ABSTRACT
Objective: To explore the relationship between extracellular enzymes activity and virulence of Candida glabrata clinical isolates based on the infection model of Galleria mellonella larvae. Methods: Using experimental research methods, 71 strains of non-repetitive Candida glabrata were collected from Qinghai Provincial People's Hospital from June 2021 to January 2022. Bovine serum protein agar medium, egg yolk agar medium, sheep blood agar medium, Tween-80 agar medium and triglyceride agar medium were used to detect the aspartyl protease activity, phospholipase activity, hemolysis activity, esterase activity and lipase activity of Candida glabrata. Median lethal concentration (LC50) was calculated by using 1.25×108 CFU/ml,2.50×108 CFU/ml,3.75×108 CFU/ml,5.00×108 CFU/ml suspension of Candida glabrata ATCC2001 to infect Galleria mellonella larvae. Histopathological and etiological analysis was performed to determine whether the infection model was successfully established. The clinical isolates of Candida glabrata were configured to infect Galleria mellonella larvae with LC50 concentration to detect the pathogenicity of Galleria mellonella larvae.Spearman test or Pearson test were used to analyze the correlation between the extracellular enzyme activity of Candida glabrata clinical isolates and the pathogenicity of Galleria mellonella larvae. Results: 71 strains of Candida glabrata isolated clinically were detected to have low hemolytic activity after 2 days of culture. Aspartyl protease was detected after 4 days of culture, among which 7 strains (9.86%), 19 strains (26.76%) and 45 strains (63.38%) showed low, medium and high aspartyl protease activity. After 7 days of culture, 71 strains did not detect phospholipase, esterase and lipase activities. Candida glabrata on Galleria mellonella larvae of LC50=2.5×108 CFU/ml Fungal spore were found in the intestinal tissue pathological section of Galleria mellonella larvae in the experimental group, and Candida glabrata was identified by the microbial Mass Spectrometry after culture, while no fungi were found in the pathological section and culture of the control group. Spearman test shows that, there was a linear positive correlation between aspartyl protease activity and the survival rate of Galleria mellonella larvae (r = 0.73, P<0.01), the difference was statistically significant.Pearson test shows that, there was no significant linear relationship between hemolytic activity and survival rate of Galleria mellonella larvae (r = 0.16, P = 0.34), the difference was not statistically significant. Conclusion: The clinical isolates of Candida glabrata in this study had aspartyl protease activity and low hemolytic activity, but no phospholipase, esterase and lipase activity. The activity of aspartyl aspartyl protease of Candida glabrata was positively correlated with the pathogenicity of Galleria mellonella larvae.