We have used bulked segregant analysis to screen the strain-specific DNA marker associated thermophilic strain of Pleurotus eryngii. Bulked genomic DNAs of Pleurotus eryngii were amplified by randomly amplified polymorphic DNA (RAPD) using OP-A, OP-B, OP-L, OP-P, OP-R and OP-S primers to screen the strain-specific DNA marker. A unique DNA fragment of 500 bp was amplified with OP-A11 primer from the psychrophilic strain and sequenced. A sequence characterized amplified region (SCAR) marker was designed on the basis of the determined sequence and named as OP-A11-1. The PCR analysis with the OP-A11-1 primer showed that this SCAR marker clearly distinguish the psychrophilic strains from the control strains.

Spent mushroom substrate (SMS) is a by-product remaining after a crop of mushrooms. The fresh SMS was sample within 24 hours of removal from the production facility in DOJUN farm located in Jinju. About 11 bacterial species were isolated from fresh SMS on TSA medium. Among of them, one isolate, designated YJ02, showed the antifungal activity against Aspergillus flavus and Aspergillus ochraceous producing mycotoxin on PDA medium, potentially. The strain YJ02 was produced cellulase, xylanase, mannanase as hydrolase. The strain YJ02 was identified as members of the genus Bacillus by biochemical characteristics using Bacillus ID kit and VITEK 2 system. Comparative 16S rDNA gene sequence analysis showed that strain YJ02 formed a distinct phylogenetic tree within the genus Bacillus and was most closely related to Bacillus amyloliquefaciens with 16S rDNA gene sequence similarity of 98.7%. On the basis of its physiological properties, biochemical characteristics and phylogenetic distinctiveness, strain YJ02 was classified within the genus Bacillus, for which the name Bacillus amyloliquefaciens YJ02 is proposed.

Spent mushroom substrate(SMS) is a by-product remaining after a crop of mushrooms. About 9 thermophilic strains were isolated from SMS(Flammulina velvtipes). Among of them, one isolate, designated UJ09, showed the antifungal activity against Aspergillus flavus and Aspergillus ochraceous producing mycotoxin on PDA medium, potentially. The strain UJ09 was produced cellulase and xylanase as hydrolase. The strain UJ21 was identified as members of the genus Bacillus by biochemical characteristics using Bacillus ID kit and VITEK 2 system. Comparative 16S rDNA gene sequence analysis showed that strain UJ09 formed a distinct phylogenetic tree within the genus Bacillus and was most closely related to Bacillus amyloliquefaciens with 16S rDNA gene sequence similarity of 99.2%. On the basis of its physiological properties, biochemical characteristics and phylogenetic distinctiveness, strain UJ09 was classified within the genus Bacillus, for which the name Bacillus amyloliquefaciens UJ09 is proposed.

The mannanase, cellulase and xylanase-producing thermophilic bacteria, designated EJ3, was isolated from fresh spent mushroom (Pleurotus eryngii) substrates taken from the DOJUN farm located in Keyongnam, Korea. The isolate EJ3 was facultatively anaerobic and grew at temperature ranging from 20℃ to 60℃ with an optimal temperature of 37℃. The DNA G+C content of the isolate EJ3 was 45 mlo%. The major fatty acids were anteiso-15:0 (38.9%), 17:0 (7.4%), and iso-15:0 (38.2%). The 16S rRNA gene sequence similarity between the isolate EJ3 and other Bacillus strains varied from 97% to 99%. In the phylogenetic analysis based on these sequences, the isolate EJ3 and Bacillus subtilis clustered within a group and separated from other species of Bacillus. Based on the physiological and molecular properties, the isolate EJ3 was classified within the genus Bacillus as Bacillus subtilis EJ3. The optimal growth condition of B. subtilis EJ3 were pH 6.0 and 55℃, respectively.

This study was conducted to evaluate the feeding value of the spent mushroom (Pleurotus eryngii) substrates (SMS) in laying hens (Hy-Line Brown). The fresh spent mushroom (Pleurotus eryngii) substrates collected from the DOJUN farm were fermented with Bacillus subtilis EJ3 for 2 weeks. A total of twenty-four laying hens were fed corn-soy based experimental diets containing 0% (control), 5% (T1), 10% (T2) and 15%(T3) fermented SMS for 7 weeks. There were no significant differences among the treatments in egg production, egg weight, egg mass, feed conversion and viability during the experimental period. Feed intake was significantly lowered in control (118.3 g) than T1 (121.9 g), T2 (120.3 g) and T3 (122.4 g). There were no significant differences among the treatments eggshell breaking strength, thickness and haugh unit, whereas the yolk color of T1, T2 and T3 were significantly heavy than T0. The palatability of boiled meat was significantly better in the T3 laying hens than in the T0 laying hens. In conclusion, fermented SMS can be used as resource of feed in laying hen feed at 5.0-15% level without effect on performance and egg qualify.