Chapter 2

REVIEW OF LITERATURE

The Chinese mushroom has very old history. This mushroom has been used as food for human beings since Cho dynasty about 3000 years ago in China.. It was introduced in southeast Asian countries by overseas Chinese, (Baker, 1934 and Benemertio, 1936), since then, its cultivation has been conducted in various countries outside the China, like Philippines, (Clora, 1937; Go, 1959), Malaysia (Baker 1934; Sands 1935), Burma (Seth, 1944) and Thailand (Jalavicharama, 1950;Hashioka, 1962). The history of the Chinese mushroom cultivation is very old. As far as its artificial cultivation is concerned it is believed that, it was begun in Nanhua temple of Chaohsi, Kwantung province in southern China, almost 200 years ago, (Chang, 1977) 2.1. METHODS OF CULTIVATION 2.1.1. Fungus culturing Munjal, (1973) reported that the productivity of spawn culture was related to the formation of chlamydospores. He described that culture beds with dens chlamydospores always produced a high yield of mushroom. Hence paddy straw pieces mixed with 4% chalk powder were best suited for spawn production of Chinese mushroom. Delmas and Sun, (1984) reported the traditional method of Chinese mushroom culture in humid tropical climate. For modern culture, a high cellulose
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substrate having C/N ratio was required. It was sterilized after fermentation at 60C for 24 hours. After spawning the temperature was required to be maintained at 35C-38C for about 4 days, then reduced 30C to encourage flushing and raised again after flush but casing is not necessary. Qumio, (1988) made attempts to recycle the spent rice straw residues both from Volvariella spp. and Pleurotus spp. either producing another crop of the same mushroom or producing another mushroom from same substrate, thus fully utilizing the straw for production of edible mushrooms before using the spent compost for feed. Luh, (1996) described the cultural methods developed in Taiwan. He reported that rice straw covered by compost was used as the medium for beds and production started 10-12 days after inoculation. 2.1.2. Cultivation forms Ho, (1972) suggested an economical plastic house with a bamboo frame structure that for indoor cultivation of Chinese mushroom consisting of polythene film of 0.1mm was lined inside this mushroom house and has a layer of sugarcane leaves on top, to block solar radiation and minimize heat penetration during hot summer .An electric blower with a one forth horse power and a polythene air duct was used to provide ventilation.

Grahum and Yaung, (1974) observed that beds of 32 cm depth gave the highest yield of 2.25Kg /100Kg of waste and maintained at a temperature of 35C38C slightly below the optimum from mycelial growth of Volvariella volvacaea, throughout the cropping season . Purkayastha et al., (1981) reported that orientation of paddy straw beds has a profound effect on the fruiting bodies production of Volvariella volvacea helix and tyre types of beds ere found to be more productive than the conventional crisscross type. Several wastes for Chinese mushroom were used as substrates. Among them cotton and jute wastes were most and least productive, respectively, supplementation gram flour up to 600g / bed augmented the production. Climatic conditions during June and July in west Bengal appeared to be the most favorable for the production of Volvariella volvacea. Qumio, (1986) reported that water hyacinth could be used for growing not only Volvariella mushroom but also Pleurouts and Auricularia. The mushroom grows fast and yield more if grown in this substrate than on rice straw. It could also be used as substrate for preparation of the spawn either alone or in combination with other substrates such as rice straw and saw dust. All parts of the plant including roots could be used in making beds for Volvariella mushrooms or in the preparation of mushroom bags for Pleurotus and Auricularia. Li, (1989) studied the cultivation of Volvariella volvacea on wheat straw in fields after the wheat harvest. High yield of Volvariella volvacea and 20% increase in yield of the subsequent wheat crop was achieved. He also described the techniques
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concerning the treatment of wheat straw compost, cultivation management and pest control. Wang, (1990) observed the key points in the management of Volvariella volvacea grown on rice straw in open fields in relation to the stages of spawn production, mycelial growth and fruiting bodies formation. Pan and Li, (1990) studied the construction and design of semi-underground sheds. Cultivation techniques including the selection of high quality spores and substrates, bed construction and the control of growth conditions were discussed. Hua, (1990) grew Volvariella volvacaea strain V2 on a rice straw substrate in an apple orchard or in shelters with conventional management. The former method gave slightly higher yields and better quality fruiting bodies than the later. Soil samples taken from around the trees 2 years after Volvariella volvacaea cultivation showed improved organic matter, P and K contents. The orchard environment which provided shade and high air humidity appeared to be favourable for Volvariella volvacaea cultivation. Nayak et al., (1990) reported the feasibility of growing paddy straw mushroom Volvariella volvacea inside plastic tunnels was. Tunnels (4m x 1.7m x 0.085m) were prepared with transparent UV stabilized film (200) in 2 beds (0.9m x 0.9m) were made one of the beds was covered with black LDPE (Low density polyethylene) film (150) under the UV stabilized film. The average mushroom yield at the end of 41 days after spawning was 2.7 kg/m2.

2.2. SOLID STATE FERMENTATION Garo, (1964) used different substrates such as paddy straw dried banana stalks and leaves, water hyacinth, wheat straw and sugarcane baggasse for mushroom production. He observed only the beds made of sugarcane baggasse did not produce mushrooms. The yield obtained from the beds of banana leaves was superior to the yield obtained from any other substrate. Gupta et al., (1970) tried wheat maize barley oat pearl millet, and sorghum straw but the yield was very low as compared to that produced on paddy straw. Before, 1970, paddy or rice straw was practically the only material used for preparing the medium for commercial cultivation of mushroom under natural condition. Chen and Graham, (1973) grew Volvariella volvacea successfully on oil palm pericarp waste. French material composed for five days before spawning gave the highest yield of about 1.54Kg/100 Kg of waste. Bed temperature may be one of the most important factors affecting the incubation period and duration of crop. Chang, (1974) studied the cultivation of Volvariella volvacea on cotton waste compost in plastic green houses. The compost was prepared by adding 4% rice or wheat bran and 4-6% lime stone to cotton waste, after which it was soaked in water and fermented for to 4 days .Production by this method was high and cropping was also regular. Graham, (1974) studied the performance of three field isolates collected from oil palm bunch waste in Selangor and four isolates from Sarawak, Hong Kong,
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Indonesia and Singapore, which were compared in two experiments. The Hong Kong isolate yielded 5-6 Kg/100Kg of substrate and gave a maximum bed yield of over 3.1 Kg or 7.4 Kg/100 Kg of waste. The performance approached and surpassed yield from paddy straw. Madane et al., (1974) produced spawn of Volvariella volvacaea on paddy straw mixed with 2.5% by weight of oat meal. Same results were obtained when powered wheat or gram was substituted for the oat meal and when wheat or sorghum straw or sugarcane bagasse was used instead of paddy straw. Granhum, (1975) found a wide range of cultural characteristics and yielding ability in single spore isolation from five cultures of Volvariella volvacaea. A selected isolate from Hong Kong culture out yielded the parent culture by about 125% and one from a Sarawak culture out yielded its parent by about 199%. Jablonsky, (1981) studied the substrates consisting of standard mushroom compost prepared with 11 days at stage 1, and 8 days at stage II, horse manure composed for 3 days and wheat straw treated in various ways. Half of each substrate was treated for 4 hours at 900C (pasteurized) and the other half at 1200C for 1.5 hour sterilized. Pasteurization of substrates produced higher yield than sterilization. The highest yield was with chopped wheat straw. The addition of 3% CaCO3 reduced the yield. Qumio, (1981) reported that the following substrates supported very well mycelial growth of Volvariella volvacea, rice straw, Ipil-Ipil leaves, sigadillas leaves, new paper prints, coconut coir dust and banana bracts. It took 8 days for mycelium to
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fill up the entire diameter of the Petri dish containing the substrate. This only confirmed the fact Volvariella, unlike Agaricus species, could grow directly on un composted substrates and therefore could be considered less specific in growth requirements than the later mushroom. Devi and Nair, (1987) observed that Volvariella volvacaea spawn prepared out of 3-10 days old culture supported maximum sporocarp formation, when used for spawning the beds. Maize and wheat grains supported good mycelial growth and found to be suitable substrate for spawn preparation. At room temperature of (28+4OC) the spawn remained viable with out a reduced in yield for about 20-60 days. Alam and Khan, (1989) calculated growth percentage index of locally crushed fresh mill bagasse with 25%, 10% and 5% molasses. The growth percentage index in the first flush ranged between 305- 425%, in the second flush between 283300%, in the third flush between 220-240% and in the fourth flush it was 180%. Adewusi et al., (1993) determined the biological value of 5 mushrooms Chlorophyllum molybditis, Psathyrella atroumbonata, Termitomyces robustus, Termitomyces striatus and Volvariella esculenta by using weanling rats and observed that T. robustus and V. esculenta did not support growth at all. Khan et al., (1994) tried dried water hyacinth for the cultivation of Volvariella volvacaea and concluded that water hyacinth + cotton waste at the rate 1:1 gave maximum yield of Chinese mushroom followed by dried water hyacinth alone and cotton waste alone.
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Chiu et al., (1996) described that the cultivated strains of shiitake in China were genetically very homogeneous, very like to the cultivated strains of Agaricus bisporus and Volvariella volvacea. However, their collection of L. edodes, covers an enormous geographical area, (approx. 1700 km N to S, 700 km E to W) and results demonstrated that the shiitake industry in China depends on an extremely small gene pool. Salmones et al., (1996) reported the mycelial growth of two Mexican strains of Volvariella volvacea (Bull.: Fr.) Sing., in 13 agro industrial wastes. He used, banana leaves, bracts of pineapple crown, coconut fiber, coffee bran, coffee pulp, corn cob, corn stover, orange peel, rice bran, rice straw, sisal bagasse, sugarcane bagasse and wheat straw as substrate and evaluated mycelial growth, mycelial thickness and pinhead formations. Fruiting bodies were obtained only from one strain growing in bracts of pineapple crown, coffee pulp, rice straw and sisal bagasse. Primordia were developed between 13 and 15 days. He recorded that the highest biological efficiency was achieved on rice straw, 33.8%, while the results obtained for coffee pulp, sisal bagasse and bracts of pineapple crown were 15, 7.8 and 6.2%, respectively. Chemical analyses of the substrates registered C/N ratios of 33:1 to 80:1. Cheung, (1997) evaluated the feasibility of using food waste, such as soya milk residue, to produce nutritive fungal biomass. They produced edible mushroom mycelia of Volvariella bombycina, Lyophyllum ulmarius and Pleurotus citrinopileatus, were produced in liquid culture containing soya milk waste. They
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observed similarities in the crude protein, lipid, ash and nucleic acid contents between the mycelia and fruiting bodies. Differences were observed in the amount of total dietary fiber and amino acid composition. Reyes et al., (1998) described the nutritional and physical requirements for the efficient mycelial colonization of Volvariella volvacea (Bull. ex. Fr.). Singer. The investigation was limited to the evaluation of two commercial strains (designated Vvc1 and Vvc2) and two wild strains (designated EAAC-0001 and EAAC-0002) of V. volvacea from the Philippines. The four strains of V. volvacea had varying preferences for carbon. Vvc1 preferred polysaccharides (starch and cellulose), whereas Vvc2 grew luxuriantly at a relatively rapid rate in sugar alcohol (sorbitol). The two wild strains preferred starch as a carbon source. In terms of nitrogen utilization, soytone, peptone, and glycine supported efficient mycelial colonization of the four strains. Efficient colonization of Vvc1, Vvc2, and EAAC-0002 with dense mycelial growth was noted in mycological agar. EAAC-0001, on the other hand, grew more efficiently in malt extract agar. The Philippine strains of V. volvacea grew luxuriantly when incubated at 35C and pH 8.0 under dark and sealed conditions. They concluded that under optimum physiological conditions, Vvc1, Vvc2, and EAAC-0002 were fast-growing strains, whereas EAAC-0001 was a moderately growing type. Tonial et al., (2000) used industrial residues from cassava and potato starch processing as substrates to produce the edible mushroom Volvariella volvacea (Bull. Fr.) Singer. Three strains of V. volvacea (LPB 08, 59, 77) were grown in a
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medium containing 2.4% (w/v) declassified potato flour and 1.6% (w/v) cassava bagasse (PFCB) in petri dishes. Growth performance of cultures was evaluated by measuring their rate of radial growth and biomass production. Strain LPB77 showed highest growth on PFCB agar medium. The liquid PFCB medium was optimized with regard to residue composition, nitrogen source and pH The best results were obtained after 8, days of fermentation in a medium containing 4.8% (w/v) declassified potato flour and 1.2% (w/v) cassava bagasse, pH5, supplemented with 0.1% (w/v) of KNO3, giving a C/N ratio of 30. Philippoussis et al., (2001) cultivated ten selected wild and commercial strains of Pleurotus ostreatus, Pleurotus eryngii, Pleurotus pulmonarius, Agrocybe aegerita and Volvariella volvacea on three agricultural wastes, i.e. wheat straw (WS), cotton waste (CW) and peanut shells (PS). They observed that one commercial strain of V. volvacea presented higher growth rates when the composted CW medium was used. Furthermore, earliness in the fructification of P. ostreatus, P. pulmonarius and V. volvacea strains was promoted in CW substrates. They detected positive correlation between cellulose content and mushroom yield for V. volvacea strains. Obodai et al., (2003) evaluated the biological efficiencies (yield of the mushroom against the dry matter of the substrates) of two strains of the mushroom Volvariella volvacea, V99 and VVO, by using banana leaves, cocoyam peelings and oil-palm pericarp as substrates. They observed that primodia were after11-12 days on banana leaves. Both strains showed their highest production on banana
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leaves, with biological efficiencies of 43 and 72%. V99 fruited on all the substrates but VVO fruited only on banana leaves as it had mycogone infection on the other substrates. Akinyele and Akinyosoye, (2005) cultivated the mushroom, Volvariella volvacea on various agro wastes and observed Maximum mycelia extensions in cotton waste (98.23 0.1 mm) and a combination of rice husk and cotton waste (101.87 0.4 mm). A decrease in moisture content resulted in significant increase percentage crude protein content of mushroom-treated waste compared to the untreated they concluded that changes in crude fiber and ash content of treated and untreated wastes were not significant, mineral contents were observed to increase. Phosphorus and potassium ion content also increased in mushroom-treated samples. Belewu and Belewu, (2005) studied the solid state fermentation of banana leaves by lignin degrading mushroom (Volvariella volvacea) ,yield of fruiting body and compositional changes of substrates were evaluated .The biological efficiency was 5.21while the total weight of fruit yield was 2.5 kg. 2.3. PHYSIOLOGY 2.3.1. Effect of temperature Alicbusan and Ela, (1967) reported that the bed should be pressed lightly because it must reach to the temperature of 40C to 45C because this temperature favourable for the mushroom production but not for the organisms present in the bed.
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Chang, 1972 reported the best temperature for the growth and fructification of Chinese mushroom is 26C to 30C and needs bed temperature of 34 to 37C. Agarwala, (1973) reported that tropical mushrooms can grow at a temperature up to 45C or even more and the minimum temperature should not go below 25C in any case even for a short period. He concluded that at higher the temperature faster growth and higher yield will be obtained. The cropping period lasted 25-30 days under poor temperature conditions. Samajpati et al., (1977) studied the cultivation of V. volvacea on paddy straw beds where growth conditions were satisfactory from April to August but highest yield was obtained in July when the optimum temperature was 32C and humidity was about 85%. Khan and Kusar, (1981) reported prospects and potential of mushroom as a cottage industry in Pakistan. Variety of climatic conditions such as northern hilly areas with high humidity and low temperature were suitable for temperature kind of mushrooms where as plains of Punjab, Sindh, and Balochistan were suitable for cultivation of tropical mushrooms in summer. Many agricultural wastes and industrial waste in the form of straws, leaves and cotton and corn wastes were available in large quantities. Ramakrishnan et al., (1986) observed that use of transparent polythene sheet to cover the bed showed higher yield than with black polythene sheet and he also reported the optimum temperature for Chinese mushroom cultivation was 35 to 37.
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Morris et al., (1988) observed the changes in morphology and viability of 20 species of fungi including Volvariella volvacea during freezing were examined in relation to cooling rate and the presence of glycerol. They observed that the morphological response of Phytophthora, Aschersonia and Volvariella differed from other genera, with shrinkage occurring at all rates of cooling. Frank, (1989) recommended measures to obtain high yield of V. volvacaea. The best planting date was when both day and night air temperatures were 23C. Lime was sprayed on the straw to increase the pH value from 7.5 to 8.5. The spawn was planted at an age of 18-20 days. Wang and Li, (1989) studied the various deformities occurred in cultures of V. volavcea including the production of empty mycelium, abnormal bodies formed from the hyphae, deformed fruiting bodies and withering of young mycelium. They suggested that these deformities could be avoided by the maintenance of a constant growing temperature in the range of 22-28C. 2.3.2. Effect of light Yau and Chang, (1970) obtained fruiting under 12 hours light. They reported the light intensity of 50lux was optimum. Antonio and Fordyce, (1972) reported an appreciable quantity of light (15 minutes full sunlight) was required for the initiation of fruiting bodies of Volvariella volvacea. The average yield was 84g/Kg dry weight compost.

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Khan, (1976) described that the light is not required for the Chinese mushroom during the period of spawn running, it respond favorably to weak light condition and ventilation Chakravarty and Mallick, (1979) reported that growth of V. diplasia was vigorous in complete darkness, intermediate in diffused light and slowest in full light. Light passing through red or blue filters produced good growth but light from green or yellow filters inhibited the growth. Singh and Saksena, (1983) mentioned that in case of V. volvacea light intensity showed no effect on mushroom yield but colour, texture and shelf life of mushroom grown under dark and diffused light conditions was better than the mushroom produced under bright light conditions. Fasidi, (1996) described that Volvariella esculenta (Mass) Singer, is able to grow at a temperature range of 20-40C (optimum = 35C) and pH range of 3-10 (optimum = 6.0) and a wide range of agricultural wastes for growth. Of these the rice straw and induced the widest mycelial extension and rice bran produced the highest mycelial density. The unfermented cotton waste compost produced the highest fruit body yield. Banik and Nandi, (2000) described that V. volvacea known as paddy straw mushroom grows well in humid and tropical environment, the biological efficiency of this was very low in comparison to oyster mushroom. Low productivity was a big hurdle for its commercial exploitation but the traditional substrate for this mushroom cultivation, with biogas residual slurry manure
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increased its production by producing fruit bodies bigger in size and higher in number and also increased the protein content significantly. The mineral nutrients viz., P, Ca, K, Fe, Cu, Zn and Mn were also increased. So, they concluded that supplementation of this bio manure for cultivation of tropical mushroom Volvariella volvacea may be a step towards its successful commercial exploitation. Zervakis et al., (2001) determined the influence of environmental parameters on mycelial linear growth of different mushroom crops including Volvariella volvacea in two different nutrient media in a wide range of temperature. V. volvacea grew faster at 35C, Wheat straw, peanut shells and particularly cotton gin-trash supported fast growth of V. volvacea. Jonathan et al., (2004) studies were conducted on the effects of temperature, pH, vitamins and plant hormones on the vegetative growth of Volvariella esculenta (Mass) Singer. They observed that, this mushroom had its optimum radial growth at 35C with mycelial extension of 85.0 mm and the pH that supported best growth was 6.0. Pyridoxine was the most utilizable vitamin with mycelial dry weight of 123 mg/30 cm3. They also observed that among the tested phytoharmones, 2, 4-D (10.0 ppm) stimulated the best growth of 150 mg/30 cm3 but, 0.1 ppm of GA3 supported poor growth (53.0 mg/30 cm3). Akinyele and Adetuyi, (2005) studied the effect of temperature variations on the growth of V. volvacea cultivated on various agricultural wastes singly and in various combinations. A pH range of 5.5 to 8.5 recorded the maximum mycelial
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yield and the highest mycelia weight was recorded at pH6.5 while poor mycelia growth of the mushroom and the least mycelia weight was recorded at pH 2.0. Highest mycelia growth of mushroom was recorded between 25C and 30C and the least mycelial dry weight of 0.5 mg obtained at 10C. 2.4. LIQUID FERMENTATION 2.4.1. Background Anon, (1954) while working in the Syracuse University, New York, conculede that it is possible to grow the highly priced mushroom Morchella hortensis, without any problem and at very low cost in water culture under constant motion. Atacador, (1967) reported that out of five edible mushrooms cultivated in liquid medium, V. volvacea gave the highest mycelial yield. The best medium for its production had pH 5, and contained 4% urea, 4% sucrose and best propagation period was 5 days to get maximum mycelial yield. Kostadinov et al., (1972) described a method of producing Pleurotus ostreatus mycelium in sub-merged culture which may be used as inoculation material (spawn) for production of sporophores on a substrate of crushed corn cobs. They also reported the most suitable nutritive medium was a combination of cane molasses with supplementation of NH4NO3, NH4SO4 and Potassium dihydrogen phosphate. Lin and Hu, (1972) reported that sugarcane baggase was found to be suitable and more economical for mushroom cultivation as straw compost. The
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dried product was amended by the wheat bran (10%), ammonium dihydrogen phisohate (1%), calcium chloride (5%) and water (60%) followed by heat treatment at 55C for 72 hours. Bukhalo, (1982) selected some species and strains of edible fungi for biomass production in submerged culture. Similarly Khan et al., (1982) reported that Pleurotus can be can grown in cane molasses medium and resulting biomass can be used as a source of protein rich food. Garo and Neelakanton, (1982) investigated the production of single cell protein by Aspergillus terrens by using alkali treated baggase as potential substrate. They obtained maximum biomass protein content of 20% by continuously shaking of culture for seven days at pH 4. Quimio, (1984) mentioned potential of using coconut water, first time as a liquid medium for mycelial production of Lentinus sajor caju and other edible mushroom, secondly as a routine agar medium for the growth of edible mushrooms. He observed that coconut water both from young and mature nuts can support the mycelial growth of edible mushrooms. After first week of incubation, 12gm of dry weight of Lentinus sajor- caju mycelia was harvested from 100ml of sterilized coconut water. When the coconut water was incorporated with agar medium it performed almost equally to the PDA medium. Trials with V. volvacea and some other mushrooms showed that coconut water agar medium can be used as a routine laboratory medium in place of potato dextrose agar medium.

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Mahmood, (1986) optimized the concentration of inorganic growth nutrients for the production of mycelial protein from rice polishing with Arachniotus spp. Maximum crude protein percentage was obtained in the shake medium containing calcium chloride 0.001%, magnesium sulphate 0.001%, and potassium dihydrogen phosphate 0.08%, after 72 hours of incubation at pH 4 and 30C temperature. Dey et al., (1995) investigated biosorptions of Pb2+, Cr6+, Cd2+ and Ni2+ by using live and dead fungal mycelia. Of the four fungi, namely Polyporus ostreiformis, Volvariella volvacea, Pleurotus sajor-caju and Phanerochaete chrysosporium, they observed total biosorption was effected in 6 days up to the Pb2+ concentration of 6 mg/l, with a specific uptake of 1.33 mg Pb2+ /g dry cell mass. The removal of other three metals varied between 28.8-73.3% from a medium containing 4 mg/l of each of the metals. Cai, et al., (1998) studied that the edible straw mushroom, Volvariella volvacea (V-14), produced -glucosidase when grown in liquid culture on a variety of carbon sources including cellulose, cellobiose, salicin, sorbose, lactose, esculin, cotton wool, and filter paper. They purified two cell-associated glucosidases, BGL-I and BGL-II, 32-fold and 23-fold, respectively, from extracts of cellulose-grown mycelium. The enzymes were found to be homogeneous and to have native molecular weights of 158 kDa (BGL-I) and 256 kDa (BGL-II) by gel filtration. Both isozymes displayed relatively broad pH optima with maximum reaction velocities recorded at pH 7.0 for BGL-I and pH 6.2 for BGL-II, and were
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rapidly denatured at temperatures of 60C and above. Isozyme activities were adversely affected by several reported -glucosidase inhibitors, various metal ions, and lignin-derived aromatic acids and aldehydes. Glucose production from microcrystalline cellulose by a commercial cellulase preparation was enhanced by 9.7% in reaction mixtures supplemented with BGL-II. Chen et al., (2004) isolated a Laccase (lac1) from culture fluid of Volvariella volvacea, grown in a defined medium containing 150 micro m CuSO4 subscript by ion exchange and gel filtration chromatography. RT-PCR analysis of gene transcription in fungal mycelia grown on rice straw revealed that, apart from during the early stages of substrate colonization, lac1 was expressed at every stage of mushroom developmental cycle defined in their study, although the levels of transcription varied considerably depending upon the developmental phase. 2.4.2. Liquid media used for fermentation Edwards, (1954) observed the growth of edible fungi in sub-merged culture, consisted of a liquid medium through which air was constantly bubbled. The rapid growth of mycelium was obtained from one cream and one white variety of cultivated mushroom Agricus compestris. He concluded that submerged culture might be much cheaper than ordinary mushroom culture. The mycelial biomass produced could be used for soups or flavouring of food items. Block, (1959) concluded that yields were high and no special production problems were encountered when mushroom mycelium were grown in submerged liquid culture with carbohydrates and nitrogen compounds with mineral salts.
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Torver, (1968) worked out a technology for the cultivation of mycelium of different species of higher fungi in liquid nutritive medium. He assumed that mycelium of some mushroom species produced in liquid culture would be used as spawn in mushroom cultivation. Anthony, (1977) reported that fungi can be successfully propagated on a wide variety of substrates. He found that protein contents of fungi were strain dependant and influenced by the growth conditions. Bukhalo et al., (1978) studied that Pl was successfully grown in liquid nutrient medium containing 10% red clover extract, sucrose, peptone and mineral salts on commercial scale. The fungus was cultivated on complex media containing potato waste and molasses. The rate of mycelial growth was greater on sub-merged culture than that in surface culture. Nagaso and Yoshikawa, (1978) observed that mycelial growth and yield was best with shaking at 100 cycles/minute rather than shaking at 90 or 120 cycles/ minute. 2.5. BIOCHEMICAL ANALYSIS 2.5.1. Nutrients contents Tumwasorn et al., (1980) reported 11.5% to 12% ash in the straw of different paddy varieties and crude fat contents of paddy straw before and after mushroom production were reported to be 0.5% and 0.2%, respectively.

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Vijaya nad Pandaya (1981) reported proximate composition of paddy straw mushroom. The value for moisture fat, crude protein, carbohydrates, crude fiber and ash were 92%, 0.21%, 2.19%, 1.15% and .99% respectively. Khowala and Sengupta, (1985) purified the enzyme, endo-alphamannanase, from culture filtrate of a mushroom Volvariella volvacea by acetone precipitation, ion-exchange chromatography (DEAE-Sephadex), and gel-

permeation chromatographies on Bio-Gel P-300 and on Sephacryl S-200 columns. They observed single protein band on sodium dodecyl sulfate-disc gel electrophoresis at pH 6.8 and has a molecular weight of approx. 56,000. It has no alpha- or beta-mannosidase activity and does not act on beta-gluco-or galactomannan. The enzyme shows maximum activity on baker's yeast alphamannan at pH 5.0 and at 55C, and is fairly stable between pH 3 and 6 and temperatures up to 50C. Enzyme activity is inhibited by Hg2+, sodium azide, iodoacetic acid, EDTA, and Ag+, in decreasing order. Kalisz et al., (1986) concluded that Agaricus bisporus was grown on defined liquid media with protein as sole source of carbon, nitrogen or sulpher and with these nutrients supplied in the form of glucose, ammonium or sulphate. The culture filtrates of Agaricus were tested for extracellular laccase activity. Constitutive laccase production was observed under all conditions tested. However, Agaricus laccase, though constitutive, was induced by protein and repressed by ammonium. No detectable extracellular laccase activity was found in tested cultures of Coprinus cinereus or Volvariella volvacea.
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Kishida et al., (1989) concluded that A (1--3)-beta-D-glucan branched by O-6 substitution (FCAP), obtained from the cold-alkali extract of the fruiting body of V. volvacea, exhibited potent growth-inhibitory activity against implanted tumors in mice. They suggested that the Volvariella glucan was structurally heterogeneous with regard to the distribution of branches, having less branched, moderately branched, and highly branched segments. Cheung (1996) analysed the mycelia caps and stalks of fruiting bodies of four edible mushrooms (Lentinus edodes, Lycophyllum shimeji, Pleurotus sajor-caju, and Volvariella volvacea) for their total dietary fiber contents.The TDF cotents of all of the mushrooms were considerably greater than those determined by using the Uppsala method. Mushroom mycelia had higher TDF values than did the fruiting bodies .The TDF composition of the mycelia and the TDF composition of the caps and stalks of the mushroom fruiting bodies were similar. Sugar composition reflected that -glucans were the major fiber polysaccharide with chitin, hemicelluloses, and polyuronides as minor ones. Phutela et al., (1996) screened two strains of Volvariella diplasia (Vd IIHR and Vd TNAU) and three of V. volvacea (Vv IARI, Vv MU and Vv TNAU) for the production of cellulases and xylanases. They observed Vd IIHR exhibited maximum activity of cellulases. This strain also showed maximum biomass production (5.8 g I- broth) and yield potential (BE 8.6%). The xylanase enzyme showed maximum enzyme activity (9.73 U mg-1 protein) in Vd TNAU strain. All the enzyme activities were the maximum in culture filtrates after 8 d of growth.
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Lin et al., (1996) crystallized Volvatoxin A2; an ion channel disturbed cardiotoxic and hemolytic protein from the edible mushroom, V. volvacea, by the vapor diffusion method using polyethylene glycol 4000 and ammonium sulfate in sodium acetate buffer pH 4.6. Zhi et al., (1998) purified a novel single chained ribosome inactivating protein (RIP) from fruiting bodies of the edible mushroom Volvariella volvacea. The mushroom RIP, designated volvarin, exhibited a potent inhibitory action on protein synthesis in the rabbit reticulocyte lysate system. It also exerted a deoxyribonuclease activity on super coiled SV-40 and demonstrated a strong abortifacient effect in mice. Wang et al., (1998) summarized existing information about mushroom lectins, with an emphasis on those from the species which have been most extensively characterized including various Agaricus species, Areanita

pantherina, Boletus satanas, Coprinus cinereus, Ganoderma lucidum, Flammulina velutipes, Grifola frondosa, Hericium erinaceum, Ischnoderma resinosum, Lactarius delerrimus, Laetiporus sulphureus, Tricholoma mongolicum and Volvariella volvacea. Immunomodulatory and antitumour/cytotoxic activities have been carried out on lectins from Agaricus bisporus, Boletus satanas, Flammulina velutipes, Ganoderma lucidum, Grifola frondosa, Tricholoma mongolicum and Volvariella volvacea. Chiu et al., (1998) explained that recalcitrant nature, persistence and toxicities of chlorophenols make them priority pollutants for treatment. He
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compared the ability of various fungi including (Armillaria gallica, A. mellea, Ganoderma lucidum, Lentinula edodes, Phanerochaete chrysosporium, Pleurotus pulmonarius, a Polyporus sp., Coprinus cinereus and Volvariella volvacea), and the spent mushroom substrate of P. pulmonarius (SMS) to remove pentachlorophenol (PCP) using a batch cultivation system. All these fungi showed active breakdown in addition to bio sorption as their PCP removal mechanisms. Mau, (1998) irradiated fresh common (Agaricus bisporus) and hightemperature mushrooms (A. bitorquis) with ultraviolet-C (UV-C) for 0, 0.5, 1, and 2 h at 12 C. Fresh common, shiitake (Lentinula edodes), and straw mushrooms (Volvariella volvacea) were irradiated with UV-B for 0, 0.5, 1, and 2 h at 12 C. After UV-C irradiation for 2 h, vitamin D2 contents in common and hightemperature mushrooms increased from 2.20 and 4.01 g/g of dry weight to 7.30 and 5.32 g/g, respectively. After UV-B irradiation for 2 h, vitamin D2 contents in shiitake and straw mushrooms increased from 2.16 and 3.86g/g to 6.58 and 7.58 g/g, respectively. The increase rates in shiitake and straw mushrooms were not as high as in common mushrooms. Yao et al., (1998) purified a novel single-chained ribosome-inactivating protein (RIP) with a molecular weight of 29,000 from fruiting bodies of the edible mushroom V. volvacea. The mushroom RIP, designated volvarin, exhibited a potent inhibitory action on protein synthesis in the rabbit reticulocyte lysate system with an IC50 value of 0.5 nM. It also exerted a deoxyribonuclease activity on supercoiled SV-40 DNA and demonstrated a strong abortifacient effect in mice.
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She et al., (1998) purified a novel lectin from the fruiting bodies as well as cultured mycelia of the edible mushroom V. volvacea. They observed that the lectin, designated as VVL, was a homodimeric protein and had no carbohydrate moiety, and its hemagglutinating activity was inhibited by thyroglobulin. The immunomodulatory activity was demonstrated by its potent stimulatory activity toward murine splenic lymphocytes. VVL also enhanced the transcriptional expression of interleukin-2 and interferon- by reverse transcriptase-polymerase chain reaction. VVL possessed a molecular structure distinct from other immunomodulatory proteins previously reported in the same fungus. Cai et al., (1999) described that the edible straw mushroom, Volvariella volvacea produced a multi component enzyme system consisting of endo-1, 4-bglucanase, cellobiohydrolyse, and glucosidase for the conversion of cellulose to glucose. Biochemical analysis of different culture fractions in cultures exhibiting peak enzyme production revealed that most of the endoglucase was present either in the culture filtrate (45.8%) of the total or associated with the insoluble pellet fraction remaining after centrifugation of homogenized mycelia (32.6%). Cellobiohydrolyse distributed with 58.9% of the total enzyme present in cultural filtrates and 31 % associated with the pellet fraction. Whiteford, (2000) described the types, economic significance and methods of production of the principal cultivated mushrooms are described in outline. These organisms are all less than ideal for conventional genetic analysis and breeding, so molecular methods afford a particular opportunity to advance our
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understanding of their biology and potentially give the prospect of improvement by gene manipulation. They also described the gene sequences isolated from the paddy straw mushroom V. volvacea and many other mushrooms. Isiloglu et al., (2001) determined the concentrations of Cu, Cd, Co, Ni, Mn, Pb, Zn and Fe in 66 samples of mushroom fruiting bodies, representing seven species, mainly all edible, were determined by atomic absorption

spectrophotometry. The mushrooms were collected from near roads and inner parts of forest and lawns in Balikesir in the north western part of Turkey. The results indicated that the Fe level in the species Volvariella speciosa (Fr.) Sing. from near the road was the highest with a mean of 6990 mg/kg. The Cd was accumulated mostly by Lactarius sanguifluus (Paulet: Fr.) Fr. and V. speciosa from near road with a mean of 1.60 mg/kg. Liu et al., (2001) studied the antiproliferative activity of a fungal lectin (VVL) isolated from the mushroom, V. volvacea. It was observed that VVL did not exert ribosome-inactivating activity or induce any changes in the expression of cyclins A, D1, and E. However, it did activate the expression of cyclin kinase inhibitors, namely p21, p27, p53, and Rb, in a dose-dependent manner. Flow cytometric analysis demonstrated an accumulation of cells in the G2/M phase in a time- and dose-dependent manner, indicating that VVL arrested cell proliferation by blocking cell cycle progression in the G2/M phase. Fu et al., (2002) studied the antioxidative potency of commercially available mushrooms in Taiwan. The order suggested by him of inhibitory activity
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of mushroom extracts on oxidation in emulsion system was Agaricus bisporus > Hypsizigus marmoreus > Volvariella volvacea > Flammulina velutipes > Pleurotus eryngii > Pleurotus ostreatus > Hericium erinaceus > Lentinula edodes. In the thermal oxidative stability test, using lard, the order of antioxidative activity of test materials showed similar tendencies, except for the extract of Lentinula edodes. Wang and Liu, (2002) isolated the lectin from the dried fruiting bodies of the mushroom Agrocybe cylindracea a heterodimeric lectin with a molecular weight of 31.5 kDa and displaying high hemagglutinating activity. The larger and the smaller subunits resembled Agaricus bisporus lectin and fungal

immunomodulatory protein from Volvariella volvacea respectively in N-terminal sequence. The lectin exhibited potent mitogenic activity toward mouse splenocytes. The hemagglutinating activity of the lectin was inhibited by lactose, salicyclic acid and inulin. Ahlawat et al.,(2005) isolated and evaluated the six parent strains and 11 monosporous isolates from two strains of V. volvacea for their enzymes induction level, substrate colonization and yield potential. They observed that all parent strains preferred wild grass over wheat straw and paddy straw for vegetative growth along with highest level of induction of -glucosidase and xylanase. Mushroom yield in parent strains was related to the level of various lingocellulolytic enzymes produced on a substrate. Monosporous isolates and

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parent strains produced more cellulases at 8 and 10 d, while xylanase, laccase and polyphenol oxidase were more at 10 or 13 d growth on paddy straw. 2.5.2. Medical effects Lin et al., (1974) isolated the cardiotoxic protein volvatoxin from Volvariella volvacea and another cardio toxic protein isolated from the Flammulina velutipes (Curt. ex Fries Sing), which is widely eaten in the Orient. This protein is called flammutoxin. Flammutoxin has three biological activities similar to those of volvatoxin, direct hemolytic action against human group O blood cells; ability to cause a writhing reaction with a delay before onset; and effect on the electrocardiogram at a dose of 0.25 mg per kg body weight, causing depression of the ST segment and inversion of the T wave. Fassold et al., (1976) isolated Volvatoxin A, present in the mushroom Volvariella volvacea, causes a competitive, dose and time dependent inhibition of the Ca2+ accumulating activity of a sarcoplasmic reticulum rich microsomal fraction isolated from guinea pig ventricular muscle. They explained why volvatoxin A increases the diastolic resting tension in heart muscle. Kishida et al., (1992) purified a potent antitumor-active branched (1 3)-D-glucan (VVG) from fruiting body of Volvariella volvacea. They observed that conversion of the glucosyl groups substituted at 0-6 atoms of the (1 3)-linked D-glucose residues into the corresponding polyhydroxyl groups gave significant enhancement of the original activities, whereas deletion of the polyhydroxyl groups resulted in a great reduction of the activity. When D-glucose residues of
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the branches were modified to the 3,6-anhydro D-glucose residues. They observed that the previous findings that, besides the conformation of (1 3)--glucan backbone, the molecular shape and the distribution pattern of the substituted groups located outside the backbone chains, must also play an important role in exhibiting anti tumor action. Chiu et al., (1995) described that the straw mushroom Volvariella volvacea was one of the common edible mushrooms cultivated in Southeast Asian countries. It has been reported to produce a hypotensive response in animals including humans. An aqueous extract of the mushroom (SME) was prepared and given through intravenous injections to normotensive rats. They examined the effects of SME on the kidney function of water-loaded rats and on isolated tissue preparations of the tail artery and right atrium. An IV injection of SME produced a hypotensive effect in rats with an ED50 of 25 mg dry weight/kg body weight. This hypotensive effect of SME was attenuated or blunted in the presence of hexamethonium, phentolamine, pyrilamine and cimetidine suggesting the involvement of the -adrenergic component of the autonomic system and/or histaminergic stimulation. They also observed that SME did not increase urinary excretion nor sodium diuresis. It produced positive chronotropic and inotropic effects on isolated right atria and induced contraction of isolated tail artery strips. This latter contractile response was inhibited by antagonists of serotonin and adrenoceptor, ketanserin and phentolamine respectively. Partial purification using dialysis and liquid chromatography revealed that the hypotensive active
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substances had molecular masses between 8000 and 12000 dalton. These substances were heat stable and resistant to trypsin digestion. In view of the similarity in blood pressure and cardiovascular response, SME might contain serotonin-like substances. Fasidi and Kadiri, (1995) studied the toxicological aspects of seven Nigerian mushrooms, namely, Chlorophyllum molybditis (Mayer ex. fr.) Masse, Cortinarius melliolens Fries, Lentinus subnudus Berk, Pleurotus tuber-regium (Fries) Singer, Termitomyces robustus (Beeli) Heim, Tricholoma lobayensis Heim and Volvariella esculenta (Mass) Singer. Amatoxin spot test and chromatographic screening of the mushrooms revealed the absence of amatoxins and phallotoxins because none of the mushroom extracts tested killed the experimental rats. Cheung, (1996) fed male Sprague-Dawley rats with two semisynthetic diets supplemented with 2% cholesterol and 1% -glucan type extracellular polysaccharide isolated from two liquid cultures of straw mushroom (Volvariella volvacea) mycelium containing different carbon sources. They concluded that both mycelial extracellular polysaccharides exhibited hypocholesterolemic activity in rats with alimentary-induced hypercholesterolemia. Fasidi and Akwakwa, (1996) studied the growth requirements of Volvariella speciosa (Fr. ex. Fr.) Sing. were studied. They observed all the tested carbohydrates except cellulose significantly enhanced mycelial growth and Mannitol was the most utilized, followed in order by fructose and maltose. All the organic and inorganic nitrogen sources investigated significantly improved
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growth. Calcium, magnesium, sodium, potassium and zinc significantly enhanced growth whereas hormones and vitamins did not. 2.5.3. Bioconversion of lignocellulosic wastes Buswell et al., (1996) described that edible mushroom cultivation was one of the most economically-viable processes for the bioconversion of many types of lignocellulosic wastes. According to him Lentinula edodes, Volvariella volvacea and Pleurotus sajor-caju were three important commercially cultivated mushrooms which exhibit varying ability to utilize different lignocellulosics as growth substrate he explained that V. volvacea, which preferred high cellulose-, low lignin-containing substrates produces a family of cellulolytic enzymes including at least five endoglucanases, five cellobiohydrolases and two glucosidases, but none of the recognized lignin-degrading enzymes. Rajor, (1996) described that the Sawdust, a bulky waste generated by wood processing industries, has very few profitable and eco friendly uses and has a problem of proper disposal. They observed that treatment with the fungus V. volvacea and a dilute solution of urea converted sawdust from a phytoinhibitory material to a phytostimulatory soil conditioner. Analyses of the major biopolymers of sawdust after fungal treatment indicated the decrease in the levels of cellulose, hemicellulose and lignin. Chiu et al., (2000) described that mushroom cultivation was a direct utilization of their ecological role in the bioconversion of solid wastes generated from industry and agriculture into edible biomass, which could also be regarded as
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a functional food or as a source of drugs and pharmaceuticals. They concluded that, this was very true for Lentinula edodes, Volvariella volvacea, and Ganoderma lucidum, which were commonly consumed in Asian communities but are now gaining popularity worldwide. Besides the conventional method, strain improvement could also be exploited by protoplast fusion and transformation. Biodiversity is the key contribution to the genetic resource for breeding programs to fulfill different consumer demands. Spent mushroom compost, a bulky solid waste generated from the mushroom industry, however, could be exploited as a soil fertilizer and as a prospective bioremediating agent. Datta and Chakravarty, (2001) studied comparative utilization of lignocellulosic componnts of paddy straw by Tricholoma lobayense and Volvariella volvacea. They observed that T. lobayense degraded the lignin more actively till the end of spawn run phase. However, V. volvacea could utilize cellulose and hemicellulose throughout spawn run and cropping phases, but was unable to utilize lignin at any stage. Yadav et al., (2002) devised an optimized protocol for the bioconversion of eucalyptus bark. It comprised: (i) mechanical reduction in bark size to 0.5-3.0 cm, (ii) moistening to 60-65%, (iii) fortification with ligninase-rich fungus Volvariella sp. (S-1) and 2% urea and (iv) maintenance of this composting mix under aerobic and ambient condition for 14-15 weeks. The resulting bark soil conditioner (BSC), with physico-chemical and microbial properties which would enrich soil fertility/productivity.
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2.5.4. Uptake of heavy metals Purkayastha and Mitra, (1992) observed the uptake of a few metals by V. volvacea during submerged growth of the organism in sub lethal concentration of each metal salt. They concluded that the uptake of Pb2+ and Hg2+ was 5 and 5.23 micrograms g-1 respectively while that of Cu2+ was 500 micrograms g-1 under experimental conditions. Treatment of spawned substrate separately with different metal salts showed maximum and minimum uptake of Pb2+ (100 micrograms g-1) and Cd2+ (2.93 micrograms g-1) respectively by sporocarps. All metal salts at test concentrations reduced biological efficiency of sporocarp production but markedly by Co2+. Cd2+ and Co2+ were highly toxic to mycelia and sporocarps respectively and the uptake of Cu2+ by mycelia and Pb2+ by sporocarps were highest among the five metals tested. 2.6. GENETICS Boekhout and Enderle, (1996) designated a neotype for Volvariella gloiocephala (DC. Fr.) Boekhout & Enderle, to serve as a representative collection for the current concept of this species, that generally was considered conspecific with V. speciosa (Fr.: Fr.) Kummer. Chiu and Moore, (1999) determined the electrophoretic karyotype of the Chinese straw mushroom, Volvariella volvacea. They observed that haploid strain V34 of V. volvacea has 15 chromosomes ranging in size from 1.4 to 5.1 Mb. No chromosomal polymorphism in terms of size and number was seen in either of two growth stages: vegetative mycelia or fruit-body gill tissues. They also prepared
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DNA fingerprints by the arbitrarily-primed polymerase chain reaction. Variation in DNA fingerprints was evident in protoplast regenerants derived from the same vegetative mycelium. Thus the haploid mycelium of strain V34 was heterokaryotic but the bulk genotype is stable during fruit body development. They overviewed the known mechanisms to generate genetic variation and proposed a novel mechanism that could account for the 1:1 segregation ratio of self-fertile to selfsterile progeny regularly obtained from selfed Volvariella volvacea fruit bodies. Ding at el., (2001) isolated an endoglucanase, EGI, from fluid of Volvariella volvacea grown on crystalline cellulose by ion exchange and gel filtration chromatography and preparative PAGE .Their work was aimed at generating improved strains of V. volvacea with enhanced growth, substrate conversion capacity and higher production capacity. Guo et al., (2002) used PCR technique for amplifying THP gene in an unknown vector with primer AFP1 and AFP2. Then THP gene was ligated to pGEM T-Vector to be the plasmid pGTHP4. Then isolated and purified big fragment containing promoter with the Agarose Gel DNA Extraction Kit, and also purified the small fragment containing THP gene from 1% agarose gel with the Agarose Gel DNA Extraction Kit. The big fragment and the small fragment were ligated at Nco I digested cohesive-end. The ligation product was re-ligated to be cyclic plasmid by addition to a specific adapter, resulting in the pCTH823, a expression vectorof V. volvacea.

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Shi et al., (2002) assessed aqueous extracts of the sporophores of eight mushroom species for their ability to prevent H2O2-induced oxidative damage to cellular DNA using the single-cell gel electrophoresis assay. They observed the highest genoprotective effects with cold (20C) and hot (100C) water extracts of Agaricus bisporus and Ganoderma lucidum fruit bodies, respectively. No protective effects were observed with mushroom derived preparations (MDPs) from Lentinula edodes, Pleurotus sajor-caju, and Volvariella volvacea. They concluded that some edible mushrooms represent a valuable source of biologically active compounds with potential for protecting cellular DNA from oxidative damage.

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