the biofertilizer ' azotobacter ' inoculant and the strain of pleurotus eous ( apk-1 ) were obtained from agriculture college , tnau , madurai . in seed the substrate with 60% moisture was mixed with caco3 ( 2% w / w ) , rice bran ( 0.6% w / w ) and filled in spawn bottles . the mouths of the spawn bottles were plugged with cotton and they were sterilized in an autoclave at 121 for 2 h. after sterilization , the spawn bottles were inoculated with varying volumes of azotobacter broth culture ( 2 ml , 5 ml , 10 ml broth culture per bottle ) aseptically and mixed thoroughly with a sterile glass rod , followed by inoculation with a 7 mm block from an actively growing mother spawn of pleurotus eous . azotobacter preparation on jensen 's medium ( jensen , 1942 ) was used for separation of pure and single colonies . well - separated and distinct colonies were selected for study . single 7 mm bits from actively growing cultures of the mushroom , pleurotus eous or the competitor moulds , trichoderma viride or trichoderma reesi , were used to inoculate individual pda plates . the bacterial broth culture in log phase of growth was streaked on either side of the fungal agar block simultaneously with the inoculation of the fungus . the microbial inoculant as log phase broth culture was thoroughly mixed with the spawn substrate using a sterile glass rod in different volumes of 2 ml , 5 ml and 10 ml per bottle and the spawn bottles were inoculated with a 7 mm agar block from an actively growing p.eous plate . after plugging the mouth with cotton , the spawn bottles were incubated at 252. the mycelial proliferation was recorded on the 7 day post - incubation . carboxyl methyl cellulase ( cmcase ) ( padmaja and balagopal , 1985 ) , filter paper degrading enzyme ( fpase ) , -glucosidase ( ray et al . , 1993 ) and total nitrogen content ( jayaraman , 1988 ) of the substrate were estimated at 7th day and 14th day after incubation . the biofertilizer ' azotobacter ' inoculant and the strain of pleurotus eous ( apk-1 ) were obtained from agriculture college , tnau , madurai . in seed the substrate with 60% moisture was mixed with caco3 ( 2% w / w ) , rice bran ( 0.6% w / w ) and filled in spawn bottles . the mouths of the spawn bottles were plugged with cotton and they were sterilized in an autoclave at 121 for 2 h. after sterilization , the spawn bottles were inoculated with varying volumes of azotobacter broth culture ( 2 ml , 5 ml , 10 ml broth culture per bottle ) aseptically and mixed thoroughly with a sterile glass rod , followed by inoculation with a 7 mm block from an actively growing mother spawn of pleurotus eous . azotobacter preparation on jensen 's medium ( jensen , 1942 ) was used for separation of pure and single colonies . well - separated and distinct colonies were selected for study . single 7 mm bits from actively growing cultures of the mushroom , pleurotus eous or the competitor moulds , trichoderma viride or trichoderma reesi , were used to inoculate individual pda plates . the bacterial broth culture in log phase of growth was streaked on either side of the fungal agar block simultaneously with the inoculation of the fungus . the microbial inoculant as log phase broth culture was thoroughly mixed with the spawn substrate using a sterile glass rod in different volumes of 2 ml , 5 ml and 10 ml per bottle and the spawn bottles were inoculated with a 7 mm agar block from an actively growing p.eous plate . after plugging the mouth with cotton , the spawn bottles were incubated at 252. the mycelial proliferation was recorded on the 7 day post - incubation . carboxyl methyl cellulase ( cmcase ) ( padmaja and balagopal , 1985 ) , filter paper degrading enzyme ( fpase ) , -glucosidase ( ray et al . , 1993 ) and total nitrogen content ( jayaraman , 1988 ) of the substrate were estimated at 7th day and 14th day after incubation . the isolate from azotobacter bioinoculant was synergistic to the growth of pleurotus eous , but showed antagonism towards trichoderma viride and trichoderma reesi in plate cultures ( fig . broth culture at 2 ml per bottle concentration increased the linear growth of p. eous mushroom . however the increase was neutralized and was brought on par with the control at 5 ml per bottle concentration . at 10 ml per bottle concentration , there was a decrease in linear growth of the mushroom indicating that the biofertilizer isolate was inhibitory to mycelial growth at higher concentrations ( table 1 ) . ( 1993 ) , which reported the role of biofertilizers in suppressing the weed fungi and increasing the mycelial growth of agaricus bisporus . the activities of the crude cellulolytic enzymes , viz . , cmcase , filter paper degrading enzyme and -glucosidase in the p. eous seed spawn substrate were promoted by adding azotobacter isolate ( fig . cmcase activity in the bioinoculant added to substrate was significantly higher than the control on both the 7 and 14 day after inoculation and the quantity of biofertilizer inoculum proportionally increased the cellulolytic activity up to 5 ml per bottle concentration . fpase and -glucosidase activities were significantly increased over control upon addition of azotobacter isolate on the 14 day but the effect of concentration of the isolate on the enzyme activity was insignificant . the cellulase enzyme activities were lower in the 10 ml per bottle treatment than in the 5 ml per bottle treatment . ( 1994 ) who observed that azotobacter chroococcum in dual culture with the basidiomycete , coprinus fimetarius , enriched the growth of the fungus by degrading cellulose prior to fungus . tan and wahab ( 1997 ) observed that most white rot fungi exhibit low cellulolytic activity in the anamorphic stage and it is known that cultivation conditions influence activity . the presence of a nitrogen - fixing microbe in the spawn substrate might have improved the nutrient requirements of the mushroom , which would have a positive influence on the activity of cellulolytic enzymes . by promoting the cellulolytic activity of the mushroom , the microbial inoculant could direct it towards the production of enzymes targeted at lignin depolymerization , which is coupled to primordia formation . thus the microbial inoculants at lower concentration might help in earlier pinning and increasing the mushroom yield . royse and bahlar ( 1988 ) reported that the nature and amount of nitrogen present in the substrate influenced the degree of cellulose degradation . the total nitrogen content of pleurotus eous seed spawn substrate increased during the course of spawn growth as well as in the presence of azotobacter biofertilizer isolate ( table 1 ) . there was a significant increase in total nitrogen content of the substrate up to 5 ml per bottle azotobacter isolate treatment indicating its promotive effect on the mushroom growth . at 10 ml per bottle azotobacter inoculum concentration , the total nitrogen content of the substrate was lower than that of the other treatments . the linear growth of the mushroom and its cellulolytic activities decreased at the same concentration . this apparent lower nitrogen content of the substrate may be due to the inhibition of substrate mineralization by the fungus at the above - optimal nitrogen content of the substrate at the higher bioinoculant concentration . the change in nitrogen content of the substrate in pure culture mushroom bags was attributed to the degree of substrate mineralization and colonization by the mushroom fungi and the subsequent organic matter loss ( theradimani and marimuthu , 1992 ) . the incidence of contamination by molds like rhizopus sp . and aspergillus niger were noticed to be higher in the p. eous spawn bottles receiving the microbial inoculants at the higher concentration ( 10 ml per bottle ) during visual examination . this is expected , as nitrogen availability in the form of biomass nitrogen increases , the potential for development of infesting weed molds increases . carroll and schisler ( 1976 ) studying an ideal supplement of mushroom substrate mentioned that excessive heating and stimulation of competitor molds might limit the rate of supplementation and corresponding benefit . the results of the study indicated that azotobacter bioinoculant was efficient for increasing the growth and substrate utilization potential of the seed spawn of p. eous and controlling weed fungal infestation at lower inoculum concentrations . further work is in progress to study the effect of this microbial inoculant on the yield of p. eous .
we investigated the effect of nitrogen fixing azotobacter bioinoculant on the mycelial growth and the rate of substrate utilization by pleurotus eous . 
 the synergistic or antagonistic role of the microorganism during dual culturing with the mushroom or the competitor molds trichoderma viride , and trichoderma reesi was studied . 
 azotobacter was inhibitory to the molds , which are competitive to the mushroom in the seed spawn substrate , but was synergistic towards the mushroom . 
 the growth , substrate utilization potential as total nitrogen content and cellulase enzyme activities of the mushroom in the seed spawn substrate were also enhanced in the presence of the bioinoculant at lower inoculum concentrations , upto 5 ml broth culture per spawn bottle .