Engineering Gut Symbionts: A Way to Promote Bee Growth?

Bees play a crucial role as pollinators, contributing significantly to ecosystems. However, the honeybee population faces challenges such as global warming, pesticide use, and pathogenic microorganisms. Promoting bee growth using several approaches is therefore crucial for maintaining their roles. To this end, the bacterial microbiota is well-known for its native role in supporting bee growth in several respects. Maximizing the capabilities of these microorganisms holds the theoretical potential to promote the growth of bees. Recent advancements have made it feasible to achieve this enhancement through the application of genetic engineering. In this review, we present the roles of gut symbionts in promoting bee growth and collectively summarize the engineering approaches that would be needed for future applications. Particularly, as the engineering of bee gut symbionts has not been advanced, the dominant gut symbiotic bacteria Snodgrassella alvi and Gilliamella apicola are the main focus of the paper, along with other dominant species. Moreover, we propose engineering strategies that will allow for the improvement in bee growth with listed gene targets for modification to further encourage the use of engineered gut symbionts to promote bee growth.

Characterization and in-vitro plant-based control of hindgut bacteria isolated from Odontotermes obesus Rambur (Termitidae) and Heterotermes indicola Wasmann (Rhinotermitidae).

Termites cause a serious menace to wooden structures all over the world. They rely mostly on entozoic fauna residing in their hindgut for the digestion of cellulosic and hemicellulosic materials. One of the ways to control termites is through their gut symbionts. The present study was designed to characterize the hindgut bacteria isolated from Odontotermes obesus and Heterotermes indicola. Furthermore, the growth inhibitory effect of eight tropical plant extracts was investigated to find out potential control agents for these bacterial isolates. The characterization of bacteria was carried out based on their morphology, Gram staining, biochemical and amplification of 16SrRNA gene. Amplified products were sequenced to confirm their relationship with bacterial isolates from termites of other regions. The growth inhibitory effect of ethanolic leaf extracts of eight plants was evaluated in an invitro agar well diffusion method. Qualitative and quantitative phytochemical analysis of the most effective plant was carried out to learn about bioactive agents. The results confirmed the presence of five bacteria from each termite species. The Bacillus cereus, Escherichia coli, and Lysinibacillus fusiformis were common to both termites whereas Lysinibacillus xylanilyticus and Lysinibacillus macrolides were found in O. obesus only and H. indicola harbor Bacillus subtilis and Shigella sonnei in addition to common three ones. Among the plant extracts of Carica papaya, Eucalyptus camaldulensis, Osmium basilicum, Grevillea robusta, Eucalyptus globulus, Pongamia pinnata, Mentha longifolia, and Melia azedarach, the G. robusta > E. camaldulensis > O. basilicum were found to have growth inhibitory effects with increasing concentrations from 100 to 2000 µg/mL. The biodiversity of the bacterial fauna is important for the biological control of termites. Leaf extracts of these medicinal plants can be used to control termite infestation in an environment-friendly manner to save huge economic loss.

The impact of early-life exposure to three agrochemicals on survival, behavior, and gut microbiota of stingless bees (Partamona helleri).

Over the last few decades, agrochemicals have been partially associated with a global reduction in bees' population. Toxicological assessment is therefore crucial for understanding the overall agrochemical risks to stingless bees. Therefore, the lethal and sublethal effects of agrochemicals commonly used in crops (copper sulfate, glyphosate, and spinosad) on the behavior and gut microbiota of the stingless bee, Partamona helleri, were assessed using chronic exposure during the larval stage. When used at the field-recommended rates, both copper sulfate (200 µg of active ingredient/bee; a.i µg bee-1) and spinosad (8.16 a.i µg bee-1) caused a decrease in bee survival, while glyphosate (148 a.i µg bee-1) did not show any significant effects. No significant adverse effects on bee development were observed in any treatment with CuSO4 or glyphosate, but spinosad (0.08 or 0.03 a.i µg bee -1) increased the number of deformed bees and reduced their body mass. Agrochemicals changed the behavior of bees and composition of the gut microbiota of adult bees, and metals such as copper accumulated in the bees' bodies. The response of bees to agrochemicals depends on the class or dose of the ingested compound. In vitro rearing of stingless bees' larvae is a useful tool to elucidate the sublethal effects of agrochemicals.

Friend or Foe: Symbiotic Bacteria in Bactrocera dorsalis-Parasitoid Associations.

Parasitoids are promising biocontrol agents of the devastating fruit fly, Bactrocera dorsalis. However, parasitoid performance is a function of several factors, including host-associated symbiotic bacteria. Providencia alcalifaciens, Citrobacter freundii, and Lactococcus lactis are among the symbiotic bacteria commonly associated with B. dorsalis, and they influence the eco-physiological functioning of this pest. However, whether these bacteria influence the interaction between this pest and its parasitoids is unknown. This study sought to elucidate the nature of the interaction of the parasitoids, Fopius arisanus, Diachasmimorpha longicaudata, and Psyttlia cosyrae with B. dorsalis as mediated by symbiotic bacteria. Three types of fly lines were used: axenic, symbiotic, and bacteria-mono-associated (Lactococcus lactis, Providencia alcalifaciens, and Citrobacter freundii). The suitable stages of each fly line were exposed to the respective parasitoid species and reared until the emergence of adult flies/parasitoids. Thereafter, data on the emergence and parasitoid fitness traits were recorded. No wasps emerged from the fly lines exposed to P. cosyrae. The highest emergence of F. arisanus and D. longicaudata was recorded in the L. lactis fly lines. The parasitoid progeny from the L. lactis and P. alcalifaciens fly lines had the longest developmental time and the largest body size. Conversely, parasitoid fecundity was significantly lower in the L. lactis lines, whereas the P. alcalifaciens lines significantly improved fecundity. These results elucidate some effects of bacterial symbionts on host-parasitoid interactions and their potential in enhancing parasitoid-oriented management strategies against B. dorsalis.

Xylan Prebiotics and the Gut Microbiome Promote Health and Wellbeing: Potential Novel Roles for Pentosan Polysulfate.

This narrative review highlights the complexities of the gut microbiome and health-promoting properties of prebiotic xylans metabolized by the gut microbiome. In animal husbandry, prebiotic xylans aid in the maintenance of a healthy gut microbiome. This prevents the colonization of the gut by pathogenic organisms obviating the need for dietary antibiotic supplementation, a practice which has been used to maintain animal productivity but which has led to the emergence of antibiotic resistant bacteria that are passed up the food chain to humans. Seaweed xylan-based animal foodstuffs have been developed to eliminate ruminant green-house gas emissions by gut methanogens in ruminant animals, contributing to atmospheric pollution. Biotransformation of pentosan polysulfate by the gut microbiome converts this semi-synthetic sulfated disease-modifying anti-osteoarthritic heparinoid drug to a prebiotic metabolite that promotes gut health, further extending the therapeutic profile and utility of this therapeutic molecule. Xylans are prominent dietary cereal components of the human diet which travel through the gastrointestinal tract as non-digested dietary fibre since the human genome does not contain xylanolytic enzymes. The gut microbiota however digest xylans as a food source. Xylo-oligosaccharides generated in this digestive process have prebiotic health-promoting properties. Engineered commensal probiotic bacteria also have been developed which have been engineered to produce growth factors and other bioactive factors. A xylan protein induction system controls the secretion of these compounds by the commensal bacteria which can promote gut health or, if these prebiotic compounds are transported by the vagal nervous system, may also regulate the health of linked organ systems via the gut-brain, gut-lung and gut-stomach axes. Dietary xylans are thus emerging therapeutic compounds warranting further study in novel disease prevention protocols.

Could termites be hiding a goldmine of obscure yet promising yeasts for energy crisis solutions based on aromatic wastes? A critical state-of-the-art review.

Biodiesel is a renewable fuel that can be produced from a range of organic and renewable feedstock including fresh or vegetable oils, animal fats, and oilseed plants. In recent years, the lignin-based aromatic wastes, such as various aromatic waste polymers from agriculture, or organic dye wastewater from textile industry, have attracted much attention in academia, which can be uniquely selected as a potential renewable feedstock for biodiesel product converted by yeast cell factory technology. This current investigation indicated that the highest percentage of lipid accumulation can be achieved as high as 47.25% by an oleaginous yeast strain, Meyerozyma caribbica SSA1654, isolated from a wood-feeding termite gut system, where its synthetic oil conversion ability can reach up to 0.08 (g/l/h) and the fatty acid composition in yeast cells represents over 95% of total fatty acids that are similar to that of vegetable oils. Clearly, the use of oleaginous yeasts, isolated from wood-feeding termites, for synthesizing lipids from aromatics is a clean, efficient, and competitive path to achieve "a sustainable development" towards biodiesel production. However, the lacking of potent oleaginous yeasts to transform lipids from various aromatics, and an unknown metabolic regulation mechanism presented in the natural oleaginous yeast cells are the fundamental challenge we have to face for a potential cell factory development. Under this scope, this review has proposed a novel concept and approach strategy in utilization of oleaginous yeasts as the cell factory to convert aromatic wastes to lipids as the substrate for biodiesel transformation. Therefore, screening robust oleaginous yeast strain(s) from wood-feeding termite gut system with a set of the desirable specific tolerance characteristics is essential. In addition, to reconstruct a desirable metabolic pathway/network to maximize the lipid transformation and accumulation rate from the aromatic wastes with the applications of various "omics" technologies or a synthetic biology approach, where the work agenda will also include to analyze the genome characteristics, to develop a new base mutation gene editing technology, as well as to clarify the influence of the insertion position of aromatic compounds and other biosynthetic pathways in the industrial chassis genome on the expressional level and genome stability. With these unique designs running with a set of the advanced biotech approaches, a novel metabolic pathway using robust oleaginous yeast developed as a cell factory concept can be potentially constructed, integrated and optimized, suggesting that the hypothesis we proposed in utilizing aromatic wastes as a feedstock towards biodiesel product is technically promising and potentially applicable in the near future.

Development of Common Leaf-Footed Bug Pests Depends on the Presence and Identity of Their Environmentally Acquired Symbionts.

Many beneficial symbioses between bacteria and their terrestrial arthropod hosts are vertically transmitted from mother to offspring, ensuring that the progeny acquire necessary partners. Unusually, in several families of coreoid and lygaeoid bugs (Hemiptera), nymphs must instead ingest the beneficial symbiont, Burkholderia (sensu lato), from the environment early in development. We studied the effects of Burkholderia on development of two species of leaf-footed bug (Coreidae) in the genus Leptoglossus, Leptoglossus zonatus and Leptoglossus phyllopus. We found no evidence for vertical transmission of the symbiont but found stark differences in performance between symbiotic and aposymbiotic individuals. Symbiotic nymphs grew more rapidly, were approximately four times more likely to survive to adulthood than aposymbiotic bugs, and were two times larger. These findings suggest that Burkholderia is an obligate symbiont for the Leptoglossus species. We also tested for variation in fitness effects conferred by four symbiont isolates representing different species within the Burkholderia insect-associated stinkbug beneficial and environmental (SBE) clade. While three isolates conferred similar benefits to hosts, nymphs associated with the fourth isolate grew more slowly and weighed significantly less as adults. The effects of the four isolates were similar for both Leptoglossus species. This work indicates that both Burkholderia acquisition and isolate identity play critical roles in the growth and development of Leptoglossus. IMPORTANCELeptoglossus zonatus and L. phyllopus are important polyphagous pests, and both species have been well-studied but generally without regard to their dependance on a bacterial symbiont. Our results indicate that the central role of Burkholderia in the biology of these insects, as well as in other leaf-footed bugs, should be considered in future studies of coreid life history, ecology, and pest management. Our work suggests that acquisition of Burkholderia is critical for the growth and development of Leptoglossus species. Further, we found that there was variation in performance outcomes according to symbiont identity, even among members of the stinkbug beneficial and environmental clade. This suggests that although environmental acquisition of a symbiont can provide extraordinary flexibility in partner associations, it also carries a risk if the partner is suboptimal.

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.

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.

BACKGROUND: The 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. RESULTS: Out 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. CONCLUSION: The 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.

Regulation of host phenotypic plasticity by gut symbiont communities in the eastern subterranean termite (Reticulitermes flavipes).

Termites are eusocial insects that host a range of prokaryotic and eukaryotic gut symbionts and can differentiate into a range of caste phenotypes. Soldier caste differentiation from termite workers follows two successive molts (worker-presoldier-soldier) that are driven at the endocrine level by juvenile hormone (JH). Although physiological and eusocial mechanisms tied to JH signaling have been studied, the role of gut symbionts in the caste differentiation process is poorly understood. Here, we used the JH analog methoprene in combination with the antibiotic kanamycin to manipulate caste differentiation and gut bacterial loads in Reticulitermes flavipes termites via four bioassay treatments: kanamycin, methoprene, kanamycin+methoprene, and an untreated (negative) control. Bioassay results demonstrated a significantly higher number of presoldiers in the methoprene treatment, highest mortality in kanamycin+methoprene treatment, and significantly reduced protist numbers in all treatments except the untreated control. Bacterial 16S rRNA gene sequencing provided alpha and beta diversity results that mirrored bioassay findings. From ANCOM analysis, we found that several bacterial genera were differentially abundant among treatments. Finally, follow-up experiments showed that if methoprene and kanamycin or untreated termites are placed together, zero or rescued presoldier initiation, respectively, occurs. These findings reveal that endogenous JH selects for symbiont compositions required to successfully complete presoldier differentiation. However, if the gut is voided before the influx of JH, it cannot select for the necessary symbionts that are crucial for molting. Based on these results, we are able to provide a novel example of linkages between gut microbial communities and host phenotypic plasticity.

Unlocking the potential of insect and ruminant host symbionts for recycling of lignocellulosic carbon with a biorefinery approach: a review.

Uprising fossil fuel depletion and deterioration of ecological reserves supply have led to the search for alternative renewable and sustainable energy sources and chemicals. Although first generation biorefinery is quite successful commercially in generating bulk of biofuels globally, the food versus fuel debate has necessitated the use of non-edible feedstocks, majorly waste biomass, for second generation production of biofuels and chemicals. A diverse class of microbes and enzymes are being exploited for biofuels production for a series of treatment process, however, the conversion efficiency of wide range of lignocellulosic biomass (LCB) and consolidated way of processing remains challenging. There were lot of research efforts in the past decade to scour for potential microbial candidate. In this context, evolution has developed the gut microbiota of several insects and ruminants that are potential LCB degraders host eco-system to overcome its host nutritional constraints, where LCB processed by microbiomes pretends to be a promising candidate. Synergistic microbial symbionts could make a significant contribution towards recycling the renewable carbon from distinctly abundant recalcitrant LCB. Several studies have assessed the bioprospection of innumerable gut symbionts and their lignocellulolytic enzymes for LCB degradation. Though, some reviews exist on molecular characterization of gut microbes, but none of them has enlightened the microbial community design coupled with various LCB valorization which intensifies the microbial diversity in biofuels application. This review provides a deep insight into the significant breakthroughs attained in enrichment strategy of gut microbial community and its molecular characterization techniques which aids in understanding the holistic microbial community dynamics. Special emphasis is placed on gut microbial role in LCB depolymerization strategies to lignocellulolytic enzymes production and its functional metagenomic data mining eventually generating the sugar platform for biofuels and renewable chemicals production.

The Gut Microbiota of the Insect Infraorder Pentatomomorpha (Hemiptera: Heteroptera) for the Light of Ecology and Evolution.

The stinkbugs of the infraorder Pentatomomorpha are a group of important plant sap-feeding insects, which host diverse microorganisms. Some are located in their complex morphological midgut compartments, while some within the specialized bacteriomes of insect hosts. This perpetuation of symbioses through host generations is reinforced via the diverse routes of vertical transmission or environmental acquisition of the symbionts. These symbiotic partners, reside either through the extracellular associations in midgut or intracellular associations in specialized cells, not only have contributed nutritional benefits to the insect hosts but also shaped their ecological and evolutionary basis. The stinkbugs and gut microbe symbioses present a valuable model that provides insights into symbiotic interactions between agricultural insects and microorganisms and may become potential agents for insect pest management.

Flexibility and resilience of great tit (Parus major) gut microbiomes to changing diets.

BACKGROUND: Gut microbial communities play important roles in nutrient management and can change in response to host diets. The extent of this flexibility and the concomitant resilience is largely unknown in wild animals. To untangle the dynamics of avian-gut microbiome symbiosis associated with diet changes, we exposed Parus major (Great tits) fed with a standard diet (seeds and mealworms) to either a mixed (seeds, mealworms and fruits), a seed, or a mealworm diet for 4 weeks, and examined the flexibility of gut microbiomes to these compositionally different diets. To assess microbiome resilience (recovery potential), all individuals were subsequently reversed to a standard diet for another 4 weeks. Cloacal microbiomes were collected weekly and characterised through sequencing the v4 region of the 16S rRNA gene using Illumina MiSeq. RESULTS: Initial microbiomes changed significantly with the diet manipulation, but the communities did not differ significantly between the three diet groups (mixed, seed and mealworm), despite multiple diet-specific changes in certain bacterial genera. Reverting birds to the standard diet led only to a partial recovery in gut community compositions. The majority of the bacterial taxa that increased significantly during diet manipulation decreased in relative abundance after reversion to the standard diet; however, bacterial taxa that decreased during the manipulation rarely increased after diet reversal CONCLUSIONS: The gut microbial response and partial resilience to dietary changes support that gut bacterial communities of P. major play a role in accommodating dietary changes experienced by wild avian hosts. This may be a contributing factor to the relaxed association between microbiome composition and the bird phylogeny. Our findings further imply that interpretations of wild bird gut microbiome analyses from single-time point sampling, especially for omnivorous species or species with seasonally changing diets, should be done with caution. The partial community recovery implies that ecologically relevant diet changes (e.g., seasonality and migration) open up gut niches that may be filled by previously abundant microbes or replaced by different symbiont lineages, which has important implications for the integrity and specificity of long-term avian-symbiont associations.

Evolution of Termite Symbiosis Informed by Transcriptome-Based Phylogenies.

Termitidae comprises ∼80% of all termite species [1] that play dominant decomposer roles in tropical ecosystems [2, 3]. Two major events during termite evolution were the loss of cellulolytic gut protozoans in the ancestor of Termitidae and the subsequent gain in the termitid subfamily Macrotermitinae of fungal symbionts cultivated externally in "combs" constructed within the nest [4, 5]. How these symbiotic transitions occurred remains unresolved. Phylogenetic analyses of mitochondrial data previously suggested that Macrotermitinae is the earliest branching termitid lineage, followed soon after by Sphaerotermitinae [6], which cultivates bacterial symbionts on combs inside its nests [7]. This has led to the hypothesis that comb building was an important evolutionary step in the loss of gut protozoa in ancestral termitids [8]. We sequenced genomes and transcriptomes of 55 termite species and reconstructed phylogenetic trees from up to 4,065 orthologous genes of 68 species. We found strong support for a novel sister-group relationship between the bacterial comb-building Sphaerotermitinae and fungus comb-building Macrotermitinae. This key finding indicates that comb building is a derived trait within Termitidae and that the creation of a comb-like "external rumen" involving bacteria or fungi may not have driven the loss of protozoa from ancestral termitids, as previously hypothesized. Instead, associations with gut prokaryotic symbionts, combined with dietary shifts from wood to other plant-based substrates, may have played a more important role in this symbiotic transition. Our phylogenetic tree provides a platform for future studies of comparative termite evolution and the evolution of symbiosis in this taxon.

Symbiotic bacteria affect oviposition behavior in the olive fruit fly Bactrocera oleae.

Microbial associations are widespread across the insects. In the olive fruit fly Bactrocera oleae (Diptera: Tephritidae), vertically transmitted gut symbionts contribute to larval development inside the olive host, and to adult nutrition. Nevertheless, their effect on behavioural decisions of adults is unknown. In this study, we show that symbiotic bacteria affect oviposition behaviour in B. oleae. We studied the effect of different fruits as hosts and different gut-bacteria as gut-symbionts on oviposition attempts and fly development in B. oleae. Untreated flies that had native gut-symbionts attempted oviposition significantly more times than axenic flies as well as flies treated with medfly-associated Pantoea or Klebsiella bacteria. Axenic flies provided with a diet containing the homogenized gut of symbiotic flies recovered the same number of oviposition attempts as their symbiotic counterparts. As for as the different hosts, green olives (unripe) and grapes were preferred while black olives (ripe) elicited the least number of oviposition attempts, with an interactive effect of host and bacterial treatments. It appears that both the host attributes and the native gut-symbionts drive oviposition preference towards green olives in B. oleae. Moreover, both bacterial treatments and hosts significantly affected the development of B. oleae larvae. Though grapes elicited as many oviposition attempts as green olives, they yielded no pupae. Taken together, our results suggest that the intimate association between B. oleae and their gut-microbes, extends beyond nutritional support to behaviour.

Behavior and gut bacteria of Partamona helleri under sublethal exposure to a bioinsecticide and a leaf fertilizer.

The exposure of bees to agrochemicals during foraging and feeding has been associated with their population decline. Sublethal exposure to agrochemicals can affect behavior and the microbiota. Gut microbiota is associated with insect nutritional health, immunocompetence, and is essential for neutralizing the damage caused by pathogens and xenobiotics. Research on the effect of the bioinsecticides and fertilizers on the microbiota of bees remains neglected. In this study, we assessed the sublethal effect of both bioinsecticide spinosad and the fertilizer copper sulfate (CuSO4) on the behavior and gut microbiota in forager adults of the stingless bee Partamona helleri (Friese), which is an important pollinator in the Neotropical region. Behavioral assays and gut microbiota profiles were assessed on bees orally exposed to estimated LC5 values for spinosad and CuSO4. The microbiota were characterized through 16S rRNA gene target sequencing. Acute and oral sublethal exposure to spinosad and CuSO4 did not affect the overall activity, flight take-off, and food consumption. However, CuSO4 decreased bee respiration rate and copper accumulated in exposed bees. Exposure to spinosad increased the proportional abundance of the genus Gilliamella, but CuSO4 did not alter the composition of the gut microbiota. In conclusion, sublethal exposure to CuSO4 induces changes in respiration, and spinosad changes the abundance of gut microorganisms of P. helleri.

Extended mutualism between termites and gut microbes: nutritional symbionts contribute to nest hygiene.

All higher eukaryotes have established symbiotic relationships with diverse microorganisms. One of the most well-characterized symbiotic systems is that of termites and their intestinal microorganisms, which digest cellulose. Recently, diverse types of symbioses between gut microbes and host organisms including humans have received growing attention for various features of their complex interactions beyond nutrition. In termites, researchers are beginning to explore such function of gut symbionts, but only the contribution to internal immunity against entomopathogen is known in a few species. Here, we report that gut symbionts of the dampwood termite Zootermopsis nevadensis protect nests from the spread of the commensal bacterium Serratia marcescens, which has pathogenic potential. Defaunated termites dispersed S. marcescens in the surrounding environment by feeding on the bacteria, which then survived passage through their alimentary tracts, while non-defaunated termites did not. Loss of gut symbionts caused a significant reduction in intestinal acetate, which is an important carbon source for termites. Culture experiments showed that acetate had significant inhibitory effects on S. marcescens at a concentration as low as 12 mM, which indicated that the intestinal acetate of non-defaunated termites (40-130 mM) was capable of suppressing this bacterium. These results suggest that digestive derivatives produced by intestinal symbionts play an essential role in nest hygiene in addition to their nutritional function for termites. Our study provides a better understanding of the multifunctionality of symbiotic relationships in diverse organisms beyond nutrition.

Geographically widespread honeybee-gut symbiont subgroups show locally distinct antibiotic-resistant patterns.

How long-term antibiotic treatment affects host bacterial associations is still largely unknown. The honeybee-gut microbiota has a simple composition, so we used this gut community to investigate how long-term antibiotic treatment affects host-associated microbiota. We investigated the phylogenetic relatedness, genomic content (GC percentage, genome size, number of genes and CRISPR) and antibiotic-resistant genes (ARG) for strains from two abundant members of the honeybee core gut microbiota (Gilliamella apicola and Snodgrassella alvi). Domesticated honeybees are subjected to geographically different management policies, so we used two research apiaries, representing different antibiotic treatment regimens in their apiculture: low antibiotic usage (Norway) and high antibiotic usage (Arizona, USA). We applied whole-genome shotgun sequencing on 48 G. apicola and 22 S. alvi. We identified three predominating subgroups of G. apicola in honeybees from both Norway and Arizona. For G. apicola, genetic content substantially varied between subgroups and distance similarity calculations showed similarity discrepancy between subgroups. Functional differences between subgroups, such as pectin-degrading enzymes (G. apicola), were also identified. In addition, we identified horizontal gene transfer (HGT) of transposon (Tn10)-associated tetracycline resistance (Tet B) across the G. apicola subgroups in the Arizonan honeybees, using interspace polymorphisms in the Tet B determinant. Our results support that honeybee-gut symbiont subgroups can resist long-term antibiotic treatment and maintain functionality through acquisition of geographically distinct antibiotic-resistant genes by HGT.

Symbiogenesis: Beyond the endosymbiosis theory?

Symbiogenesis, literally 'becoming by living together', refers to the crucial role of symbiosis in major evolutionary innovations. The term usually is reserved for the major transition to eukaryotes and to photosynthesising eukaryotic algae and plants by endosymbiosis. However, in some eukaryote lineages endosymbionts have been lost secondarily, showing that symbiosis can trigger a major evolutionary innovation, even if symbionts were lost secondarily. This leads to the intriguing possibility that symbiosis has played a role in other major evolutionary innovations as well, even if not all extant representatives of such groups still have the symbiotic association. We evaluate this hypothesis for two innovations in termites (Termitoidae, also known informally as "Isoptera"): i) the role of flagellate gut protist symbionts in the transition to eusociality from cockroach-like ancestors, and ii) the role of non-gut associated symbionts in the transition to 'higher' termites, characterized by the absence of flagellate gut protists. In both cases we identify a crucial role for symbionts, even though in both cases, subsequently, symbionts were lost again in some lineages. We also briefly discuss additional possible examples of symbiogenesis. We conclude that symbiogenesis is more broadly applicable than just for the endosymbiotic origin of eukaryotes and photosynthetic eukaryotes, and may be a useful concept to acknowledge the important role of symbiosis for evolutionary innovation. However, we do not accept Lynn Margulis's view that symbiogenesis will lead to a paradigm shift from neoDarwinism, as the role of symbiosis in evolutionary change can be integrated with existing theory perfectly.