Abstract
Background: Polyporus umbellatus sclerotia have been used as a diuretic agent in China for over two thousand years. A
shortage of the natural P. umbellatus has prompted researchers to induce sclerotial formation in the laboratory.
Methodology/Principal Finding: P. umbellatus c*tion in a sawdust-based substrate was investigated to evaluate the
effect of low temperature conditions on sclerotial formation. A phenol-sulfuric acid method was employed to determine the
polysaccharide content of wild P. umbellatus sclerotia and mycelia and sclerotia grown in low-temperature treatments. In
addition, reactive oxygen species (ROS) content, expressed as the fluorescence intensity of mycelia during sclerotial
differentiation was determined. Analysis of ROS generation and sclerotial formation in mycelia after treatment with the
antioxidants such as diphenyleneiodonium chloride (DPI), apocynin (Apo), or vitamin C were studied. Furthermore,
macroscopic and microscopic characteristics of sclerotial differentiation were observed. Sclerotia were not induced by
continuous c*tion at 25uC. The polysaccharide content of the artificial sclerotia is 78% of that of wild sclerotia. In the
low-temperature treatment group, the fluorescent intensity of ROS was higher than that of the room temperature (25uC)
group which did not induce sclerotial formation all through the c*tion. The antioxidants DPI and Apo reduced ROS
levels and did not induce sclerotial formation. Although the concentration-dependent effects of vitamin C (5–15 mg mL21)
also reduced ROS generation and inhibited sclerotial formation, using a low concentration of vitamin C (1 mg mL21)
successfully induced sclerotial differentiation and increased ROS production.
Conclusions/Significance: Exposure to low temperatures induced P. umbellatus sclerotial morphogenesis during c*tion.
Low temperature treatment enhanced ROS in mycelia, which may be important in triggering sclerotial differentiation in P.
umbellatus. Moreover, the application of antioxidants impaired ROS generation and inhibited sclerotial formation. Our
findings may help to provide new insights into the biological mechanisms underlying sclerotial morphogenesis in P.
umbellatus.
Citation: Xing Y-M, Zhang L-C, Liang H-Q, Lv J, Song C, et al. (2013) Sclerotial Formation of Polyporus umbellatus by Low Temperature Treatment under Artificial
Conditions. PLoS ONE 8(2): e56190. doi:10.1371/journal.pone.0056190
Editor: Pratibha V. Nerurkar, College of Tropical Agriculture and Human Resources, University of Hawaii, United States of America
Received August 16, 2012; Accepted January 7, 2013; Published February 20, 2013
Copyright: 2013 Xing et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits
unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Funding: The research was financially supported by the National Natural Sciences Foundation of China (No.30830117, 31201666), the International Science and
Technology Cooperation Projects of China (No. 2011DFA31260) and the National Research Foundation (NRF 2011–0016999). The funders had no role in study
design, data collection and analysis, decision to publish, or preparation of the manuscript.
Competing Interests: The authors have declared that no competing interests exist.
* : sxguo2006.cn (SXG); clwang@implad.ac.cn (CLW)