http://informahealthcare.com/bty ISSN: 0738-8551 (print), 1549-7801 (electronic) Crit Rev Biotechnol, Early Online: 1–11 ! 2013 Informa Healthcare USA, Inc. DOI: 10.3109/07388551.2013.791245 REVIEW ARTICLE Advances in research of the artificial cultivation of Ophiocordyceps sinensis in China Xuan-Wei Zhou1,2, Lin-Jun Li3, and En-Wei Tian4 1Plant Biotechnology Research Centre, School of Agriculture and Biology, Shanghai Jiao Tong University, Shanghai 200240, People’s Republic of China, 2Tibet Shengniu Biotechnology Co., Ltd., Lhasa, Tibet 85000, People’s Republic of China, 3Key Laboratory for Urban Habitat Environment Science and Technology, School of Environment and Energy, Shenzhen Graduate School of Peking University, Shenzhen 518005, People’s Republic of China, and 4Key Laboratory of Plant Resources Conservation and Sustainable Utilization, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou 510650, People’s Republic of China Abstract Keywords Ophiocordyceps sinensis (syn. Cordyceps sinensis), a traditional Chinese medicine called Artificial cultivation, environmental DongChongXiaCao (DCXC) in Chinese, is well known and has been used in Asia countries protection, Hepialus larvae, Hirsutella since the fifteenth century, and it contains some valuable medicinal component defined by sinensis, Ophiocordyceps sinensis modern pharmacological science. DCXC only appears at high altitudes on the Qinghai-Tibetan Plateau. Consequently, it is difficult to find and harvest. Because of its rarity and medicinal History value, DCXC has always been one of the most expensive medicines known. As the price of DCXC has risen in recent years, thousands of migrants have entered into the various grasslands Received 26 April 2012 to search for them in season, which makes ecological environments of the grassland more Revised 21 August 2012 fragile. In order to relieve the environmental pressures and protect this valuable resource, the Accepted 12 February 2013 artificial cultivation of DCXC involving two aspects of the genus Hepialus and the fungi of Published online 22 July 2013 the host larvae should be employed and applied at the first available time point. In this article, the reproduction of moth larvae of the genus Hepialus is first described, which includes their ecological characteristics and the methods of artificial feeding. Second, the generation and isolation method of the fungi from DCXC are subsequently summarized, and then the For personal use only. mechanism of fungal spores to attack the moth larvae are restated. Finally, the basic model of artificial cultivation of DCXC is introduced; meanwhile, the potential application of modern biotechnology to the artificial cultivation is analyzed in prospect. This review article will not only expand people’s knowledge regarding the artificial cultivation of DCXC, but also hopefully provide an informative reference for the development of this valuable resource and the environmental protection of alpine meadows. Introduction Xin’’ written by Yiluo Wu in the Qing dynasty (1757 AD) and ‘‘Ben Cao Gang Mu Shi Yi’’ written by Xueming Zhao in Ophiocordyceps sinensis (also known as Cordyceps sinensis) 1765 AD. In 1843, Berkeley (1843) described the DCXC as is a parasitic complex of a fungus (Hirsutella sinensis) and the Sphaeria sinensis, and then Saccardo (1878) classified caterpillar which belongs to Thitarodes, Hepialidae, Critical Reviews in Biotechnology Downloaded from informahealthcare.com by Fudan University on 07/22/13 them into the genus of Cordyceps. In western countries, the Lepidoptera. The Chinese name, DongChongXiaCao 2013 first person introducing how to use the DCXC was a priest, (DCXC), originates from the growth process of O. sinensis: doctor Du Halde, who described its Chinese name as ‘‘Hia the larva of some species. Hepialidae is initially infected by [sic] Tsao Tong Tchong’’ (Winkler, 2009). The name fungi in the spring and summer seasons, and turn into ‘‘stiff Cordyceps sinensis came from the Latin words, i.e., Cord worm’’ in winter, so it is called as DongChong (winter denotes ‘‘club’’, ceps is ‘‘head’’ and sinensis means ‘‘made worm). In the next spring and summer seasons, the stroma in China’’. In 2007, molecular biological methods were germinates and then grows from the head of the larva, which used to modify the classification of Cordycipitaceae and is known as XiaCao (summer grass). This Chinese name is Clavicipitaceae, and some Cordyceps spp. were classified into translated from its Tibetan name Yartsa gunbu first recorded a new family, Ophiocordycipitaceae (Sung et al., 2007). As by a Tibetan doctor Zurkhar Namnyi Dorje in the fifteenth Ben Cao Cong Xin (New Compilation of Materia Medica) century, and then is found in the books of ‘‘Bei Cao Cong depicted, the DCXC lives as an old silkworm in the winter soil, and its body hair grows out of soil and turns into grass in Address for correspondence: Xuan-Wei Zhou, School of Agriculture and the next summer. It tastes ‘‘Gan Ping’’ (sweet flat), and its Biology, Shanghai Jiao Tong University, Shanghai 200240, People’s Republic of China. Tel: +86 21 34205778. Fax: +86 21 65642425. functions were ‘‘beneficial for lung and kidney, activating E-mail: [email protected], [email protected] blood circulation to dissipate blood stasis and treating virtual 2 X.-W. Zhou et al. Crit Rev Biotechnol, Early Online: 1–11 cough and haemoptysis’’. Based on the description of (Liu et al., 1995; Yan, 2000). Nowadays, the biological Pharmacopoeia of the People’s Republic of China (2005 characteristics of more than 10 species of genus insects have ed.), the DCXC is also functional on curing impotence, been well studied in China, which include H. armodcamu spermatorrhea and the resolving pain in the knee and waist. Oberthar, H. minyuancus (Wang et al., 2001a), H. yushusis Besides, modern medicines have proved that it possesses Chuet Wang, H. lagii Yan (Xu, 2004), H. gonggaensis (Yin some functions, such as anti-tumor, anti-hyperglycemia et al., 1995), H. guidera Yan, H. oblifurcus Chuet Wang, (Lo et al., 2006) and significant improvement of the body’s H. renzhiensis, H. deginensis, H. alttdcoh W, H. biru immune function (Paterson, 2008; Zhou et al., 2009; (Yin et al., 2004), H. pui sp. nov. and Thitarodes pui Zhu et al., 1998). Up to now, the medicinal and economic (Chen et al., 2004; Li et al., 2011a; Sasaki et al., 2008; Wang values of DCXC have been wildly realized and studied in & Yao, 2011; Yang et al., 2009; Zhang et al., 1989, 2007). the world (Weckerle et al., 2010; Winkler, 2008, 2010; The host insects of the DCXC are mainly located in Zhang et al., 2009b). the zones of the Qinghai-Tibetan Plateau, which have the In recent years, the price of the DCXC has continuously characteristics of regional and vertical distribution. increased and has gradually become the main income source Each insect population has specific geographical location of local farmers and herdsmen. During the harvesting and distribution pattern. The vertical distribution of the genus season of every year, thousands of locals and migrants Hepialus ranges from 2200 to 5000 m above sea level. Most travel to the Sichuan-Tibet and Qinghai-Tibet Plateau of them are located around the altitude of 4000 m. Among and search inch-by-inch for the DCXC. This has caused different zones, the range of vertical distribution is the widest serious eco-environmental damage to the local grassland. in Tibet province, followed by Sichuan and Yunnan provinces, Furthermore, the increase of population and human activities, and the narrowest in Qinghai and Gansu provinces. Except for accompanied by some natural factors like drought and water the adult period, the host insects live in the soil, whose eco- resources shortage, has made the degradation and desertifi- distribution is in respect of many natural factors, such as cation amount of grassland of this area up to 40% of the total terrain, landform, altitude, climate, vegetation and agrotype. grassland area, and meanwhile it is increasing at an annual Among them, the soil microclimate and foodstuffs are the rate of 3–5%. The enormous environmental pressure and most important factors closely related to the distribution of ecological problems have attracted the attention of many host insects. The host insects of Hepialus distribute every- scientists and governments (Bian et al., 2008; Li et al., where in the alpine, sub-alpine meadow and alpine shrubs, the 2011b). How to implement sustainable utilization of the soil types of which are the alpine meadow soil and sub-alpine DCXC resource on the Qinghai-Tibet Plateau has become an meadow soil (Boesi & Cardi, 2009; Li et al., 2011c; Liu et al., urgent issue to be solved. In the recent 20 years, some 2007; Yan, 2001; Yang et al., 1992, 1996). research institutes and universities from China have paid a lot of efforts and made some progress under the support of the Biological characteristics For personal use only. Chinese government, such as the Major State Basic Research Development Program of China (973 Program), the National The host insects Hepialus of the DCXC are of complete High-tech R&D Program of China (863 Program) and the metamorphosis. There are four stages in its life cycle, i.e. egg, National Natural Science Foundation of China. Beside some larva, pupa and adult Chen et al. (1973). Although there are rules and regulations on the measures in policy and law, all some differences on the biological and ecological characters attentions have been focused on the following aspects: among the different Hepialus, they all have a long larva stage artificial generation of the moth larvae of Hepialus, artificial and an obvious alternate of generation as shown in Figure generation of O. sinensis fungal strains, how the fungal spores 1(a). The insects of Hepialus need to take 3–4 years or even attack the moth larvae and artificial cultivation of DCXC. 45 years in general to complete a life cycle: Meanwhile, a conclusion has been made that the eco- (1) About 500 eggs were laid by each female from June to environmental pressure of the grasslands can be relieved by August.