Termite Gut Research Articles

Harnessing the potential use of cellulolytic Klebsiella oxytoca (M21WG) and Klebsiella sp. (Z6WG) isolated from the guts of termites (Isoptera)

BackgroundFor many years, denim-heavy quality cotton twill colored with indigo colors and with a well-worn/faded look has held a lot of appeal. Machine damage, drainage system blockage, and other issues come with the conventional usage of pumice stones for “stone-washing” denims. In view of the abovementioned information, a range of works has been done to investigate the economic prospects of bacterial cellulase enzymes for use in industrial processes, including biopolishing in the textile sector. Ethiopia has excellent termite diversity to isolate bacterial gut-associated cellulose enzymes for biostoning applications. The main purpose of this study was, therfore, to decipher how to isolate and characterize cellulase enzymes from termite (Isoptera) gut bacteria with the intention of employing it for biostoning of textiles.PurposeTo use cellulolytic enzymes of Klebsiella oxytoca (M21WG) and Klebsiella sp. (Z6WG) isolated from termite guts in biostoning of textiles and improving garment quality.MethodsCellulase enzyme-producing bacteria were isolated and screened from the guts of worker termites sampled from Meki and Zeway termite mounds in the Central Rift Valley region of Ethiopia. Bacterial screening, biochemical, morphological, and 16S rRNA sequence identification techniques were employed to characterize the bacterial strains. In addition, the production, optimization, and purification of the associated cellulase enzymes were employed, and the potential application of the enzymes for biostoning of a textile was demonstrated.ResultThe isolated M21WG was found to be 99% identical to the Klebsiella oxytoca (MT104573.1) strain, while the isolated Z6WG showed 97.3% identity to the Klebsiella sp. strain (MN629242.1). At an ideal pH of 7, a temperature of 37 °C, a 72-h incubation time, and a substrate concentration of 1.5% carboxymethylcellulose sodium, the maximum activity of the crude cellulase extract from these bacteria was assessed. These bacteria produced cellulase enzymes that were moderately efficient. Consequently, it was determined that the cellulase enzymes were effective for biostoning of denim cloth.ConclusionIt was determined that Klebsiella oxytoca (M21WG) and Klebsiella sp. (Z6WG) could be used as a doorway to better understand harnessing the use of these cellulase-producing bacteria from termite (Isoptera) guts. In this study, it was also attempted to assess the effectiveness of the two bacterial isolates in biostoning in anticipation of their potential application in the textile realm.

Simultaneous Single-Cell Genome and Transcriptome Sequencing of Termite Hindgut Protists Reveals Metabolic and Evolutionary Traits of Their Endosymbionts.

ABSTRACTSome of the protist species which colonize the hindguts of wood-feeding Reticulitermes termites are associated with endosymbiotic bacteria belonging to the genus Endomicrobium. In this study, we focused on the endosymbionts of three protist species from Reticulitermes flavipes, as follows: Pyrsonympha vertens, Trichonympha agilis, and Dinenympha species II. Since these protist hosts represented members of different taxa which colonize separate niches within the hindguts of their termite hosts, we investigated if these differences translated to differential gene content and expression in their endosymbionts. Following assembly and comparative genome and transcriptome analyses, we discovered that these endosymbionts differed with respect to some possible niche-specific traits, such as carbon metabolism. Our analyses suggest that species-specific genes related to carbon metabolism were acquired by horizontal gene transfer (HGT) and may have come from taxa which are common in the termite hind gut. In addition, our analyses suggested that these endosymbionts contain and express genes related to natural transformation (competence) and recombination. Taken together, the presence of genes acquired by HGT and a putative competence pathway suggest that these endosymbionts are not cut off from gene flow and that competence may be a mechanism by which members of Endomicrobium can acquire new traits.IMPORTANCE The composition and structure of wood, which contains cellulose, hemicellulose, and lignin, prevent most organisms from using this common food source. Termites are a rare exception among animals, and they rely on a complex microbiota housed in their hindguts to use wood as a source of food. The lower termite, Reticulitermes flavipes, houses a variety of protists and prokaryotes that are the key players in the disassembly of lignocellulose. Here, we describe the genomes and the gene expression profiles of five Endomicrobium endosymbionts living inside three different protist species from R. flavipes. Data from these genomes suggest that these Endomicrobium species have different mechanisms for using carbon. In addition, they harbor genes that may be used to import DNA from their environment. This process of DNA uptake may contribute to the high levels of horizontal gene transfer noted previously in Endomicrobium species.

Thermal tolerance of an invasive drywood termite, Cryptotermes brevis (Blattodea: Kalotermitidae)

West Indian drywood termites (Cryptotermes brevis, Blattodea: Kalotermitidae) are an important invasive termite in many countries including Australia where they are spreading across two eastern states. Fumigation is often used to eliminate infestations, but it is costly, has negative environmental effects and does not prevent reinfestation. Heat treatment has been suggested as an alternative. Many insect pest mitigation strategies recommend 30 min exposure at 56 °C, but this may be difficult to achieve in structural applications. The potential for heating at lower temperatures was explored to determine the effect on termite survival and gut fauna. Exposure to 40 °C up to an hour did not kill the termites; however, 1-h exposure at 45 °C was lethal. Exposure for little as 3 min at 50 °C or 2 min at 55 °C was lethal. Protozoa levels were lower in termites that survived shorter exposures, but there appeared to be some recovery over time. The results suggest that short term exposures to 50 or 55 °C could be used to eliminate infestations, creating an opportunity for localized spot heating as a mitigation measure.

Oxidative cleavage of polysaccharides by a termite-derived superoxide dismutase boosts the degradation of biomass by glycoside hydrolases.

Wood-feeding termites effectively degrade plant biomass through enzymatic degradation. Despite their high efficiencies, however, individual glycoside hydrolases isolated from termites and their symbionts exhibit anomalously low effectiveness in lignocellulose degradation, suggesting hereto unknown enzymatic activities in their digestome. Herein, we demonstrate that an ancient redox-active enzyme encoded by the lower termite Coptotermes gestroi, a Cu/Zn superoxide dismutase (CgSOD-1), plays a previously unknown role in plant biomass degradation. We show that CgSOD-1 transcripts and peptides are up-regulated in response to an increased level of lignocellulose recalcitrance and that CgSOD-1 localizes in the lumen of the fore- and midguts of C. gestroi together with termite main cellulase, CgEG-1-GH9. CgSOD-1 boosts the saccharification of polysaccharides by CgEG-1-GH9. We show that the boosting effect of CgSOD-1 involves an oxidative mechanism of action in which CgSOD-1 generates reactive oxygen species that subsequently cleave the polysaccharide. SOD-type enzymes constitute a new addition to the growing family of oxidases, ones which are up-regulated when exposed to recalcitrant polysaccharides, and that are used by Nature for biomass degradation.

Saccharification of agricultural lignocellulosic feedstocks by endogenous and symbiotic cellulases from the subterranean termites

This study investigated the protein fraction-level in vitro and in vivo saccharification potential by the guts of two geographically different lower termites’ species Reticulitermes flavipes and Heterotermes indicola using biofuel producing feedstocks sugarcane bagasse, pinewood, cottonwood, corn stover, and rice husk. The gut protein extract from both termite species saccharified all lignocellulose material regardless of feedstock loading. However, R. flavipes showed significantly (p < 0.05) higher saccharification potential than H. indicola. Consumption of the sugarcane bagasse, pinewood, and corn stover was maximum by R. flavipes, whereas H. indicola exposed cottonwood and sugarcane feedstocks as the most palatable food source with 100% survivability. This study also hypothesized that feeding on various feedstocks will show variable impacts on enzyme activities. An adaptive mechanism enables the termites and their endosymbionts for optimized consumption of variable food sources. Results supported "diet adaptation" hypothesis by indicating β-glucosidase, exoglucanase, and xylanase activities, which were significantly the highest for sugarcane bagasse, pinewood, and cottonwood to rice husk and control diets in both termite species. These results are important in physiological changes in termites that lead to gut microbial environment changes and 100% survivability on exclusive sugarcane bagasse, pinewood, and cottonwood feedstocks. Both termite species' guts exhibit the clear saccharification for all feedstocks, which validate the H indicola and R. flavipes systems as a potential source for lignocellulose biomass digesting enzymes particularly β-glucosidase, endoglucanase, exoglucanase, and xylanases.

Amylolytic microorganisms from diverse tropical environments: Isolation, identification, and amylase production

AbstractThis study aimed to identify microbes producing amylases from diverse unexplored environments in Kenya. Screening based on microorganisms' ability to grow on minimal media (M9) supplemented with starch resulted in the selection of 70 amylolytic isolates. The isolates were then subjected to primary screening based on starch hydrolysis ratio (SHR) obtained by halo formation on starch agar plates after flooding plates with Lugol's iodine solution. SHR narrowed down the isolates' number to 17 (SHR &gt; 1.5), which were used for subsequent experiments. The 17 isolates were grown on M9 starch broth for the production of amylases. Crude amylases and commercial enzyme (control) were assayed for amylase activity using the 3,5‐dinitrosalicylic acid method. Specific enzyme activities of crude enzymes ranged between 0.0079 and 0.0629 U g−1, which were lower than 1.7188 U g−1 of the control. Five enzymes with the highest specific enzyme activity were used for cassava hydrolysis. All crude enzymes exhibited higher specific enzyme activity (0.0707–0.1853 U g−1) than the control (0.0052 U g−1) in hydrolyzing cassava. Alkaline lakes had the highest number (4) of isolates whose enzymes had the highest specific enzyme activity, while isolates from potato processing plant waste, termite gut, and black soldier fly gut had a representative each. Isolates with enzymes having the highest specific enzyme activity were identified by DNA sequencing and belonged to Lysinibacillus sp. (four isolates), Alternaria sp. (one isolate), and Bacillus sp. (two isolates). The study demonstrated the presence of microorganisms in tropical environments with amylase activity that can be scaled and optimized for practical applications.

Fine-scale genetic diversity and putative ecotypes of oxymonad protists coinhabiting the hindgut of Reticulitermes speratus.

The hindgut of lower termites is generally coinhabited by multiple morphologically identifiable protist species. However, it is unclear how many protist species truly coexist in this miniaturized environment, and moreover, it is difficult to define the fundamental unit of protist diversity. Species delineation of termite gut protists has therefore been guided without a theory-based concept of species. Here, we focused on the hindgut of the termite Reticulitermes speratus, where 10 or 11morphologically distinct oxymonad cell types, that is, morphospecies, coexist. We elucidated the phylogenetic structure of all co-occurring oxymonads and addressed whether their diversity can be explained by the "ecotype" hypothesis. Oxymonad-specific 18S rRNA gene amplicon sequencing analyses of whole-gut samples, combined with single-cell 18S rRNA sequencing of the oxymonad morphospecies, identified 210 one-nucleotide-level variants. The phylogenetic analysis of these variants revealed the presence of microdiverse clusters typically within 1% sequence divergence. Each known oxymonad morphospecies comprised one to several monophyletic or paraphyletic microdiverse clusters. Using these sequence data sets, we conducted computational simulation to predict the rates of ecotype formation and periodic selection, and to demarcate putative ecotypes. Our simulations suggested that the oxymonad genetic divergence is constrained primarily by strong selection, in spite of limited population size and possible bottlenecks during intergenerational transmission. A total of 33 oxymonad ecotypes were predicted, and most of the putative ecotypes were consistently detected among different colonies and host individuals. These findings provide a possible theoretical basis for species diversity and underlying mechanisms of coexistence of termite gut protists.

Wood-feeding termite gut symbionts as an obscure yet promising source of novel manganese peroxidase-producing oleaginous yeasts intended for azo dye decolorization and biodiesel production

BackgroundThe ability of oxidative enzyme-producing micro-organisms to efficiently valorize organic pollutants is critical in this context. Yeasts are promising enzyme producers with potential applications in waste management, while lipid accumulation offers significant bioenergy production opportunities. The aim of this study was to explore manganese peroxidase-producing oleaginous yeasts inhabiting the guts of wood-feeding termites for azo dye decolorization, tolerating lignocellulose degradation inhibitors, and biodiesel production.ResultsOut of 38 yeast isolates screened from wood-feeding termite gut symbionts, nine isolates exhibited high levels of extracellular manganese peroxidase (MnP) activity ranged between 23 and 27 U/mL after 5 days of incubation in an optimal substrate. Of these MnP-producing yeasts, four strains had lipid accumulation greater than 20% (oleaginous nature), with Meyerozyma caribbica SSA1654 having the highest lipid content (47.25%, w/w). In terms of tolerance to lignocellulose degradation inhibitors, the four MnP-producing oleaginous yeast strains could grow in the presence of furfural, 5-hydroxymethyl furfural, acetic acid, vanillin, and formic acid in the tested range. M. caribbica SSA1654 showed the highest tolerance to furfural (1.0 g/L), 5-hydroxymethyl furfural (2.5 g/L) and vanillin (2.0 g/L). Furthermore, M. caribbica SSA1654 could grow in the presence of 2.5 g/L acetic acid but grew moderately. Furfural and formic acid had a significant inhibitory effect on lipid accumulation by M. caribbica SSA1654, compared to the other lignocellulose degradation inhibitors tested. On the other hand, a new MnP-producing oleaginous yeast consortium designated as NYC-1 was constructed. This consortium demonstrated effective decolorization of all individual azo dyes tested within 24 h, up to a dye concentration of 250 mg/L. The NYC-1 consortium's decolorization performance against Acid Orange 7 (AO7) was investigated under the influence of several parameters, such as temperature, pH, salt concentration, and co-substrates (e.g., carbon, nitrogen, or agricultural wastes). The main physicochemical properties of biodiesel produced by AO7-degraded NYC-1 consortium were estimated and the results were compared to those obtained from international standards.ConclusionThe findings of this study open up a new avenue for using peroxidase-producing oleaginous yeasts inhabiting wood-feeding termite gut symbionts, which hold great promise for the remediation of recalcitrant azo dye wastewater and lignocellulosic biomass for biofuel production.

Microbial Response to Fungal Infection in a Fungus-Growing Termite, Odontotermes formosanus (Shiraki).

The crosstalk between gut microbiota and host immunity has emerged as one of the research foci of microbiome studies in recent years. The purpose of this study was to determine how gut microbes respond to fungal infection in termites, given their reliance on gut symbionts for food intake as well as maintaining host health. Here, we used Metarhizium robertsii, an entomopathogenic fungus, to infect Odontotermes formosanus, a fungus-growing termite in the family Termitidae, and documented changes in host gut microbiota via a combination of bacterial 16S rDNA sequencing, metagenomic shotgun sequencing, and transmission electron microscopy. Our analyses found that when challenged with Metarhizium, the termite gut showed reduced microbial diversity within the first 12 h of fungal infection and then recovered and even surpassed pre-infection flora levels. These combined results shed light on the role of gut flora in maintaining homeostasis and immune homeostasis in the host, and the impact of gut flora dysbiosis on host susceptibility to infection.

Cellulose Degrading Ability of Bacterial Strains Isolated from Gut of Termites in Vinhlong Province - Vietnam

Cellulose are the most abundant renewable biomass. The bioconversion of cellulose plays an important role for sustainable development and application in alcohol production, compost production and waste treatment. Termites were known as the most successful wood-degrading species on the earth, can degrade in large quantities of cellulosic biomass by activities of enzymes secreted by the termites and/or their gut. A total of 70 bacterial isolates were isolated from the gut of termites obtained from thirteen termite nests in TraOn district, VinhLong province, Mekong Delta, Vietnam. Of the 70 bacterial isolates, 60 (86%) are able to degrade CMC (Carboxyl Methyl Cellulose) after culturing in media containing CMC agar and halo formation after staining with Congo red. Among of them, 5 isolates having high CMC degradation efficiency are chosen to evaluate the potential ability to liberate glucose from cellulose of straw for 5, 10, 15 and 20 days respectively at room temperatures. The results showed that all of the 5 isolates have the cellulose degradation ability and produce the highest glucose contents after 15 days. Out of the 5 isolates, 2 produced glucose in large quantities are 2T1 isolate (0.73 g/L) and 5T6 isolate (0.79 g/L) in hydrolysis solutions. The 2T1, 5T6 strains were 98.99%, 99.01% of identity Paenibacillus humicus strain T20C, Psychrobacter faecalis strain AP3Ka respectively by 16S rRNA genes sequencing and could be potential sources of enzymes for cellulose hydrolysis from cellulosic biomass in future.

Isolation and Identification of Cellulase Producing and Sugar Fermenting Bacteria for Second-Generation Bioethanol Production

Over the last decades, the negative impacts of fossil fuel on the environment and increasing demand for energy due to the unavoidable depletion of fossil fuels, has transformed the world’s interests towards alternative fuels. In particular, bioethanol production from cellulosic biomass for the transportation sector has been incrementing since the last decade. The bacterial pathway for bioethanol production is a relatively novel concept and the present study focused on the isolation of potential “cellulase-producing” bacteria from cow dung, compost soil, and termite gut and isolating sugar fermenting bacteria from palm wine. To select potential candidates for cellulase enzyme production, primary and secondary assays were conducted using the Gram’s iodine stain in Carboxy Methyl Cellulose (CMC) medium and the Dinitrosalicylic acid (DNS) assays, respectively. Durham tube assay and Solid-Phase Micro-Extraction (SPME) coupled with Gas Chromatography-Mass Spectrometry (GC-MS) was used to evaluate the sugar fermenting efficiency of the isolated bacteria. Out of 48 bacterial isolates, 27 showed cellulase activity where Nocardiopsis sp. (S-6) demonstrated the highest extracellular crude enzyme activity of endoglucanase (1.56±0.021 U) and total cellulase activity (0.93±0.012 U). The second-highest extracellular crude enzyme activity of endoglucanase (0.21±0.021 U) and total cellulase activity (0.35±0.021 U) was recorded by Bacillus sp. (T-4). Out of a total of 8 bacterial isolates, Achromobacter sp. (PW-7) was positive for sugar fermentation resulting in 3.07% of ethanol in broth medium at 48 h incubation. The results of the study revealed that Nocardiopsis sp. (S-6) had the highest cellulase enzyme activity. However, the highest ethanol percentage was achieved with by having both Bacillus sp. (T-4) and Achromobacter sp. (PW-7) for the simultaneous saccharification and fermentation (SSF) method, as compared to separate hydrolysis and fermentation (SHF) methodologies.

Isolation and Identification Cellulolytic Bacteria from Termite Gut Obtained from Indralaya Peatland area

The production of second-generation bioethanol as renewable energy has developed very rapidly and has become a promising alternative energy source. Bioethanol production using biomass can be obtained alternatively from cellulose in wood, sawdust, organic waste, and agricultural waste. This research used termites obtained from Indralaya peatland area as organisms that can decompose cellulose into glucose with the cellulase enzymes produced by bacteria in their digestive tract. Cellulases are enzymes capable of hydrolyzing lignocellulose into glucose. The study aimed to isolate and identify of cellulolytic bacteria from termite gut obtained from Indralaya peatland area. The bacterial isolates were classified by using morphological and biochemical standard methods, and identification based on Bergey’s Manual of Determinative Bacteriology. Cellulolytic bacteria of termite gut were isolated and cultured on CMC (Carboxymethyl cellulose) agar medium. The activity of cellulolytic bacterial was conducted based on halo area and cellulolytic index on CMC agar medium. Among 64 isolates of bacteria, 24 isolates were identified as cellulolytic bacteria. Futhermore, our isolates with higher cellulolytic index were identified as the Staphylococcus, Microbacterium, Bacillus, and Brevibacterium genus.

Performance of Meyerozyma caribbica as a novel manganese peroxidase-producing yeast inhabiting wood-feeding termite gut symbionts for azo dye decolorization and detoxification

For hazardous toxic pollutants such as textile wastewater and azo dyes, microbial-based and peroxidase-assisted remediation represents a highly promising and environmentally friendly alternative. Under this scope, gut symbionts of the wood-feeding termites Coptotermes formosanus and Reticulitermes chinenesis were used for the screening of manganese peroxidase (MnP) producing yeasts intended for decolorization and detoxification of textile azo dyes, such as Acid Orange 7 (AO7). To this end, nine out of 38 yeast isolates exhibited high levels of extracellular MnP activity ranging from 23 to 27 U/mL. The isolate PPY-27, which had the highest MnP activity, was able to decolorize various azo dyes with an efficiency ranging from 87.2 to 98.8%. This isolate, which represents the molecularly identified species Meyerozyma caribbica, was successfully characterized in terms of morphological and physiological traits, as well as enzymatic activities. Almost complete decolorization was achieved by the MnP-producing M. caribbica strain SSA1654 after 6 h of incubation with 50 mg/L of the sulfonated azo dye AO7 at 28 °C with an agitation speed of 150 rpm. The maximum decolorization efficiency of AO7 reached 93.8% at 400 mg/L. The decolorization of AO7 was confirmed by Fourier transform infrared (FTIR) and UV–Vis spectral analysis. High performance liquid chromatography (HPLC) and gas chromatography–mass spectrometry (GC–MS) were used to identify AO7 decomposition intermediates. Based on UV–Vis spectra, FTIR, HPLC, and GC–MS analyses, a plausible AO7 biodegradation mechanism pathway was explored, showing azo bond (-N=N-) cleavage and toxic aromatic amines mineralization CO2 and H2O. Microtox® and phytotoxicity assays confirmed that the AO7 metabolites produced by the strain SSA1654 were almost non-toxic compared to the original sulfonated azo dye.

Ancestral predisposition toward a domesticated lifestyle in the termite-cultivated fungus Termitomyces

The ancestor of termites relied on gut symbionts for degradation of plant material, an association that persists in all termite families.1,2 However, the single-lineage Macrotermitinae has additionally acquired a fungal symbiont that complements digestion of food outside the termite gut.3 Phylogenetic analysis has shown that fungi grown by these termites form a clade-the genus Termitomyces-but the events leading toward domestication remain unclear.4 To address this, we reconstructed the lifestyle of the common ancestor of Termitomyces using a combination of ecological data with a phylogenomic analysis of 21 related non-domesticated species and 25 species of Termitomyces. We show that the closely related genera Blastosporella and Arthromyces also contain insect-associated species. Furthermore, the genus Arthromyces produces asexual spores on the mycelium, which may facilitate insect dispersal when growing on aggregated subterranean fecal pellets of a plant-feeding insect. The sister-group relationship between Arthromyces and Termitomyces implies that insect association and asexual sporulation, present in both genera, preceded the domestication of Termitomyces and did not follow domestication as has been proposed previously. Specialization of the common ancestor of these two genera on an insect-fecal substrate is further supported by similar carbohydrate-degrading profiles between Arthromyces and Termitomyces. We describe a set of traits that may have predisposed the ancestor of Termitomyces toward domestication, with each trait found scattered in related taxa outside of the termite-domesticated clade. This pattern indicates that the origin of the termite-fungus symbiosis may not have required large-scale changes of the fungal partner.

Characterization of cellulose-degrading microbiota from the eastern subterranean termite and soil

Background: While there have been a lot of studies on the termite gut microbiota, there has been very little research directly on the cellulose-degrading microbiota in termites or their soil environment. This study addresses this problem by profiling cellulose-degrading bacteria and archaea in the selective cellulose cultures of two samples of the eastern subterranean termite (Reticulitermes flavipes) and one soil sample collected at the same location as one of the termite samples. Methods: All the cultures were examined for cell concentration and remaining cellulose after the culture was completed. The 16S rRNA pyrotag sequencing method was used to identify the prokaryotic microbiota for the three cultures and one termite colony without culture. The MOTHUR, SSU-ALIGN, RDPTools, phyloseq, and other R packages were used for sequence and statistical analyses. Results: Biochemical analyses of the cultures suggested high efficiency of cellulose degradation. Comparative analyses between the cultured and uncultured termite gut microbiota revealed a significant difference. Proteobacteria and Firmicutes were found to be the two most abundant phyla of cellulose-degrading bacteria from the three cultures, but different classes within each phylum dominated the different samples. Shared and sample-specific cellulose-degrading taxa, including a core set of taxa across all the cultures, were identified. Conclusions: Our study demonstrates the importance of using selective cellulose culture to study the cellulose-degrading microbial community. It also revealed that the cellulose-degrading microbiota in the eastern subterranean termite is significantly influenced by the microbiota in the surrounding soil environment. Biochemical analyses showed that the microbial communities enriched from all the selective cultures were efficient in degrading cellulose, and a core set of bacteria have been identified as targets for further functional analyses.

Diversity and cellulolytic activity of culturable bacteria isolated from the gut of higher termites (Odontotermes sp.) in eastern Thailand

Abstract. Boontanom P, Chantarasiri A. 2021. Diversity and cellulolytic activity of culturable bacteria isolated from the gut of higher termites (Odontotermes sp.) in eastern Thailand. Biodiversitas 22: 3349-3357. Cellulolytic bacteria are vital symbionts associated with the gut of all higher termites. Odontotermes termites are a higher termite widely found in Thailand. However, information concerning the diversity of cellulolytic bacteria in this termite gut remains inadequate. The aim of this study is to isolate and identify the culturable cellulolytic bacteria from the Odontotermes gut collected from eastern Thailand. The crude cellulases produced from the most active cellulolytic bacterium were further characterized. Thirty-two cellulolytic bacteria were isolated and subsequently classified by PCR-RFLP of the 16S rRNA gene. A total of 10 different RFLP patterns were obtained belonging to five bacterial genera, namely Acinetobacter, Bacillus, Citrobacter, Paenibacillus, and Serratia. The B. cereus strain TWV503 was considered to be the most active cellulolytic bacterium based on the CMC agar method. B. cereus strain TWV503 showed CMCase activity at 2.190 ± 0.063 U/mL of CMCase and 0.276 ± 0.031 U/mL of FPase. The optimum temperature and pH for CMCase activity were 50°C and the neutral pH ranging from 7.0 to 8.0, respectively. CMCase activity remained stable at up to 70°C and neutral pH ranging from 7.0 to 8.0 for 24 hours of incubation. This study revealed novel information related to cellulolytic bacteria isolated from the gut of Odontotermes termites collected from Thailand.

A genomic perspective on the potential of termite-associated Cellulosimicrobium cellulans MP1 as producer of plant biomass-acting enzymes and exopolysaccharides.

BackgroundLignocellulose is a renewable and enormous biomass resource, which can be degraded efficiently by a range of cocktails of carbohydrate-active enzymes secreted by termite gut symbiotic bacteria. There is an urgent need to find enzymes with novel characteristics for improving the conversion processes in the production of lignocellulosic-based products. Although various studies dedicated to the genus Cellulosimicrobium as gut symbiont, genetic potential related to plant biomass-acting enzymes and exopolysaccharides production has been fully untapped to date.MethodsThe cellulolytic bacterial strain MP1 was isolated from termite guts and identified to the species level by phenotypic, phylogenetic, and genomic analysis. To further explore genes related to cellulose and hemicellulose degradation, the draft genome of strain MP1 was obtained by using whole-genome sequencing, assembly, and annotation through the Illumina platform. Lignocellulose degrading enzymes and levan production in the liquid medium were also examined to shed light on bacterial activities.ResultsAmong 65 isolates obtained, the strain MP1 was the most efficient cellulase producer with cellulase activity of 0.65 ± 0.02 IU/ml. The whole genome analysis depicted that strain MP1 consists of a circular chromosome that contained 4,580,223 bp with an average GC content of 73.9%. The genome comprises 23 contigs including 67 rRNA genes, three tRNA genes, a single tmRNA gene, and 4,046 protein-coding sequences. In support of the phenotypic identification, the 16S rRNA gene sequence, average nucleotide identity, and whole-genome-based taxonomic analysis demonstrated that the strain MP1 belongs to the species Cellulosimicrobium cellulans. A total of 30 genes related to the degradation of cellulases and hemicellulases were identified in the C. cellulans MP1 genome. Of note, the presence of sacC1-levB-sacC2-ls operon responsible for levan and levan-type fructooligosaccharides biosynthesis was detected in strain MP1 genome, but not with closely related C. cellulans strains, proving this strain to be a potential candidate for further studies. Endoglucanases, exoglucanases, and xylanase were achieved by using cheaply available agro-residues such as rice bran and sugar cane bagasse. The maximum levan production by C. cellulans MP1 was 14.8 ± 1.2 g/l after 20 h of cultivation in media containing 200 g/l sucrose. To the best of our knowledge, the present study is the first genome-based analysis of a Cellulosimicrobium species which focuses on lignocellulosic enzymes and levan biosynthesis, illustrating that the C. cellulans MP1 has a great potential to be an efficient platform for basic research and industrial exploitation.

Termite gas emissions select for hydrogenotrophic microbial communities in termite mounds

Organoheterotrophs are the dominant bacteria in most soils worldwide. While many of these bacteria can subsist on atmospheric hydrogen (H2), levels of this gas are generally insufficient to sustain hydrogenotrophic growth. In contrast, bacteria residing within soil-derived termite mounds are exposed to high fluxes of H2 due to fermentative production within termite guts. Here, we show through community, metagenomic, and biogeochemical profiling that termite emissions select for a community dominated by diverse hydrogenotrophic Actinobacteriota and Dormibacterota. Based on metagenomic short reads and derived genomes, uptake hydrogenase and chemosynthetic RuBisCO genes were significantly enriched in mounds compared to surrounding soils. In situ and ex situ measurements confirmed that high- and low-affinity H2-oxidizing bacteria were highly active in the mounds, such that they efficiently consumed all termite-derived H2 emissions and served as net sinks of atmospheric H2 Concordant findings were observed across the mounds of three different Australian termite species, with termite activity strongly predicting H2 oxidation rates (R2 = 0.82). Cell-specific power calculations confirmed the potential for hydrogenotrophic growth in the mounds with most termite activity. In contrast, while methane is produced at similar rates to H2 by termites, mounds contained few methanotrophs and were net sources of methane. Altogether, these findings provide further evidence of a highly responsive terrestrial sink for H2 but not methane and suggest H2 availability shapes composition and activity of microbial communities. They also reveal a unique arthropod-bacteria interaction dependent on H2 transfer between host-associated and free-living microbial communities.

Characterization of Lignocellulolytic Bacteria from Gut of Termite (Isoptera: Rhinotermitidae and Termitidae)

A total of 10 bacterial isolates have been isolated from the gut of termites (Isoptera: Rhinotermitidae and Termitidae) and are known to have the ability to produce lignocellulolytic enzymes consisting of cellulase, laccase, and lignin peroxidase. The enzymatic ability allows these bacteria to be used as a source of new enzymes in the industrial world. However, further research on the character of bacteria to support identification has not been carried out. This study aims to characterize lignocellulolytic bacteria in the gut of termites morphologically and biochemically. Morphological observations were carried out including colony shape, colony edge, colony color, growth type, bacterial cell shape, and gram staining. Meanwhile, the biochemical characterization carried out included glucose fermentation test, indole formation, starch hydrolysis, catalase test, and nitrate reduction. The results showed that ten isolates of lignocellulolytic bacteria from the gut of termites (Isoptera: Rhinotermitidae and Termitidae) grew facultatively anaerobic, had almost similar morphological characters, with colony shapes including irregular and filamentous, colony edges in the form of lobate, undulate, and filamentous, and coloration. Colonies are white to yellowish white. The result of gram staining showed that most of the bacteria were gram positive bacteria with coccus and bacillus shaped bacterial cells. Biochemical analysis showed that these bacteria have the ability to ferment glucose, hydrolyze starch, reduce hydrogen peroxide (Hâ‚‚Oâ‚‚ 30%) and reduce nitrate.ÂÂ

Qualitative and Molecular Screening of Potential Ligninolytic Microbes from Termite (Coptotermes curvignathus) Gut

Ligninolytic microbes have great potential in converting high lignin by-products to more utilisable products by decomposing the lignin-rich agricultural and industrial wastes. Thus, the aim of this study are to screen and identify the potential ligninolytic microbes from the termite (Coptotermes curvignathus) gut. The study was conducted at Universiti Putra Malaysia Bintulu Sarawak Campus, Malaysia. Twenty-seven microbes isolated from termite gut obtained from the Microbiology Laboratory, Faculty of Agricultural Science and Forestry, were used for the ligninolytic activity screening. Media with four different ligninolytic indicator dyes (Azure B, phenol red, methylene blue, and Remazol Brilliant Blue) were streaked with microbial isolates and incubated at 37 °C for 48 h. Out of twenty-seven microbe isolates, only three (CH2, CH5, and CH9) isolates showed decolourisation zone indicating the positive presence of ligninolytic activity. The 16S rRNA gene sequence data indicated the isolates are highly homologous to Bacillus spp.