Isolation and Identification of Cellulolytic Bacteria from the Gut of Holotrichia parallela Larvae (Coleoptera: Scarabaeidae)

Abstract

In this study, 207 strains of aerobic and facultatively anaerobic cellulolytic bacteria were isolated from the gut of Holotrichia parallela larvae. These bacterial isolates were assigned to 21 genotypes by amplified ribosomal DNA restriction analysis (ARDRA). A partial 16S rDNA sequence analysis and standard biochemical and physiological tests were used for the assignment of the 21 representative isolates. Our results show that the cellulolytic bacterial community is dominated by the Proteobacteria (70.05%), followed by the Actinobacteria (24.15%), the Firmicutes (4.35%), and the Bacteroidetes (1.45%). At the genus level, Gram-negative bacteria including Pseudomonas, Ochrobactrum, Rhizobium, Cellulosimicrobium, and Microbacterium were the predominant groups, but members of Bacillus, Dyadobacter, Siphonobacter, Paracoccus, Kaistia, Devosia, Labrys, Ensifer, Variovorax, Shinella, Citrobacter, and Stenotrophomonas were also found. Furthermore, our results suggest that a significant amount of bacterial diversity exists among the cellulolytic bacteria, and that Siphonobacter aquaeclarae, Cellulosimicrobium funkei, Paracoccus sulfuroxidans, Ochrobactrum cytisi, Ochrobactrum haematophilum, Kaistia adipata, Devosia riboflavina, Labrys neptuniae, Ensifer adhaerens, Shinella zoogloeoides, Citrobacter freundii, and Pseudomonas nitroreducens are reported to be cellulolytic for the first time in this study. Our results indicate that the scarab gut is an attractive source for the study of novel cellulolytic microorganisms and enzymes useful for cellulose degradation.

Optimization of Endoglucanase Production from a Novel Bacterial Isolate, Arthrobacter sp. HPG166 and Characterization of Its Properties

“…The present results showed that CMC had a positive influence on the cellulase production and 1.19% (W/V) CMC was the most effective level. Sheng et al (2012) found that CMC was the most effective inducer for cellulase production among eight different carbon sources tested, and CMC 1.5% (w/v) led to maximal enzyme yield. Furthermore, an interesting phenomenon was found that the interaction effects of CMC and beef extract had significant effects on cellulase production (p=0.0163, Table 4).…”

“…BCCS A3, while the pH optimum of endoglucanase production was pH 9.0. Sheng et al (2012) reported that the optimal temperature for endoglucanase production by Pseudomonas sp. HP207 was 30°C, while Adeleke (2013) found that the Penicillium atrovenetum exhibited maximum endoglucanase activity at 40ºC.…”

“…Third-instar larvae of H. parallela were collected from a peanut field in Xuzhou, Jiangsu Province and dissected as described by Huang et al (2012). Then, each hindgut (from the pyloric sphincter to the rectum including the modified fermentation sac) was placed into a 1.5 mL eppendorf tube, which contained 0.5 mL of phosphate buffered saline.…”

“…Then DNA fragment of 16S rRNA gene was amplified as previously described in Huang et al (2012). The PCR product was cloned into pEASY-T1 cloning kit (Beijing TransGen Biotech Co., Ltd., Beijing, China) with blue-white screening.…”

“…More importantly, bacterial cellulases can survive in harsh conditions of bioconversion processes, and also be less inhibited by the presence of material that has already been hydrolyzed (feedback inhibition) (Kadarmoidheen et al 2012). Therefore, the isolation and characterization of novel glycoside hydrolases from bacteria are now becoming widely exploited (Maki et al 2009;Abdel-Fatah et al 2012), and the cellulolytic property of some bacterial genera such as Clostridium (Yuan et al 2015), Cellulomonas (Kojima et al 2013), Pseudomonas (Huang et al 2012;Sheng et al 2012), Bacillus (Gaur and Tiwari 2015), and Streptomyces (Azzeddine et al 2013) has been widely reported. Athrobacter species are predominant members of culturable soil bacteria and can utilize a wide variety of organic compounds, such as aromatic hydrocarbons, nucleic acids and their degradation products, herbicides (Crocker et al 2000).…”