A Method for Measuring the Degree of Fermentation of the Edible Mushroom Cultivation Substrate

In the study, eight treated substrates were designed to explore the possibility to determine the degree of fermentation of the substrate by the mycelial growth rate, whose main raw material includes composted pine sawdust, oil tea shell and hickory shell respectively, and auxiliary materials contain rice bran, soybean powder, etc. The result showed that the shiitake mushroom grew well in 7 treatments whose mycelial growth rate could be measured on 3rd days when the mycelial growth rates of P1, C1 and H1 were 5.0 mm/d, 9.66 m/s, 13.33 m/s. Auricularia cornea var. Li exhibited the fastest growth on P1 substrate. And mycelial growth rates of P1, P0 and CK1 were 5.8 mm/d, 3.66 mm/d, and 4.66 mm/d on 3rd day, respectively. The growth rates of Pleurotusgeesteranus of C1, CK2 and P0 were 9.0 mm/d, 11.66 mm/d, and 4.00 mm/d on 3rd day, respectively. So the degree of fermentation of the substrate could be determined within 3 days according to the mycelial growth rate. As the growth of edible fungi is affected by degree of fermentation of the substrate and there is little literature on degree of fermentation of edible fungi substrate, the study will provide theoretical and technical basis for determination of substrate fermentation.


Introduction
The cultivation substrate of edible fungi provides the nutrients for their growth. Therefore, even under the same conditions, the composition, formula, and DOI: 10.4236/nr.2018.911022 356 Natural Resources treatment of the substrate can significantly affect the mycelial growth rate, as well as the yield and flavor of the fruiting body [1]. A mushroom cultivation substrate is usually composed of a main component and auxiliary materials.
Wood, herbs, feces, fiber, etc. with abundant carbons can be used as the main component of a mushroom cultivation substrate [1]. Organic nitrogen sources, phosphorus and potassium fertilizers, inorganic salts, etc. have been used as the auxiliary materials for the cultivation of mushrooms [1]. The mixed substrate can be used directly without any treatment, which makes the cultivation easy and time saving. However, the fermentation of the substrate can burn some mushroom spores, resulting in low germination rates, especially in summer when the substrate composts to acids [1]. Therefore, directly using raw substrates for mushroom cultivation requires larger amounts of spores. Another option is to use clinker substrates, which is usually applied to fungi with low decomposing abilities. In general, bagged substrates are sterilized under atmospheric pressure or autoclaved to kill harmful microorganisms, pests, and bacteria, to reduce contaminations, promote yield, and improve the stability of production [1].
The effects of the degree of fermentation of substrate on the mycelial growth rates of edible fungi have been reported by many researchers. For example, Xiao and Liu [2] cultivated Pleurotus cystidiosus using the defatted pine and fir sawdust, and evaluated the substrates with the mycelial growth rate. They found that the formulation of substrate significantly affected the mycelial growth rate and the bagged cultivation substrate was outgrown in 42 -47 days. Lin and Zhang [3] investigated the spawning and mycelial growth rates of edible fungi on the substrates containing different ratios of crushed cassava stems to sawdust. Zhang [4] treated pine sawdust by the lime water soaking method and stacking fermentation method as the cultivation substrate of fungi. They determined the degree of fermentation of the substrate with the aromatic scent. Tang [5] treated pine sawdust by the detoxification method and used the pine oil scent and alkaline odor as the indicators of the degree of fermentation. Qiu [6] evaluated the degree of fermentation of pine sawdust with the aromatic scent, color (yellow-brown), and feels (soft) during the natural composting.
In the study, substrates composed of pine sawdust, oil tea shell or hickory shell that were used as the main raw materials, and rice bran, soy flour, corn flour, and gypsum that were used as auxiliary materials with the moisture contents of 55% -60% were inoculated with shiitake mushroom, Auricularia cornea var. Li and Pleurotus geesteranus spores, respectively, in test tubes. The mycelial length was used to evaluate the growth rate of mycelium and the degree of fermentation of the substrate. Our work has provided a simple and rapid method to determine the degree of fermentation of substrate, as well as a theoretical and technical foundation for studying the mycelial growth and substrate fermentation of edible fungi.
This study used the mycelial growth rates of edible fungi to determine the de-

Materials
The shiitake mushroom, Auricularia cornea var. Li and Pleurotus geesteranus spores were provided by Wuhan SuisuiFeng Agricultural Technology Development Co., Ltd., (Hubei Province, China). The contents of terpenes in the untreated and composted Pinus massoniana sawdust were measured to be 0.396% and 0.096%, respectively. The contents of saponin in the untreated and composted oil tea shell were 4.8% and 2.06%, respectively.

Experimental Design
Eight substrates were prepared with different raw materials, the specific treatments were shown in

Preparation of Substrate
The components of each substrate were weighted and mixed, and the moisture content of the substrate was adjusted to 55% -60% with water. The substrate was further mixed thoroughly, and filled in test tubes of the same size. Each tube was loaded with the same amount of substrate, and sterilized in an autoclave at 121˚C for 20 min.

Inoculation
The sterilized substrates in test tubes were cooled to room temperature on a

Cultivation and Measurement
The inoculated substrates in tubes were cultured in an incubator at 24˚C and the air humidity of ~70%. The lengths of mycelia were measured every 3 days to calculate the mycelial growth rate.

Effects of Substrate on the Mycelial Growth Rate of Shiitake Mushroom
As shown in Figure 1, all of the shiitake groups grew well, yet with different mycelial growth rates.

Growth of Auricularia cornea var. Li on Pinus massoniana Sawdust Substrates
The Auricularia cornea var. Li mycelia could geminate and grow in the three treatment substrates with the growth rates in the order of P1 > CK1 > P0 ( Figure   2). The daily mycelial growth rate using P1 substrate was measured to be 3.0 mm/d which was similar to that of shiitake mushroom. And the growth rate on day 3 and day 18 was 6.0 mm/d and 5.8 mm/d respectively. The mycelium were all over the tubes with P0 and CK1 as substrates on day 27 and 22 with the growth rates of 3.66 mm/d and 4.66 mm/d on day 3 and the growth rate of 5.11 mm/d on day 18. These results suggest that the content of terpenes in the Pinus massoniana sawdust, as well as its fermentation degree, promoted the growth of Auricularia cornea var. Li.

Growth of Pleurotus Geesteranus on Oil Tea Shell Substrates
Pleurotus geesteranus was geminated and grown only in C1, C0, and CK2 substrates ( Figure 3), with the growth rates on day 18 of 3.55 mm/d, 3.44 mm/d,

Conclusion
In summary, the composted Pinus massoniana sawdust, oil tea shell and hickory shell are conducive to the germinations and growths of edible fungi. The mycelial growth rate can be used to determine the degree of fermentation of substrate in three days with appropriate amounts of substrate and liquid spawn. In addition, the degradation degree of substance in conducive to the growth of edible fungi can also be evaluated with the mycelial growth rate. The looseness and the content of hemicellulose of the substrate are the major factors affecting the growth of mycelia.