inhibitory effect of double gene deficiency on bacteria growth was similar to that of htrA alone. MTT test could reflect the number of live bacteria of standard strain and three defective strains in a certain range. Compared with the standard strain of Streptococcus mutans, the deletion of htrAgna clpP gene alone or at the same time decreased the acid-tolerance ability of the strain to varying degrees.
【学位授予单位】：天津医科大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R378
本文编号：2181841
[Abstract]:Objective to study the effect of htrA. clpP gene deletion on the growth and acid tolerance of Streptococcus mutans. Methods using the standard strain of Streptococcus mutans and the successful construction of htrA single gene deficiency strain and htrA-clpP double gene deficiency strain, the absorbance of four strains were measured under normal culture conditions. MTT test was used to determine the absorbance changes of the four strains in different time periods and to explore the relationship between them and the live bacteria quantity obtained by the method of counting live bacteria on a flat plate, and the four strains were cultured to the middle of logarithmic period and divided into two groups. One group was treated with 5-hour preacidification (pH=5.5) as experimental group, the other group was treated in normal TPY culture medium for 5 hours as control group, then the two groups were treated in lethal acid environment (pH=3.0) for 3 hours. The number of viable bacteria was determined by plate counting method every 1 hour. The survival rate was calculated and analyzed statistically. Result 1. Under normal nutrition condition, the growth of the standard strain was faster than that of the htrAgna htrA-clpP deficiency strain (P0.05). There was no significant difference between the standard strain and the clpP deficient strain except 3-4 hours (P0.05), but there was no significant difference between the htrA deficient strain and the htrA-clpP deficient strain (P0.05). 2. The survival rate of the four strains without preacidification was not significantly different from that of the standard strain within 3 hours (P0.05), but that of the htrA-clpP double gene deficient strain was significantly different from that of the standard strain (P0.05), while the survival rate of the standard strain was not significantly different from that of the htrA-clpP strain (P0.05). The difference was statistically significant (P0.05). Compared with the standard strain, the survival rate of clpP single gene deficient strain and htrA-clpP double gene deficient strain decreased after preacidification (P0.05). There was no significant difference in survival rate among the three defective strains (P0.05). The survival rates of standard strains and htrA single-gene deficient strains were significantly higher than those of non-preacidified strains (P0.05). The effect of preacidification on survival rate was significant. The noise was greater than that of two gene deletions (P0.05). Conclusion 1. In this experiment, the growth of Streptococcus mutans could be inhibited by the deletion of htrAfion clpP, and the inhibitory effect of double gene deficiency on bacteria growth was similar to that of htrA alone. MTT test could reflect the number of live bacteria of standard strain and three defective strains in a certain range. Compared with the standard strain of Streptococcus mutans, the deletion of htrAgna clpP gene alone or at the same time decreased the acid-tolerance ability of the strain to varying degrees.
【学位授予单位】：天津医科大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R378
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