支原体菌株低温保存活力稳定性研究
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摘要
目的研究低温冻存时间及温度对支原体菌种保存稳定性和菌种活力的影响。方法用固体平板计数法计算不同冻存时间、温度下的口腔支原体、肺炎支原体菌落数差异,分析这2个因素对口腔支原体、肺炎支原体菌种活力的影响。结果口腔支原体在生长48 h后达到生长高峰,肺炎支原体生长较为缓慢,在103 h到达生长高峰。口腔支原体在-70℃冻存1个月时,菌落数由10~(12) cfu/mL下降至10~(11) cfu/mL,冻存1个月的菌落数和冻存3个月、12个月的菌落数比较,差异均无统计学意义(P>0.05)。肺炎支原体在-70℃的冻存条件下,菌落数一直处于稳定水平;比较-35℃与-70℃的冻存条件下肺炎支原体的菌落数,差异无统计学意义(P>0.05),口腔支原体在-70℃冻存条件下的菌种活力明显高于-35℃(P<0.05)。结论口腔支原体生长迅速,但对于储存温度的敏感性较高,肺炎支原体相较口腔支原体生长缓慢,停滞期较长,菌株对冻存温度和时间的敏感度较低,稳定性高。菌种冻存温度应选择较低温度,冻存周期在1年内较为稳定。
Objective To investigate the effects of cryopreservation time and temperature on the stability and activities of some Mycoplasma strains. Methods The differences of CFUs(colony-forming units) of Mycoplasma pneumonia and Mycoplasma orale were calculated under different freezing time and temperature by a solid plate counting method. Results Mycoplasma orale reached the growth peak after 48 hours in culture, and the growth peak of Mycoplasma pneumonia reached in slower than Mycoplasma orale, with 103 hours in culture. After frozen for one month in -70 ℃, the counting of Mycoplasma orale decreased from 10~(12) cfu/mL to 10~(11) cfu/mL, but there was no a statistical significance(P>0.05) in stored for one month with for three and 12 months, respectively. The counting of Mycoplasma pneumoniae matained in a stable level in -70 ℃, and no a statistical significance under the -35 ℃ and-70 ℃ condition(P>0.05), but the activty of Mycoplasma orale strain in -70 ℃ was significantly higher than that in -35 ℃(P<0.05). Conclusion The growth trend of Mycoplasma orale was rapid, but the sensitivity to storage temperature was higher than that of Mycoplasma pneumoniae. Compared with Mycoplasma orale, Mycoplasma pneumoniae grew slowly and stagnated for a longer time. The freezing temperature of Mycoplasma strains should be in a lower state and the freezing period preferably within one year.
Objective To investigate the effects of cryopreservation time and temperature on the stability and activities of some Mycoplasma strains. Methods The differences of CFUs(colony-forming units) of Mycoplasma pneumonia and Mycoplasma orale were calculated under different freezing time and temperature by a solid plate counting method. Results Mycoplasma orale reached the growth peak after 48 hours in culture, and the growth peak of Mycoplasma pneumonia reached in slower than Mycoplasma orale, with 103 hours in culture. After frozen for one month in -70 ℃, the counting of Mycoplasma orale decreased from 10~(12) cfu/mL to 10~(11) cfu/mL, but there was no a statistical significance(P>0.05) in stored for one month with for three and 12 months, respectively. The counting of Mycoplasma pneumoniae matained in a stable level in -70 ℃, and no a statistical significance under the -35 ℃ and-70 ℃ condition(P>0.05), but the activty of Mycoplasma orale strain in -70 ℃ was significantly higher than that in -35 ℃(P<0.05). Conclusion The growth trend of Mycoplasma orale was rapid, but the sensitivity to storage temperature was higher than that of Mycoplasma pneumoniae. Compared with Mycoplasma orale, Mycoplasma pneumoniae grew slowly and stagnated for a longer time. The freezing temperature of Mycoplasma strains should be in a lower state and the freezing period preferably within one year.
引文
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[7] 杨阳, 杨智琪, 周运恒, 等. 支原体固体-液体联合培养与分子检测的比较评价[J]. 检验医学, 2017, 32(9): 813-817.
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[9] European Pharmacopoeia Commission. European Pharmacopoeia 8.0[S]. Strasbourg, France: Directorate for the Quality of Medicines& HealthCare of the Council of Europe, 2013, 01:178-182.
[2] 武昱孜, 张旭, 华利忠, 等. 支原体对细胞培养污染的研究概况[J]. 动物医学进展, 2013, 34(9): 112-117.
[3] 朱真, 万建青, 杨挺英, 等. 支原体培养基质控菌株菌种代次与生长曲线研究[J]. 中国兽药杂志, 2015, 49(8): 10-14.
[4] BENISHEVA T, S0VOVA V, IVANOY I, et al. Comparison of methods used for detection of mycoplasma contamination in cell cultures, sera, and live-virus vaccines[J]. Folia Biologica,1993, 39(5): 214-217.
[5] 国家药典委员会. 中华人民共和国药典(三部)[S]. 北京: 中国医药科技出版社, 2015: 通则98-99.
[6] 赵宏大, 谢文, 范文平, 等. 不同来源药品支原体检查用培养基的促生长能力研究[J]. 中国药师, 2015, 18(12): 2062-2066.
[7] 杨阳, 杨智琪, 周运恒, 等. 支原体固体-液体联合培养与分子检测的比较评价[J]. 检验医学, 2017, 32(9): 813-817.
[8] 范文平, 赵宏大, 谢文. 中国药典与欧美药典对支原体检查培养基规定的差异[J]. 中国药品标准, 2014,15(2): 85-87.
[9] European Pharmacopoeia Commission. European Pharmacopoeia 8.0[S]. Strasbourg, France: Directorate for the Quality of Medicines& HealthCare of the Council of Europe, 2013, 01:178-182.