Preliminary Study Of The Microbial Diversity And Key Functional Microbes Of The Effective Cellulolytic Microbial Consotrium RXS

The use of microbial co-culture (that is, cellulolytic microbial consortium) is able toeffectively degrade lignocellulose，which could produce the more multiplex enzymes andcome to no substrate and feedback inhibition constantly occurring when single bacteriadegraded lignocellulose. The studies on the degradation of lignocellulose using microbialconsortium to enhance the utilization of cellulosic wastes had been a hot topic, currently.However, it was rather difficult to demonstrate the mechanism why cellulolytic microbialconsortium could maintain its own structural and functional stability owing to the diversity ofmicrobial species and the complicate interaction among cellulolytic microbial consortium.This paper mainly intended to analyse the microbial diversity and find out the keyfunctional bacteria strains of the cellulolytic microbial consortium RXS through thetraditional cultivation technique combined with the culture-independent technique(PCR-DGGE).In this study,14strains were isolated in the PCS medium by the traditional plateseparation technique, belonging to Bacillus sp., Petrobacter sp., Chelatococcus sp. andTepidiphilus sp. respectively, however, they had no cellulose-degrading abilities. We have thusisolated a thermophilic cellulolytic facultative anaerobic bacterium named FLJ1with efficientfilter paper degradation power. The results of phylogenetic tree construction indicate that theclosest phylogenetic relative product is the Caloramator sp. LSA2(AY770516) with99%ofsequential similarity.Strain Caloramator sp. FLJ1could degrade the filter paper effectively under theanaerobic conditions, with a degrading ratio of46.25%in the third day. The xylanase, fpaseand CMCase activity during the degrading process reached68.90,35.09and29.23U/mL,respectively. Correspondingly, the molecular weight of xylanase activated proteins secretedby the strain Caloramator sp. FLJ1were about160,180and190kDa and those of CMCaseactivated proteins were80,85and130kDa.The analysis of enzymatic properties about the CMCase and xylanase secreted byCaloramator sp. FLJ1indicated that their optimum temperature and optimum pH were60℃and6.0, respectively. Both CMCase and xylanase has good thermal stability and pH stability.In addition, strain Caloramator sp. FLJ1produced the xylanase using corn stalk ascarbon resource under the micro-aerobic condition, whose activity was only890.41U/mL. Sowe designed the response surface to optimize the fermentation medium. Results were asfollows (g/L): corn stalk8.68, beef extract6.0, NaCl9.21, KH2PO40.92, CaCO33.0, pH8.1.The xylanase activity produced by Caloramator sp. FLJ1reached1273.02U/mL, whichincreased by33.91%compared to the original activity, with a innoculum size of2%at55℃under the micro-aerobic condition.Gradient dilution for microbial consortium RXS were used to degrade cassava residuesand filter paper and we found that10-5was the critical dilution gradient of microbialconsortium RXS. Filter paper and cassava residues were treated by the RXS in differentgradient dilution. Using the DGGE technique for analysis, one of the non-culture techniques,we eventually recovered26trips, which belonged to Clostridium sp., Bacillus sp., Petrobacter sp., Thermotogae sp. and Uncultured bacterium via sequence analysis. It was preliminarilyconducted that Clostridium clariflavum DSM19732and Uncultured Paenibacillus sp. in themicrobial consortium RXS play a critical role in cellulose degradation.