TY - JOUR
T1 - Protocol for identifying natural agents that selectively affect adhesion, thickness, architecture, cellular phenotypes, extracellular matrix, and human white blood cell impenetrability of candida albicans biofilms
AU - Park, Yang Nim
AU - Srikantha, Thyagarajan
AU - Daniels, Karla J.
AU - Jacob, Melissa R.
AU - Agarwal, Ameeta K.
AU - Li, Xing Cong
AU - Solla, David R.
N1 - Funding Information:
This work was supported in part by the Developmental Studies Hybridoma Bank, a National Resource created by NIH and housed at the University of Iowa, and a USDA-ARS cooperative agreement (no. 58-6408-2-0009) in association with the National Center for Natural Products Research at the University of Mississippi. We are indebted to Claude Pujol for his help with statistical analyses.
Publisher Copyright:
Copyright © 2017 American Society for Microbiology. All Rights Reserved.
PY - 2017/11
Y1 - 2017/11
N2 - In the screening of natural plant extracts for antifungal activity, assessment of their effects on the growth of cells in suspension or in the wells of microtiter plates is expedient. However, microorganisms, including Candida albicans, grow in nature as biofilms, which are organized cellular communities with a complex architecture capable of conditioning their microenvironment, communicating, and excluding low- and high-molecular-weight molecules and white blood cells. Here, a confocal laser scanning microscopy (CLSM) protocol for testing the effects of large numbers of agents on biofilm development is described. The protocol assessed nine parameters from a single z-stack series of CLSM scans for each individual biofilm analyzed. The parameters included adhesion, thickness, formation of a basal yeast cell polylayer, hypha formation, the vertical orientation of hyphae, the hyphal bend point, pseudohypha formation, calcofluor white staining of the extracellular matrix (ECM), and human white blood cell impenetrability. The protocol was applied first to five plant extracts and derivative compounds and then to a collection of 88 previously untested plant extracts. They were found to cause a variety of phenotypic profiles, as was the case for 64 of the 88 extracts (73%). Half of the 46 extracts that did not affect biofilm thickness affected other biofilm parameters. Correlations between specific effects were revealed. The protocol will be useful not only in the screening of chemical libraries but also in the analysis of compounds with known effects and mutations.
AB - In the screening of natural plant extracts for antifungal activity, assessment of their effects on the growth of cells in suspension or in the wells of microtiter plates is expedient. However, microorganisms, including Candida albicans, grow in nature as biofilms, which are organized cellular communities with a complex architecture capable of conditioning their microenvironment, communicating, and excluding low- and high-molecular-weight molecules and white blood cells. Here, a confocal laser scanning microscopy (CLSM) protocol for testing the effects of large numbers of agents on biofilm development is described. The protocol assessed nine parameters from a single z-stack series of CLSM scans for each individual biofilm analyzed. The parameters included adhesion, thickness, formation of a basal yeast cell polylayer, hypha formation, the vertical orientation of hyphae, the hyphal bend point, pseudohypha formation, calcofluor white staining of the extracellular matrix (ECM), and human white blood cell impenetrability. The protocol was applied first to five plant extracts and derivative compounds and then to a collection of 88 previously untested plant extracts. They were found to cause a variety of phenotypic profiles, as was the case for 64 of the 88 extracts (73%). Half of the 46 extracts that did not affect biofilm thickness affected other biofilm parameters. Correlations between specific effects were revealed. The protocol will be useful not only in the screening of chemical libraries but also in the analysis of compounds with known effects and mutations.
KW - Antibiofilm agents
KW - Candida albicans
KW - Natural agents
UR - http://www.scopus.com/inward/record.url?scp=85032490944&partnerID=8YFLogxK
U2 - 10.1128/AAC.01319-17
DO - 10.1128/AAC.01319-17
M3 - Article
C2 - 28893778
AN - SCOPUS:85032490944
SN - 0066-4804
VL - 61
JO - Antimicrobial Agents and Chemotherapy
JF - Antimicrobial Agents and Chemotherapy
IS - 11
M1 - e01319-17
ER -