Abstract
Fungus-growing termites play an important role in lignocellulose degradation and carbon mineralization in tropical and subtropical regions, but the degradation potentiality of their gut microbiota has long been neglected. The high quality and quantity of intestinal microbial DNA are indispensable for exploring new cellulose genes from termites by function-based screening. Here, using a refined intestinal microbial DNA extraction method followed by multiple-displacement amplification (MDA), a fosmid library was constructed from the total microbial DNA isolated from the gut of a termite growing in fungi. Functional screening for endoglucanase, cellobiohydrolase, β-glucosidase, and xylanase resulted in 12 β-glucosidase-positive clones and one xylanase-positive clone. The sequencing result of the xylanase-positive clone revealed an 1,818-bp open reading frame (ORF) encoding a 64.5-kDa multidomain endo-1,4-β-xylanase, designated Xyl6E7, which consisted of an N-terminal GH11 family catalytic domain, a CBM_4_9 domain, and a Listeria-Bacteroides repeat domain. Xyl6E7 was a highly active, substrate-specific, and endo-acting alkaline xylanase with considerably wide pH tolerance and stability but extremely low thermostability.
Highlights
Fungus-growing termites play an important role in lignocellulose degradation and carbon mineralization in tropical and subtropical regions, but the degradation potentiality of their gut microbiota has long been neglected
As the guts of M. annandalei termites contained a large proportion of soil and humic acid, approximately 450 ng of DNA was obtained from about 100 M. annandalei worker termite guts, compared to several micrograms of DNA from equal numbers of other termite guts
The refined extraction method used in this study limited the contamination from termites to the total microbial DNA
Summary
Fungus-growing termites play an important role in lignocellulose degradation and carbon mineralization in tropical and subtropical regions, but the degradation potentiality of their gut microbiota has long been neglected. Increasing attention has been focused on exploring lignocellulase genes from diverse organisms, including free-living bacteria and fungi [45] as well as gut microbiomes of lignocellulose-consuming ruminants and insects [14, 18, 24, 46]. To date, little attention has been paid to the hydrolyzing capabilities of these intestinal microorganisms, except for fermentation and acetogenesis or methanogenesis [6] We believe it possible that intestinal microorganisms may play a certain role in the overall process of lignocellulose digestion for their hosts. As the majority of microorganisms in termite guts are unculturable, metagenomics becomes an ideal tool for exploring these presently inaccessible metagenome reservoirs [17], just as it has played significant roles in Downloaded from https://journals.asm.org/journal/aem on 08 November 2021 by 3.86.85.159
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