Int J Pharm Pharm Sci, Vol 8, Issue 2, 326-328Short Communication


CULTURING AND SPAWNING STRATEGIES FOR CULTIVATION OF GANODERMA LUCIDUM (CURTIS) P. KARST

MADHAVI JOSHI*, ANAND SAGAR

Department of Biosciences, Himachal Pradesh University, Shimla 171005, (Himachal Pradesh), India
Email: madhavi.joshi1986@gmail.com

Received: 11 Aug 2015 Revised and Accepted: 12 Dec 2015

ABSTRACT

Objective: To access the vegetative growth of Ganoderma lucidum on ten solid media and four liquid media. Spawn development on different grains viz., wheat, barley, maize, soybean and pearl millet was also assessed for lab scale artificial cultivation of this mushroom.

Methods: Culture was raised on Potato Dextrose Medium and growth parameters were assessed on different solid and liquid media at 25±2 °C. Various grains were analyzed for best spawn development of the mushroom mycelium. Oak sawdust and wheat bran mixture was used as a substrate for cultivation of the mushroom.

Results: Malt Agar Extract medium and Glucose Asparagine Solution supported the best mycelial growth among the solid and liquid media screened for the purpose. Barley grains were the best for the spawn development. Maximum yield was observed on the Oak sawdust and the wheat bran (4:1 w/w) mixture used as a substrate.

Conclusion: The study revealed that this pharmacologically important mushroom can be grown under lab conditions, and more yield of the mushroom can be obtained using barley based spawn.

Keywords: Cultivation, Fruiting bodies, Ganoderma lucidum, Spawn, Substrates.

© 2016 The Authors. Published by Innovare Academic Sciences Pvt Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/)

Ganoderma lucidum (Curtis) P. Karst. is a species of basidiomycetes which belongs to Polyporaceae (or Ganodermaceae) of Aphyllo-phorales. Its fruiting body is called “Reishi” in Japanese and “Lingzhi” in Chinese [1, 2]. Ganoderma lucidum is one of the most famous traditional Chinese medicinal herbs and used as a healthy food and medicine in the Far East for more than 2000 y [3]. This mushroom contains various chemical substances, including more than 119 different triterpenes and several types of polysaccharides [4]. A successful artificial cultivation of G. lucidum has been reported on most broadleaf hardwood trees and commonly used species include oak, pecan, elder, choke, cherry and plum [5, 6]. Moreover, Ganoderma species can also be cultivated on the sawdust which may originate from different kinds of trees [7, 8]. Artificial cultivation of G. lucidum is still in its early stages in India as compared to China, Japan, Taiwan, Korea, and North America, where it is being cultivated on the large commercial scale. Few studies have been carried out on the feasibility of cultivation of G. lucidum in India [9-11]. The scope of the market for Ganoderma based products is enormous in national and international markets both but the paucity of knowledge on its cultivation practices is the main constraints.

The aim of this paper is to collect knowledge on mycelial growth using various media and also different attempts regarding spawn production on various substrates were made to find out the best substrate for spawn production and cultivation.

The materials used in the study were the fruiting bodies of G. lucidum collected from Glen forest of District Shimla (Himachal Pradesh) in North-West India. It was taxonomically identified following Lincoff [12] and the specimen has been deposited in the Herbarium of Biosciences Department, Himachal Pradesh University, Shimla. The pure culture was raised on the Potato Dextrose Agar medium by tissue culture method.

Both solid as well as liquid media were evaluated for the vegetative growth of G. lucidum. The composition of each media used was based on Tuite [13]. Ten solid media, used for evaluation were Yeastal Potato Dextrose Agar (YPDA), Potato Dextrose Agar (PDA), Coriander Extract (CE), Maize Grain Extract (MGE), Wheat Grain Extract (WGE), Malt Yeast Agar Extract (MYAE), Pridham Yeast Malt Dextrose Medium (PYMD), Czapek Dox Agar (CDA), Malt Agar Extract (MAE) and Dimmick Agar Extract (DAE). Four liquid media, Glucose Asparagine Solution (GAS), Czapek’s Solution (CS), Dimmicks Solution (DS) and Asthana and Hawker’s Solution (AHS) were used during experimentation for making a comparative observation on vegetative growth of G. lucidum.

To measure the growth rate of mycelium in various solid media, the diameters of mycelial colonies were measured in cm scale and the average growth of mycelium was calculated. The mycelial growth rate in various liquid media was calculated by harvesting mycelial mat from each flask, washed and dried at 65 °C for 24 h. The dry weight of mycelium was recorded for two subsequent days and an average was taken as the actual weight.

Different types of grain substrates were evaluated to see their effect on spawn development of G. lucidum. Spawn was prepared using cereal grains (maize, barley, wheat, pearl millet and soybean) as substrates. The grains were washed, boiled and filled in polypropylene bags plugged with non-absorbent cotton, and sterilized in an autoclave. Thereafter, these bags were inoculated with equal sized mycelial bit of pure culture, and incubated at 25±2 °C and observations were recorded when the mycelium covered the entire grains in every treatment. Three replications were kept in each treatment.

The cultivation experiment was done using oak sawdust and wheat bran mixture (4:1 w/w). The substrate media were mixed homogeneously. The pH was stabilized by gypsum and CaCO3 at 5.5-6.5 [14]. After inoculating with the barley and pearl millet based spawn the bags were kept in the dark at 25 °C for spawn run with 80-85% relative humidity. After few days of spawning, small buttons appear, at this stage, polyethene top is cut at the level of the substrate totally exposing the top side. Proper conditions for fruiting were provided in the cropping chamber. Relative humidity was maintained at 95 % and temperature of the chamber was kept around 28-30 °C with a 10 h light exposure. The cap formation of G. lucidum initiated in 2-3 d. Fruiting bodies were harvested when the caps become completely red and the white margins disappeared [15]. Fresh weight and dry weight of mushrooms harvested in each flush were recorded. The mean colony diameter of the mycelium (±SD) in different solid media is graphically represented in fig. 1. The mean colony diameter of the mycelium was maximum on Malt Agar Extract (8.1±0.06 cm), followed by Maize Grain Extract, Pridham Yeast Malt Dextrose Medium, Yeastal Potato Dextrose Agar, Potato Dextrose Agar Medium, Coriander Extract, Malt Yeast Agar Extract, Wheat Grain Extract, Dimmick Agar Extract and Czapek Dox Agar Medium respectively. The average dry weight of the mycelium on Glucose Asparagine Solution (145.60±2.64 mg) was maximum, followed by Czapek’s Solution, Dimmick Solution and Asthana and Hawker’s Solution respectively (fig. 2).

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Fig. 1: Average colony diameter of G. lucidum on different solid media


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Fig. 2: Average weight of mycelium of G. lucidum in different liquid media

Among the five grain substrates used for spawn production, minimum (8 d) period was recorded in barley grains where grains were completely covered by the mycelium and tightly held, followed by pearl millet, wheat and maize. However, soybean grains took maximum (20 d) with poor mycelia growth and grains were not fully covered (table 1, fig. 3).

On sawdust and wheat bran mixture as a substrate while using pearl millet and barley as a spawn source, the spawn run was completed in 23.3 d and 23.8 d respectively. Pinheads were developed in 32.8 and 32.0 d respectively. The highest yield was obtained on the substrate having barley as spawn source (65.24 g/kg) (table 2).

Table 1: Evaluation of different grain substrates for spawn development of G. lucidum

Grain substrate

Spawn  development

(days)*

Mycelial characters

Wheat

10

White cottony compact mycelia growth, all grains were completely

colonised by the mycelium and tightly held with each other

Pearl Millet

9

White cottony compact mycelia growth, all grains were completely

colonised by the mycelium and tightly held with each other

Barley

8

White cottony compact mycelial growth, all grains were completely

colonised by the mycelium and tightly held with each other

Maize

15

Mycelial growth was poor, and grains were not fully covered

Soybean

20

Very poor mycelial growth and grains were not fully covered

* Average of three replications


Table 2: Effect of cereal grains on spawn run and yield of G. lucidum on Oak sawdust

Spawn source

Spawn run (days)**

Pinhead stage (days)**

Yield (g/kg)

Pearl millet

23.3

32.8

59.72

Barley

23.8

32.0

65.24

**Average of three replications


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Fig. 3: Spawn growth on various grain substrates. A. Wheat, B. Pearl millet, C. Barley, D. Maize, E. Soybean

On comparing the previous work on other mushrooms, Potato Dextrose Agar and Yeast Malt Agar found the best medium for Ganoderma applanatum [16]. Malt extract medium was found best for radial growth of Lentinus cladopus [17].

Maximum growth of L. squarrosulus was observed on yeast glucose medium [18] and malt broth was found best for L. cladopus [17]. In our results, maximum growth of mycelium in Glucose Asparagine Solution could be attributed to asparagine, which as a nitrogen source can enhance the growth of mycelium in culture whereas other synthetic media do not contain asparagine, hence show poor growth of mycelium.

Cereal grains and millets are generally used as spawn substrates. They act as a reservoir of carbohydrates; which offer sufficient nutrition for mycelia growth and provide the vehicle for eventual distribution of mushroom inoculants [19]. Barley grains were best for the spawn development and took only 8 d for spawn development as compared to other substrates.

Days required for complete colonization of the substrate was minimum with pearl millet. After seeding the substrate, pearl millet grain spawn allows a quick spreading of mycelium from a small propagation centre because, small grains provide more points of inoculum per gram of spawn [19, 20]. The present findings are very close to the results obtained by earlier workers [21-25]. Oak sawdust and wheat bran combination was used in the present study and yield of the mushroom was also in conformity with some other authors who reported hardwood sawdust as a preferred substrate for the commercial production [14, 15].

Based on the investigations, mushroom growers are advised to maintain the pure culture of G. lucidum on Malt Agar Extract Medium and develop spawn on barley grains. Cultivation of G. lucidum with enormous pharmacological value is highly recommended in India. The authors express their sincere thanks to the Chairman, Department of Biosciences, HPU, Shimla for providing laboratory facilities. Financial support from UGC, (F.17-40/08 SA-1), New Delhi is also gratefully acknowledged.

CONFLICT OF INTERESTS

The authors declare no conflict of interest.

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