| Objective:Fungi are an important part of biology.Pathogenic fungi can infect human body and cause fungal infectious diseases.The pathogenicity of fungi is closely related to many factors,and the hydrolase produced by fungi is one of the important factors.Some fungi can cause both superficial infection and deep infection.Among them,Candida and Aspergillus are the main pathogenic bacteria causing deep and superficial mycosis in human body.In order to adapt to different external environment,the activities and types of hydrolases secreted by fungi may be different.The purpose of this study was to explore the differences on the activity and types of secreting hydrolases between superficial and deep mycosis isolates by simulating the shallow and deep environment of fungal infection,and to further explore the relationship between fungal extracellular hydrolase and fungal pathogenicity.Methods:1.From 2019 to November 2020,211 fungal strains from patients with positive fungal culture were collected from the Department of Dermatology and Laboratory of the affiliated Hospital of Chengde Medical College.Among them,there were 161 superficial fungal strains and 50 deep fungal strains.2.The collected fungal strains were inoculated on potato glucose Agar medium(PDA)for subculture and preservation.3.Some filamentous fungal strains were cultured and made into slide specimens,which were preliminarily identified as Aspergillus collected images and retained;all Candida strains identified by Candida chromogenic medium were retained.4.The preliminary identification of similar fungal strains from superficial and deep fungal strains were sequenced.Among them,there were 6 strains of Candida and 9 strains of Aspergillus.5.The fungal strains with the same results of shallow and deep sequencing were inoculated in the potato liquid medium containing nail shavings and scurf and the potato liquid medium containing mouse intestinal mucosa respectively.The potato liquid culture medium containing nail shavings and scurf was placed in a 27℃ incubator to simulate the shallow environment of fungal survival,and the potato liquid culture medium containing mouse intestinal mucosa was placed in a 37℃ incubator to simulate the deep environment of fungal survival.6.The extracellular hydrolase activity was determined by using the enzyme activity kit API ZYM of Bio-Merier Company in France,and the corresponding color depth was marked with 0-5.First of all,the quality control experiment was carried out with α-chymotrypsin according to the kit instructions;then,the two fungal strains cultured in the simulated environment were centrifuged,ground,and the bacterial suspensions were prepared,and the extracellular hydrolase activity was determined according to the kit instructions.7.SPSS 26.0 software was used to test the normality,if it conformed to the normal distribution,t-test was used to compare the two groups,and rank sum test was used if it did not conform to the normal distribution.Results:1.Candida albicans of the same strain were cultured in simulated shallow environment and deep environment respectively,and the activity expression of esterase lipase(t=5.20,P<0.05),acid phosphatase(t=5.20,P<0.05)and N-acetyl-β-glucosaminidase(t=5.20,P<0.05)were statistically significant.Aspergillus Niger of the same strain was cultured in simulated shallow environment and deep environment respectively,and the activity expression of esterase lipase(t=6.93,P<0.05),acid phosphatase(t=6.93,P<0.05)andα-glucosidase(t=5.20,P<0.05)produced by Aspergillus Niger of the same strain was statistically significant.Aspergillus oryzae of the same strain was cultured in simulated shallow environment and deep environment respectively,the activity expression of α-galactosidase(t=5.20,P<0.05)and P-galactosidase(t=5.20,P<0.05)was statistically significant.2.After cultured in the simulated shallow environment,there was a statistically significant difference in the activity expression of acid phosphatase(t=5.20,P<0.05)and α-glucosidase(t=5.20,P<0.05)in Aspergillus Niger from the deep and shallow parts respectively,and after cultured in the simulated shallow environment,there was a statistically significant difference in the activity expression of esterase lipase(t=6.93,P<0.05)from Aspergillus oryzae from the deep and shallow parts,respectively.3.After cultured in simulated deep environment,there was a statistically significant difference in the activity expression of acid phosphatase(t=6.93,P<0.05)in Candida albicans from deep and shallow parts,respectively.After cultured in the simulated deep environment,there was a significant difference in the expression of esterase lipase(t=6.93,P<0.05)activity in Aspergillus Niger from the deep and shallow parts respectively.After cultured in simulated deep environment,there was significant difference in the expression of estrease lipase(t=5.20,P<0.05)andβ-galactosidase(t=5.20,P<0.05)activity in Aspergillus oryzae from deep and shallow parts respectively.4.Compared with the shallow Candida albicans cultured in the simulated shallow environment,the estrease lipase(t=5.20,P<0.05)activity expression of the deep Candida albicans in the simulated deep environment was statistically significant.Compared with Aspergillus Niger cultured in the simulated shallow environment,there was a statistically significant difference in the expression of estrease lipase(t=6.93,P<0.05)activity from Aspergillus Niger in the simulated deep environment.Compared with the shallow Aspergillus oryzae cultured in the simulated shallow environment,there was a statistically significant difference in the activity expression of estrease lipase(t=4.44,P<0.01)and α-galactosidase(t=4.47,P<0.01)from Aspergillus oryzae in the simulated deep environment.5.There was no significant difference in the expression of extracellular hydrolase activity among the mentioned strains.Conclusion:1.The activities of estrease lipase,acid phosphatase and N-acetyl-β-aminoglucosidase of Candida albicans were different after being cultured in simulated deep environment and shallow environment respectively.The activities of estrease lipase,acid phosphatase and α-glucosidase of Aspergillus Niger were differentafter after being cultured in simulated deep environment and shallow environment respectively.The activities ofα-galactosidase and β-galactosidase of Aspergillus oryzae were differentafte afterr being cultured in simulated deep environment and shallow environment respectively.2.In the simulated deep environment,the changes of acid phosphatase activity of Candida albicans from deep and shallow parts were different,and the estrease lipase activities of Aspergillus Niger from deep and shallow parts were different.The changes of estrease lipase and β-galactosidase activities of Aspergillus oryzae from deep and shallow parts were different.In the simulated shallow environment,the changes of acid phosphatase andα-glucosidase activities of Aspergillus Niger from deep and shallow parts were different,and the estrease lipase activities of Aspergillus oryzae from deep and shallow parts were different.3.After the culture of the same strains from different parts,it was found that there were differences in the activity of estrease lipase between Candida albicans and Aspergillus Niger,and in the activities of estrease lipase andα-galactosidase in Aspergillus oryzae. |
PDF Full Text Request