OYSTER MUSHROOM CULTIVATION

Introduction


‘Oyster mushroom’or ‘Dhingri’ as refered in India is a basidiomycetes and belongs to the genus ‘Pleurotus’. It is lignocellulolytic fungus that grows naturally in the temperate and tropical forest on dead, decaying wooden logs, sometimes on drying trunks of deciduous or coniferous woods. It can also grow on decaying organic matter. The fruit bodies of this mushroom are distinctly shell, fan or spatula shaped with different shades of white, cream, grey, yellow, pink or light brown depending upon the species.   However, the colour of the sporophores is extremely variable character influenced by the temperature, light intensity and nutrients present in the substrate.  The name Pleurotus has its origin from   Greek word, ‘Pleuro’ means formed laterally or lateral position of the stalk or stem. The oyster mushroom is one of the most suitable fungal organism for producing protein rich food from various agrowastes without composting. 

This mushroom is cultivated in about 25 countries of far-east Asia, Europe and America.  It is the 3rd largest cultivated mushroom in the world. China alone contributes 88% of the total world production. The other major producing countries are South Korea, Japan, Italy, Taiwan, Thailand and Philippines. At present India produces annually 10,000 tones of this mushroom. It is popularly grown in the states of Orissa, Karnataka, Maharashtra, Andhra Pradesh, Madhya Pradesh, West Bengal and in the North-Eastern States of Meghalaya, Tripura Manipur, Mizoram and Assam.

A.  Advantages of growing oyster mushroom 
  1.  Variety of substrates Pleurotus mushroom can degrade and grow on any kind of agricultural or forest wastes, which contain lignin, cellulose and hemicellulose. 
  2. Choice of species Among all the cultivated mushrooms Pleurotus has maximum number of commercially cultivated species   suitable for round the year cultivation.  Moreover, variation in shape, colour, texture, and aroma are also available as per   consumer’s choice. 
  3. Simple Cultivation Technology Pleurotus mycelium can grow on fresh and fermented straw and it does not require composted   substrate for growth. Substrate preparation for oyster mushroom is very simple. Further this mushroom does not require controlled environmental conditions like A.bisporus as most of the species have very wide temperature, relatively humidity   and CO2 tolerance. 
  4. Longer shelf life Unlike white button mushroom, the oyster mushroom fruit bodies can be easily dried and stored. Dried oyster mushrooms can be instantly used after soaking in hot water for 5 to 10 minutes or it can be used in powdered form for several preparations.  Fresh mushrooms have a shelf life of 24-48 h even at room temperature. 
  5. High productivity The productivity of oyster mushroom per unit time is very high as compared to all other cultivated mushrooms.  One can harvest minimum of about 500 to 700 kg of fresh oyster mushroom from one ton of dry wheat or paddy straw in 45-60 days, while with the same quantity of straw only about 400-500 kg of white button mushrooms are obtained in 80-100 days (including period needed for compost preparation).  Yield of this mushroom can further be increased by supplementing the substrate with suitable nitrogen source viz., Soybean and cottonseed meal or by introducing high yielding cultures/strains. The present day cultivation technology of oyster mushroom is a result of various successive steps evolved throughout the world during   20th century.  A very primitive form of growing Pleurotus spp. was adopted by Lumberman in Europe during 19th century that involved collection of wood logs and stumps showing fructification in nature and keeping them in cool and moist places. 
First successful experimental cultivation of Pleurotus ostreatus was achieved in Germany by Falck in 1917. In India cultivation of P.flabellatus on paddy straw was reported by Bano & Srivastava in 1962 at CFTRI, Mysore.  Kaul and Janardhanan (1970) cultivated a white form of P. ostreatus on dried Euphorbia royleana (Thor) stems. Jandaik and Kapoor in 1974 could grow P. sajor-caju on various substrates including wheat and banana pseudostems.

B. The biology of oyster mushroom 

Visually the basidiocarps or fruit bodies of an oyster mushroom have three distinct parts - a fleshy shell or spatula shaped cap  (pileus), a short or long lateral or central stalk called stipe and long ridges and furrows underneath the pileus called gills or lamellae.  The gills stretch from the edge of the cap down to the stalk and bear the spores.  If a fruit body is kept on a paper directly (gills facing the paper) a dirty white or lilac deposition of powdery spores can be seen.   The spore print colour may be whitish, pinkish, lilac or grey.  The spores are hyaline, smooth and cylindric. The spores are heterothallic and germinate very easily on any kind of mycological media and within 48-96 h whitish thread like colonies could be seen.   The mycelium of most Pleurotus sp. is pure white in colour.  P. cystidiosus and P. columbinus forms coremia like stalked structures (asexual spores). Basidiospores on germination forms primary mycelium.  Fusion between two compatible primary mycelia develops into secondary mycelium, which is having clamp connections and it is fertile.  Primary mycelium is clampless and non fertile.

C. Varieties of oyster mushroom 

All the varieties or species of oyster mushroom are edible except P. olearius and P.nidiformis which are reported to be poisonous.  There are 38 species of the genus recorded throughout the world (Singer).  In recent years 25 species are commercially cultivated in   different  parts  of  the  world  which  are   as   follows: P. ostreatus, P. flabellatus, P. florida,  P.sajor-caju,  P. sapidus,  P. cystidiosus, P. eryngii,  P. fossulatus,  P. opuntiae, P. cornucopiae,   P.yuccae,  P. platypus, P. djamor,  P. tuber-regium,  P. australis, P. purpureo-olivaceus, P. populinus, P. levis, P. columbinus, P. membranaceus etc.(Fig. 1)

D. Cultivation 

The procedure for oyster mushroom cultivation can be divided into following four steps

  1. Preparation or procurement of spawn.
  2. Substrate preparation. 
  3. Spawning of substrate  
  4. Crop management.
1. Preparation or procurement of spawn 
The spawn preparation technique for oyster mushroom is similar to white button mushroom (A.bisporus).  One should have a pure culture of Pleurotus spp. for inoculation on sterilized wheat grain.  It takes 10-15 days for mycelial growth on grains.  It has been reported that Jowar and Bajra grains are   superior over wheat grains.  The mycelium of oyster mushroom grows very
fast on wheat grains and 25-30 days old spawn starts forming fruitbodies in the bottle itself.  It is therefore, suggested that the schedule for spawn preparation or spawn procurement should be planned accordingly.  Sometimes the mushroom farmers are using active mycelium growing on substrate for spawning fresh new oyster mushroom bags.  This method can be used on a small scale.   There are always chances of spread of contamination through infested straw by active mycelium spawning method so it is not advisable on large scale commercial cultivation.

2. Substrate preparation 
a.
Substrates for oyster mushroom and their nutrition quality  A large number of agricultural, forest and agro-industrial by-products are useful for growing oyster mushroom.  These by-products   or   wastes are rich in   cellulose, lignin   and hemicellulose. However, yield of oyster mushroom largely depends on the nutrition and nature of the substrate.  The substrate should be fresh, dry, free from mould infestation and properly stored.  The substrates exposed to rain and harvested immature with green chlorophyll patches inhibit the growth of Pleurotus mycelium due to the presence of competitor moulds.  Oyster mushroom can utilize a number of agro wastes including straws of wheat, paddy and ragi, stalks and leaves of maize, jowar, bajra and cotton, sugarcane bagasse, jute and cotton waste, dehulled corncobs, pea nut shells, dried grasses, sunflower stalks, used tea leaf waste, discarded   waste paper and synthetic   compost   of    button mushroom.   It can also be cultivated using industrial wastes like paper mill sludges, coffee byproducts, tobacco waste, apple pomace etc.   The cellulose and lignin contents are important components of any substrate as far as yield is concerned.   Cellulose rich substrates like cotton waste give better yields. Cellulose helps in more enzyme production, which is correlated, with higher yield.
b.
Methods of substrate preparation
The mycelium of Pleurotus is saprophytic in nature and it does not require selective substrate for its growth.   The mycelial growth can take place on a simple water treated straw but there are number of other cellulolytic moulds already present on straw which compete with Pleurotus mycelium during spawn run and also toxic metabolites secreted by these competitors hampers its growth. There are various methods to kill undesirable microorganism present in the straw to favour the growth of Pleurotus mycelium.  The substrate can be prepared by adopting different methods like steam pasteurization, hot water treatment, chemical sterilization technique, sterile technique and fermentation or composting. The choice of method will depend upon the scale of cultivation envisages and the facilities available. The growers may adopt any one of these method depending upon their need. The details of different methods are given below:

i. Steam pasteurization 
In this method prewetted straw is packed in wooden trays or boxes and then kept in a pasteurization room at 58-620C for four hours. Temperature of the pasteurization room is manipulated with the help of steam through a boiler.  Substrate after cooling at room temperature is seeded with spawn.  The entire process takes around 3-5 days.  This method is
adopted on a commercial scale by Zadrazil and Schneidereit in Germany.  There are various minor variations of this methods adopted in Europe. The tunnel prepared for pasteurizing compost/casing of button mushroom can be used for pasteurizing the straw for oyster. However, adequate boiler facility will be must.

ii. Hot water treatment 
The substrate after chopping (5-10 cm) as such in case of wheat straw is soaked in cold water overnight. The substrate is taken out and excess water is drained. Thereafter the straw is soaked in hot water for one hour where the temperature may be in the range of 65 to 70C. normally soaking the substrate in boiling water helps in achieving this temperature and no further heating may be necessary. It will be appropriate to check the temperature and standardize the conditions as per your location. Over boiling or over heating may not lead to proper result. Hot water treatment makes the hard substrate like maize cobs, stems etc. soft so the growth of mycelial takes place very easily. This method is not suitable for large-scale commercial cultivation.

iii. Chemical sterilization technique 
Various species of Trichoderma, Gliocladium Penicillium, Aspergillus and Doratomycs spp. are the common competitor fungi on the straw during oyster mushroom cultivation. If present on the straw during spawn run, they do not allow the growth of mushroom mycelium resulting in yield loss or complete crop failure.  When wheat straw or paddy straw is treated by steeping in a chemical solution of carbendazim 50% WP (37.5 ppm) and formaldehyde (500 ppm) for a period of 16-18 h, most of the competitor moulds are either killed or their growth is suppressed for 2540 days after spawning.  The technique, which was standardized at DMR, Solan in 1987, is as follows:   Ninety litres of water is taken in a rust proof drum (preferably of galvanized sheet) or G.I. tub of 200 litres capacity.  Ten to twelve kg of wheat straw is slowly steeped in water.  In another plastic bucket, Bavistin 7.5 g and 125 ml formaldehyde (37-40%) is dissolved and slowly poured on the already soaked wheat straw. Straw is pressed and covered with a polythene sheet. After 15 to 18 hour the straw is taken out and excess water drained. One can use a larger container or cemented tank of 1000-2000 liters for soaking more straw. The chemicals to be added can be calculated accordingly. The remaining solution can be used at least once again for chemical sterilization of straw without any further addition of chemicals. Some of the farmers fill the prewetted substrate in nylon net bags and press these bags in to the cemented tank containing chemical solution. This makes the process of taking out of substrate easier.

iv. Sterile technique 
The chopped substrate after soaking in cold water is put in heat resistant polypropylene bags and sterilize in an autoclave at 22 lb. pressure for 1-2 hours (depending upon the size of the bags) followed by spawning under aseptic conditions. This method is more suitable for research work rather then on large-scale commercial production. Some species of oyster like king oyster can be cultivated only by following this method of substrate preparation. Most commonly used bag size is 20 x 40 cm for 3 kg wet substrate.

v. Fermentation or composting without pasteurisation 
This method is a modification of composting technique used for white button mushroom.   It is most suitable for hard substrates like cotton stalks, maize stalks and leguminous stubbles, etc.   Composting should be done on a covered area or shed. Chop the substrate into 5-6 cm long pieces. Add ammonium sulphate or urea (0.5-1%) and lime (1%) on dry weight basis of the ingredients. Horse manure or chicken manure (10% dry weight basis) can also be used instead of nitrogenous fertilizers. Addition of lime improves the physical structure and pH of the compost. After wetting of straw, other ingredients are mixed to make a pile. Prepare a heap 75-90 cm high, about one meter wide. After 2 days of fermentation, turning of pile is done and 1% superphosphate and 0.5% lime is added. The compost will be ready after 6 days with three turnings.

vi   Fermentation or composting with steam pasteurization
Straw is prewetted and made into pile as described above. One per cent lime is added to adjust the pH at the time of stacking.  After two turning at two days interval, the substrate is filled in the tunnel and steam pasteurized as described in the steam pasteurization section above.

c. Substrate supplementation 
The nitrogen content in most of the substrates ranges between 0.5 to 0.8% and hence addition of organic nitrogen in the straw helps in getting higher yields. Some of the common supplements are wheat bran, rice bran, cottonseed meal, soybean cake, etc. Wheat bran and rice bran should be used at the rate of 10% while cottonseed meal, soybean cake and groundnut cake should be used at the rate of 3-6% on dry weight basis of the substrate. The supplements should be treated with 25 ppm carbendazim (250 mg in 10 litre water) + 500 ppm of formaldehyde for 48 hour. Supplements are thoroughly mixed with straw while spawning. Addition of supplements increases substrate temperature by 2-3C or even more and hence supplementation during summer season is not advisable.  However, during winter months though increased temperature is observed, which helps in quick spawn run. Excess nitrogen can attract mould infestation, which should be taken care of. In many cases, addition of supplements is counter productive due to the diseases. The better results of supplementation can be obtained in sterile techniques.   

3 .Spawning of substrate  
Freshly prepared (20-30 days old) grain spawn is best for spawning. The spawning should be done in a pre-fumigated room (48 h with 2% formaldehyde). The spawn should be mixed @ 2 to 3% of the wet weight of the substrate. One bag spawn of 500 g is sufficient for 20 kg of wet substrate or 5 kg dry substrate. Spawn can be mixed thoroughly or mixed in layers. Spawned substrates can be filled in polythene bags (60 x 45 cm) of 125-150 gauze thickness. Ten to 15 small holes (0.5-1.0 cm dia) should be made on all sides especially two to four holes in the bottom for draining excess water. Perforated bags give higher and early crop (4-6 days) than nonperforated bags. One can also use empty fruit packing cartons or boxes for filling substrate.  We can also make a block of the substrate by using compression machine. Polythene sheets of 200300 gauze thickness of 1.25 x 1.25 m are spread in rectangular wooden or metal box.  Spawned substrate is filled and the polythene sheet is folded from all the four sides and compressed to
make a compact rectangular block. It is taken out of the box and tied with a nylon rope. The block is incubated as such and after mycelium growth polythene sheet is removed.

4.  Crop management 
The spawned bags or blocks are kept in incubation room for mycelial growth at desirable temperature. Some of the Pleurotus species fruit at low temperature around 150C whereas other species fruit between 20-300C. The incubation temperature for mycelial spread, however, is around 250C for most of the species.

a. Incubation
Spawn bags can be kept on a raised platform or shelves or can be hanged in cropping room for mycelial colonization of the substrate. Although mycelium can grow between 10-30°C but the optimum temperature lies between 22-26°C. Higher temperature (more than 30C) in the cropping room will inhibit the growth and kill the mycelium. Daily maximum and minimum temperature of cropping rooms and beds should be recorded. The bed temperature is generally 24°C higher from the room temperature. Mycelium can tolerate very high CO2 concentration of 1.5-2.0%. During mycelial growth the bags are not opened and no ventilation is needed. Moreover, there is no need for any high relative humidity, so no water should be sprayed. However, some chemicals for control of flies can be sprayed on the walls. Similarly, water can be sprayed in the room or on the wall in case the environmental temperature is more than required.

b.Fruit body induction
Once the mycelium has fully colonized the substrate, it forms a thick mycelial mat and is ready for fruiting. Contaminated bags with mould infestation should be discarded while bags with patchy mycelial growth may be left for few more days to complete the spawn run. In no case bags should be opened before 16-18 days except in case of P. membranaceus and P. djamore var.roseus which forms fruit bodies within 10 days even in closed bags from small holes. Casing is not required in oyster mushroom cultivation. All the bundles, cubes or blocks are arranged on wooden platforms or shelves with a minimum distance of 15-20 cm between each bag in the tier. They can also be hanged. In case, small long bags are used these can be stacked horizontally or in a inclined manner one above the other. This method helps to accommodate more substrate in less space and therefore getting more production from the same area. Some workers have also used long cylinder for mushroom production. The poly bags can be tied at the base to get a circular base or alternatively bags with side in-folds can be used. Various cultural conditions required for fruiting are as follows.

i.  Temperature
Mycelial growth of all the Pleurotus spp. can take place between 20-30°C.  However, for fruiting different species have different   temperature   requirement. Depending upon the temperature requirement of a species they can be categorized into two groups-winter or low temperature requiring species (10-20°C) and summer or moderate temperature requiring species (16-30°C). Summer varieties can fruit at low temperature but the winter varieties will not fruit at higher temperature. They need a low temperature shock for inducing fruit body formation.
Commercial varieties which can be cultivated during summer are P. flabellatus, P. sapidus, P. citrinopileatus,  P. sajor-caju and P. eous.  Low temperature   requiring species are  P. ostreatus, P. florida, P. eryngii, P. fossulatus and P. cornucopiae. The growing temperature not only affects the yield but also the quality of produce. The pileus or cap colour of P. florida is light brown when cultivated at low temperature (10-15°C) but changes to white pale to yellowish at 20-25°C.  Similarly fruit body colour of P.sajor-caju when cultivated at 15-19°C is white to dull white with high dry matter content while at 25-30°C it is grayish brown.

ii. Relative humidity 
All the Pleurotus species require high relative humidity (70-80%) during fruiting. To maintain relative humidity, water spraying is to be done in the cropping rooms. During hot and dry weather conditions, daily 2-3 spray are recommended while in hot and humid conditions (monsoon) one light spray will be sufficient. The requirement of water spray can be judged by touching the surface of the substrate.  Spraying should be done with a fine nozzle to create a mist or fog in the cropping room.  It is desirable that mushrooms are harvested before water spray.  Ventilators and exhausts fans should be operated for air circulation so that the excess moisture from the cap surface evaporates. Sometimes fruit bodies give offensive smell due to the growth   of saprophytic bacteria on the wet cap surface; under such conditions 0.05% bleaching powder spray at weekly interval is recommended.

iii. Oxygen and carbon dioxide requirements The oyster mushroom mycelium can tolerate high carbon dioxide concentration during spawn run (up to 20,000 ppm or 2%) while it should be less than 600 ppm or 0.06% during cropping. Therefore sufficient ventilation should be provided during fructification. If the CO2 concentration is high the, mushrooms will have long stipe and small pileus. Mushrooms will appear like a mouth of trumpet.

iv. Light
Unlike green plants mushrooms do not require light for the synthesis of food.  They grow on dead organic plant material.  Light is, however, required to initiate fruit body formation. For primordia formation light requirement is 200 lux intensity for 8-12 hour. Inadequate light conditions can be judged by long stalk (stipe), small cap and poor yield. The colour of the pileus is also influenced by the light intensity and its duration. Fruit bodies raised in bright light are dark brown, grey or blackish coloured. If the light intensity is less than 100 lux the mushrooms will be pale yellowish. Thus both light and fresh air is necessary for formation of normal fruit body. It is not necessary to give the light and fresh air simultaneously but the required CO2 concentration and light requirement must be met for getting normal fruit body. It may be a good idea to give the fresh air just after water spray as it helps in removal of excess water from the surface of fruit bodies.

v. Hydrogen ion concentration (pH)
The optimum pH during mycelial colonization should be between 7.0 and 8.0. The pH of the water for spraying should be neither too acidic nor alkaline. Water should not contain harmful
salts or heavy metals. The mushrooms tend to accumulate various metals if present in substrate or water used. Rusted iron drums and tubs used for substrate treatment or storing water for spraying delay fructification due to presence of excess iron in the water.

F. Post Harvest Practices 
Mushrooms should   always be harvested before water spray.   The right stage for picking can be judged by the shape and size of fruit body.  In young mushrooms the edge of the cap is thick and cap margin is enrolled while the cap of mature mushroom as flat and inward curling starts.   It is advisable to harvest all the mushrooms at one time from a bag so that the next crop of mushrooms starts early.  After harvesting lower portion of the stalk with adhering debris should be cut using a knife. Stipe is kept short or almost non-existent, as it is hard and not liked by many consumers. Fresh mushrooms should be packed in perforated polythene bags for marketing. They can also be sun dried by spreading thinly on a cotton cloth in bright sunlight or diffused light. The dried produce with 2-4% moisture can be stored for 3 to 4 months after sealing properly.