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The effects of different chemical oxygen demand (COD) concentrations on the anammox granular sludge with Bamboo Charcoal (BC) addition were evaluated in UASB reactor. The results showed that the average total nitrogen (TN) removal efficiency was reduced from 85.9% to 81.4% when COD concentration was increased from 50 to 150 mg/L. However, the TN removal efficiency of BC addition reactors was dramatically 3.1%–6.4% higher than that without BC under different COD concentrations. The average diameter of granular sludge was 0.13 mm higher than that without BC. The settling velocity was increased by elevated COD concentration, while the EPS and VSS/SS were increased with BC addition. The high-throughput Miseq sequencing analyses revealed that the bacterial diversity and richness were decreased under COD addition, and the Planctomycetes related to anammox bacteria were Candidatus Brocadia and Candidatus Kuenenia. The Metagenomic sequencing indicated that the abundance of denitrification related functional genes all increased with elevated COD, while the abundance of anammox related functional genes of decreased. The functional genes related to anammox was hydrazine synthase encoding genes (hzsA, hzsB and hzsB). The average relative abundance of hzs genes in the reactor with BC addition was higher than the control at COD concentrations of 50 mg/L and 150 mg/L. The functional genes of denitrification mediated by BC were higher than those without BC throughout the operation phase. It is interesting to note that BC addition greatly enriched the related functional genes of denitrification and anammox.

Ethanol-type sludge fermentation has recently attracted much attention because it can enhance direct interspecies electron transfer and thus improve the anaerobic digestion of waste activated sludge (WAS). In this paper, the enhancement of short-term ethanol-type fermentation of WAS via adding Saccharomyces was investigated. The experimental results show that the maximum ethanol production of 1030.8 ± 20.6 mg/L was achieved, with the optimum fermentation conditions of a pH of 5.1, temperature of 26.0 ℃ and time of 8.0 hr. Although the content of volatile fatty acid (VFA) increased within 10 hr, it is one order of magnitude lower than the content of ethanol, indicating that the VFA generation did not affect the efficient production of ethanol. The analyses of changes in the microbial community during the fermentation process demonstrate that the greatest Saccharomyces activity occurred in the first 8 hr and it can play an important role in ethanol production even at a very low relative abundance. Meanwhile, most typical acid-producing bacteria were inhibited, but the hydrogenotrophic methanogens (i.e., Methanobacterium) were enriched to a certain extent. Further statistical analyses reveal that the Rhodobacter, Thermomonas, Terrimonas and Saccharomyces are responsible for ethanol production during the fermentation. However, these findings not only provide a reference for the development of enhancing ethanol-type fermentation of sludge, but also are expected to provide a new way of thinking for the efficient bioenergy and resource recovery from sludge.

Aims The study of microbial inactivation rates in aquifer systems has most often been determined in aerobic and oxidized systems. This study examined the inactivation (i.e. loss of infectivity) of MS2, poliovirus type 1 (PV1) and Cryptosporidium parvum in an anaerobic and reduced groundwater system that has been identified as storage zones for aquifer storage and recovery (ASR) facilities. Methods and Results Anaerobic and reduced (ORP < −250 mV) groundwater from an artesian well was diverted to an above-ground, flow-through mesocosm that contained diffusion chambers filled with MS2, PV1 or Cryptosporidium parvum. The respective infectivity assays were performed on microorganisms recovered from the diffusion chambers during 30- to 58-day experiments. The net reduction in infectivity was 5.73 log10 over 30 days for MS2, 5.00 log10 over 58 days for PV1 and 4.07 log10 over 37 days for C. parvum. The best fit inactivation model for PV1 was the log-linear model and the Weibull model for MS2 and C. parvum, with respective inactivation rates (95% confidence interval) of 0.19 (0.17–0.21) log10 day−1, 0.31 (0.19–0.89) log10 day−1 and 0.20 (0.14–0.37) log10 day−1. Conclusions The groundwater geochemical conditions in this aquifer enhanced the inactivation of MS2, PV1, and C. parvum at rates approximately 2.0–5.3-fold, 1.2–17.0-fold, and 4.5–5.6-fold greater, respectively, than those from published studies that used diffusion chambers in aerobic-to-anoxic groundwater systems, with positive redox potentials. Significance and Impact of the Study Geochemical conditions like those in the aquifer zone in this study can naturally and significantly reduce concentrations of microbial indicators and pathogens of human health concern in injected surface water. Appropriate storage times for injected surface water could complement above-ground engineered processes for microorganism removal and inactivation (e.g. filtration, disinfection) by naturally increasing overall microorganism log-inactivation rates of ASR facilities.

To compare deep infection rate and causative organisms in open fractures of the lower extremity from agricultural trauma with similar injuries in nonagricultural trauma.Retrospective.Two tertiary care institutions.Open lower extremity fractures sustained between 2003 and 2018 caused by agriculture-related trauma in adult patients were reviewed. A nonagriculture open fracture control group (NAG) was identified for comparison. Patient demographics and injury characteristics were assessed. Outcomes included occurrence of deep infection and causative organisms.One hundred seventy-eight patients were identified in the agriculture (AG) (n = 89) and control (NAG) (n = 89) groups. Among patients with agricultural injury, farm machinery was the most common mechanism in 69 (77.5%) of them. Open injuries of the foot (38.2%) and tibial shaft injuries (25.8%) were the most common in the AG and NAG groups, respectively. Deep infection was seen in 21% of patients in the AG group compared with 10% in the NAG group (P0.05). In the AG group, anaerobic infection occurred in 44% of patients with deep infection compared with 9.1% in the NAG group (P0.05). The most common anaerobic organisms included enterococci, Pseudomonas aeruginosa, and Clostridium perfringens.This study supports that open fractures due to agricultural trauma have a high infection rate, with anaerobic infection occurring at higher rates than in injuries due to nonagricultural trauma. Prophylactic treatment with antibiotics directed against anaerobes is indicated in these injuries.Prognostic Level III. See Instructions for Authors for a complete description of levels of evidence.

The enormous chemical diversity and strain variability of prokaryotic protein glycosylation makes a large-scale exploration exceptionally challenging. Therefore, despite the universal relevance of protein glycosylation across all domains of life, the understanding of their biological significance and the evolutionary forces shaping oligosaccharide structures remains highly limited.Here, we report on a newly established mass binning glycoproteomics approach that establishes the chemical identity of the carbohydrate components and performs untargeted exploration of prokaryotic oligosaccharides from large-scale proteomics data directly. We demonstrate our approach by exploring an enrichment culture of the globally relevant anaerobic ammonium-oxidizing bacterium Ca. Kuenenia stuttgartiensis. By doing so we resolved a remarkable array of oligosaccharides, produced by two entirely unrelated glycosylation machineries targeting the same surface-layer protein (SLP) simultaneously. More intriguingly, the investigated strain also accomplished modulation of highly specialized sugars, supposedly in response to its energy metabolism—the anaerobic oxidation of ammonium —which depends on the acquisition of substrates of opposite charge. Ultimately, we provide a systematic approach for the compositional exploration of prokaryotic protein glycosylation, and reveal for the first time a remarkable balance between maximising cellular protection through a complex array of oligosaccharides and adhering to the requirements of the ‘metabolic lifestyle’.

Two anaerobic membrane bioreactors (AnMBRs) equipped with different membrane pore size (0.4 or 0.05 µm) were operated at 25˚C and fed with domestic wastewater. The hydraulic retention time (HRT) of the reactors was shortened. The microbial communities of the two AnMBRs were investigated by 16S rRNA gene amplicon sequencing to see the effects of HRT. The predominant Archaea was an aceticlastic methanogen Methanosaeta. The composition of hydrogenotrophic methanogens changed with the HRTs: the population of Methanobacterium was higher for longer HRTs, whereas the population of unclassified Methanoregulaceae was higher for shorter HRTs. The Anaerolineae, Bacteroidia and Clostridia bacteria were dominant in both of the reactors, with a combined relative abundance of over 55%. The relative abundance of Anaerolineae was proportional to the biogas production performance. The change in the population of hydrogenotrophic methanogens or Anaerolineae can be used as an indicator for process monitoring. The sum of the relative abundance of Anaerolineae and Clostridia fluctuated slightly with changes in the HRT in both AnMBRs when the reactor was stably operated. The co-occurrence analysis revealed the relative abundance of the operational taxonomic units belonging to Anaerolineae and Clostridia was functionally equivalent during the treatment of real domestic sewage. A principal coordination analysis revealed that the changes in the microbial community in each reactor were consistent with the change of HRT. In addition, both the HRT and the stability of the process are important factors for maintaining microbial community structures.

It is unadvisable to discuss the antibiotic resistance genes (ARGs) reduction in anaerobic digestion (AD) system neglecting its main purpose-methane production. The methane production improvement coupling with antibiotic resistance genes (ARGs) reduction in anaerobic digestion (AD) by zero valent iron (ZVI) were simultaneously investigated. Whether the role of ZVI on the ARGs fate was random or specific was clarified through the high-throughput qPCR (HT-qPCR). Results indicated that ZVI improved methane production and ARGs reduction by 23.9% and 25.0%, respectively. The improved methane production was associated with chemical reaction and variances of microbial community caused by ZVI, where DIET between Petrimonas, Clostridium and Syntrophomonas, Methanosarcina was established along with ACAS being enriched. ZVI specifically, not randomly, facilitated the reduction of aminoglycoside resistance genes of antibiotic inactivation and tetracycline resistance genes of ribosomal protection proteins. The specifical reduction could be attributed to enzyme activity inhibition and intracellular ionic disturbance caused by higher amounts of ZVI, although most of ARGs fate could be well explained by microbial community which contributed the most to ARGs dynamics as a whole. ZVI-based AD was a promising way for the improvement of methane production coupling ARGs reduction.

Euglena gracilis produces ATP in the anaerobic mitochondria with concomitant wax ester formation, and NADH is essential for ATP formation and fatty acid synthesis in the mitochondria. This study demonstrated that mitochondrial cofactor conversion by nicotinamide nucleotide transhydrogenase (NNT), converting NADPH/NAD+ to NADP+ /NADH, is indispensable for sustaining anaerobic metabolism. Silencing of NNT genes significantly decreased wax ester production and cellular viability during anaerobiosis but had no such marked effects under aerobic conditions. An analogous phenotype was observed in the silencing of the gene encoding a mitochondrial NADP+ -dependent malic enzyme. These results suggest that the reducing equivalents produced in glycolysis are shuttled to the mitochondria as malate, where cytosolic NAD+ regeneration is coupled with mitochondrial NADPH generation.

Transitions in microbial community structure in response to increasing ammonia concentrations were determined by monitoring mesophilic anaerobic digesters seeded with a predominantly acetoclastic methanogenic community from a sewage sludge digester. Ammonia concentration was raised by switching the feed to source segregated domestic food waste and applying two organic loading rates (OLR) and hydraulic retention times (HRT) in paired digesters. One of each pair was dosed with trace elements (TE) known to be essential to the transition, with the other unsupplemented digester acting as a control. Samples taken during the trial were used to determine the metabolic pathway to methanogenesis using 14C labelled acetate. Partitioning of 14C between the product gases was interpreted via an equation to indicate the proportion produced by acetoclastic and hydrogenotrophic routes. Archaeal and selected bacterial groups were identified by 16S rRNA sequencing, to determine relative abundance and diversity. Acclimatisation for digesters with TE was relatively smooth, but OLR and HRT influenced both metabolic route and community structure. The 14C ratio could be used quantitatively and, when interpreted alongside archaeal community structure, showed that at longer HRT and lower loading Methanobacteriaceae were dominant and hydrogenotrophic activity accounted for 77% of methane production. At the higher OLR and shorter HRT, Methanosarcinaceae were dominant with the 14C ratio indicating simultaneous production of methane by acetoclastic and hydrogenotrophic pathways: the first reported observation of this in digestion under mesophilic conditions. Digesters without TE supplementation showed similar initial changes but, as expected failed to complete the transition to stable operation.

Oxygen is one of the important substances for the survival of most life systems on the earth, and plateau and underground burrow systems are two typical hypoxic environments. Small mammals living in hypoxic environments have evolved different adaptation strategies, which include increased oxygen delivery, metabolic regulation of physiological responses and other physiological responses that change tissue oxygen utilization. Multi‐omics predictions have also shown that these animals have evolved different adaptations to extreme environments. In particular, vascular endothelial growth factor (VEGF) and erythropoietin (EPO), which have specific functions in the control of O2 delivery, have evolved adaptively in small mammals in hypoxic environments. Naked mole‐rats and blind mole‐rats are typical hypoxic model animals as they have some resistance to cancer. This review primarily summarizes the main living environment of hypoxia tolerant small mammals, as well as the changes of phenotype, physiochemical characteristics and gene expression mode of their long‐term living in hypoxia environment., Small mammals have evolved mechanisms of hypoxia adaptation living under plateau and underground burrow conditions.

The exopolysaccharide (EPS) from the mycelial fermentation of a medicinal fungus Cordyceps sinensis Cs-HK1 had shown significant anti-inflammatory activity previously, and EPS-LM was a highly active fraction with a relatively low molecular weight (MW) isolated from the Cs-HK1 EPS. This study was to assess the effects of Bifidobacterial fermentation in anaerobic conditions on the molecular properties and anti-inflammatory activity of EPS-LM. In both Bifidobacterial cultures (B. breve and B. longum), EPS-LM was fractionally consumed as a carbon source, increasing the bacterial growth and acetic acid production. Analytical results from the fermentation digesta (supernatant) suggested that EPS-LM was partially degraded to lower molecular weight (MW) products with modified structures during the Bifidobacterial fermentation. More interestingly, the higher MW digesta fraction containing the partially degraded EPS-LM showed even stronger inhibiting activity than the original EPS-LM on the LPS-induced pro-inflammatory responses in THP-1 cell culture, including NF-κB activation, release of NO, TNF-α and IL-8. The study has shown that the fermentation by selected Bifidobacterial strains is effective to modify natural polysaccharides with enhanced bioactivities.

The lack of real-time reporters in obligate anaerobes has limited studies in gene expression, promoter characterization, library screening, population dynamics, and cell biology in these organisms. While the use of enzymatic, colorimetric, and luminescent reporters has been reported, the need for reliable anaerobic fluorescent proteins is widely acknowledged. Recently, the fluorescent proteins HaloTag, SNAP-tag and FAST have been established as reliable reporters in Clostridium spp., thus suggesting that these reporters can be adopted widely for many obligate anaerobes. With a multitude of labeling options, these anaerobic fluorescent proteins hold a great potential for screening promoters, terminators, and RBS sites, tracking population dynamics in complex multi-species co-cultures, such as microbiomes, screening libraries, and in cell biology studies of protein localization and interactions using high-resolution microscopy.

The microbiome of the anaerobic digester (AD) regulates the level of energy production. To assess the microbiome diversity and composition in different stages of anaerobic digestion, we collected 16 samples from the AD of cow dung (CD) origin. The samples were categorized into four groups (Group-I, Group-II, Group-III and Group-IV) based on the level of energy production (CH4%), and sequenced through whole metagenome sequencing (WMS). Group-I (n = 2) belonged to initial time of energy production whereas Group-II (n = 5), Group-III (n = 5), and Group-IV (n = 4) had 21–34%, 47–58% and 71–74% of CH4, respectively. The physicochemical analysis revealed that level of energy production (CH4%) had significant positive correlation with digester pH (r = 0.92, p 2 level (%) (r = 0.54, p Firmicutes, Bacteroidetes, Proteobacteria, Actinobacteria, Spirochaetes and Fibrobacteres phyla (> 93.0% of the total abundances). Simultaneously, 343 archaeal strains were detected, of which 95.90% strains shared across four metagenomes. We identified 43 dominant species including 31 bacterial and 12 archaeal species in AD microbiomes, of which only archaea showed positive correlation with digester pH, CH4 concentration, pressure and temperature (Spearman correlation; r > 0.6, p Methanosarcina vacuolate, Dehalococcoides mccartyi, Methanosarcina sp. Kolksee and Methanosarcina barkeri were highly specific for energy production. The correlation network analysis showed that different strains of Euryarcheota and Firmicutes phyla exhibited significant correlation (p = 0.021, Kruskal–Wallis test; with a cutoff of 1.0) with the highest level (74.1%) of energy production (Group-IV). In addition, top CH4 producing microbiomes showed increased genomic functional activities related to one carbon and biotin metabolism, oxidative stress, proteolytic pathways, membrane-type-1-matrix-metalloproteinase (MT1-MMP) pericellular network, acetyl-CoA production, motility and chemotaxis. Importantly, the physicochemical properties of the AD including pH, CH4 concentration (%), pressure, temperature and environmental temperature were found to be positively correlated with these genomic functional potentials and distribution of ARGs and metal resistance pathways (Spearman correlation; r > 0.5, p

Uropathogenic Escherichia coli (UPEC) is the primary causative agent of urinary tract infections (UTIs). Successful urinary tract colonization requires appropriate expression of virulence factors in response to host environmental cues, such as limited oxygen and iron availability. Hemolysin is a pore-forming toxin, and its expression correlates with the severity of UPEC infection. Previously, we showed that hemolysin expression is enhanced under anaerobic conditions; however, the genetic basis and regulatory mechanisms involved remain undefined. Here, a transposon-based forward screen identified bis-molybdopterin guanine dinucleotide cofactor (bis-MGD) biosynthesis as an important factor for a full transcription of hemolysin under anaerobiosis but not under aerobiosis. bis-MGD positively influences hemolysin transcription via c3566-c3568, an operon immediately upstream of and cotranscribed with hlyCABD. Furthermore, suppressor mutation analysis identified the nitrogen regulator NtrC as a direct repressor of c3566-c3568-hlyCABD expression, and intact bis-MGD biosynthesis downregulated ntrC expression, thus at least partially explaining the positive role of bis-MGD in modulating hemolysin expression. Finally, bis-MGD is involved in hemolysin-mediated uroepithelial cell death and contributes to the competitive fitness of UPEC in a murine model of UTI. Collectively, our data establish that bis-MGD biosynthesis plays a crucial role in UPEC fitness in vivo, thus providing a potential target for combatting UTIs.

Anaerobic oxidation of methane (AOM) can be microbially mediated by the reduction of different terminal electron acceptors. AOM coupled to reduction of sulfate, manganese/iron oxides, humic substances, selenate, arsenic and other artificial extracellular electron acceptors are recognized as processes associated with microbial extracellular respiration. In these processes, methane-oxidizing archaea transfer electrons to external electron acceptors or to interdependent microbial species, which are mechanistically dependent on versatile extracellular electron transfer (EET) pathways. This review compiles recent progress in the research of electromicrobiology of AOM based on the catalogue of different electron acceptors. Naturally distributed and artificially constructed EET-mediated AOM is summarized, with the discussion of their environmental importance and application potentials. The diversity of responsible microorganisms involved in EET-mediated AOM is discussed with both methane-oxidizing archaea and their putative bacterial partners. More importantly, the review highlights progress and deficiencies in our understanding of EET pathways in EET-mediated AOM, raising open research questions for future research.

Bacterial lipids are well-preserved in ancient rocks and certain ones have been used as indicators of specific bacterial metabolisms or environmental conditions existing at the time of rock deposition. Here we show that an anaerobic bacterium produces 3-methylhopanoids, pentacyclic lipids previously detected only in aerobic bacteria and widely used as biomarkers for methane-oxidizing bacteria. Both Rhodopila globiformis, a phototrophic purple nonsulfur bacterium isolated from an acidic warm spring in Yellowstone, and a newly isolated Rhodopila species from a geochemically similar spring in Lassen Volcanic National Park (USA), synthesized 3-methylhopanoids and a suite of related hopanoids and contained the genes encoding the necessary biosynthetic enzymes. Our results show that 3-methylhopanoids can be produced under anoxic conditions and challenges the use of 3-methylhopanoids as biomarkers of oxic conditions in ancient rocks and as prima facie evidence that methanotrophic bacteria were active when the rocks were deposited.

Consortia of aerobic heterotrophic bacteria (AHB) are appealing as sustainable alternative protein ingredient for aquaculture given their high nutritional qualities, and their production potential on feed-grade industrial wastewater. Today, the impacts of pre-treatment, bioprocess choice and key parameter settings on AHB productivity and nutritional properties are unknown. This study investigated for the first time AHB microbial protein production effects based on (i) raw vs anaerobically fermented brewery wastewater, (ii) high-rate activated sludge (HRAS) without vs with feast-famine conditions, and (iii) three short solid retention time (SRT): 0.25, 0.50 and 1.00 d. High biomass (4.4–8.0 g TSS/L/d) and protein productivities (1.9–3.2 g protein/L/d) were obtained while achieving COD removal efficiencies up to 98 % at SRT 0.50 d. The AHB essential amino acid (EAA) profiles were above rainbow trout requirements, excluding the S-containing EAA, highlighting the AHB biomass replacement potential for unsustainable fishmeal in salmonid diets.

A quite intriguing subject being intensively researched in the forensic toxicology field is the source of postmortem determined blood ethanol concentration: antemortem ingestion or postmortem microbial production. Our previous research on microbial ethanol production has reported a quantitative relationship between the ethanol and the higher alcohols and 1-butanol produced by Escherichia coli, Clostridium perfrigens, and Clostridium sporogenes. In this contribution, we continue our research reporting on the following: (i) the patterns of ethanol, higher alcohols, and 1-butanol production by the microbes Klebsiella pneumoniae, Staphylococcus aureus, and Enterococcus faecalis (all being aerobic/facultative anaerobic species, common corpse’s colonizers, and ethanol producers), under controlled laboratory conditions, (ii) the mathematical modeling, with simple mathematical equations, of the correlation between ethanol concentration and the other studied alcohols’ concentrations, by performing multiple linear regression analysis of the results, and (iii) the applicability of the constructed models in microbial cultures developed under different temperature than that used to build the models, in denatured blood cultures and in real postmortem cases. The aforementioned alcohols were proved to be all indicators of ethanol production, both in qualitative and quantitative terms. 1-Propanol was the most significant alcohol in modeling microbial ethanol production, followed by methyl-butanol. The K. pneumoniae’s models achieved the best scoring in applicability (E

Denitrifying Betaproteobacteria play a key role in the anaerobic degradation of monoaromatic hydrocarbons. We performed a multi-omics study to better understand the metabolism of the representative organism Georgfuchsia toluolica strain G5G6 known as a strict anaerobe coupling toluene oxidation with dissimilatory nitrate and Fe(III) reduction. Despite the genomic potential for degradation of different carbon sources, we did not find sugar or organic acid transporters, in line with the inability of strain G5G6 to use these substrates. Using a proteomics analysis, we detected proteins of fumarate-dependent toluene activation, membrane-bound nitrate reductase, and key components of the metal-reducing (Mtr) pathway under both nitrate- and Fe(III)-reducing conditions. High abundance of the multiheme cytochrome MtrC implied that a porincytochrome complex was used for respiratory Fe(III) reduction. Remarkably, strain G5G6 contains a full set of genes for aerobic toluene degradation, and we detected enzymes of aerobic toluene degradation under both nitrate- and Fe(III)-reducing conditions. We further detected an ATP-dependent benzoyl-CoA reductase, reactive oxygen species detoxification proteins, and cytochrome c oxidase indicating a facultative anaerobic lifestyle of strain G5G6. Correspondingly, we found diffusion through the septa a substantial source of oxygen in the cultures enabling concurrent aerobic and anaerobic toluene degradation by strain G5G6., This work was supported by Wageningen University & Research through its investment theme Resilience, the Technology Foundation (STW), the Applied Science Division of the Dutch Research Council (NWO; project 08053), NWO grant 016.Vidi.189.050, and a Gravitation grant of the Netherlands Ministry of Education, Culture and Science and NWO (project 024.002.002 SIAM). B.K. was supported by the Villum foundation, Denmark (VYI Grant 25491)., info:eu-repo/semantics/publishedVersion

Corynebacterium glutamicum, a gram-positive and facultative anaerobic bacterium, is widely used for the industrial production of amino acids, such as L-glutamate and L-lysine. C. glutamicum grows and produces amino acids under aerobic conditions. When restricted under anaerobic conditions, it produces organic acids, such as L-lactate and succinate, through metabolic shift. With the increasing threat of global warming, these organic acids have drawn considerable attention as bio-based plastic monomers. In addition to the organic acids, the anaerobic bioprocess is also used to produce other value-added compounds, including isobutanol, ethanol, 3-methyl-1-butanol, 2,3-butanediol, L-alanine, and L-valine. Therefore, C. glutamicum is now a versatile cell factory for producing a wide variety of useful chemicals under both aerobic and anaerobic conditions. The growth and metabolism of the bacterium depend on the oxygen levels, which modulate the rearrangement of the carbon flux by reprogramming gene expression patterns and intracellular redox states. Anaerobic cell growth and L-lysine production as well as aerobic succinate production have been demonstrated by engineering the metabolic pathways or supplying a terminal electron acceptor instead of oxygen. In this review, we discuss the physiological and metabolic changes in C. glutamicum associated with its application as a cell factory under different oxygen states. Physiological switching in bacteria is initiated with the sensing of oxygen availability. While such a sensor has not been identified in C. glutamicum yet, the molecular mechanism for oxygen sensing in related bacteria is also discussed. KEY POINTS: • C. glutamicum produces a wide variety of useful compounds under anaerobic conditions. • C. glutamicum is a versatile cell factory under both aerobic and anaerobic conditions. • Metabolic fate can be overcome by engineering metabolic pathways.

Quinones and humus are ubiquitous in the biosphere and play an important role in the anaerobic biodegradation and biotransformation of organic acids, poisonous compounds as well as inorganic compounds. The impact of humic model compound, anthraquinone-2, 6-disulfonate (AQDS) on anaerobic phenol and p-cresol degradation were studied. Four methanogenic AQDS-free phenol and p-cresol enrichments and two phenol-AQDS enrichments were obtained using two sludges with potential biodegradability of phenol and cresol isomers as inoculum. 16S rRNA gene-cloning analysis combined with fluorescence in situ hybridization revealed that syntrophic aromatic compound degrading bacterium Syntrophorhabdus aromaticivorans was dominant in four AQDS-free enrichments, whereas phenol degrading Cryptanaerobacter phenolicus was dominant in two phenol-AQDS enrichments. Neither co-culture of S. aromaticivorans with Methanospirillum hungatei nor two phenol-AQDS enrichments could metabolize phenol using AQDS as the terminal electron acceptor. Further degradation experiments suggested that C. phenolicus related microbes in two phenol-AQDS enrichments were responsible for the conversion of phenol to benzoate, and benzoate was further degraded by benzoate degraders of Syntrophus aciditrophicus or Sporotomaculum syntrophicum to acetate. HIGHLIGHTS Anaerobic degradation of phenol and p-cresol were faster than o- and m-cresols.; 2 mM AQDS mitigated phenol and completely inhibited p-cresol degradation.; The dominant bacteria in AQDS-free and phenol-AQDS enrichments were different.; Neither S. aromaticivorans nor C. phenolicus affinities could use AQDS as TEA.; The characteristic of key phenol degraders in phenol-AQDS enrichments was described.

Biogas (methane) as a source of renewable energy, was produced in the anaerobic co-digestion of blackwater (BW, municipal toilet wastewater) and organic kitchen waste (KW). The impact on methane production of various BW to KW mixing ratios, with and without the addition of granular activated carbon (GAC), were studied under thermophilic (55 °C) temperatures. GAC is reported to enhance methane production in such digestions through direct interspecies electron transfer. The results showed that the co-digestion of BW and KW under the 1:2 VS ratio significantly improved the biomethane potential (BMP). In the absence of GAC, an optimal BW:KW ratio was found to be 1:2, achieving a BMP of 0.76 g CH4-COD/g feed-COD. With GAC addition, the BMP increased to 0.81 g CH4-COD/g feed-COD, the lag phase in the digestion was significantly reduced, and the methane production rate increased. Microbial communities in the BW-KW anaerobic digestion were analyzed with and without the addition of GAC. Methanothermobacter and Methanosarcina were predominant archaea in BW-KW digests, with and without GAC amendment, while a third methanogen, Methanomassiliicoccus, was enriched with the addition of GAC to the digest. Further, through SEM image, the enrichment of pili-like stucture was observed in GAC surface.

Sulfonates include diverse natural products and anthropogenic chemicals and are widespread in the environment. Many bacteria can degrade sulfonates and obtain sulfur, carbon, and energy for growth, playing important roles in the biogeochemical sulfur cycle. Cleavage of the inert sulfonate C–S bond involves a variety of enzymes, cofactors, and oxygen-dependent and oxygen-independent catalytic mechanisms. Sulfonate degradation by strictly anaerobic bacteria was recently found to involve C–S bond cleavage through O2-sensitive free radical chemistry, catalyzed by glycyl radical enzymes (GREs). The associated discoveries of new enzymes and metabolic pathways for sulfonate metabolism in diverse anaerobic bacteria have enriched our understanding of sulfonate chemistry in the anaerobic biosphere. An anaerobic environment of particular interest is the human gut microbiome, where sulfonate degradation by sulfate- and sulfite-reducing bacteria (SSRB) produces H2S, a process linked to certain chronic diseases and conditions.

Little is known about the cell physiology of anammox bacteria growing at extremely low growth rates. Here, “Candidatus Brocadia sinica” and “Candidatus Scalindua sp.” were grown in continuous anaerobic membrane bioreactors (MBRs) with complete biomass retention to determine maintenance energy (i.e., power) requirements at near-zero growth rates. After prolonged retentostat cultivations, the specific growth rates (μ) of “Ca. B. sinica” and “Ca. Scalindua sp.” decreased to 0.000023 h(−1) (doubling time of 1255 days) and 0.000157 h(−1) (184 days), respectively. Under these near-zero growth conditions, substrate was continuously utilized to meet maintenance energy demands (m(e)) of 6.7 ± 0.7 and 4.3 ± 0.7 kJ mole of biomass-C(−1) h(−1) for “Ca. B. sinica” and “Ca. Scalindua sp.”, which accorded with the theoretically predicted values of all anaerobic microorganisms (9.7 and 4.4 kJ mole of biomass-C(−1) h(−1)at 37 °C and 28 °C, respectively). These m(e) values correspond to 13.4 × 10(−15) and 8.6 × 10(−15) watts cell(−1) for “Ca. B. sinica” and “Ca. Scalindua sp.”, which were five orders of magnitude higher than the basal power limit for natural settings (1.9 × 10(−19) watts cells(−1)). Furthermore, the minimum substrate concentrations required for growth (S(min)) were calculated to be 3.69 ± 0.21 and 0.09 ± 0.05 μM NO(2)(−) for “Ca. B. sinica” and “Ca. Scalindua sp.”, respectively. These results match the evidence that “Ca. Scalindua sp.” with lower maintenance power requirement and S(min) are better adapted to energy-limited natural environments than “Ca. B. sinica”, suggesting the importance of these parameters on ecological niche differentiation in natural environments.

A novel strictly anaerobic, Gram-negative bacterium, designated as strain FXJYN30E22T, was isolated from the feces of a healthy woman in Yining county, Xinjiang province, China. This strain was non-spore-forming, bile-resistant, non-motile and rod-shaped. It was found to belong to a single separate group in the Phocaeicola genus based on its 16 S ribosomal RNA (rRNA) gene sequence. Alignments of 16 S rRNA gene sequences showed only a low sequence identity (≤ 95.5 %) between strain FXJYN30E22T and all other Phocaeicola strains in public data bases. The genome (43.0% GC) of strain FXJYN30E22T was sequenced, and used for phylogenetic analysis which showed that strain FXJYN30E22T was most closely related to the type strain Phocaeicola massiliensis JCM 13223T. The average nucleotide identity (ANI) value and digital DNA–DNA hybridization (dDDH) between FXJYN30E22T and P. massiliensis JCM 13223T were 90.4 and 41.9 %, which were lower than the generally accepted species boundaries (94.0 and 70 %, respectively). The major cellular fatty acids and polar lipids were anteiso-branched C15:0 and phosphatidylethanolamine, respectively. The result of genome annotation and KEGG analysis showed that strain FXJYN30E22T contains a number of genes in polysaccharide and fatty acid synthesis that indicated adaptation to the human gut system. Furthermore, a pbpE (penicillin-binding protein) gene was found in the genome of strain FXJYN30E22T but in no other Phocaeicola species, which suggested this gene might be contribute to the adaptive capacity of strain FXJYN30E22T. Based on our data, strain FXJYN30E22T (= CGMCC1.17870T/KCTC25195T) was classified as a novel Phocaeicola species, and the name Phocaeicola faecalis sp. nov., was proposed.

For a successful granulation process in activated sludge systems, the stimulation of slow growing organisms such as glycogen accumulating microorganisms (GAOs) is a key factor. Here we show that the introduction of an anaerobic feast followed by an aerobic famine phase successfully transforms bulking sludge, caused by the abundance of genus Kouleothrix, to a hybrid floccular-granular sludge. Two sequencing batch reactors (SBRs) were operated for 228 days treating the same industrial wastewater derived from the cleaning of trucks transporting liquid food (the cargo consists of approximately 70% chocolate and 30% beer). By respectively applying a fast and slow feeding in two parallel SBRs, different degrees of substrate build-up were achieved in the two reactors during the feast phase. The F/M ratio over the feeding time was 1.41 ± 0.48 and 0.57 ± 0.16 kg COD·(kg VSS*d)−1 for the fast-fed and the slow-fed SBR respectively. Our results demonstrate that substrate build-up during the anaerobic selection step is not necessary to obtain well-settling granular-like sludge.

Primary metabolism in cyanobacteria is built on the Calvin-Benson-Bassham (CBB) cycle, oxidative pentose phosphate (OPP) pathway, Embden-Meyerhof-Parnas (EMP) pathway, and the tricarboxylic acid (TCA) cycle. Phosphoketolase (Xpk), commonly found in cyanobacteria, is an enzyme that is linked to all these pathways. However, little is known about its physiological role. Here, we show that most of the cyanobacterial Xpk surveyed are inhibited by ATP, and both copies of Xpk in nitrogen-fixing Cyanothece ATCC51142 are further activated by ADP, suggesting their role in energy regulation. Moreover, Xpk in Synechococcus elongatus PCC7942 and Cyanothece ATCC51142 show that their expressions are dusk-peaked, suggesting their roles in dark conditions. Finally, we find that Xpk in S. elongatus PCC7942 is responsible for survival using ATP produced from the glycogen-to-acetate pathway under dark, anaerobic condition. Interestingly, under this condition, xpk deletion causes glucose secretion in response to osmotic shock such as NaHCO3, KHCO3 and NaCl as part of incomplete glycogen degradation. These findings unveiled the role of this widespread enzyme and open the possibility for enhanced glucose secretion from cyanobacteria.

The capability to design microbiomes with predictable functions would enable new technologies for applications in health, agriculture, and bioprocessing. Towards this goal, we develop a model-guided approach to design synthetic human gut microbiomes for production of the health-relevant metabolite butyrate. Our data-driven model quantifies microbial interactions impacting growth and butyrate production separately, providing key insights into ecological mechanisms driving butyrate production. We use our model to explore a vast community design space using a design-test-learn cycle to identify high butyrate-producing communities. Our model can accurately predict community assembly and butyrate production across a wide range of species richness. Guided by the model, we identify constraints on butyrate production by high species richness and key molecular factors driving butyrate production, including hydrogen sulfide, environmental pH, and resource competition. In sum, our model-guided approach provides a flexible and generalizable framework for understanding and accurately predicting community assembly and metabolic functions., Microbiomes designed with predictable functions could enable broad applications in health, agriculture and bioprocessing. Here the authors use a model-guided approach to design diverse synthetic human gut communities for production of the health-relevant metabolite butyrate.

Entodinium caudatum is an anaerobic binucleated ciliate representing the most dominant protozoal species in the rumen. However, its biological features are largely unknown due to the inability to establish an axenic culture. In this study, we primally sequenced its macronucleus (MAC) genome to aid the understanding of its metabolism, physiology, ecology. We isolated the MAC of E. caudatum strain MZG-1 and sequenced the MAC genome using Illumina MiSeq, MinION, and PacBio RSII systems. De novo assembly of the MiSeq sequence reads followed with subsequent scaffolding with MinION and PacBio reads resulted in a draft MAC genome about 117 Mbp. A large number of carbohydrate-active enzymes were likely acquired through horizontal gene transfer. About 8.74% of the E. caudatum predicted proteome was predicted as proteases. The MAC genome of E. caudatum will help better understand its important roles in rumen carbohydrate metabolism, and interaction with other members of the rumen microbiome.

Biobased production of 5-aminovalerate (5AVA) from biomass can support a sustainable and economic biorefinery process to produce bio-based nylon 5 for food packaging materials. Cost-competitive production of 5AVA from biomass is a key factor in the successful commercialization of nylon 5. Bioproduction of 5AVA is a promising candidate for the industrial process to the current petrochemical route. In this study, we developed an artificial 2-keto-6-aminocaproate-mediated pathway for cost-competitive and high efficiency production of 5AVA in engineered Escherichia coli. Firstly, the combination of native l -lysine α-oxidase (RaiP) from Scomber japonicas, α-ketoacid decarboxylase (KivD) from Lactococcus lactis and aldehyde dehydrogenase (PadA) from Escherichia coli could efficiently convert l -lysine into 5AVA. Moreover, the engineered strains ML03-PnirB-RKP, ML03-PPL-PR-RKP, ML03-PM1-93-RKP induced by anaerobic condition, temperature-induced, constitutive expression instead of expensive isopropyl β-D-thiogalactoside were constructed, respectively. The use of nirB promoter induced by anaerobic condition not only could attain a higher titer of 5AVA than PL-PR and M1-93 promoters, but omit cost of expensive exogenous inducers. After the replacement of industrial materials, 5AVA titer successfully reached 33.68 g/L in engineered strain ML03-PnirB-RKP via biotransformation. This biotransformation process conduces to the cosmically industrial 5AVA bioproduction.

Parageobacillus thermoglucosidasius represents a thermophilic, facultative anaerobic bacterial chassis, with several desirable traits for metabolic engineering and industrial production. To further optimize strain productivity, a systems level understanding of its metabolism is needed, which can be facilitated by a genome-scale metabolic model. Here, we present p-thermo, the most complete, curated and validated genome-scale model (to date) of Parageobacillus thermoglucosidasius NCIMB 11955. It spans a total of 890 metabolites, 1175 reactions and 917 metabolic genes, forming an extensive knowledge base for P. thermoglucosidasius NCIMB 11955 metabolism. The model accurately predicts aerobic utilization of 22 carbon sources, and the predictive quality of internal fluxes was validated with previously published 13C-fluxomics data. In an application case, p-thermo was used to facilitate more in-depth analysis of reported metabolic engineering efforts, giving additional insight into fermentative metabolism. Finally, p-thermo was used to resolve a previously uncharacterised bottleneck in anaerobic metabolism, by identifying the minimal required supplemented nutrients (thiamin, biotin and iron(III)) needed to sustain anaerobic growth. This highlights the usefulness of p-thermo for guiding the generation of experimental hypotheses and for facilitating data-driven metabolic engineering, expanding the use of P. thermoglucosidasius as a high yield production platform.

Resistant starches (RSs) with different structural features were isolated from both native and pullulanase-debranched and acid-hydrolyzed pea starches. Their microscopic changes, short-chain fatty acids (SCFA) composition, microbiota communities, and structural characteristics of the corresponding fermenta residues by the end of 24 h of the in vitro fermentation period were investigated. The microbial fermentation clearly caused numerous cracks and erosion on the RS granule surface. In comparison to the positive control, significantly higher levels of butyrate, propionate, and total SCFA were produced after 24 h of in vitro fecal fermentation when resistant starches were used as substrates. The RS substrates with different structural characteristics enabled varying growth of Bifidobacterium spp., Eubacterium spp., and Faecalibacterium spp. The discrepancy in microbiota communities associated with the differences in SCFA from the fermentation of RS with different structural features would be critical toward the rational design of foods containing resistant starch with targeted health benefits.

In this study, aimed at investigating and characterizing river sediment bacteria, we isolated a Gram-stain-positive, rod-shaped, obligate anaerobic bacterium, strain CBA3637T, from the sediment of the Geum River. This strain grew at 10–40 °C (optimum, 30 °C), 0–1% NaCl (optimum, 0%), and pH 7–8 (optimum, pH 7). The 16S rRNA gene sequence comparison revealed Aminipila butyrica DSM 103574T to be the closest relative of strain CBA3637T (96.6–96.7% similarity); and both strains clustered together in phylogenetic analysis. The genome of strain CBA3637T was found to consist of a single chromosome (3.51 Mbp; 36.98% G + C content). Comparative genomic analysis of the strain CBA3637T with A. butyrica DSM 103574T revealed that strain CBA3637T possessed five unique pathways related to polyamine biosynthesis, lipopolysaccharide metabolism, pyrimidine metabolism, and cofactor and vitamin metabolism. Strain CBA3637T contained C14:0, C16:0, and C18:1 ω9c as the major fatty acids, and diphosphatidylglycerol as the major polar lipid. No respiratory quinone was observed. Biochemical, chemotaxonomic, and genotypic data revealed that the strain CBA3637T is a representative of a novel species within the genus Aminipila, for which the name Aminipila terrae is proposed. The type strain is CBA3637T (= KACC 21651T = DSM 110662T).

Anaerobic digestion is promising for waste activated sludge (WAS) degradation. However, conventional processes were generally stuck with limited hydrolysis and poor pathogen destruction. Hyperthermophilic digestion at 70 °C has drawn attention in overcoming those issues at a relatively low energy requirement and operating difficulties. In order to illuminate its operation characteristics, a single-stage hyperthermophilic digester was controlled at 70 °C and operated continuously to degrade WAS. 88.7 mL/g VSadded of methane yield could be achieved in the hyperthermophilic system, fourfold higher than that in the mesophilic system. Kinetic analysis revealed that hyperthermophilic digestion was advantageous in converting the non-degradable fraction. Consequently, hydrolysis under the hyperthermophilic condition was able to be significantly improved. Above 10 d was necessary for the hyperthermophilic system to gain such a high methane production. In the case of stability, the organic loading of higher than 10.2 g VS/L/d resulted in increasing limitation from methanogenesis and accumulation of propionic, butyric and valeric acids. In addition to the dominant acetoclastic genus Methanothrix for methane production in the hyperthermophilic system, two hydrogenotrophic methanogens Methanospirillum and Methanothermobacter reached 18.84% and 8.31%, respectively. The genus Coprothermobacter, affiliated with the phylum Firmicutes, made more contribution to protein hydrolysis in the hyperthermophilic digester.

The microbial electrolysis cell assisted anaerobic digestion (MEC-AD) holds great promises over conventional anaerobic digestion. This article reports an experimental investigation of extracellular polymeric substances (EPS), reactive oxygen species (ROS), and the expression of genes associated with extracellular electron transfer (EET) in methanogenic biocathodes. The MEC-AD systems were examined using two cathode materials: carbon fibers and stainless-steel mesh. A higher abundance of hydrogenotrophic Methanobacterium sp. and homoacetogenic Acetobacterium sp. appeared to play a major role in superior methanogenesis from stainless steel biocathode than carbon fibers. Moreover, the higher secretion of EPS accompanied by the lower ROS level in stainless steel biocathode indicated that higher EPS perhaps protected cells from harsh metabolic conditions (possibly unfavorable local pH) induced by faster catalysis of hydrogen evolution reaction. In contrast, EET-associated gene expression patterns were comparable in both biocathodes. Thus, these results indicated hydrogenotrophic methanogenesis is the key mechanism, while cathodic EET has a trivial role in distinguishing performances between two cathode electrodes. These results provide new insights into the efficient methanogenic biocathode development.

How the acetate and propionate accumulation impact anaerobic syntrophy during methane formation is not well understood. To investigate such effect, continuous acetate (35 g/L), propionate (11.25 g/L) and bicarbonate (30 g/L) supplementation were used during mesophilic anaerobic digestion. The high throughput sequencing (16S rRNA and mcrA), Real-Time quantitative PCR, and stable carbon isotope fingerprinting were applied to investigate the structure and activity of microbial community members. The results demonstrated that the abundance of syntrophic acetate oxidizing bacteria exhibited a gradual decrease coupled with heavier stable carbon isotopic signature of methane (δ 13CH4) in the three reagents impacted reactors. The increased acetate and propionate concentrations exerted negative influence on biogas production but the relatively stable hydrogenotrophic methanogens together with syntrophic acetate/propionate oxidizing bacteria kept the stable methane formation facing acetate and propionate accumulation. The functional genes copy number of the hydrogenotrophic Methanocellaceae and Methanomicrobiaceae correlated significantly with δ 13CH4 (R2 > 0.74), but only the abundance of Methanocellaceae fitted well with δ 13CH4 (p

The low growth rate of anammox bacteria is one of the main obstacles to its broad use in removing biological nitrogen in sewage treatment. Free cells show the characteristics of relatively rapid growth. To increase the growth rate, this study cultivated free-living anammox bacteria using a membrane bioreactor. The particle size distributions showed that more than 95.48% of biomass was less than 50 μm in size, and the growth rate of free-living anammox bacteria was shortened to 5.68 days after cultivation. Candidatus Brocadia was the most important anammox genus, with a relative abundance of 8.83%. The key functional genes, including hzs, hdh, and hao, were identified and expressed using metagenomic and metaproteomic analyses. After the rapid cultivation of the free-living anammox bacteria, a potential method shortening the start-up time was proposed to rapidly form anammox biofilm by attaching to carriers. PRACTITIONER POINTS: The free-living anammox bacteria are cultivated in a membrane bioreactor. More than 95.48% of biomass size is less than 50 μm. The doubling time of these free anammox bacteria was 5.68 days. The key functional genes of anammox bacteria were identified and expressed. A novel method is proposed to rapidly form anammox biofilm.

Although polyvinyl alcohol and sodium alginate gel (PVA/SA) cell immobilization technology has been successfully applied in anaerobic ammonium oxidation (anammox) processes, there is no comprehensive evaluation of the PVA:SA ratio in PVA/SA gel beads. Therefore, to determine the optimal PVA:SA ratio, the nitrogen removal performance and structure of PVA/SA anammox gel beads under different PVA:SA ratios were studied through batch experiments. The results suggested that cell immobilization technology could significantly improve the nitrogen removal rate (NRR). PVA concentration was positively correlated with the proportion of -macropore in the gel beads but negatively correlated with mechanical strength. Despite having poor mechanical strength, PVA/SA (12%/2%) gel beads had the highest NRR owing to the increased pore size and were experimentally determined to be the most suitable concentration of immobilized carrier. UASB reactor tests showed that compared with anammox granular sludge, the response time of anammox PVA/SA (12%/2%) beads to increased nitrogen load was shorter and the specific anammox activity was higher. Candidatus "Jettenia" was the dominant bacterium in anammox gel beads, accounting for 37.96% of the community. This study provides a reference for preparing PVA/SA cell immobilization. PRACTITIONER POINTS: Increasing the concentration of PVA can reduce the apoptosis of microorganisms during the gel process. The macropore of PVA/SA beads increased with the increase of the PVA:SA ratio. This study provides a reference for preparing PVA/SA gel beads immobilized anammox bacteria.

Rhodopseudomonas palustris is a model microorganism for studying the anaerobic metabolism of aromatic compounds. While it is well documented which aromatics can serve as sole organic carbon sources, co-metabolism of other aromatics is poorly understood. This study used kinetic modeling to analyze the simultaneous degradation of aromatic compounds present in corn stover hydrolysates and model the co-metabolism of aromatics not known to support growth of R. palustris as sole organic substrates. The simulation predicted that p-coumaroyl amide and feruloyl amide were hydrolyzed to p-coumaric acid and ferulic acid, respectively, and further transformed via p-coumaroyl-CoA and feruloyl-CoA. The modeling also suggested that metabolism of p-hydroxyphenyl aromatics was slowed by substrate inhibition, whereas the transformation of guaiacyl aromatics was inhibited by their p-hydroxyphenyl counterparts. It also predicted that substrate channeling may occur during degradation of p-coumaroyl-CoA and feruloyl-CoA, resulting in no detectable accumulation of p-hydroxybenzaldehyde and vanillin, during the transformation of these CoA ligated compounds to p-hydroxybenzoic acid and vanillic acid, respectively. While the simulation correctly represented the known transformation of p-hydroxybenzoic acid via the benzoyl-CoA pathway, it also suggested co-metabolism of vanillic acid and syringic acid, which are known not to serve as photoheterotrophic growth substrate for R. palustris.

It has been previously shown that magnetite (Fe3O4) nanoparticles stimulate the anaerobic digestion process in several anaerobic reactors. Here we evaluate the effect of magnetite nanoparticles on the efficiency of anaerobic digestion of distillers grains with solubles and sugar beet pulp in mesophilic batch experiments. The addition of magnetite nanopowder had a positive effect on the anaerobic digestion process. CH4 was produced faster in the presence of 50 mg of Fe3O4 per 1 g of added total solids than from treatments without addition of Fe3O4. These results demonstrate that the addition of magnetite enhances the methanogenic decomposition of organic acids. Microbial community structure and dynamics were investigated based on bacterial and archaeal 16S rRNA genes, as well as mcrA genes encoding the methyl-CoM reductase. Depending on the reactor, Bacteroides, midas_1138, Petrimonas, unclassified Rikenellaceae (class Bacteroidia), Ruminiclostridium, Proteiniclasticum, Herbinix, and Intestinibacter (class Clostridia) were the main representatives of the bacterial communities. The archaeal communities in well-performed anaerobic reactors were mainly represented by representatives of the genera Methanosarcina and Methanobacterium. Based on our findings, Fe3O4 nanoparticles, when used properly, will improve biomethane production.

Background There is a dearth of sequenced and closed microbial genomes from environments that exceed > 500 m below level terrestrial surface. Coupled with even fewer cultured isolates, study and understanding of how life endures in the extreme oligotrophic subsurface environments is greatly hindered. Using a de novo hybrid assembly of Illumina and Oxford Nanopore sequences we produced a circular genome with corresponding methylome profile of the recently characterized thermophilic, anaerobic, and fumarate-respiring subsurface bacterium, Thermanaerosceptrum fracticalcis, strain DRI-13T to understand how this microorganism survives the deep subsurface. Results The hybrid assembly produced a single circular genome of 3.8 Mb in length with an overall GC content of 45%. Out of the total 4022 annotated genes, 3884 are protein coding, 87 are RNA encoding genes, and the remaining 51 genes were associated with regulatory features of the genome including riboswitches and T-box leader sequences. Approximately 24% of the protein coding genes were hypothetical. Analysis of strain DRI-13T genome revealed: 1) energy conservation by bifurcation hydrogenase when growing on fumarate, 2) four novel bacterial prophages, 3) methylation profile including 76.4% N6-methyladenine and 3.81% 5-methylcytosine corresponding to novel DNA methyltransferase motifs. As well a cluster of 45 genes of unknown protein families that have enriched DNA mCpG proximal to the transcription start sites, and 4) discovery of a putative core of bacteriophage exclusion (BREX) genes surrounded by hypothetical proteins, with predicted functions as helicases, nucleases, and exonucleases. Conclusions The de novo hybrid assembly of strain DRI-13T genome has provided a more contiguous and accurate view of the subsurface bacterium T. fracticalcis, strain DRI-13T. This genome analysis reveals a physiological focus supporting syntrophy, non-homologous double stranded DNA repair, mobility/adherence/chemotaxis, unique methylome profile/recognized motifs, and a BREX defense system. The key to microbial subsurface survival may not rest on genetic diversity, but rather through specific syntrophy niches and novel methylation strategies.

OBJECTIVE The aim of this study was to examine the effect of osteopathic manipulative treatment (OMT) on anaerobic performance and lactate clearance in male athletes. METHODS This study was a double-blind, randomized, sham-controlled and crossover trial. Fourteen male athletes were volunteered to participate this study. All subjects visited to laboratory 3 times in total: familiarization session, test session 1, and test session 2, respectively. At the beginning of the study, the subjects were randomly divided into 2 groups: In sessions 1 and 2, (a) 30-minute OMT or sham treatment before Wingate anaerobic cycling test (WAnT), (b) 30-second WAnT test, and (c) 10-minute OMT or sham therapy between 5th and 15th minutes of passive rest after WAnT was applied to all subjects, respectively. In both groups blood samples were taken at rest and 5, 15 and 30 minute after the WAnT for the determination of lactate concentrations. RESULTS There was no significant differences in WAnT parameters such as peak power, mean power and fatigue index between the OMT and sham treatment. Blood lactate levels were significantly higher 5, 15 and 30 minute after the WAnT when compared to the rest and were lower 15 and 30 minute after the WAnT when compared to 5 minute after the WAnT in both groups (P < 0.05). In addition, blood lactate concentration was significantly lower in OMT than sham treatment at 15 and 30 minute after the WAnT (P < 0.05). CONCLUSIONS This study suggests that OMT may improve lactate clearance while not affecting anaerobic performance in athletes.

Rice (Oryza sativa L.) is an important crop and a staple food for half of the population around the world. The recent water and labor shortages are encouraging farmers to shift from traditional transplanting to direct-seeding. However, poor germination and slow elongation of the coleoptile constrains large-scale application of direct-seeding. This study was aimed to investigate the genetic basis of the anaerobic germination (AG) potential using a set of Oryza nivara (O. nivara) introgression lines (ILs). In this study, a total of 131 ILs were developed by introducing O. nivara chromosome segments into the elite indica rice variety 93-11 through advanced backcrossing and repeated selfing. A high-density genetic map has been previously constructed with 1,070 bin-markers. The seeds of ILs were germinated and used to measure coleoptile length under normal and anaerobic conditions. QTLs associated with AG potential were determined in rice. Based on the high-density genetic map of the IL population, two QTLs, qAGP1 and qAGP3 associated with AG tolerance were characterized and located on chromosomes 1 and 3, respectively. Each QTL explained 15% of the phenotypic variance. Specifically, the O. nivara-derived chromosome segments of the two QTLs were positively tolerance to anaerobic condition by increasing coleoptile length. In a further analysis of public transcriptome data, a total of 26 and 36 genes within qAGP1 and qAGP3 were transcriptionally induced by anaerobic stress, respectively. Utilization of O. nivara-derived alleles at qAGP1 and qAGP3 can potentially enhance tolerance to anaerobic stress at the germination stage in rice, thereby accelerating breeding of rice varieties to be more adaptative for direct-seeding.

The anaerobic digestion performance correlates with the functional microbial community. Mesophilic and thermophilic digestions of vegetable waste were conducted, and dynamics of the microbial community were investigated. The mesophilic and thermophilic collapsed stages occurred at organic loading rates of 1.5 and 2.0 g VS/(L d) due to the accumulation of volatile fatty acids with final concentrations of 2276 and 6476 mg/L, respectively. A high concentration of volatile fatty acids caused the severe inhibition of methanogens, which finally led to the imbalance between acetogenesis and methanogenesis. The mesophilic digestion exhibited a higher microbial diversity and richness than the thermophilic digestion. Syntrophic acetate-oxidizing coupled with hydrogenotrophic methanogenesis was the dominant pathway in the thermophilic stable system, and acetoclastic methanogenesis in the mesophilic stable system. The dominant acidogens, syntrophus, and methanogens were unclassified_f__Anaerolineaceae (8.68%), Candidatus_Cloacamonas (19.70%), Methanosaeta (6.10%), and Methanosarcina (4.08%) in the mesophilic stable stage, and Anaerobaculum (12.59%), Syntrophaceticus (4.84%), Methanosarcina (30.58%), and Methanothermobacter (3.17%) in thermophilic stable stage. Spirochaetae and Thermotogae phyla were the characteristic microorganisms in the mesophilic and thermophilic collapsed stages, respectively. These findings provided valuable information for the deep understanding of the difference of the microbial community and methane-producing mechanism between mesophilic and thermophilic digestion of vegetable waste.

The γ isomer of hexachlorocyclohexane (HCH), also known as lindane, is a carcinogenic persistent organic pollutant. Lindane was used worldwide as an agricultural insecticide. Legacy soil and groundwater contamination with lindane and other HCH isomers is still a big concern. The biotic reductive dechlorination of HCH to nondesirable and toxic lower chlorinated compounds such as monochlorobenzene (MCB) and benzene, among others, has been broadly documented. Here, we demonstrate that complete biodegradation of lindane to nontoxic end products is attainable using a sequential treatment approach with three mixed anaerobic microbial cultures referred to as culture I, II, and III. Biaugmentation with culture I achieved dechlorination of lindane to MCB and benzene. Culture II was able to dechlorinate MCB to benzene, and finally, culture III carried out methanogenic benzene degradation. Distinct Dehalobacter populations, corresponding to different 16S rRNA amplicon sequence variants in culture I and culture II, were responsible for lindane and MCB dechlorination, respectively. This study continues to highlight key roles of Dehalobacter as chlorobenzene- and HCH -respiring bacteria and demonstrates that sequential treatment with specialized anaerobic cultures may be explored at field sites in order to address legacy soil and groundwater contamination with HCH.

Chain elongation is a growth-dependent anaerobic metabolism that combines acetate and ethanol into butyrate, hexanoate, and octanoate. While the model microorganism for chain elongation, Clostridium kluyveri, was isolated from a saturated soil sample in the 1940s, chain elongation has remained unexplored in soil environments. During soil fermentative events, simple carboxylates and alcohols can transiently accumulate up to low mM concentrations, suggesting in situ possibility of microbial chain elongation. Here, we examined the occurrence and microbial ecology of chain elongation in four soil types in microcosms and enrichments amended with chain elongation substrates. All soils showed evidence of chain elongation activity with several days of incubation at high (100 mM) and environmentally relevant (2.5 mM) concentrations of acetate and ethanol. Three soils showed substantial activity in soil microcosms with high substrate concentrations, converting 58% or more of the added carbon as acetate and ethanol to butyrate, butanol, and hexanoate. Semi-batch enrichment yielded hexanoate and octanoate as the most elongated products and microbial communities predominated by C. kluyveri and other Firmicutes genera not known to undergo chain elongation. Collectively, these results strongly suggest a niche for chain elongation in anaerobic soils that should not be overlooked in soil microbial ecology studies.

Artículo escrito por un elevado número de autores, solo se referencian el que aparece en primer lugar, el nombre del grupo de colaboración, si le hubiere, y los autores pertenecientes a la UAM, Background: Extreme terrestrial, analogue environments are widely used models to study the limits of life and to infer habitability of extraterrestrial settings. In contrast to Earth’s ecosystems, potential extraterrestrial biotopes are usually characterized by a lack of oxygen. Methods: In the MASE project (Mars Analogues for Space Exploration), we selected representative anoxic analogue environments (permafrost, salt-mine, acidic lake and river, sulfur springs) for the comprehensive analysis of their microbial communities. We assessed the microbiome profile of intact cells by propidium monoazide-based amplicon and shotgun metagenome sequencing, supplemented with an extensive cultivation effort. Results: The information retrieved from microbiome analyses on the intact microbial community thriving in the MASE sites, together with the isolation of 31 model microorganisms and successful binning of 15 high-quality genomes allowed us to observe principle pathways, which pinpoint specific microbial functions in the MASE sites compared to moderate environments. The microorganisms were characterized by an impressive machinery to withstand physical and chemical pressures. All levels of our analyses revealed the strong and omnipresent dependency of the microbial communities on complex organic matter. Moreover, we identified an extremotolerant cosmopolitan group of 34 poly-extremophiles thriving in all sites. Conclusions: Our results reveal the presence of a core microbiome and microbial taxonomic similarities between saline and acidic anoxic environments. Our work further emphasizes the importance of the environmental, terrestrial parameters for the functionality of a microbial community, but also reveals a high proportion of living microorganisms in extreme environments with a high adaptation potential within habitability borders. Keywords: Extreme environments, Microbiomes, Archaea, Bacteria, Propidium monoazide, Astrobiology, Spaceanalogue, Extremophiles, Extraterrestrial life, Metagenomics