Cultivation of Pleurotus ostreatus on weed plants

Bioresource Technology 98 (2007) 2723–2726 Cultivation of Pleurotus ostreatus on weed plants Nirmalendu Das a,¤ , Mina Mukherjee b a b Department of Botany, Durgapur Government College, Durgapur 14, India Indian Institute of Chemical Biology, 4, Raja S.C. Mullick Road, Kolkata 700 032, India Received 1 September 2006; received in revised form 18 September 2006; accepted 18 September 2006 Available online 11 December 2006 Abstract Oyster mushroom, Pleurotus ostreatus (Jacq.:Fr.) Kumm. ITCC 3308 (collected from Indian Type Culture Collection, IARI, New Delhi, India, 110012) was grown on dry weed plants, Leonotis sp, Sida acuta, Parthenium argentatum, Ageratum conyzoides, Cassia sophera, Tephrosia purpurea and Lantana camara. Leonotis sp. was the best substrate in fruit body production of P. ostreatus when it was mixed with rice straw (1:1, wet wt/wet wt) for mushroom cultivation. The fruiting time for P. ostreatus was also less on Leonotis sp. than on any other weed substrates tested in the present investigation. T. purpurea was the least suited weed for oyster mushroom cultivation. The main problem of oyster mushroom cultivation on weed substrates was found to be low yield in the second Xush that could be overcome by blending weed plants with rice straw. The protein contents of the fruit bodies obtained from Cassia sophera, Parthenium argentatum and Leonotis sp. were not only better than rice straw but also from the rice straw supplemented weeds.  2006 Elsevier Ltd. All rights reserved. Keywords: Cultivation; Mushroom; Pleurotus ostreatus; Rice straw; Weed 1. Introduction Oyster mushroom (Pleurotus spp.) cultivation has increased tremendously throughout the world during the last few decades (Chang, 1999; Royse, 2002). This mushroom accounted for 14.2% of the total world production of edible mushroom in 1997 (Chang, 1999). Although commonly grown on pasteurized straw of wheat or rice, they can be cultivated on wide variety of substrates that contain lignin and cellulose. Oyster mushroom cultivation can play an important role in managing organic wastes which have become problematic for disposal. The word ‘weed’ is generally used for the undesirable plants. Suitable utilization of weeds is a subject of interest as most weeds are not used even as fodder due to the presence of lignin and anti-metabolites like phenolics, glycosides, Xavonoides and other compounds (Fianu et al., 1981). Disposal of these plants through burning causes environ- mental pollution as they release high level of CO2 as well as it is the cause of unnecessary wasting of large amount of organic materials (Croan, 2000). Oyster mushrooms (Pleurotus spp.) can produce fruit bodies on straws of rice (Oryza sativa), wheat (Triticum vulgare), ragi (Elucine coracana), bazra (Pennisetum typhoides), sorghum (Sorghum vulgare), maize (Zea mays) (Bano et al., 1987; Goswami et al., 1987), woods of poplar (Populus robusta), oak (Quercus leucotrichopora), horse chest nut (Aesculus indica), Acasia sp. (Pant et al., 1987), chopped banana pseudostem (Singh and Tandon, 1987), cotton stalk, pea shells and poplar saw dust (Philippoussis et al., 2001; Zervakis et al., 2001). In this paper we report that weeds can be used as the substrate for cultivation of oyster mushroom (Pleurotus ostreatus). 2. Methods 2.1. Mushroom strain * Corresponding author. Present address: A.B.N. Seal College, Cooch Behar, West Bengal, 736 101, India. Tel.: +91 358 2226112. E-mail address: nirmalendus@yahoo.co.uk (N. Das). 0960-8524/$ - see front matter  2006 Elsevier Ltd. All rights reserved. doi:10.1016/j.biortech.2006.09.061 Pleurotus ostreatus (Jacq.:Fr.) Kumm. (ITCC 3308) was collected from Indian Type Culture Collection, Division of 2724 N. Das, M. Mukherjee / Bioresource Technology 98 (2007) 2723–2726 Mycology and Plant Pathology, Indian Agricultural Research Institute, New Delhi, 110012, India and was maintained on potato-dextrose agar (pH 7.0) containing 20% potato extract; 2% dextrose; 2% agar as reported earlier (Das and Mukherjee, 1996). 2.2. Substrate preparations Weed plants were collected after seed formation from local Welds of Durgapur, West Bengal, India. The plants were Leonotis sp. (Lamiaceae), Sida acuta (Malvaceae), Parthenium argentatum (Asteraceae), Ageratum conyzoides (Asteraceae), Cassia sophera (Caesalpiniaceae), Tephrosia purpurea (Papilionaceae) and Lantana camara (Verbenaceae). The plants were completely dried in the sun after harvesting. Rice straw (var. MINIKIT) was obtained from a local farm at Durgapur, West Bengal, India and it was approximately three months in storage after harvesting. The branches of weeds without leaves or rice straw were chopped into small pieces (1–2 cm), weighed and soaked in water for overnight. Extra water present in the substrates was drained oV and the substrates were spread on the surface of a clean blotting paper and air dried for 15 min to remove the excess water. No heat treatment of substrates was done. Wet (600 g) substrate (»85% moisture), which was either individual weed or combination of weed and rice straw (1:1) was mixed with 20% spawn (wet wt/ wet wt). The spawned substrates were then put into 30 cm £ 42 cm polythene bags. The bags were tightly closed and pin holes were made on the surfaces. The bags were subsequently kept in a spawn running room at 25 § 1 °C under dark condition until primordia was formed. After primordia formation, large holes were made in the polythene bags to allow the normal development of fruit bodies. The bags were then kept at 22 § 1 °C with a 12 h photoperiod (1500–2000 Lux) and 85–90% relative humidity. Adequate ventilation was provided to prevent increase of CO2 concentration. Mushrooms were harvested in clusters from the substrates manually, three days after primordia initiation. Biological eYciencies (BE) of mushrooms were calculated from the ratios of weight (kg) of fresh mushrooms harvested per kg of dry substrates. 2.3. Preparation of fruit body extract and measurement of protein Fresh fruit bodies (28 g) after third day of primordia formation were disrupted by being crushed with acidwashed sand in mortar and pestle. The tissue was then extracted with 100 ml 20 mM imidazole buVer containing 1 mM EDTA, 2 mM 2-mercaptoethanol, 2 mM PMSF (pH 7.8). Unbroken cells and cell debris were removed by centrifugation at 32,000g for 30 min and the supernatant was used for analysis of protein. Protein concentrations were determined by the Bradford method (Bradford, 1976). 2.4. Statistical analysis of experimental data The data obtained were analyzed using SPSS 13.0 version Software. Mean and standard deviations of 15 replications (3 sets £ 5 batches) were calculated in each of the substrates. The means were sorted in decreasing order to show the absolute ranking of biological eYciencies of diVerent substrates. ‘Student’s t-test’ was performed at 5% level to check the level of signiWcance of the diVerences between two consecutive substrates. A “statistical rank” data column has been provided to indicate whether there is statistically signiWcant diVerences between the results in each set of substrates. Data followed by the same letter in statistical rank column indicates that there is no signiWcant diVerence at 5% level according to ‘Student’s t-test’. All experiments were carried out using 15 replicates for each substrate. 3. Results and discussion 3.1. Fruiting eYciency of P. ostreatus on weeds Biological eYciency (BE) of mushroom production and time for primordia initiation on diVerent substrates are given in Tables 1 and 2. In the Wrst Xush of production, biological eYciency was better on Leonotis sp. than that with rice straw. However, the diVerence was apparent, not statistically signiWcant as both the substrates belong to same statistical rank (Table 1). Biological eYciency increased signiWcantly (P < 0.05) in comparison to rice straw or Leonotis sp. when both the substrates were mixed in 1:1 proportion for mushroom cultivation. Biological eYciency of the mushroom was found to be highest on combination substrates of rice straw and Leonotis sp. than on any weed or blended substrates not only in individual Xush but also in accumulated conditions (Tables 1 and 2). Among the seven weeds tested for mushroom growth, T. purpurea was found to be the least responsive in terms of biological eYciency (Tables 1 and 2). However, addition of rice straw to T. purpurea showed an ameliorating eVect on fruit body production when used as mushroom substrate. Here the biological eYciency of combination substrate uplifted to statistical rank ‘C’ from ‘J’ of the individual T. purpurea (Table 2). Other weeds showed a diVerential response but the trend of response in terms of biological eYciency was obvious when those were combined with rice straw (Table 2). In second Xush the combination of weed and rice straw blends showed higher biological eYciency than any of the individual weed (Table 1). All the combined substrates in second Xush belong to same statistical rank along with rice straw indicate that higher biological eYciency was due to the blending of rice straw with weeds. The increase of yields in the second Xush might be due to higher water retention capacity of the combined substrates (particularly for rice straw) than that of individual weed substrate (data not shown). Among the diVerent substrates primordia was formed most rapidly (9.66 days) either on Leonotis sp. or 2725 N. Das, M. Mukherjee / Bioresource Technology 98 (2007) 2723–2726 Table 1 Biological eYciency of the cultivation of Pleurotus ostreatus mushroom (fresh mushroom in kg/kg dry substrate) on non-heat treated rice straw and weed species Biological eYciency in Wrst Xush (kg/kg substrate) Biological eYciency in second Xush (kg/kg substrate) Substrate Mean Std. p-value Statistical Substrate Deviation rank Meana Std. p-value Statistical Deviation rank Rice straw + Leonotis sp (1:1) Leonotis sp. Rice straw Rice straw + Sida acuta (1:1) Rice straw + Cassia sophera (1:1) Ageratum conyzoides 0.943 0.802 0.771 0.749 0.737 0.0250 0.0349 0.0405 0.0240 0.0324 <0.05 >0.05 >0.05 >0.05 A B B B B Rice straw + Leonotis sp. (1:1) Rice straw Rice straw + Cassia sophera (1:1) Rice straw + Tephrosia purpurea (1:1) Rice straw + Sida acuta (1:1) 0.447 0.437 0.435 0.431 0.421 0.0226 0.0226 0.0119 0.0329 0.0181 >0.05 >0.05 >0.05 >0.05 A A A A A 0.696 0.0159 <0.05 C 0.411 0.0146 >0.05 A Sida acuta 0.687 0.0247 >0.05 C 0.411 0.0128 >0.05 A Rice straw + Ageratum conyzoides (1:1) Rice straw + Lantana camara (1:1) Rice straw + Tephrosia purpurea (1:1) Lantana camara Rice straw + Parthenium argentatum (1:1) Cassia sophera Parthenium argentatum Tephrosia purpurea 0.649 0.0285 <0.05 D 0.405 0.0125 >0.05 A 0.647 0.0229 >0.05 D Rice straw + Ageratum conyzoides (1:1) Rice straw + Lantana camara (1:1) Rice straw + Parthenium argentatum (1:1) Leonotis sp. 0.222 0.0231 <0.05 B 0.642 0.0224 >0.05 D Sida acuta 0.219 0.0133 >0.05 B 0.638 0.605 0.0218 0.0196 >0.05 <0.05 D E Ageratum conyzoides Cassia sophera 0.189 0.183 0.0110 0.0171 <0.05 >0.05 C C 0.515 0.480 0.229 0.0229 0.0248 0.0299 <0.05 <0.05 <0.05 F G H Lantana camara Parthenium argentatum Tephrosia purpurea 0.168 0.113 0.000 0.0101 0.0111 0.0000 <0.05 <0.05 <0.05 D E F a a Means of 15 replications of each substrate were ranked according to Student’s t-test at 5% level. Table 2 Biological eYciency of the cultivation of Pleurotus ostreatus mushroom (fresh mushroom in kg/kg dry substrate) on non-heat treated rice straw and weed species Substrate Rice straw + Leonotis sp. (1:1) Rice straw Rice straw + Cassia sophera (1:1) Rice straw + Sida acuta (1:1) Rice straw + Tephrosia purpurea (1:1) Rice straw + Ageratum conyzoides (1:1) Rice straw + Lantana camara (1:1) Leonotis sp. Rice straw + Parthenium argentatum (1:1) Sida acuta Ageratum conyzoides Lantana camara Cassia sophera Parthenium argentatum Tephrosia purpurea Biological eYciency of mushroom accumulating 1st and 2nd Xush (kg/kg substrate) Time to primordia initiation Meana Std. Deviation Statistical rank 1st Flush 2nd Flush 1.390 1.208 1.171 1.170 1.073 0.0307 0.0544 0.0374 0.0299 0.0213 <0.05 >0.05 >0.05 <0.05 A B B B C 9.66 15.33 17.66 10.00 13.66 16.66 22.33 25.66 19.00 21.66 1.060 0.0378 >0.05 C 11.66 18.33 1.057 1.024 1.01 0.0324 0.0485 0.0275 >0.05 <0.05 >0.05 C D D 13.66 9.66 15.00 20.66 17.66 22.00 0.906 0.885 0.806 0.696 0.593 0.229 0.0287 0.0223 0.0256 0.0294 0.0304 0.0299 <0.05 <0.05 <0.05 <0.05 <0.05 <0.05 E F G H I J 10.00 11.33 15.66 18.66 16.66 14.33 18.00 18.33 22.66 24.33 24.66 – p-value – No fruit body was detected. a Means of 15 replications of each substrate were ranked according to Student’s t-test at 5% level. Leonotis sp. supplemented with rice straw substrate in the Wrst Xush. Primordia formations on those two substrates were also found to take less time in the second Xush as compared to other substrates used for mushroom cultivation (Table 2). 3.2. Protein concentrations in fruit bodies of P. ostreatus Protein concentration in the fruit body was highest when mushrooms were cultivated on C. sophera and P. argentatum (Table 3). However, supplementation of those two 2726 N. Das, M. Mukherjee / Bioresource Technology 98 (2007) 2723–2726 Table 3 Protein values of Pleurotus ostreatus fruit bodies in two Xushes grown on non-heat treated rice straw and weed species and harvested after three days of primordia formation Substrate Protein mg/g of fresh fruit body Mean Std. Deviation p-Value Statistical rank Cassia sophera Parthenium argentatum Leonotis sp. Rice straw + Leonotis sp. Ageratum conyzoides Lantana camara Rice straw Rice straw + Ageratum conyzoides Rice straw + Sida acuta Rice straw + Cassia sophera Rice straw + Tephrosia purpurea Sida acuta Rice straw + Lantana camara Rice straw + Parthenium argentatum Tephrosia purpurea 10.85 10.13 8.44 7.52 7.23 6.94 6.7 6.52 6.34 6.09 6.08 5.8 5.79 5.75 5.2 0.96 1.05 0.82 0.12 0.84 0.73 0.82 0.59 0.23 0.68 0.58 0.68 0.42 0.6 0.72 >0.05 <0.05 <0.05 <0.05 >0.05 >0.05 >0.05 <0.05 <0.05 >0.05 >0.05 >0.05 >0.05 <0.05 A A B C D D D D E F F F F F G weeds with rice straw resulted in decrease in protein level of the fruit bodies. The protein concentration of the fruit bodies derived from Leonotis sp. singly showed better results than the rice supplemented weeds. Except T. purpurea, the protein content of the fruit bodies obtained from diVerent weeds were better than that obtained from rice straw supplemented weeds (Table 3). 4. Conclusion Selective weeds can be used successfully as substrates for oyster mushroom cultivation. Weeds are not only proved as the alternative substrate for oyster mushroom cultivation, they also can signiWcantly increase the protein content and reduce the production time. Supplementation of weed substrate with rice straw increases the accumulated biological eYciency of mushroom, stimulating mainly the production in the second Xush. In the present investigation Leonotis sp. has been identiWed as the best substrate for oyster mushroom cultivation with respect to biological eYciency and fruiting time. Therefore, oyster mushroom cultivation proves to be a highly eYcient method for disposing of weed plants as well as producing protein-rich food. Acknowledgements This work was Wnancially supported by University Grants Commission, New Delhi, India. We are grateful to Prof. S. 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