Abstract
BACKGROUND: Cotton bollworm, Helicoverpa armigera is a widely distributed, devastating pest of over 200 crop plants that mainly consist of some cellulosic materials. Despite its economic importance as a pest, little is known about the diversity and community structure of gut symbiotic bacteria potentially functioned in cellulose digestion in different gut-sections of H. armigera. In view of this lacuna, we attempted to evaluate and characterize cellulose-degrading bacteria (CDB) from foregut, midgut, and hindgut -regions of H. armigera by using a culture-dependent approach. METHODOLOGY: The symbiotic bacteria were isolated from different gut-systems of H. armigera by enrichment techniques using Carboxymethyl cellulose sodium salt (CMC) as carbon source. The isolated bacteria were purified and subsequently screened for cellulose-degradation by plate-based method to display the zones of CMC clearance around the colonies. The identification and phylogeny of the gut-bacteria were reconstructed by using 16S rRNA gene sequencing. Different enzymes such as endoglucanase, exoglucanase, β-glucosidase, and xylanase were assayed to determine the cellulolytic repertoire of the isolated bacteria. RESULTS: The enrichment of CDB and subsequent plate based screening methods resulted in isolation of 71 bacteria among which 54% of the bacteria were obtained from foregut. Among the isolated bacteria, 25 isolates showed discernible cellulose-degradation potential on CMC-agar plates. The phylogenetic analysis based on 16S rRNA gene amplification and sequencing affiliated these cellulolytic bacteria to two major phyla viz., Firmicutes and Proteobacteria. The members of the genus Klebsiella accounted for 39.43% of the total isolated bacteria while 31% of the Bacillus strains were enriched from hindgut region. The principal component analysis (PCA) further suggested that the members of Bacillus and Klebsiella together dominated the foregut and hindgut regions as they accounted for 68% of the total CDB. The four potential isolates selected on the basis of plate-based activities were further evaluated for their lignocellulases production by using various agricultural wastes as substrates. The PCA of the enzyme activities demonstrated that potential isolates majorly secreted endoglucanase and xylanase enzymes. Among the agro-wastes, multivariate analysis validated wheat husk (WH) and sugarcane bagasse (SCB) as most favorable substrates for xylanase and endoglucanase productions respectively. The overall findings suggest that H. armigera harbors diverse bacterial communities in different gut-sections that could assist the host in digestion processes, which may potentially serve as a valuable reservoir of some unique symbionts applied for biomass conversion in biofuel industry.