Symbiotic Gut Bacteria Research Articles

Valorization Potential of a Novel Bacterial Strain, Bacillus altitudinis RSP75, towards Lignocellulose Bioconversion: An Assessment of Symbiotic Bacteria from the Stored Grain Pest, Tribolium castaneum.

Bioconversion of lignocellulose into renewable energy and commodity products faces a major obstacle of inefficient saccharification due to its recalcitrant structure. In nature, lignocellulose is efficiently degraded by some insects, including termites and beetles, potentially due to the contribution from symbiotic gut bacteria. To this end, the presented investigation reports the isolation and characterization of cellulolytic bacteria from the gut system of red flour beetle, Tribolium castaneum. Out of the 15 isolated bacteria, strain RSP75 showed the highest cellulolytic activities by forming a clearance zone of 28 mm in diameter with a hydrolytic capacity of ~4.7. The MALDI-TOF biotyping and 16S rRNA gene sequencing revealed that the strain RSP75 belongs to Bacillus altitudinis. Among the tested enzymes, B. altitudinis RSP75 showed maximum activity of 63.2 IU/mL extract for xylanase followed by β-glucosidase (47.1 ± 3 IU/mL extract) which were manifold higher than previously reported activities. The highest substrate degradation was achieved with wheat husk and corn cob powder which accounted for 69.2% and 54.5%, respectively. The scanning electron microscopy showed adhesion of the bacterial cells with the substrate which was further substantiated by FTIR analysis that depicted the absence of the characteristic cellulose bands at wave numbers 1247, 1375, and 1735 cm−1 due to hydrolysis by the bacterium. Furthermore, B. altitudinis RSP75 showed co-culturing competence with Saccharomyces cerevisiae for bioethanol production from lignocellulose as revealed by GC-MS analysis. The overall observations signify the gut of T. castaneum as a unique and impressive reservoir to prospect for lignocellulose-degrading bacteria that can have many biotechnological applications, including biofuels and biorefinery.

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.

A natural symbiotic bacterium drives mosquito refractoriness to Plasmodium infection via secretion of an antimalarial lipase.

The stalling global progress in the fight against malaria prompts the urgent need to develop new intervention strategies. Whilst engineered symbiotic bacteria have been shown to confer mosquito resistance to parasite infection, a major challenge for field implementation is to address regulatory concerns. Here, we report the identification of a Plasmodium-blocking symbiotic bacterium, Serratia ureilytica Su_YN1, isolated from the midgut of wild Anopheles sinensis in China that inhibits malaria parasites via secretion of an antimalarial lipase. Analysis of Plasmodium vivax epidemic data indicates that local malaria cases in Tengchong (Yunnan province, China) are significantly lower than imported cases and importantly, that the local vector A. sinensis is more resistant to infection by P. vivax than A. sinensis from other regions. Analysis of the gut symbiotic bacteria of mosquitoes from Yunnan province led to the identification of S. ureilytica Su_YN1. This bacterium renders mosquitoes resistant to infection by the human parasite Plasmodium falciparum or the rodent parasite Plasmodium berghei via secretion of a lipase that selectively kills parasites at various stages. Importantly, Su_YN1 rapidly disseminates through mosquito populations by vertical and horizontal transmission, providing a potential tool for blocking malaria transmission in the field.

Evaluation of cellulose degrading bacteria isolated from the gut-system of cotton bollworm, Helicoverpa armigera and their potential values in biomass conversion

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.

Antibiotic Treatment Decrease the Fitness of Honeybee (Apis mellifera) Larvae.

Simple SummaryTo determine the biologic function of gut bacteria with no host specificity in honeybee larvae, honeybee larvae were treated with antibiotics for disrupting the gut bacteria. Then, the body weight, development time, and expression of nutrient metabolism genes and immune genes of honeybee larvae were investigated. The results demonstrated that the disruption of gut microbiota by antibiotics weakened the nutrient metabolism, decreased the body weight, extended the development process, and decrease the immune competence of honeybee larvae, indicating the vital roles of gut bacteria in bee larvae fitness.Symbiotic bacteria could increase the nutrient provision, regulate the physiological state, and promote immunity in their insect host. Honeybee larvae harbor plenty of bacteria in their gut, but their functions are not well studied. To determine their effect on honeybee larvae, the 1-day-old larvae were grafted on to 24-well plates from the comb and artificially reared in the lab. They were treated with penicillin–streptomycin to remove the gut symbiotic bacteria. Then, the 5-day-old larvae and the newly emerged adults were weighted. The developmental periods to pupae and eclosion were investigated, respectively. The bacterial amount, expression of developmental regulation genes (ecr and usp), nutrient metabolism genes (ilp1, ilp2, hex 70a, hex 70b, hex 70c, and hex 110), and immunity genes (apidaecin, abaecin, defensin-1, and hymenoptaecin) were determined by qRT-PCR. The result showed that the antibiotics-treated larvae have significantly lower body weights in the 5-day-old larvae and the emerged bees. The expression of ilp2 and hex 70c in 5-day-old larvae was down-regulated. The usp was down-regulated in 5-day-old larvae, but increased in 7-day-old larvae, which disturbed the normal developmental process and caused the extension of eclosion. Moreover, antibiotics treatment significantly decreased the expression of apidaecin and abaecin in 5-day-old larvae, and defensin-1 and hymenoptaecin in 7-day-old larvae, respectively. These results showed that antibiotics could weaken the nutrient metabolism, disturb the development process, and decrease the immune competence of honeybee larvae, indicating the vital roles of gut bacteria in bee larvae fitness, so the antibiotics should be avoided to control microbial disease in honeybee larvae.

Hindgut microbiota reflects different digestive strategies in dung beetles (Coleoptera: Scarabaeidae: Scarabaeinae).

Gut microbes play an important role in the biology and evolution of insects. Australian native dung beetles (Scarabaeinae) present an opportunity to study gut microbiota in an evolutionary context as they come from two distinct phylogenetic lineages and some species in each lineage have secondarily adapted to alternative or broader diets. In this study, we characterised the hindgut bacterial communities found in 21 species of dung beetles across two lineages using 16S rRNA sequencing. We found that gut microbial diversity was more dependent on host phylogeny and gut morphology than specific dietary preferences or environment. In particular, gut microbial diversity was highest in the endemic, flightless genus Cephalodesmius that feeds on a broad range of composted organic matter. The hindgut of Cephalodesmius harbours a highly conserved core set of bacteria suggesting that the bacteria are symbiotic. Symbiosis is supported by the persistence of the core microbiota across isolated beetle populations and between species in the genus. A co-evolutionary relationship is supported by the expansion of the hindgut to form a fermentation chamber and the fermentative nature of the core microbes. In contrast, Australian species of the widespread dung beetle genus Onthophagus, specialise on a single food resource such as dung or fungus, exhibit minimal food processing behaviour, have a short, narrow hindgut and a variable gut microbiota with relatively few core bacterial taxa. A conserved, complex gut microbiota is hypothesised to be unnecessary for this highly mobile genus.IMPORTANCE Dung beetles are a very important part of an ecosystem because of their role in the removal and decomposition of vertebrate dung. It has been suspected that symbiotic gut bacteria facilitate this role, a hypothesis that we have explored with high throughput barcoding. We found that differences in hindgut morphology had the greatest effect on the bacterial community composition. Species with a hindgut fermentation chamber harboured a distinctly different hindgut community compared to those species with a narrow, undifferentiated hindgut. Diet and phylogeny were also associated with differences in gut community. Further understanding of the relationships between dung beetles and their gut microbes will provide insights into the evolution of their behaviours and how gut communities contribute to their fitness.

Composition and Diversity of Gut Bacteria Associated with the Eri Silk Moth, Samia ricini, (Lepidoptera: Saturniidae) as Revealed by Culture-Dependent and Metagenomics Analysis.

The polyphagous eri silk moth, Samia ricini, is associated with various symbiotic gut bacteria believed to provide several benefits to the host. The larvae of S. ricini were subjected to isolation of gut bacteria using culture-dependent 16S rRNA generic characterization, metagenomics analysis and qualitative enzymatic assays. Sixty culturable aerobic gut bacterial isolates comprising Firmicutes (54%) and Proteobacteria (46%); and twelve culturable facultative anaerobic bacteria comprising Proteobacteria (92%) and Firmicutes (8%) were identified inhabiting the gut of S. ricini. The results of metagenomics analysis revealed the presence of a diverse community of both culturable and un-culturable gut bacteria belonging to Proteobacteria (60%) and Firmicutes (20%) associated with seven orders. An analysis of the results of culturable isolation indicates that these bacterial isolates inhabited all the three compartments of the gut. Investigation on persistence of bacteria coupled with metagenomics analysis of the fifth instar suggested that bacteria persist in the gut across the different instar stages. In addition, enzymatic assays indicated that 48 and 75% of culturable aerobic, and 75% of anaerobic gut bacterial isolates had cellulolytic, lipolytic and nitrate reductase activities, thus suggesting that they may be involved in food digestion and nutritional provision to the host. These bacterial isolates may be good sources for profiling novel genes and biomolecules for biotechnological application.

Gut microbiota promotes host resistance to low-temperature stress by stimulating its arginine and proline metabolism pathway in adult Bactrocera dorsalis.

Gut symbiotic bacteria have a substantial impact on host physiology and ecology. However, the contribution of gut microbes to host fitness during long-term low-temperature stress is still unclear. This study examined the role of gut microbiota in host low-temperature stress resistance at molecular and biochemical levels in the oriental fruit fly Bactrocera dorsalis. The results showed that after the gut bacteria of flies were removed via antibiotic treatment, the median survival time was significantly decreased to approximately 68% of that in conventional flies following exposure to a temperature stress of 10°C. Furthermore, we found that Klebsiella michiganensis BD177 is a key symbiotic bacterium, whose recolonization in antibiotic treated (ABX) flies significantly extended the median survival time to 160% of that in the ABX control, and restored their lifespan to the level of conventional flies. Notably, the relative levels of proline and arginine metabolites were significantly downregulated by 34- and 10-fold, respectively, in ABX flies compared with those in the hemolymph of conventional flies after exposure to a temperature stress of 10°C whereas recolonization of ABX flies by K. michiganensis BD177 significantly upregulated the levels of proline and arginine by 13- and 10- fold, respectively, compared with those found in the hemolymph of ABX flies. qPCR analysis also confirmed that K. michiganensis-recolonized flies significantly stimulated the expression of transcripts from the arginine and proline metabolism pathway compared with the ABX controls, and RNAi mediated silencing of two key genes Pro-C and ASS significantly reduced the survival time of conventional flies, postexposure low-temperature stress. We show that microinjection of L-arginine and L-proline into ABX flies significantly increased their survival time following exposure to temperature stress of 10°C. Transmission electron microscopy (TEM) analysis further revealed that low-temperature stress caused severe destruction in cristae structures and thus resulted in abnormal circular shapes of mitochondria in ABX flies gut, while the recolonization of live K. michiganensis helped the ABX flies to maintain mitochondrial functionality to a normal status, which is important for the arginine and proline induction. Our results suggest that gut microbiota plays a vital role in promoting the host resistance to low-temperature stress in B. dorsalis by stimulating its arginine and proline metabolism pathway.

Sublethal Doses of Essential Oils Impact Growth and Esterase Levels in German Cockroach (Blattodea: Ectobiidae) with and without Symbiotic Bacteria

The inhibition by three essential oils of the growth and esterase production of German cockroaches, Blattella germanica (L.) (Blattodea: Ectobiidae), with and without symbiotic bacteria, was determined in laboratory assays. Essential oils extracted from Lilium brownii var. viridulum Baker, Croton tiglium L., and Lonicera japonica Thunberg were mixed with a powdered rat food at a 10% concentration and fed to cockroach nymphs. Body mass and body length were measured in adults in all three treatments. Acetylcholinesterase and nonspecific esterase activities were compared between the Lilium and Croton essential oil treatments. The nonlethal doses of these essential oils reduced cockroach growth, while the growth of the females lacking symbiotic bacteria was significantly reduced when fed the diet containing Lilium essential oil. Lilium and Croton essential oils significantly inhibited esterase activity. Our results indicate that symbiotic gut bacteria might be involved in mediating detoxification in the German cockroach.

Modulation of microbially derived short-chain fatty acids on intestinal homeostasis, metabolism, and neuropsychiatric disorder.

A diverse range of symbiotic gut bacteria codevelops with the host and is considered a metabolic "organ" that not only facilitates harvesting of nutrients from the dietary components but also produces a class of metabolites. Many metabolites of gut microbes have an important impact on host health. For example, an inventory of metabolic intermediates derived from bacterial protein fermentation may affect host physiology and pathophysiology. Additionally, gut microbiota can convert cholesterol to bile acids and further into secondary bile acids which can conversely modulate microbial community. Moreover, new research identifies that microbes synthesize vitamins for us in the colon. Here, we will review data implicating a major class of bacterial metabolites through breaking down dietary fiber we cannot process, short-chain fatty acids (SCFAs), as crucial executors of alteration of immune mechanisms, regulation of metabolic homeostasis, and neuroprotective effects to combat disease and improve health.

A Gut Symbiotic Bacterium Serratia marcescens Renders Mosquito Resistance to Plasmodium Infection Through Activation of Mosquito Immune Responses.

The malaria development in the mosquito midgut is a complex process that results in considerable parasite losses. The mosquito gut microbiota influences the outcome of pathogen infection in mosquitoes, but the underlying mechanisms through which gut symbiotic bacteria affect vector competence remain elusive. Here, we identified two Serratia strains (Y1 and J1) isolated from field-caught female Anopheles sinensis from China and assessed their effect on Plasmodium development in An. stephensi. Colonization of An. stephensi midgut by Serratia Y1 significantly renders the mosquito resistant to Plasmodium berghei infection, while Serratia J1 has no impact on parasite development. Parasite inhibition by Serratia Y1 is induced by the activation of the mosquito immune system. Genome-wide transcriptomic analysis by RNA-seq shows a similar pattern of midgut gene expression in response to Serratia Y1 and J1 in sugar-fed mosquitoes. However, 24 h after blood ingestion, Serratia Y1 modulates more midgut genes than Serratia J1 including the c-type lectins (CTLs), CLIP serine proteases and other immune effectors. Furthermore, silencing of several Serratia Y1-induced anti-Plasmodium factors like the thioester-containing protein 1 (TEP1), fibrinogen immunolectin 9 (FBN9) or leucine-rich repeat protein LRRD7 can rescue parasite oocyst development in the presence of Serratia Y1, suggesting that these factors modulate the Serratia Y1-mediated anti-Plasmodium effect. This study enhances our understanding of how gut bacteria influence mosquito-Plasmodium interactions.

Diversity and evolution of bacterial symbionts in the gut symbiotic organ of jewel stinkbugs (Hemiptera: Scutelleridae)

The majority of plant-sucking stinkbugs of the superfamily Pentatomoidea possess numerous crypts in the midgut as the symbiotic organ, where specific and beneficial symbiotic bacteria are harbored extracellularly. The host–symbiont relationships are co-speciating in the families Plataspidae, Acanthosomatidae and Urostylididae, but promiscuous in the families Pentatomidae and Cydnidae. As for the family Scutelleridae, only a few species have been examined for their gut symbiotic bacteria. Here, we comprehensively investigated Japanese scutellerid stinkbugs representing 6 genera, 8 species, and 24 individuals. Molecular phylogenetic analysis revealed that the scutellerid gut symbionts are polyphyletic, consisting of at least seven distinct phylogenetic groups in the Gammaproteobacteria. In four of the seven groups, the symbionts were closely related to each other, to the pentatomid gut symbionts, and to environmental bacteria of the genus Pantoea. These results strongly suggest that the scutellerid gut symbionts are of multiple evolutionary origins, presumably entailing repeated symbiont acquisitions, replacements and/or horizontal transfers in a promiscuous manner. Elimination of the gut symbionts from Lampromicra miyakona resulted in high mortality without adult emergence, confirming that the gut symbionts are of beneficial nature. In conclusion, the host–symbiont associations in the Scutelleridae seem to be similar to those in the Pentatomidae and Cydnidae.

Abundance and diversity of gut-symbiotic bacteria, the genus Burkholderia in overwintering Riptortus pedestris (Hemiptera: Alydidae) populations and soil in South Korea.

Riptortus pedestris is a major agricultural pest on leguminous plants in South Korea and Japan. Recent studies have revealed that R. pedestris can form beneficial symbiosis with bacteria belonging to genus Burkholderia acquired from soil newly for every generation. Although their physiological interactions are relatively well-understood, infection rate and abundance of the Burkholderia in overwintering natural populations of R. pedestris remain unknown. Therefore, the objective of this study was to characterize Burkholderia infection ratio and clade composition of overwintering R. pedestris populations as well as prevalence and diversity of the genus Burkholderia in soil by conducting a two-year field survey. From the field survey, we found 29 overwintering R. pedestris adults in forested areas nearby soybean fields. Diagnostic PCR analysis revealed that overall infection rate of the symbiotic Burkholderia was 93.1% from overwintering adults. Among the Burkholderia-infected R. pedestris, 70.4% of individuals harbored unclassified Burkholderia clades whereas 22.2% and 7.4% of R. pedestris harbor stinkbug-associated beneficial and environmental (SBE) group and Burkholderia cepacia and complex (BCC), respectively. All R. pedestris were infected with a single clade of Burkholderia. In soil, 56.2% of soil samples were Burkholderia positive, and unlike R. pedestris, multiple Burkholderia clades were detected from 62.2% of those samples. Clade composition of the genus Burkholderia in the samples with the bacteria was 91.1%, 60.0%, 31.1% and 8.8% for plant-associated beneficial and environment (PBE), BCC, SBE and unclassified clade, respectively.

Burkholderia gut symbiont modulates titer of specific juvenile hormone in the bean bug Riptortus pedestris

Recent studies have provided molecular evidence that gut symbiotic bacteria modulate host insect development, fitness and reproduction. However, the molecular mechanisms through which gut symbionts regulate these aspects of host physiology remain elusive. To address these questions, we prepared two different Riptortus-Burkholderia insect models, Burkholderia gut symbiont-colonized (Sym) Riptortus pedestris insects and gut symbiont-noncolonized (Apo) insects. Upon LC-MS analyses, juvenile hormone III skipped bisepoxide (JHSB3) was newly identified from Riptortus Apo- and Sym-female and male adults’ insect hemolymph and JHSB3 titer in the Apo- and Sym-female insects were measured because JH is important for regulating reproduction in adult insects. The JHSB3 titer in the Sym-females were consistently higher compared to those of Apo-females. Since previous studies reported that Riptortus hexamerin-α and vitellogenin proteins were upregulated by the topical abdominal application of a JH-analog, chemically synthesized JHSB3 was administered to Apo-females. As expected, the hexamerin-α and vitellogenin proteins were dramatically increased in the hemolymph of JHSB3-treated Apo-females, resulting in increased egg production compared to that in Sym-females. Taken together, these results demonstrate that colonization of Burkholderia gut symbiont in the host insect stimulates biosynthesis of the heteroptera-specific JHSB3, leading to larger number of eggs produced and enhanced fitness in Riptortus host insects.

Isolation, characterization and mode of action of a larvicidal compound, 22-hydroxyhopane from Adiantum latifolium Lam. against Oryctes rhinoceros Linn.

Oryctes rhinoceros Linn. is one of the most serious pests of coconuts and other palms. Following bioassay guided method, a larvicidal compound, 22-hydroxyhopane has been isolated for the first time from methanol extract of leaves of Adiantum latifolium Lam. against the pest (LC50 value 20.81 μg/g). It is a hopanoid triterpene with molecular mass of 442.42 g/mol. The compound exhibited antibacterial activity against symbiotic gut bacteria, caused histolysis of midgut tissues and inhibited secretion of digestive enzymes such as protease, amylase and trehalase resulting in weight loss of larvae. Enzyme immunoassay showed an elevation of 20-hydroxyecdysone level in haemolymph causing disruption of metamorphosis of larvae.

Interrogation of Human Milk Oligosaccharide Fucosylation Patterns for Antimicrobial and Antibiofilm Trends in Group B Streptococcus.

For newborns, human milk oligosaccharides (HMOs) serve as an important source of protection against bacterialpathogens. HMOs prevent infection by functioning as decoy receptors that bind pathogens to inhibit cellular adhesion. HMOs also play a protective role by acting as prebiotics that selectively promote the growth of symbiotic gut bacteria over pathogens. Fucosylated HMOs in particular are well-known for their roles as both decoy receptors and prebiotics. Recently, we discovered that HMOs possess antimicrobial activity against Group B Streptococcus (GBS) by increasing cellular permeability. HMO extracts from a single donor can contain over 100 different structures; however, studies using heterogeneous HMO mixtures do not provide insight into the specific structural requirements needed to achieve antimicrobial activity. In this study, we address this void by completing a structure activity study on the antimicrobial and antibiofilm activities of six neutral, fucosylated and five neutral, nonfucosylated HMOs against GBS. We determined that while the presence of fucose alone does not correlate to antimicrobial activity, the location and degree of fucosylation does play a key role in the antimicrobial activityof HMOs. Moreover, the antimicrobial and antibiofilm activities of single HMOs were found to be strain-specific. This further supports our vision of developing narrow-spectrum antibacterial agents against GBS.

Larvicidal efficacy of Adiantobischrysene from Adiantum latifolium against Oryctes rhinoceros through disrupting metamorphosis and impeding microbial mediated digestion.

Oryctes rhinoceros Linn. (Coleoptera: Scarabaeidae) is a serious pest of coconuts and other palms. Symbiotic gut bacteria play significant roles in the digestion of cellulosic materials as well as in some other physiological processes essential for the existence of O. rhinoceros larvae. The study was undertaken to isolate a compound with antibacterial and larvicidal activities from the leaves of Adiantum latifolium Lam. following a bioassay-guided method. Methanol extract (ME) of dry leaf powder of A. latifolium showed larvicidal activity against third-instar O. rhinoceros (LD50 , 5018 mg/kg) with antibacterial activity on its gut microbiota. An in vitro study showed the bacteria Bacillus cereus, Micrococcus lylae, Stenotrophomonas maltophilia, Kocuria rosea, Burkholderia mallei, Staphylococcus epidermidis, S. arlettae and Corynebacterium afermentans identified from the larval gut were sensitive to ME. Bioactivity-guided isolation of the compound by liquid-liquid extraction and column chromatography resulted in Adiantobischrysene which showed antibacterial and larvicidal activity (LD50 , 8.4 mg/kg) and led to weight loss and precocious metamorphosis in larvae. An enzyme immunoassay showed a large peak in 20-hydroxyecdysone that commits larvae to precocious metamorphosis. This study demonstrated that the antibacterial and metamorphosis disrupting activity of Adiantobischrysene make it a natural pesticidal compound against O. rhinoceros. © 2018 Society of Chemical Industry.

Environmental factors have a strong impact on the composition and diversity of the gut bacterial community of Chinese black honeybees

The Chinese black honeybees is an ecotype of the European honeybee that is formed by the natural hybridization of Apis mellifera mellifera and A. m. carnica. It is distributed in nature reserves in North China and has been an important breeding resource for disease resistance and other desirable traits. Compared to the areas outside of reserves, the nature reserves offer significant biodiversity benefits not only to the Chinese black honey bees but also to the other valuable plants and animals. In recent years there has been growing evidence that environmental factors including food choices play an important role in shaping the composition and activity of gut microbiota, which in turn can impact host health. In the previous studies on Chinse black honeybees, little attention has been paid to the diverse population of microbes in the gut that play a vital role in host health. In order to achieve a better understanding on the role of environmental factors in diversity and composition of gut microbiota of honey bees, in the present study, we analyzed the gut bacterial communities of Chinese black honeybees using terminal restriction fragment length polymorphism (T-RFLP). The results showed that the samples from the national nature reserves that are protected and managed so as to preserve and enrich their natural condition and resources for Chinese black honeybees had higher variety and richness of gut bacteria than that collected from unreserved regions that also harbor populations of Chinese black honeybees. The four terminal restriction fragments (T-RFs), 201, 223, 247 and 320bp, were identified to be the dominant bacteria of Chinese black honeybees. Of which 247 and 320bp had greater differences between bee groups sampled in different regions and therefore could be used as genetic markers to separate samples collected from the national nature reserves to samples collected from unreserved regions. The results clearly indicate that national nature reserve protects biological diversity and ecological and evolutionary processes which have had a significant influence on the diversity of gut bacteria of Chinese black honeybees. The ubiquity of gut symbiotic bacteria identified in Chinese black honeybee suggests that environmental factors could play an important role in diversity and composition of gut bacteria and warrant further investigation into the functional significance of these gut bacteria for the honeybee health.

Optimization of a metatranscriptomic approach to study the lignocellulolytic potential of the higher termite gut microbiome

BackgroundThanks to specific adaptations developed over millions of years, the efficiency of lignin, cellulose and hemicellulose decomposition of higher termite symbiotic system exceeds that of many other lignocellulose utilizing environments. Especially, the examination of its symbiotic microbes should reveal interesting carbohydrate-active enzymes, which are of primary interest for the industry. Previous metatranscriptomic reports (high-throughput mRNA sequencing) highlight the high representation and overexpression of cellulose and hemicelluloses degrading genes in the termite hindgut digestomes, indicating the potential of this technology in search for new enzymes. Nevertheless, several factors associated with the material sampling and library preparation steps make the metatranscriptomic studies of termite gut prokaryotic symbionts challenging.MethodsIn this study, we first examined the influence of the sampling strategy, including the whole termite gut and luminal fluid, on the diversity and the metatranscriptomic profiles of the higher termite gut symbiotic bacteria. Secondly, we evaluated different commercially available kits combined in two library preparative pipelines for the best bacterial mRNA enrichment strategy.ResultsWe showed that the sampling strategy did not significantly impact the generated results, both in terms of the representation of the microbes and their transcriptomic profiles. Nevertheless collecting luminal fluid reduces the co-amplification of unwanted RNA species of host origin. Furthermore, for the four studied higher termite species, the library preparative pipeline employing Ribo-Zero Gold rRNA Removal Kit “Epidemiology” in combination with Poly(A) Purist MAG kit resulted in a more efficient rRNA and poly-A-mRNAdepletion (up to 98.44% rRNA removed) than the pipeline utilizing MICROBExpress and MICROBEnrich kits. High correlation of both Ribo-Zero and MICROBExpresse depleted gene expression profiles with total non-depleted RNA-seq data has been shown for all studied samples, indicating no systematic skewing of the studied pipelines.ConclusionsWe have extensively evaluated the impact of the sampling strategy and library preparation steps on the metatranscriptomic profiles of the higher termite gut symbiotic bacteria. The presented methodological approach has great potential to enhance metatranscriptomic studies of the higher termite intestinal flora and to unravel novel carbohydrate-active enzymes.

Host plant species determines symbiotic bacterial community mediating suppression of plant defenses

Herbivore associated bacteria are vital mediators of plant and insect interactions. Host plants play an important role in shaping the gut bacterial community of insects. Colorado potato beetles (CPB; Leptinotarsa decemlineata) use several Solanum plants as hosts in their natural environment. We previously showed that symbiotic gut bacteria from CPB larvae suppressed jasmonate (JA)-induced defenses in tomato. However, little is known about how changes in the bacterial community may be involved in the manipulation of induced defenses in wild and cultivated Solanum plants of CPB. Here, we examined suppression of JA-mediated defense in wild and cultivated hosts of CPB by chemical elicitors and their symbiotic bacteria. Furthermore, we investigated associations between the gut bacterial community and suppression of plant defenses using 16 S rRNA amplicon sequencing. Symbiotic bacteria decreased plant defenses in all Solanum hosts and there were different gut bacterial communities in CPB fed on different host plants. When larvae were reared on different hosts, defense suppression differed among host plants. These results demonstrate that host plants influence herbivore gut bacterial communities and consequently affect the herbivore’s ability to manipulate JA-mediated plant defenses. Thus, the presence of symbiotic bacteria that suppress plant defenses might help CPB adapt to host plants.