Expression Of Nucleolar Factor RCL1 In Clinical Isolation Of Itraconazol-Resistant Candida Albicans And Under Different Stress Conditions

Objective:1.The m RNA expression level of RCL1 gene in clinical isolates of itraconazol-resistant Candida albicans was detected by RT-PCR method,and the possible role of RCL1 in the generation of Candida albicans resistance was analyzed.2.The RT-PCR method was used to detect the clinical isolation of Candida albicans and standard strains under different stress conditions（temperature,carbon dioxide concentration,PH value）,and to detect the m RNA expression level of RCL1,analyze the relationship between environmental and other stress factors and RCL1 expression,and predict its role in stress.Methods:1.Each of the 14 strains resistant to itraconazole single drug and the 14 strains sensitive to itraconazole,fluconazole and fluconazole were selected from the previous collection,culture,identification and drug sensitivity tests of the research group.Total m RNA of the two groups of strains were extracted and transcribed into c DNA.Real-time quantitative PCR was used to detect the m RNA expression level of RCL1 and analyze the relationship between RCL1 and drug resistance formation in clinical isolates of Candida albicans.2.Culture conditions（temperature,carbon dioxide concentration,PH value）of clinically isolated Candida albicans were changed,and total m RNA of the re-cultured Candida albicans was extracted again,then c DNA was transcribed and synthesized,and m RNA expression level of RCL1 was detected by real-time fluorescence quantitative PCR method.To analyze the effect of different environmental stress conditions on the m RNA expression level of RCL1 in Candida albicans.Results:1.Real-time quantitative PCR detection results of clinical isolation of Candida albicans:The m RNA expression level of RCL1 in drug-resistant strain group（0.980±0.124）was lower than that in sensitive strain group（1.329±0.263）,and the difference was statistically significant（t=-3.867,P<0.05）.2.The results of real-time quantitative PCR detection of Candida albicans cultured under different stress conditions:（1）The m RNA expression level of clinically isolated Candida albicans RCL1 cultured at 37℃（1.183±0.276）was higher than that at 25℃,with no statistical significance（t=0.600,P>0.05）;At 25℃,the m RNA expression level of RCL1 in drug-resistant strain group（1.156±0.078）was higher than that in sensitive strain group（1.136±0.170）,the difference was not statistically significant（t=0.348,P>0.05）;（2）The m RNA expression level of clinically isolated Candida albican RCL1 in0.03%CO₂ culture（1.183±0.276）was higher than that in 5%CO₂ culture,the difference was statistically significant（t=2.634,P<0.05）;Under 5%CO₂,the m RNA expression level of RCL1 in drug-resistant strain group（1.002±0.216）was lower than that in sensitive strain group（1.089±0.156）,the difference was not statistically significant（t=-1.149,P>0.05）;（3）The m RNA expression level of clinically isolated Candida albicans RCL1 under p H4 culture（1.183±0.276）was higher than that under p H7 culture,the difference was statistically significant（t=4.981,P<0.05）;Under p H7 condition,the m RNA expression level of RCL1 in drug-resistant strain group（0.744±0.188）was lower than that in sensitive strain group（1.027±0.172）,and the difference was statistically significant（Z=-3.104,P<0.05）.Conclusion:1.Low expression of RCL1 may be related to the formation of drug resistance in clinical isolation of Candida albicans ITR,and RCL1 is involved in the formation of drug resistance in Candida albicans.2.Culture at 25℃and 37℃had no effect on the expression level of RCL1m RNA,and the expression level of RCL1m RNA at these two temperatures was not correlated with drug resistance of Candida albicans.3.Cultured Candida albicans at 0.03%CO₂ and 5%CO₂ had an effect on the expression level of RCL1 m RNA,and the expression level at 0.03%CO₂was higher than that at 5%CO₂,suggesting that a certain increase in CO₂ concentration would lead to a decrease in RCL1m RNA expression.At 5%CO₂ concentration,the mrna expression level of RCL1 had nothing to do with drug resistance of Candida albicans,which may be caused by the expression change of RCL1 under stress conditions,which needs further study and verification.4.The expression level of RCL1 m RNA was affected by the culture of Candida albicans under p H4 and p H7 conditions,and the expression level was higher in p H4 than in p H7 environment,suggesting that increasing the p H value of culture medium to a certain extent would lead to the decrease of RCL1 m RNA expression.In p H7 condition,down-regulated RCL1 m RNA expression was associated with Candida albicans resistance.