Pharmaceuticals can directly inhibit the growth of gut bacteria, but the degree to which such interactions manifest in complex community settings is an open question. Here, we compared the effects of 30 drugs on a 32-species synthetic community with their effects on each community member in isolation. While most individual drug-species interactions remained the same in the community context, communal behaviors emerged in 26% of all tested cases. Cross-protection during which drug-sensitive species were protected in community was 6 times more frequent than cross-sensitization, the converse phenomenon. Cross-protection decreased and cross-sensitization increased at higher drug concentrations, suggesting that the resilience of microbial communities can collapse when perturbations get stronger. By metabolically profiling drug-treated communities, we showed that both drug biotransformation and bioaccumulation contribute mechanistically to communal protection. As a proof of principle, we molecularly dissected a prominent case: species expressing specific nitroreductases degraded niclosamide, thereby protecting both themselves and sensitive community members.

The gut microbiota operates at the interface of host-environment interactions to influence human homoeostasis and metabolic networks1-4. Environmental factors that unbalance gut microbial ecosystems can therefore shape physiological and disease-associated responses across somatic tissues5-9. However, the systemic impact of the gut microbiome on the germline-and consequently on the F1 offspring it gives rise to-is unexplored10. Here we show that the gut microbiota act as a key interface between paternal preconception environment and intergenerational health in mice. Perturbations to the gut microbiota of prospective fathers increase the probability of their offspring presenting with low birth weight, severe growth restriction and premature mortality. Transmission of disease risk occurs via the germline and is provoked by pervasive gut microbiome perturbations, including non-absorbable antibiotics or osmotic laxatives, but is rescued by restoring the paternal microbiota before conception. This effect is linked with a dynamic response to induced dysbiosis in the male reproductive system, including impaired leptin signalling, altered testicular metabolite profiles and remapped small RNA payloads in sperm. As a result, dysbiotic fathers trigger an elevated risk of in utero placental insufficiency, revealing a placental origin of mammalian intergenerational effects. Our study defines a regulatory 'gut-germline axis' in males, which is sensitive to environmental exposures and programmes offspring fitness through impacting placenta function.

Pharmaceuticals can directly inhibit the growth of gut bacteria, but the degree to which such interactions manifest in complex community settings is an open question. Here, we compared the effects of 30 drugs on a 32-species synthetic community with their effects on each community member in isolation. While most individual drug-species interactions remained the same in the community context, communal behaviors emerged in 26% of all tested cases. Cross-protection during which drug-sensitive species were protected in community was 6 times more frequent than cross-sensitization, the converse phenomenon. Cross-protection decreased and cross-sensitization increased at higher drug concentrations, suggesting that the resilience of microbial communities can collapse when perturbations get stronger. By metabolically profiling drug-treated communities, we showed that both drug biotransformation and bioaccumulation contribute mechanistically to communal protection. As a proof of principle, we molecularly dissected a prominent case: species expressing specific nitroreductases degraded niclosamide, thereby protecting both themselves and sensitive community members.

Asymmetric reduction of prochiral ketone via biocatalysis is one of the most promising and straight forward routes to prepare optically active alcohols. In this work, whole cells of Vigna radiata (also called mung) catalyzed enantioselective reduction of prochiral aromatic ketones was investigated. Effects of various parameters such as co-substrate, temperature, cell loading, substrate concentration on conversion and enantioselectivity were studied in a systematic manner by using acetophenone as the model compound. Glucose was found to be the most effective co-substrate for co-factor regeneration among different co-substrates employed. Whole cells of mung have shown excellent thermostability and high substrate tolerance (up to 200 mM). Under optimum conditions, maximum conversion of 93.44% and enantioselectivity of 98.92% were obtained in 9h. Good conversion and enantioselectivity were observed with various substituted aromatic ketones. This is a cheap, clean, and eco-friendly process for preparation of chiral secondary alcohols compared to chemical processes.

Bacteria in the gut can modulate the availability and efficacy of therapeutic drugs. However, the systematic mapping of the interactions between drugs and bacteria has only started recently1 and the main underlying mechanism proposed is the chemical transformation of drugs by microorganisms (biotransformation). Here we investigated the depletion of 15 structurally diverse drugs by 25 representative strains of gut bacteria. This revealed 70 bacteria-drug interactions, 29 of which had not to our knowledge been reported before. Over half of the new interactions can be ascribed to bioaccumulation; that is, bacteria storing the drug intracellularly without chemically modifying it, and in most cases without the growth of the bacteria being affected. As a case in point, we studied the molecular basis of bioaccumulation of the widely used antidepressant duloxetine by using click chemistry, thermal proteome profiling and metabolomics. We find that duloxetine binds to several metabolic enzymes and changes the metabolite secretion of the respective bacteria. When tested in a defined microbial community of accumulators and non-accumulators, duloxetine markedly altered the composition of the community through metabolic cross-feeding. We further validated our findings in an animal model, showing that bioaccumulating bacteria attenuate the behavioural response of Caenorhabditis elegans to duloxetine. Together, our results show that bioaccumulation by gut bacteria may be a common mechanism that alters drug availability and bacterial metabolism, with implications for microbiota composition, pharmacokinetics, side effects and drug responses, probably in an individual manner.

Adaptive laboratory evolution has proven highly effective for obtaining microorganisms with enhanced capabilities. Yet, this method is inherently restricted to the traits that are positively linked to cell fitness, such as nutrient utilization. Here, we introduce coevolution of obligatory mutualistic communities for improving secretion of fitness-costly metabolites through natural selection. In this strategy, metabolic cross-feeding connects secretion of the target metabolite, despite its cost to the secretor, to the survival and proliferation of the entire community. We thus co-evolved wild-type lactic acid bacteria and engineered auxotrophic Saccharomyces cerevisiae in a synthetic growth medium leading to bacterial isolates with enhanced secretion of two B-group vitamins, viz., riboflavin and folate. The increased production was specific to the targeted vitamin, and evident also in milk, a more complex nutrient environment that naturally contains vitamins. Genomic, proteomic and metabolomic analyses of the evolved lactic acid bacteria, in combination with flux balance analysis, showed altered metabolic regulation towards increased supply of the vitamin precursors. Together, our findings demonstrate how microbial metabolism adapts to mutualistic lifestyle through enhanced metabolite exchange.

Our knowledge about the gut microbiota of pigs is still scarce, despite the importance of these animals for biomedical research and agriculture. Here, we present a collection of cultured bacteria from the pig gut, including 110 species across 40 families and nine phyla. We provide taxonomic descriptions for 22 novel species and 16 genera. Meta-analysis of 16S rRNA amplicon sequence data and metagenome-assembled genomes reveal prevalent and pig-specific species within Lactobacillus, Streptococcus, Clostridium, Desulfovibrio, Enterococcus, Fusobacterium, and several new genera described in this study. Potentially interesting functions discovered in these organisms include a fucosyltransferase encoded in the genome of the novel species Clostridium porci, and prevalent gene clusters for biosynthesis of sactipeptide-like peptides. Many strains deconjugate primary bile acids in in vitro assays, and a Clostridium scindens strain produces secondary bile acids via dehydroxylation. In addition, cells of the novel species Bullifex porci are coccoidal or spherical under the culture conditions tested, in contrast with the usual helical shape of other members of the family Spirochaetaceae. The strain collection, called 'Pig intestinal bacterial collection' (PiBAC), is publicly available at www.dsmz.de/pibac and opens new avenues for functional studies of the pig gut microbiota.

The proinflammatory and genotoxic properties of hydrogen sulfide (H2S) have been implicated as an environmental trigger of colonic disease. Recent evidence links sulfidogenic bacteria capable of metabolizing organic sulfur substrates, like dietary sulfur amino acids and taurine conjugated bile acids, with increased colorectal cancer (CRC) and colitis risk as a consequence of a “western” type diet. Our previous work demonstrates that the sulfidogenic bacterium, Odoribacter splanchnicus, is a significant indicator of CRC in African Americans who are at higher risk of CRC incidence and death than other races and ethnicities in the United States. This observation has been corroborated by several recent studies that observed O. splanchnicus to be associated with CRC and colitis-induced CRC. While previous work has reported that O. splanchnicus may be sulfidogenic, its method of H2S production has yet to be characterized. Production of H2S by O. splanchnicus was confirmed in vitro using sulfide indole and motility media, and varying sulfur substrates. A predictive metabolic analysis of 74 O. splanchnicus metagenomes taken from publicly available CRC cohorts did not reveal complete pathways commonly associated with H2S production, however genes for a polar amino acid transporter were revealed. Therefore, a genomic analysis was performed revealing that O. splanchnicus harbors the sulfidogenic genes tryptophanase (tnaA) and cystathionine-β-synthase. The enzyme TnaA has been previously shown in Eschericia coli to produce indole or H2S from tryptophan or cysteine, respectively. However, the E. coli amino acid sequence shares only 44.85% identity with that from O. splanchnicus, thus heterologous protein expression of recombinant tnaA was performed to determine enzyme functionality. Enzyme kinetics and pH optimum were obtained using the spectrophotometric BiCl3 assay, which turns black in presence of H2S. Functional analysis of recombinant purified TnaA revealed bifunctionality of the enzyme with positive tryptophanase and cysteine desulfhydrase activity with differing substrates. Cysteine desulfhydrase activity resulted in consumption of L-cysteine with subsequent production of H2S and pyruvate. Thus, TnaA is a functional enzyme that warrants further evaluation as possibly relating to sulfidogenic O. splanchnicus being linked to CRC and colitis-associated CRC. Citation Format: Patricia G. Wolf, Vladimir Kolosslov, Zhichao Zhou, Lindsey Ly, Heidi Doden, Saravanan Devendran, Adam M. Breister, Luke Lucio, Paige Polak, Sarah Matatov, Karthik Anantharaman, Jason M. Ridlon, Rex Gaskins. The colorectal cancer associated microbe Odoribacter splanchnicus produces genotoxic hydrogen sulfide via cysteine metabolism [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3342.

Microalgae have higher productivity of biomass than the conventional crops of fuel and are therefore, considered a potential biofuel source. Lipid, an important precursor of biodiesel, can be overproduced in microalgae by nitrogen deprivation. During nitrogen deficiency, radicals are overproduced, and the antioxidant levels are insufficient to counteract the radicals. Thus, the increase in cellular oxidative stress level, consequently acts as a stimulus for lipid accumulation. Lipid accumulation requires an excess of acetyl CoA and NADPH that is made possible by the following mechanism. Glycolysis upregulation overproduces pyruvate, which could be further transformed into acetyl CoA by the pyruvate dehydrogenase complex; while the upregulation of the oxidative pentose phosphate cycle generates a high amount of NADPH. In addition to lipid overproduction, the lack of nitrogen often causes the accumulation of carbohydrates in selected species of microalgae, which could be used to generate biogas and bioethanol from the defatted biomass. By providing details on the differential regulation of the biochemical pathways leading to lipid and carbohydrate accumulation in nitrogen starved microalgae, the review opens up new possibilities in the microalgal biofuel production.

Due to their rapid growth rates, high lipid productivity, and ability to synthesize value-added products, microalgae are considered as the potential biofuel feedstocks. However, among the several bottlenecks that are hindering the commercialization of microalgal biofuel synthesis, the issue of high water consumption is the least explored. This analysis, therefore, examines the factors that decide water use for the production of microalgae biofuel. Microalgae biodiesel water footprint varies from 3.5 to 3726 kg of water per kg of biodiesel. The study further investigates the cause for large variability in the estimation of the water footprint for microalgae fuel. Various strategies, including the reuse of harvested water, the use of high density cultivation that could be adopted for low water consumption in microalgal biofuel production are discussed. Specifically, the review identified a reciprocal relationship between biomass productivity and water footprint. On the basis of which the review emphasizes the significance of high density cultivation, which can be inexpensive and feasible relative to other water-saving techniques. With the setback of water scarcity due to the rapid industrialization in developing countries, the implementation of the cultivation system with a focus on minimizing the water consumption is inevitable for a successful large scale microalgal biofuel production.

•Review reports on physiological and biochemical reaction to N-stress in microalgae.•Lipid and carbohydrate increased and protein decreased in N-stressed cells.•During nitrogen deprivation, radicals are overproduced creating oxidative stress.•The increase in cellular oxidative stress causes the accumulation of lipids.

ABSTRACTThe ability to metabolize both endogenous and exogenous compounds to a variety of metabolic products is not exclusive to our human cells. In fact, the bacterial communities that inhabit our digestive system are responsible for a network of steroid transformations that can produce hormones in the gut, which are then absorbed to act in the host. These communities have been shown to impact our health in numerous ways, affecting disease predisposition, pathogenesis, physical fitness, and dietary responsiveness. Steroid biotransformations by gut bacteria are predicted to impact the host endocrine system. A particular set of transformations facilitated by microbial enzymes has been shown to result in the formation of 11-oxy-androgens from host-derived cortisol. Since androgens have been implicated in disease and immune modulations, understanding the structure and catalytic mechanism of enzymes involved in cortisol metabolism is a key step to hasten the development of strategies that reduce the formation of disease-promoting bioactive steroids in certain individuals. Here, we combine experimental and computational techniques to describe DesC, an enzyme capable of creating 20α-dihydrocortisol and siphoning cortisol away from pathways that produce androgens. DesC diverges significantly from previously described bacterial and eukaryotic counterparts, catalyzing an NADH-dependent 20α-hydroxysteroid dehydrogenase reaction but presenting little sequence and structure similarity to them. The structural information obtained by X-ray crystallography and hybrid QM/MM simulations, validated through mutagenesis studies, show the reaction occurs through a multi-step proton relay mechanism. Free energy calculations were then used to describe the kinetics of the reaction mechanism. The mechanistic information presented here can be employed in the development of therapeutics to divert microbial pathways away from disease-promoting steroids.

The adrenal gland has traditionally been viewed as a source of "weak androgens"; however, emerging evidence indicates 11-oxy-androgens of adrenal origin are metabolized in peripheral tissues to potent androgens. Also emerging is the role of gut bacteria in the conversion of C21 glucocorticoids to 11-oxygenated C19 androgens. Clostridium scindens ATCC 35,704 is a gut microbe capable of converting cortisol into 11-oxy-androgens by cleaving the side-chain. The desA and desB genes encode steroid-17,20-desmolase. Our prior study indicated that the urinary tract bacterium, Propionimicrobium lymphophilum ACS-093-V-SCH5 encodes desAB and converts cortisol to 11β-hydroxyandrostenedione. We wanted to determine how widespread this function occurs in the human microbiome. Phylogenetic and sequence similarity network analyses indicated that the steroid-17,20-desmolase pathway is taxonomically rare and located in gut and urogenital microbiomes. Two microbes from each of these niches, C. scindens and Propionimicrobium lymphophilum, respectively, were screened for activity against endogenous (cortisol, cortisone, and allotetrahydrocortisol) and exogenous (prednisone, prednisolone, dexamethasone, and 9-fluorocortisol) glucocorticoids. LC/MS analysis showed that both microbes were able to side-chain cleave all glucocorticoids, forming 11-oxy-androgens. Pure recombinant DesAB from C. scindens showed the highest activity against prednisone, a commonly prescribed glucocorticoid. In addition, 0.1 nM 1,4-androstadiene-3,11,17-trione, bacterial side-chain cleavage product of prednisone, showed significant proliferation relative to vehicle in androgen-dependent growth LNCaP prostate cancer cells after 24 h (2.3 fold; P <  0.01) and 72 h (1.6 fold; P < 0.01). Taken together, DesAB-expressing microbes may be an overlooked source of androgens in the body, potentially contributing to various disease states, such as prostate cancer.

The cement industry generates a substantial amount of gaseous pollutants that cannot be treated efficiently and economically using standard techniques. Microalgae, a promising bioremediation and biodegradation agent used as feedstock for biofuel production, can be used for the biotreatment of cement flue gas. In specific, components of cement flue gas such as carbon dioxide, nitrogen, and sulfur oxides are shown to serve as nutrients for microalgae. Microalgae also have the capacity to sequestrate heavy metals present in cement kiln dust, adding further benefits. This work provides an extensive overview of multiple approaches taken in the inclusion of microalgae biofuel production in the cement sector. In addition, factors influencing the production of microalgal biomass are also described in such an integrated plant. In addition, process limitations such as the adverse impact of flue gas on medium pH, exhaust gas toxicity, and efficient delivery of carbon dioxide to media are also discussed. Finally, the article concludes by proposing the future potential for incorporating the microalgae biofuel plant into the cement sector.

Microalgae are a group of microorganisms considered to be a potential source of next-generation biofuel owing to high photosynthetic rate. However, biofuel compounds including lipid, carbohydrate, and hydrogen are not sufficiently produced by microalgae under normal conditions; therefore economical viability of microalgal biofuel production has not reached the ultimate stage of commercial viability. In order to overcome the above limitation, various stress conditions have been applied on microalgae to boost lipid, carbohydrate and hydrogen production. Even though stress conditions increases the production of biofuel compounds, microalgae growth is severely affected. In this regard, a two-stage cultivation mode wherein first stage involving high biomass production under media optimized heterotrophic condition and second stage involving lipid, carbohydrate and hydrogen accumulation under stress integrated phototrophic mode is considered as a potential route for biofuel production in microalgae. The present review critically evaluates various two-stage strategies adopted in microalgal biofuel production by comparing lipid, carbohydrate and hydrogen productivities with that of single stage cultivation. In addition, advantages of two-stage cultivation including co-product generation, less bacterial contamination, improved biodiesel quality, auto-flocculation, and bioremediation potential are also discussed. Finally, the paper summarizes the two-stage cultivation modes by identifying the bottlenecks and suggesting the future potentialities of the process.

The conventional method of lipid extraction mandatorily includes cell lysis which is considered an essential process for the enhancement of extraction efficiency. However, cell lysis method especially those incorporating mechanical and physical methods such as homogenization, sonication, microwave techniques, etc., are energy intensive and drastically escalates the biofuel production cost. Therefore, the development of an alternate route of extraction skipping the conventional mechanical cell lysis step remains a challenge to be accomplished. In this context, we discuss the reports involving passive techniques that are able to extract lipid from microalgae without significant cell wall shearing. This includes methods such as in-situ transesterification, direct saponification, supercritical fluid extraction, organic solvent extraction, etc. Moreover, the review discusses the employment of passive lipid extraction methods in a biorefinery set-up, outlining the various bioproducts that could be generated along with lipid. The review concludes with the analysis of the economics of microalgal lipid extraction using passive disruption methods and compares with the processes incorporating cell lysis steps.

C. scindens is one of a few identified gut bacterial species capable of converting host cholic acid into disease-associated secondary bile acids such as deoxycholic acid. The current work represents an important advance in understanding the nutritional requirements and response to bile acids of the medically important human gut bacterium, C. scindens ATCC 35704. A defined medium has been developed which will further the understanding of bile acid metabolism in the context of growth substrates, cofactors, and other metabolites in the vertebrate gut. Analysis of the complete genome supports the nutritional requirements reported here. Genome-wide transcriptomic analysis of gene expression in the presence of cholic acid and deoxycholic acid provides a unique insight into the complex response of C. scindens ATCC 35704 to primary and secondary bile acids. Also revealed are genes with the potential to function in bile acid transport and metabolism.

Anaerobic bacteria inhabiting the human gastrointestinal tract have evolved various enzymes that modify host-derived steroids. The bacterial steroid-17,20-desmolase pathway cleaves the cortisol side chain, forming pro-androgens predicted to impact host physiology. Bacterial 20 beta-hydroxysteroid dehydrogenase (20 beta-HSDH) regulates cortisol side-chain cleavage by reducing the C-20 carboxyl group on cortisol, yielding 20 beta-dihydrocortisol. Recently, the gene encoding 20 beta-HSDH in Butyricicoccus desmolans ATCC 43058 was reported, and a nonredundant protein search yielded a candidate 20 beta-HSDH gene in Bifidobacterium adolescentis strain L2-32. B. adolescentis 20 beta-HSDH could regulate cortisol side-chain cleavage by limiting pro-androgen formation in bacteria such as Clostridium scindens and 21-dehydroxylation by Eggerthella lenta. Here, the putative B. adolescentis 20 beta-HSDH was cloned, overexpressed, and purified. 20 beta-HSDH activity was confirmed through whole-cell and pure enzymatic assays, and it is specific for cortisol. Next, we solved the structures of recombinant 20 beta-HSDH in both the apo- and holo-forms at 2.0-2.2 angstrom resolutions, revealing close overlap except for rearrangements near the active site. Interestingly, the structures contain a large, flexible N-terminal region that was investigated by gel-filtration chromatography and CD spectroscopy. This extended N terminus is important for protein stability because deletions of varying lengths caused structural changes and reduced enzymatic activity. A nonconserved extended N terminus was also observed in several short-chain dehydrogenase/reductase family members. B. adolescentis strains capable of 20 beta-HSDH activity could alter glucocorticoid metabolism in the gut and thereby serve as potential probiotics for the management of androgen-dependent diseases.