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Purpose: Locomotor profiling using anaerobic speed reserve (ASR) enables insights into athletes' physiological and neuromuscular contributing factors and prescription of high-intensity training beyond maximal aerobic speed (MAS). This systematic review aimed to determine the validity and reliability of different methods to assess the characteristics of ASR, i.e., MAS and maximal sprinting speed (MSS)., Methods: A comprehensive search of the PubMed and Web of Science databases was conducted according to the PRISMA guidelines. Studies were included if they reported data on validity and/or reliability for methods to assess MAS or MSS., Results: 58 studies were included with 28 studies referring to MAS and 30 studies to MSS. Regarding MAS, different methods for cardiopulmonary exercise testing yielded different values (four out of seven studies) of MAS (Cohen's d (ES) = 0.83-2.8; Pearson's r/intraclass correlation coefficient (r/ICC) = 0.46-0.85). Criterion validity of different field tests showed heterogeneous results (ES = 0-3.57; r/ICC = 0.40-0.96). Intraday and interday reliability was mostly acceptable for the investigated methods (ICC/r>0.76; CV<16.9%). Regarding MSS, radar and laser measurements (one out of one studies), timing gates (two out of two studies), and video analysis showed mostly good criterion validity (two out of two studies) (ES = 0.02-0.53; r/ICC = 0.93-0.98) and reliability (r/ICC>0.83; CV<2.43%). Criterion validity (ES = 0.02-7.11) and reliability (r/ICC = 0.14-0.97; CV = 0.7-9.77%) for global or local positioning systems (seven out of nine studies) and treadmill sprinting (one out of one studies) was not acceptable in most studies., Conclusion: The criterion validity of incremental field tests or shuttle runs to examine MAS cannot be confirmed. Results on time trials indicate that distances adapted to the participants' sporting background, fitness, or sex might be suitable to estimate MAS. Regarding MSS, only sprints with radar or laser measures, timing gates, or video analysis provide valid and reliable results for linear sprints of 20 to 70 m., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Thron et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Mitochondrial metabolism is entirely dependent on the biosynthesis of the [4Fe-4S] clusters, which are part of the subunits of the respiratory chain. The mitochondrial late ISC pathway mediates the formation of these clusters from simpler [2Fe-2S] molecules and transfers them to client proteins. Here, we characterized the late ISC pathway in one of the simplest mitochondria, mitosomes, of the anaerobic protist Giardia intestinalis that lost the respiratory chain and other hallmarks of mitochondria. In addition to IscA2, Nfu1 and Grx5 we identified a novel BolA1 homologue in G. intestinalis mitosomes. It specifically interacts with Grx5 and according to the high-affinity pulldown also with other core mitosomal components. Using CRISPR/Cas9 we were able to establish full bolA1 knock out, the first cell line lacking a mitosomal protein. Despite the ISC pathway being the only metabolic role of the mitosome no significant changes in the mitosome biology could be observed as neither the number of the mitosomes or their capability to form [2Fe-2S] clusters in vitro was affected. We failed to identify natural client proteins that would require the [2Fe-2S] or [4Fe-4S] cluster within the mitosomes, with the exception of [2Fe-2S] ferredoxin, which is itself part of the ISC pathway. The overall uptake of iron into the cellular proteins remained unchanged as also observed for the grx5 knock out cell line. The pull-downs of all late ISC components were used to build the interactome of the pathway showing specific position of IscA2 due to its interaction with the outer mitosomal membrane proteins. Finally, the comparative analysis across Metamonada species suggested that the adaptation of the late ISC pathway identified in G. intestinalis occurred early in the evolution of this supergroup of eukaryotes., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Motyčková et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The six-component maintenance of lipid asymmetry (Mla) system is responsible for retrograde transport of phospholipids, ensuring the barrier function of the Gram-negative cell envelope. Located within the outer membrane, MlaA (VacJ) acts as a channel to shuttle phospholipids from the outer leaflet. We identified Neisseria gonorrhoeae MlaA (ngo2121) during high-throughput proteomic mining for potential therapeutic targets against this medically important human pathogen. Our follow-up phenotypic microarrays revealed that lack of MlaA results in a complex sensitivity phenome. Herein we focused on MlaA function in cell envelope biogenesis and pathogenesis. We demonstrate the existence of two MlaA classes among 21 bacterial species, characterized by the presence or lack of a lipoprotein signal peptide. Purified truncated N. gonorrhoeae MlaA elicited antibodies that cross-reacted with a panel of different Neisseria. Little is known about MlaA expression; we provide the first evidence that MlaA levels increase in stationary phase and under anaerobiosis but decrease during iron starvation. Lack of MlaA resulted in higher cell counts during conditions mimicking different host niches; however, it also significantly decreased colony size. Antimicrobial peptides such as polymyxin B exacerbated the size difference while human defensin was detrimental to mutant viability. Consistent with the proposed role of MlaA in vesicle biogenesis, the ΔmlaA mutant released 1.7-fold more membrane vesicles. Comparative proteomics of cell envelopes and native membrane vesicles derived from ΔmlaA and wild type bacteria revealed enrichment of TadA–which recodes proteins through mRNA editing–as well as increased levels of adhesins and virulence factors. MlaA-deficient gonococci significantly outcompeted (up to 16-fold) wild-type bacteria in the murine lower genital tract, suggesting the growth advantage or increased expression of virulence factors afforded by inactivation of mlaA is advantageous in vivo. Based on these results, we propose N. gonorrhoeae restricts MlaA levels to modulate cell envelope homeostasis and fine-tune virulence. [ABSTRACT FROM AUTHOR]

Sulfate-coupled anaerobic oxidation of methane (AOM) is performed by multicellular consortia of anaerobic methanotrophic archaea (ANME) in obligate syntrophic partnership with sulfate-reducing bacteria (SRB). Diverse ANME and SRB clades co-associate but the physiological basis for their adaptation and diversification is not well understood. In this work, we used comparative metagenomics and phylogenetics to investigate the metabolic adaptation among the 4 main syntrophic SRB clades (HotSeep-1, Seep-SRB2, Seep-SRB1a, and Seep-SRB1g) and identified features associated with their syntrophic lifestyle that distinguish them from their non-syntrophic evolutionary neighbors in the phylum Desulfobacterota. We show that the protein complexes involved in direct interspecies electron transfer (DIET) from ANME to the SRB outer membrane are conserved between the syntrophic lineages. In contrast, the proteins involved in electron transfer within the SRB inner membrane differ between clades, indicative of convergent evolution in the adaptation to a syntrophic lifestyle. Our analysis suggests that in most cases, this adaptation likely occurred after the acquisition of the DIET complexes in an ancestral clade and involve horizontal gene transfers within pathways for electron transfer (CbcBA) and biofilm formation (Pel). We also provide evidence for unique adaptations within syntrophic SRB clades, which vary depending on the archaeal partner. Among the most widespread syntrophic SRB, Seep-SRB1a, subclades that specifically partner ANME-2a are missing the cobalamin synthesis pathway, suggestive of nutritional dependency on its partner, while closely related Seep-SRB1a partners of ANME-2c lack nutritional auxotrophies. Our work provides insight into the features associated with DIET-based syntrophy and the adaptation of SRB towards it., Competing Interests: The authors have declared that no competing interests exist., (Copyright: This is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.)

Proteus mirabilis is an opportunistic pathogen associated with a variety of human infections, including urinary tract infections. The prevalence of P. mirabilis in foods of animal origin and in the manure by-products created in animal production is not well documented. Further, the prevalence and persistence of extended-spectrum cephalosporin (ESC) resistant P. mirabilis is largely unknown. In this study, we characterized ESC-resistant P. mirabilis recovered from various stages of dairy manure anaerobic digestion. Isolates were screened by PCR for blaCTX-M, blaCMY and blaSHV, and antimicrobial susceptibility testing was performed. Fifty-six P. mirabilis carrying CTX-M were sequenced with short and long read sequencing technologies, and the assembled chromosomes and plasmids were compared. ESC-resistant Proteus was found in four of the six manure digesters, an indication that not all digesters were colonized with resistant strains. Both CTX-M-1 and CTX-M-15 plasmids were found in P. mirabilis isolates. Transfer of plasmid DNA by conjugation was also explored, with ESC-resistance plasmids able to transfer to Escherichia coli at high frequency. We concluded that P. mirabilis can harbour and transfer ESC-resistance genes and plasmids, and may be an overlooked reservoir of antimicrobial resistance., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Chalmers et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

This study investigated the relationship between psychological skills and fitness levels among elite taekwondo athletes. A total of ten Iranian male elite taekwondo athletes (mean age of 20.6±2 years, BMI 18.78±0.62 kg/m2, and fat percentage of 8.87±1.46%) participated in the study. The Sports Emotional Intelligence Questionnaire, Sports Success Scale, Sport Mental Toughness Questionnaire, and Mindfulness Inventory for Sport were used to assess psychological factors. The Wingate test was used to determine anaerobic power, and the Bruce test to determine aerobic fitness. Descriptive statistics and Spearman rank correlation coefficients were utilised to examine any relationships between subscales. Statistically significant correlations were recorded between the evaluation of feelings (EI scale) and VO2peak (ml/kg/min) (r = -0.70, p = 0.0235) and between social skills (EI scale) and relative peak power (W/kg) (r = 0.84, p = 0.0026). Also, between optimism (EI scale) and VO2peak (ml/kg/min) (r = -0.70, p = 0.0252) and between optimism (EI scale) and HR-MAX (r = -0.75, p = 0.0123); and, finally, between control (mental toughness scale) and relative peak power (W/kg) (r = 0.67, p = 0.0360). These findings demonstrate relationships between psychological factors and the advantages of good anaerobic and aerobic capabilities. Finally, the study also demonstrated that elite taekwondo athletes have high mental performance abilities that are interrelated with anaerobic and aerobic performance., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Nabilpour et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Organisms have evolved elaborate physiological pathways that regulate growth, proliferation, metabolism, and stress response. These pathways must be properly coordinated to elicit the appropriate response to an ever-changing environment. While individual pathways have been well studied in a variety of model systems, there remains much to uncover about how pathways are integrated to produce systemic changes in a cell, especially in dynamic conditions. We previously showed that deletion of Protein Kinase A (PKA) regulatory subunit BCY1 can decouple growth and metabolism in Saccharomyces cerevisiae engineered for anaerobic xylose fermentation, allowing for robust fermentation in the absence of division. This provides an opportunity to understand how PKA signaling normally coordinates these processes. Here, we integrated transcriptomic, lipidomic, and phospho-proteomic responses upon a glucose to xylose shift across a series of strains with different genetic mutations promoting either coupled or decoupled xylose-dependent growth and metabolism. Together, results suggested that defects in lipid homeostasis limit growth in the bcy1Δ strain despite robust metabolism. To further understand this mechanism, we performed adaptive laboratory evolutions to re-evolve coupled growth and metabolism in the bcy1Δ parental strain. The evolved strain harbored mutations in PKA subunit TPK1 and lipid regulator OPI1, among other genes, and evolved changes in lipid profiles and gene expression. Deletion of the evolved opi1 gene partially reverted the strain's phenotype to the bcy1Δ parent, with reduced growth and robust xylose fermentation. We suggest several models for how cells coordinate growth, metabolism, and other responses in budding yeast and how restructuring these processes enables anaerobic xylose utilization., Competing Interests: JJC is a consultant for Thermo Fisher Scientific., (Copyright: © 2023 Wagner et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

We previously were able to predict the anaerobic mechanical power outputs using features taken from a maximal incremental cardiopulmonary exercise stress test (CPET). Since a standard aerobic exercise stress test (with electrocardiogram and blood pressure measurements) has no gas exchange measurement and is more popular than CPET, our goal, in the current paper, was to investigate whether features taken from a clinical exercise stress test (GXT), either submaximal or maximal, can predict the anaerobic mechanical power outputs to the same level as we found with CPET variables. We have used data taken from young healthy subjects undergoing CPET aerobic test and the Wingate anaerobic test, and developed a computational predictive algorithm, based on greedy heuristic multiple linear regression, which enabled the prediction of the anaerobic mechanical power outputs from a corresponding GXT measures (exercise test time, treadmill speed and slope). We found that for submaximal GXT of 85% age predicted HRmax, a combination of 3 and 4 variables produced a correlation of r = 0.93 and r = 0.92 with % error equal to 15 ± 3 and 16 ± 3 on the validation set between real and predicted values of the peak and mean anaerobic mechanical power outputs (p < 0.001), respectively. For maximal GXT (100% of age predicted HRmax), a combination of 4 and 2 variables produced a correlation of r = 0.92 and r = 0.94 with % error equal to 12 ± 2 and 14 ± 3 on the validation set between real and predicted values of the peak and mean anaerobic mechanical power outputs (p < 0.001), respectively. The newly developed model allows to accurately predict the anaerobic mechanical power outputs from a standard, submaximal and maximal GXT. Nevertheless, in the current study the subjects were healthy, normal individuals and therefore the assessment of additional subjects is desirable for the development of a test applicable to other populations., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Leopold et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Candida albicans can cause two major types of infections: superficial infection and systemic candidiasis. C. albicans infects diverse host niches, owing to a wide range of virulence factors and attributes, such as morphological transitions and phenotypic switching. C. albicans uses glycolysis, followed by alcoholic fermentation or mitochondrial respiration to rapidly generate ATP under aerobic conditions. In this study, we quantified the mRNA expression of several glycolysis-related enzymes associated with the initial phase of environmental changes using two strains: a type strain, NBRC 1385, and a strain from a patient with auto-brewery syndrome, LSEM 550. Additionally, we analyzed the regulation of a rate-limiting enzyme in glycolysis, phosphofructokinase 1 (PFK1). Our results showed that the mRNA expression of enzymes in the middle and last stages of glycolysis and alcoholic fermentation increased, and that of mitochondrial respiration enzymes decreased under short-term anaerobic conditions. Carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP) administration showed similar results under anaerobic conditions. Moreover, PFK1 maintained its regulatory effect under different conditions; no significant change was observed in its mRNA expression. Our results suggest that C. albicans obtains energy via carbohydrate catabolism in the early phase of environmental change and survives in various parts of the host., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Okabayashi et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The opportunistic human pathogen Pseudomonas aeruginosa can thrive under microaerophilic to anaerobic conditions in the lungs of cystic fibrosis patients. RNASeq based comparative RNA profiling of the clinical isolate PA14 cultured in synthetic cystic fibrosis medium was performed after planktonic growth (OD600 = 2.0; P), 30 min after shift to anaerobiosis (A-30) and after anaerobic biofilm growth for 96h (B-96) with the aim to reveal differentially regulated functions impacting on sustained anoxic biofilm formation as well as on tolerance towards different antibiotics. Most notably, functions involved in sulfur metabolism were found to be up-regulated in B-96 cells when compared to A-30 cells. Based on the transcriptome studies a set of transposon mutants were screened, which revealed novel functions involved in anoxic biofilm growth.In addition, these studies revealed a decreased and an increased abundance of the oprD and the mexCD-oprJ operon transcripts, respectively, in B-96 cells, which may explain their increased tolerance towards meropenem and to antibiotics that are expelled by the MexCD-OprD efflux pump. The OprI protein has been implicated as a target for cationic antimicrobial peptides, such as SMAP-29. The transcriptome and subsequent Northern-blot analyses showed that the abundance of the oprI transcript encoding the OprI protein is strongly decreased in B-96 cells. However, follow up studies revealed that the susceptibility of a constructed PA14ΔoprI mutant towards SMAP-29 was indistinguishable from the parental wild-type strain, which questions OprI as a target for this antimicrobial peptide in strain PA14. [ABSTRACT FROM AUTHOR]

Pseudomonas aeruginosa is an opportunistic bacterial pathogen able to thrive in highly diverse ecological niches and to infect compromised patients. Its genome exhibits a mosaic structure composed of a core genome into which accessory genes are inserted en bloc at specific sites. The size and the content of the core genome are open for debate as their estimation depends on the set of genomes considered and the pipeline of gene detection and clustering. Here, we redefined the size and the content of the core genome of P. aeruginosa from fully re-analyzed genomes of 17 reference strains. After the optimization of gene detection and clustering parameters, the core genome was defined at 5,233 orthologs, which represented ~ 88% of the average genome. Extrapolation indicated that our panel was suitable to estimate the core genome that will remain constant even if new genomes are added. The core genome contained resistance determinants to the major antibiotic families as well as most metabolic, respiratory, and virulence genes. Although some virulence genes were accessory, they often related to conserved biological functions. Long-standing prophage elements were subjected to a genetic drift to eventually display a G+C content as higher as that of the core genome. This contrasts with the low G+C content of highly conserved ribosomal genes. The conservation of metabolic and respiratory genes could guarantee the ability of the species to thrive on a variety of carbon sources for energy in aerobiosis and anaerobiosis. Virtually all the strains, of environmental or clinical origin, have the complete toolkit to become resistant to the major antipseudomonal compounds and possess basic pathogenic mechanisms to infect humans. The knowledge of the genes shared by the majority of the P. aeruginosa isolates is a prerequisite for designing effective therapeutics to combat the wide variety of human infections. [ABSTRACT FROM AUTHOR]

Background: In yaws-endemic areas, two-thirds of exudative cutaneous ulcers (CU) are associated with Treponema pallidum subsp. pertenue (TP) and Haemophilus ducreyi (HD); one-third are classified as idiopathic ulcers (IU). A yaws eradication campaign on Lihir Island in Papua New Guinea utilizing mass drug administration (MDA) of azithromycin initially reduced but failed to eradicate yaws; IU rates remained constant throughout the study. Using 16S rRNA gene sequencing, we previously determined that Streptococcus pyogenes was associated with some cases of IU. Here, we applied shotgun metagenomics to the same samples we analyzed previously by 16S rRNA sequencing to verify this result, identify additional IU-associated microorganisms, and determine why S. pyogenes-associated IU might have persisted after MDA of azithromycin., Methodology/principal Findings: We sequenced DNA extracted from 244 CU specimens separated into four groups based upon microorganism-specific PCR results (HD+, TP+, TP+HD+, and TP-HD- or IU). S. pyogenes was enriched in IU (24.71% relative abundance [RA]) specimens compared to other ulcer sub-groups, confirming our prior results. We bioinformatically identified the emm (M protein gene) types found in the S. pyogenes IU specimens and found matches to emm156 and emm166. Only ~39% of IU specimens contained detectable S. pyogenes, suggesting that additional organisms could be associated with IU. In the sub-set of S. pyogenes-negative IU specimens, Criibacterium bergeronii, a member of the Peptostreptococcaceae, and Fusobacterium necrophorum (7.07% versus 0.00% RA and 2.18% versus 0.00% RA, respectively), were enriched compared to the S. pyogenes-positive sub-set. Although a broad range of viruses were detected in the CU specimens, none were specifically associated with IU., Conclusions/significance: Our observations confirm the association of S. pyogenes with IU in yaws-endemic areas, and suggest that additional anaerobic bacteria, but not other microorganisms, may be associated with this syndrome. Our results should aid in the design of diagnostic tests and selective therapies for CU., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2022 Griesenauer et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Herein we report the use of an environmental multimetal(loid)-resistant strain, MF05, to biosynthesize single- or multi-element nanostructures under anaerobic conditions. Inorganic nanostructure synthesis typically requires methodologies and conditions that are harsh and environmentally hazardous. Thus, green/eco-friendly procedures are desirable, where the use of microorganisms and their extracts as bionanofactories is a reliable strategy. First, MF05 was entirely sequenced and identified as an Escherichia coli-related strain with some genetic differences from the traditional BW25113. Secondly, we compared the CdS nanostructure biosynthesis by whole-cell in a design defined minimal culture medium containing sulfite as the only sulfur source to obtain sulfide reduction from a low-cost chalcogen reactant. Under anaerobic conditions, this process was greatly favored, and irregular CdS (ex. 370 nm; em. 520-530 nm) was obtained. When other chalcogenites were tested (selenite and tellurite), only spherical Se0 and elongated Te0 nanostructures were observed by TEM and analyzed by SEM-EDX. In addition, enzymatic-mediated chalcogenite (sulfite, selenite, and tellurite) reduction was assessed by using MF05 crude extracts in anaerobiosis; similar results for nanostructures were obtained; however Se0 and Te0 formation were more regular in shape and cleaner (with less background). Finally, the in vitro nanostructure biosynthesis was assessed with salts of Ag, Au, Cd, and Li alone or in combination with chalcogenites. Several single or binary nanostructures were detected. Our results showed that MF05 is a versatile anaerobic bionanofactory for different types of inorganic NS. synthesis., Competing Interests: The authors have declared that no competing interests exist.

Background: Hepatic steatosis is a major risk factor for graft failure in liver transplantation. Hepatic steatosis shows a greater negative influence on graft function following prolonged cold ischaemia. As the impact of steatosis on hepatocyte metabolism during extended cold ischaemia is not well-described, we compared markers of metabolic capacity and mitochondrial function in steatotic and lean livers following clinically relevant durations of cold preservation. Methods: Livers from 10-week old leptin-deficient obese (ob/ob, n = 9) and lean C57 mice (n = 9) were preserved in ice-cold University of Wisconsin solution. Liver mitochondrial function was then assessed using high resolution respirometry after 1.5, 3, 5, 8, 12, 16 and 24 hours of storage. Metabolic marker enzymes for anaerobiosis and mitochondrial mass were also measured in conjunction with non-bicarbonate tissue pH buffering capacity. Results: Ob/ob and lean mice livers showed severe (>60%) macrovesicular and mild (<30%) microvesicular steatosis on Oil Red O staining, respectively. Ob/ob livers had lower baseline enzymatic complex I activity but similar adenosine triphosphate (ATP) levels compared to lean livers. During cold storage, the respiratory control ratio and complex I-fueled phosphorylation deteriorated approximately twice as fast in ob/ob livers compared to lean livers. Ob/ob livers also demonstrated decreased ATP production capacities at all time-points analyzed compared to lean livers. Ob/ob liver baseline lactate dehydrogenase activities and intrinsic non-bicarbonate buffering capacities were depressed by 60% and 40%, respectively compared to lean livers. Conclusions: Steatotic livers have impaired baseline aerobic and anaerobic capacities compared to lean livers, and mitochondrial function indices decrease particularly from after 5 hours of cold preservation. These data provide a mechanistic basis for the clinical recommendation of shorter cold storage durations in steatotic donor livers. [ABSTRACT FROM AUTHOR]

This study aims to evaluate whether a test protocol with standardized and individualized resistance settings leads to valid wheelchair Wingate tests (WAnT) and graded exercise tests (GXT) in healthy novices. Twenty able-bodied individuals (10M/10F, age 23 ± 2 years, body mass 72 ± 11 kg) performed an isometric strength test, sprint test, WAnT and GXT on a wheelchair ergometer. Using a previously developed set of regression equations, individuals' isometric strength outcome was used to estimate the WAnT result (P30est), from which an effective individual WAnT resistance was derived. The subsequently measured WAnT outcome (P30meas) was used to estimate the GXT outcome (POpeakest) and to scale the individual GXT resistance steps. Estimated and measured outcomes were compared. The WAnT protocol was considered valid when maximal velocity did not exceed 3 m·s-1; the GXT protocol was considered valid when test duration was 8-12 min. P30est did not significantly differ from P30meas, while one participant did not have a valid WanT, as maximal velocity exceeded 3 m·s-1. POpeakest was 10% higher than POpeakmeas, and six participants did not reach a valid GXT: five participants had a test duration under 8 min and one participant over 12 min. The isometric strength test can be used to individually scale the WAnT protocol. The WAnT outcome scaled the protocol for the GXT less accurately, resulting in mostly shorter-than-desired test durations. In conclusion, the evaluated standardized and individualized test protocol was valid for the WAnT but less valid for the GXT among a group of novices. Before implementing the standardized individual test protocol on a broader scale, e.g. among paralympic athletes, it should be evaluated among different athletic wheelchair-dependent populations., Competing Interests: The authors have declared that no competing interests exist.

FNR is a well-studied global regulator of anaerobiosis, which is widely conserved across bacteria. Despite the importance of FNR and anaerobiosis in microbial lifestyles, the factors that influence its function on a genome-wide scale are poorly understood. Here, we report a functional genomic analysis of FNR action. We find that FNR occupancy at many target sites is strongly influenced by nucleoid-associated proteins (NAPs) that restrict access to many FNR binding sites. At a genome-wide level, only a subset of predicted FNR binding sites were bound under anaerobic fermentative conditions and many appeared to be masked by the NAPs H-NS, IHF and Fis. Similar assays in cells lacking H-NS and its paralog StpA showed increased FNR occupancy at sites bound by H-NS in WT strains, indicating that large regions of the genome are not readily accessible for FNR binding. Genome accessibility may also explain our finding that genome-wide FNR occupancy did not correlate with the match to consensus at binding sites, suggesting that significant variation in ChIP signal was attributable to cross-linking or immunoprecipitation efficiency rather than differences in binding affinities for FNR sites. Correlation of FNR ChIP-seq peaks with transcriptomic data showed that less than half of the FNR-regulated operons could be attributed to direct FNR binding. Conversely, FNR bound some promoters without regulating expression presumably requiring changes in activity of condition-specific transcription factors. Such combinatorial regulation may allow Escherichia coli to respond rapidly to environmental changes and confer an ecological advantage in the anaerobic but nutrient-fluctuating environment of the mammalian gut. [ABSTRACT FROM AUTHOR]

Like a majority of photosynthetic microorganisms, the green unicellular alga Chlamydomonas reinhardtii may encounter O2 deprived conditions on a regular basis. In response to anaerobiosis or in a respiration defective context, the photosynthetic electron transport chain of Chlamydomonas is remodeled by a state transition process to a conformation that favours the photoproduction of ATP at the expense of reductant synthesis. In some unicellular green algae including Chlamydomonas, anoxia also triggers the induction of a chloroplast-located, oxygen sensitive hydrogenase, which accepts electrons from reduced ferredoxin to convert protons into molecular hydrogen. Although microalgal hydrogen evolution has received much interest for its biotechnological potential, its physiological role remains unclear. By using specific Chlamydomonas mutants, we demonstrate that the state transition ability and the hydrogenase function are both critical for induction of photosynthesis in anoxia. These two processes are thus important for survival of the cells when they are transiently placed in an anaerobic environment. [ABSTRACT FROM AUTHOR]

Ribonucleotide reductases (RNRs) use radical-based chemistry to catalyze the conversion of all four ribonucleotides to deoxyribonucleotides. The ubiquitous nature of RNRs necessitates multiple RNR classes that differ from each other in terms of the phosphorylation state of the ribonucleotide substrates, oxygen tolerance, and the nature of both the metallocofactor employed and the reducing systems. Although these differences allow RNRs to produce deoxyribonucleotides needed for DNA biosynthesis under a wide range of environmental conditions, they also present a challenge for establishment of a universal activity assay. Additionally, many current RNR assays are limited in that they only follow the conversion of one ribonucleotide substrate at a time, but in the cell, all four ribonucleotides are actively being converted into deoxyribonucleotide products as dictated by the cellular concentrations of allosteric specificity effectors. Here, we present a liquid chromatography with tandem mass spectrometry (LC-MS/MS)-based assay that can determine the activity of both aerobic and anaerobic RNRs on any combination of substrates using any combination of allosteric effectors. We demonstrate that this assay generates activity data similar to past published results with the canonical Escherichia coli aerobic class Ia RNR. We also show that this assay can be used for an anaerobic class III RNR that employs formate as the reductant, i.e. Streptococcus thermophilus RNR. We further show that this class III RNR is allosterically regulated by dATP and ATP. Lastly, we present activity data for the simultaneous reduction of all four ribonucleotide substrates by the E. coli class Ia RNR under various combinations of allosteric specificity effectors. This validated LC-MS/MS assay is higher throughput and more versatile than the historically established radioactive activity and coupled RNR activity assays as well as a number of the published HPLC-based assays. The presented assay will allow for the study of a wide range of RNR enzymes under a wide range of conditions, facilitating the study of previously uncharacterized RNRs., Competing Interests: No. The authors have declared that no competing interests exist.

In commercial large-scale aquaria, controlling levels of nitrogenous compounds is essential for macrofauna health. Naturally occurring bacteria are capable of transforming toxic nitrogen species into their more benign counterparts and play important roles in maintaining aquaria health. Nitrification, the microbially-mediated transformation of ammonium and nitrite to nitrate, is a common and encouraged process for management of both commercial and home aquaria. A potentially competing microbial process that transforms ammonium and nitrite to dinitrogen gas (anaerobic ammonium oxidation [anammox]) is mediated by some bacteria within the phylum Planctomycetes. Anammox has been harnessed for nitrogen removal during wastewater treatment, as the nitrogenous end product is released into the atmosphere rather than in aqueous discharge. Whether anammox bacteria could be similarly utilized in commercial aquaria is an open question. As a first step in assessing the viability of this practice, we (i) characterized microbial communities from water and sand filtration systems for four habitats at the Tennessee Aquarium and (ii) examined the abundance and anammox potential of Planctomycetes using culture-independent approaches. 16S rRNA gene amplicon sequencing revealed distinct, yet stable, microbial communities and the presence of Planctomycetes (~1-15% of library reads) in all sampled habitats. Preliminary metagenomic analyses identified the genetic potential for multiple complete nitrogen metabolism pathways. However, no known genes diagnostic for the anammox reaction were found in this survey. To better understand the diversity of this group of bacteria in these systems, a targeted Planctomycete-specific 16S rRNA gene-based PCR approach was used. This effort recovered amplicons that share <95% 16S rRNA gene sequence identity to previously characterized Planctomycetes, suggesting novel strains within this phylum reside within aquaria., Competing Interests: The authors have declared that no competing interests exist.

Cellular metabolism continuously processes an enormous range of external compounds into endogenous metabolites and is as such a key element in human physiology. The multifaceted physiological role of the metabolic network fulfilling the catalytic conversions can only be fully understood from a whole-body perspective where the causal interplay of the metabolic states of individual cells, the surrounding tissue and the whole organism are simultaneously considered. We here present an approach relying on dynamic flux balance analysis that allows the integration of metabolic networks at the cellular scale into standardized physiologically-based pharmacokinetic models at the whole-body level. To evaluate our approach we integrated a genome-scale network reconstruction of a human hepatocyte into the liver tissue of a physiologically-based pharmacokinetic model of a human adult. The resulting multiscale model was used to investigate hyperuricemia therapy, ammonia detoxification and paracetamol-induced toxication at a systems level. The specific models simultaneously integrate multiple layers of biological organization and offer mechanistic insights into pathology and medication. The approach presented may in future support a mechanistic understanding in diagnostics and drug development. [ABSTRACT FROM AUTHOR]

Pseudomonas aeruginosa, a gram-negative bacterium of clinical importance, forms more robust biofilm during anaerobic respiration, a mode of growth presumed to occur in abnormally thickened mucus layer lining the cystic fibrosis (CF) patient airway. However, molecular basis behind this anaerobiosis-triggered robust biofilm formation is not clearly defined yet. Here, we identified a morphological change naturally accompanied by anaerobic respiration in P. aeruginosa and investigated its effect on the biofilm formation in vitro. A standard laboratory strain, PAO1 was highly elongated during anaerobic respiration compared with bacteria grown aerobically. Microscopic analysis demonstrated that cell elongation likely occurred as a consequence of defective cell division. Cell elongation was dependent on the presence of nitrite reductase (NIR) that reduces nitrite (NO2-) to nitric oxide (NO) and was repressed in PAO1 in the presence of carboxy-PTIO, a NO antagonist, demonstrating that cell elongation involves a process to respond to NO, a spontaneous byproduct of the anaerobic respiration. Importantly, the non-elongated NIR-deficient mutant failed to form biofilm, while a mutant of nitrate reductase (NAR) and wild type PAO1, both of which were highly elongated, formed robust biofilm. Taken together, our data reveal a role of previously undescribed cell biological event in P. aeruginosa biofilm formation and suggest NIR as a key player involved in such process. [ABSTRACT FROM AUTHOR]

The article reports that Vitamin B1 is in its active form thiamine pyrophosphate (TPP), an essential cofactor for several key enzymes in the carbohydrate metabolism. It is mentioned that mammals need to salvage crucial nutrient from their diet in order to complement the deficiency of de novo synthesis. In the human pathogenic bacterium Staphylococcus aureus, two operons were identified which are involved in vitamin B1 metabolism. It is mentioned that the kinetic properties of the enzymes as well as the binding of TPP to the in vitro transcribed RNA of the proposed operons suggest that the vitamin B1 homeostasis in S. aureus is strongly regulated at transcriptional as well as enzymatic levels.

Background: The Escherichia coli heterodimeric HU protein is a small DNA-bending protein associated with the bacterial nucleoid. It can introduce negative supercoils into closed circular DNA in the presence of topoisomerase I. Cells lacking HU grow very poorly and display many phenotypes. Methodology/Principal Findings: We analyzed the transcription profile of every Escherichia coli gene in the absence of one or both HU subunits. This genome-wide in silico transcriptomic approach, performed in parallel with in vivo genetic experimentation, defined the HU regulon. This large regulon, which comprises 8% of the genome, is composed of four biologically relevant gene classes whose regulation responds to anaerobiosis, acid stress, high osmolarity, and SOS induction. Conclusions/Significance: The regulation a large number of genes encoding enzymes involved in energy metabolism and catabolism pathways by HU explains the highly pleiotropic phenotype of HU-deficient cells. The uniform chromosomal distribution of the many operons regulated by HU strongly suggests that the transcriptional and nucleoid architectural functions of HU constitute two aspects of a unique protein-DNA interaction mechanism. [ABSTRACT FROM AUTHOR]

The present study aimed to propose a new multidimensional taekwondo-specific test to estimate aerobic power, anaerobic fitness, and agility. Out of sixty-five male volunteers, forty-six, forty-eight, and fifty athletes (18-35 years; black- and red-belt level) were included in the final analysis for aerobic, anaerobic, and agility assessments, respectively. Maximum oxygen uptake (VO2max, using a graded exercise test on a treadmill), anaerobic power (using the 30-s Wingate anaerobic test, WAnT), and agility performance (using the agility T-Test) were measured via non-specific laboratory and field tests across a two-week period. The taekwondo-specific aerobic-anaerobic-agility (TAAA) test comprised six 20-s intervals of shuttle sprints over a 4-m distance, and the execution of roundhouse kicks alternating the legs at the end of each distance, with 10-s rest intervals between the sets. The multiple linear regression revealed that the difference between heart rate (HR) after and 1 minute after the TAAA test (p < 0.001), and body mass index (BMI; p = 0.006) were significant to estimate VO2max. Likewise, there was a very large (R = 0.79) and large (R = 0.55) correlation between the average and maximum number of kicks performed in the TAAA test and the WAnT mean and peak power, respectively (p < 0.001). Moreover, a linear relationship was found between the T-Test and agility performance acquired in the TAAA test (R = 0.74; p < 0.001). The TAAA test can be considered a valid simple tool for monitoring VO2max, anaerobic fitness, and agility in male taekwondo athletes., Competing Interests: The authors have declared that no competing interests exist.

The mechanisms controlling the extraordinarily slow carbon (C) mineralization rates characteristic of Sphagnum-rich peatlands ("bogs") are not fully understood, despite decades of research on this topic. Soluble phenolic compounds have been invoked as potentially significant contributors to bog peat recalcitrance due to their affinity to slow microbial metabolism and cell growth. Despite this potentially significant role, the effects of soluble phenolic compounds on bog peat C mineralization remain unclear. We analyzed this effect by manipulating the concentration of free soluble phenolics in anaerobic bog and fen peat incubations using water-soluble polyvinylpyrrolidone ("PVP"), a compound that binds with and inactivates phenolics, preventing phenolic-enzyme interactions. CO2 and CH4 production rates (end-products of anaerobic C mineralization) generally correlated positively with PVP concentration following Michaelis-Menten (M.M.) saturation functions. Using M.M. parameters, we estimated that the extent to which phenolics inhibit anaerobic CO2 production was significantly higher in the bog-62 ± 16%-than the fen-14 ± 4%. This difference was found to be more substantial with regards to methane production-wherein phenolic inhibition for the bog was estimated at 54 ± 19%, while the fen demonstrated no apparent inhibition. Consistent with this habitat difference, we observed significantly higher soluble phenolic content in bog vs. fen pore-water. Together, these findings suggest that soluble phenolics could contribute to bogs' extraordinary recalcitrance and high (relative to other peatland habitats) CO2:CH4 production ratios., Competing Interests: The authors have declared that no competing interests exist.

Inoculation dose is a key operational parameter for the solid-state anaerobic digestion (SS-AD) of lignocellulosic biomass, maximum methane recovery, and stable digester performance. The novelty of this study was the co-digestion of unamended full-strength grape marc and cheese whey for peak methane extraction at variable inoculation levels. An acclimatised digestate from a preceding anaerobic treatment was used as a downstream inoculum. The impact of inoculum size (wet weight) was evaluated at 0/10, 5/5, 7/3 and 9/1 substrate-to-inoculum (S/I) ratios, corresponding to an initial concentration of 20-30% total solids (TS) in digesters over 58 days at 45°C. The optimal 7/3 S/I produced the highest cumulative methane yield, 6.45 L CH4 kg-1 VS, coinciding with the lowest initial salinity at 11%; the highest volumetric methane productivity rate of 0.289±0.044 L CH4 LWork-1 d-1; the highest average COD/N ratio of 9.88; the highest final pH of 9.13, and a maximum 15.07% elemental carbon removal; for a lag time of 9.4 days. This study identified an optimal inoculation dose and opens up an avenue for the direct co-digestion of grape marc and cheese whey without requirements for substrate pretreatment, thus improving the overall bioenergy profile of the winery and dairy joint resource recovery operations., Competing Interests: The authors have declared that no competing interests exist.

Sequencing-based protocols for studying the human microbiome have unearthed a wealth of information about the relationship between the microbiome and human health. But these microbes cannot be leveraged as therapeutic targets without culture-based studies to phenotype species of interest and to establish culture collections for use in animal models. Traditional sample collection protocols are focused on preserving nucleic acids and metabolites and are largely inappropriate for preserving sensitive anaerobic bacteria for later culture recovery. Here we introduce a novel microbiome preservation kit (BIOME-Preserve) that facilitates recovery of anaerobic bacteria from human stool. Using a combination of culture recovery and shallow whole-genome shotgun sequencing, we characterized the anaerobes cultured from fresh human stool and from human stool held at room temperature in BIOME-Preserve for up to 120 hours. We recovered several species of interest to microbiome researchers, including Bifidobacterium spp., Bacteroides spp., Blautia spp., Eubacterium halii (now Anaerobutyricum hallii), Akkermansia muciniphila, and Faecalibacterium prausnitzii. We also demonstrated that freezing at -80°C did not adversely affect our ability to culture organisms from BIOME-Preserve, suggesting that it is appropriate both as a transport medium and as a medium for longer-term ultra-cold storage. Together, our results suggest BIOME-Preserve is practical for the collection, transport, and culture of anaerobic bacteria from human samples and can help enable researchers to better understand the link between the microbiome and human health and how to leverage that link through novel microbiome-based therapeutics., Competing Interests: I have read the journal’s policy and the authors of this manuscript have the following competing interests: H.L. is an employee of Anaerobe Systems; S.C. is the president of Anaerobe Systems. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

The anaerobic oxidation of methane coupled to sulfate reduction is a microbially mediated process requiring a syntrophic partnership between anaerobic methanotrophic (ANME) archaea and sulfate-reducing bacteria (SRB). Based on genome taxonomy, ANME lineages are polyphyletic within the phylum Halobacterota, none of which have been isolated in pure culture. Here, we reconstruct 28 ANME genomes from environmental metagenomes and flow sorted syntrophic consortia. Together with a reanalysis of previously published datasets, these genomes enable a comparative analysis of all marine ANME clades. We review the genomic features that separate ANME from their methanogenic relatives and identify what differentiates ANME clades. Large multiheme cytochromes and bioenergetic complexes predicted to be involved in novel electron bifurcation reactions are well distributed and conserved in the ANME archaea, while significant variations in the anabolic C1 pathways exists between clades. Our analysis raises the possibility that methylotrophic methanogenesis may have evolved from a methanotrophic ancestor., Competing Interests: S.J.H. is a co-founder of Koonkie Inc., a bioinformatics consulting company that designs and provides scalable algorithmic and data analytics solutions in the cloud.

Entamoeba histolytica is believed to be devoid of peroxisomes, like most anaerobic protists. In this work, we provided the first evidence that peroxisomes are present in E. histolytica, although only seven proteins responsible for peroxisome biogenesis (peroxins) were identified (Pex1, Pex6, Pex5, Pex11, Pex14, Pex16, and Pex19). Targeting matrix proteins to peroxisomes is reduced to the PTS1-dependent pathway mediated via the soluble Pex5 receptor, while the PTS2 receptor Pex7 is absent. Immunofluorescence microscopy showed that peroxisomal markers (Pex5, Pex14, Pex16, Pex19) are present in vesicles distinct from mitosomes, the endoplasmic reticulum, and the endosome/phagosome system, except Pex11, which has dual localization in peroxisomes and mitosomes. Immunoelectron microscopy revealed that Pex14 localized to vesicles of approximately 90-100 nm in diameter. Proteomic analyses of affinity-purified peroxisomes and in silico PTS1 predictions provided datasets of 655 and 56 peroxisomal candidates, respectively; however, only six proteins were shared by both datasets, including myo-inositol dehydrogenase (myo-IDH). Peroxisomal NAD-dependent myo-IDH appeared to be a dimeric enzyme with high affinity to myo-inositol (Km 0.044 mM) and can utilize also scyllo-inositol, D-glucose and D-xylose as substrates. Phylogenetic analyses revealed that orthologs of myo-IDH with PTS1 are present in E. dispar, E. nutalli and E. moshkovskii but not in E. invadens, and form a monophyletic clade of mostly peroxisomal orthologs with free-living Mastigamoeba balamuthi and Pelomyxa schiedti. The presence of peroxisomes in E. histolytica and other archamoebae breaks the paradigm of peroxisome absence in anaerobes and provides a new potential target for the development of antiparasitic drugs., Competing Interests: The authors have declared that no competing interests exist.

Antibiotic resistance genes (ARGs) are emerging contaminants causing serious global health concern. Interventions to address this concern include improving our understanding of methods for treating waste material of human and animal origin that are known to harbor ARGs. Anaerobic digestion is a commonly used process for treating dairy manure, and although effective in reducing ARGs, its mechanism of action is not clear. In this study, we used three ARGs to conducted a longitudinal bench scale anaerobic digestion experiment with various temperatures (28, 36, 44, and 52°C) in triplicate using fresh dairy manure for 30 days to evaluate the reduction of gene abundance. Three ARGs and two mobile genetic elements (MGEs) were studied: sulfonamide resistance gene (sulII), tetracycline resistance genes (tetW), macrolide-lincosamide-streptogramin B (MLSB) superfamily resistance genes (ermF), class 1 integrase gene (intI1), and transposase gene (tnpA). Genes were quantified by real-time quantitative PCR. Results show that the thermophilic anaerobic digestion (52°C) significantly reduced (p < 0.05) the absolute abundance of sulII (95%), intI1 (95%), tnpA (77%) and 16S rRNA gene (76%) after 30 days of digestion. A modified Collins-Selleck model was used to fit the decay curve, and results suggest that the gene reduction during the startup phase of anaerobic digestion (first 5 days) was faster than the later stage, and reductions in the first five days were more than 50% for most genes., Competing Interests: The authors have declared that no competing interests exist.

Oil-produced wastewater treatment plants, especially those involving biological treatment processes, harbor rich and diverse microbes. However, knowledge of microbial ecology and microbial interactions determining the efficiency of plants for oil-produced wastewater is limited. Here, we performed 16S rDNA amplicon sequencing to elucidate the microbial composition and potential microbial functions in a full-scale well-worked offshore oil-produced wastewater treatment plant. Results showed that microbes that inhabited the plant were diverse and originated from oil and marine associated environments. The upstream physical and chemical treatments resulted in low microbial diversity. Organic pollutants were digested in the anaerobic baffled reactor (ABR) dominantly through fermentation combined with sulfur compounds respiration. Three aerobic parallel reactors (APRs) harbored different microbial groups that performed similar potential functions, such as hydrocarbon degradation, acidogenesis, photosynthetic assimilation, and nitrogen removal. Microbial characteristics were important to the performance of oil-produced wastewater treatment plants with biological processes., Competing Interests: The authors have read the journal’s policy and have the following competing interests: WL is an employee of Sinopec Shengli Oilfield, and LW is an employee of CNOOC Energy Development Co. Ltd. This does not alter our adherence to PLOS ONE policies on sharing data and materials. There are no patents, products in development or marketed products associated with this research to declare.

Burkholderia pseudomallei, the etiological agent of melioidosis in humans and animals, often occupies environmental niches and infection sites characterized by limited concentrations of oxygen. Versatile genomic features enable this pathogen to maintain its physiology and virulence under hypoxia, but the crucial regulatory networks employed to switch from oxygen dependent respiration to alternative terminal electron acceptors (TEA) like nitrate, remains poorly understood. Here, we combined a Tn5 transposon mutagenesis screen and an anaerobic growth screen to identify a two-component signal transduction system with homology to RegAB. We show that RegAB is not only essential for anaerobic growth, but also for full virulence in cell lines and a mouse infection model. Further investigations of the RegAB regulon, using a global transcriptomic approach, identified 20 additional regulators under transcriptional control of RegAB, indicating a superordinate role of RegAB in the B. pseudomallei anaerobiosis regulatory network. Of the 20 identified regulators, NarX/L and a FNR homolog were selected for further analyses and a role in adaptation to anaerobic conditions was demonstrated. Growth experiments identified nitrate and intermediates of the denitrification process as the likely signal activateing RegAB, NarX/L, and probably of the downstream regulators Dnr or NsrR homologs. While deletions of individual genes involved in the denitrification process demonstrated their important role in anaerobic fitness, they showed no effect on virulence. This further highlights the central role of RegAB as the master regulator of anaerobic metabolism in B. pseudomallei and that the complete RegAB-mediated response is required to achieve full virulence. In summary, our analysis of the RegAB-dependent modulon and its interconnected regulons revealed a key role for RegAB of B. pseudomallei in the coordination of the response to hypoxic conditions and virulence, in the environment and the host., Competing Interests: The authors have declared that no competing interests exist.

Annually, agricultural activity produces an enormous amount of plant biomass by-product. Many studies have reported the biomethane potential of agro-industrial wastes, but only a few studies have investigated applying the substrates in both batch and continuous mode. Tomato is one of the most popular vegetables globally; its processing releases a substantial amount of by-product, such as stems and leaves. This study examined the BMP of tomato plant (Solanum lycopersicum Mill. L. cv. Alfred) waste. A comparative test revealed that the BMPs of corn stover, tomato waste,and their combination were approximately the same, around 280 mL methane/g Volatile Solid. In contrast, the relative biogas production decreased in the presence of tomato waste in a continuous mesophilic anaerobic digestion system; the daily biogas productions were 860 ± 80, 290 ± 50, and 570 ± 70 mL biogas/gVolatile Solid/day in the case of corn stover, tomato waste, and their mixture, respectively. The methane content of biogas was around 46-48%. The fermentation parameters of the continuous AD experiments were optimal in all cases; thus, TW might have an inhibitory effect on the microbial community. Tomato plant materials contain e.g. flavonoids, glycoalkaloids (such as tomatine and tomatidine), etc. known as antimicrobial and antifungal agents. The negative effect of tomatine on the biogas yield was confirmed in batch fermentation experiments. Metagenomic analysis revealed that the tomato plant waste caused significant rearrangements in the microbial communities in the continuously operated reactors. The results demonstrated that tomato waste could be a good mono-substrate in batch fermentations or a co-substrate with corn stover in a proper ratio in continuous anaerobic fermentations for biogas production. These results also point to the importance of running long-term continuous fermentations to test the suitability of a novel biomass substrate for industrial biogas production., Competing Interests: The authors have declared that no competing interests exist.

Fundamental ecological principles of ecosystem-level respiration are extensively applied in greenhouse gas and elemental cycle studies. A laboratory system termed CEMS (Carbon Dioxide Evolution Measurement System), developed to explore microbial biofilm growth and metabolic responses, was evaluated as an early-warning system for microbial disturbances in industrial settings: in (a) potable water system contamination, and (b) bioreactor inhibition. Respiration was detected as CO2 production, rather than O2 consumption, including aerobic and anaerobic metabolism. Design, thresholds, and benefits of the remote CO2 monitoring technology were described. Headspace CO2 correlated with contamination levels, as well as chemical (R2 > 0.83-0.96) and microbiological water quality indicators (R2 > 0.78-0.88). Detection thresholds were limiting factors in monitoring drinking water to national and international standards (0 CFU/100 mL fecal coliforms) in both open- (>1500 CFU/mL) and closed-loop CO2 measuring regimes (>100 CFU/100 mL). However, closed-loop detection thresholds allow for the detection of significant contamination events, and monitoring less stringent systems such as irrigation water (<100 CFU/mL). Whole-system respiration was effectively harnessed as an early-warning system in bioreactor performance monitoring. Models were used to deconvolute biological CO2 fluctuations from chemical CO2 dynamics, to optimize this real-time, sustainable, low-waste technology, facilitating timeous responses to biological disturbances in bioreactors., Competing Interests: The authors have declared that no competing interests exist.

Mechanisms controlling CO2 and CH4 production in wetlands are central to understanding carbon cycling and greenhouse gas exchange. However, the volatility of these respiration products complicates quantifying their rates of production in the field. Attempts to circumvent the challenges through closed system incubations, from which gases cannot escape, have been used to investigate bulk in situ geochemistry. Efforts towards mapping mechanistic linkages between geochemistry and microbiology have raised concern regarding sampling and incubation-induced perturbations. Microorganisms are impacted by oxygen exposure, increased temperatures and accumulation of metabolic products during handling, storage, and incubation. We probed the extent of these perturbations, and their influence on incubation results, using high-resolution geochemical and microbial gene-based community profiling of anaerobically incubated material from three wetland habitats across a permafrost peatland. We compared the original field samples to the material anaerobically incubated over 50 days. Bulk geochemistry and phylum-level microbiota in incubations largely reflected field observations, but divergence between field and incubations occurred in both geochemistry and lineage-level microbial composition when examined at closer resolution. Despite the changes in representative lineages over time, inferred metabolic function with regards to carbon cycling largely reproduced field results suggesting functional consistency. Habitat differences among the source materials remained the largest driver of variation in geochemical and microbial differences among the samples in both incubations and field results. While incubations may have limited usefulness for identifying specific mechanisms, they remain a viable tool for probing bulk-scale questions related to anaerobic C cycling, including CO2 and CH4 dynamics., Competing Interests: Funding for this research was provided by the Genomic Science Program of the United States Department of Energy Office of Biological and Environmental Research Grants DE-SC0010580 & DESC0016440. A portion of this research was performed using EMSL (Ringgold ID 130367), a DOE Office of Science User Facility sponsored by the Office of Biological and Environmental Research. One or more of the authors is employed by a commercial company: Ventana Medical Systems, Horsely Witten Group, Viosimo LLC. This does not alter our adherence to PLOS ONE policies on sharing data and materials.

Although a number of different methods have been used to quantify soil bacteria, identifying the optimal method(s) for soil bacterial abundance is still in question. No single method exists for undertaking an absolute microbial count using culture-dependent methods (CDMs) or even culture-independent methods (CIMs). This study investigated soil storage and pretreatment methods for optimal bacterial counts. Appropriate storage temperature (4°C) and optimal pretreatment methods (sonication time for 3 min and centrifugation at 1400 g) were necessary to preserve bacterial cell viability and eliminate interference from soil particles. To better estimate soil bacterial numbers under various cellular state and respiration, this study also evaluated three CDMs (i.e., colony forming unit, spotting, and most probable number (MPN) and three CIMs (i.e., flow cytometry (FCM), epifluorescence microscopy (EM) count, and DNA quantitation). Each counting method was tested using 72 soil samples collected from a local arable farm site at three different depths (i.e., 10-20, 90-100, and 180-190 cm). Among all CDMs, MPN was found to be rapid, simple, and reliable. However, the number of bacteria quantified by MPN was 1-2 orders lower than that quantified by CIMs, likely due to the inability of MPN to count anaerobic bacteria. The DNA quantitation method appeared to overestimate soil bacterial numbers, which may be attributed to DNA from dead bacteria and free DNA in the soil matrix. FCM was found to be ineffective in counting soil bacteria as it was difficult to separate the bacterial cells from the soil particles. Dyes used in FCM stained the bacterial DNA and clay particles. The EM count was deemed a highly effective method as it provided information on soil mineral particles, live bacteria, and dead bacteria; however, it was a time-consuming and labor-intensive process. Combining both types of methods was considered the best approach to acquire better information on the characteristics of indigenous soil microorganisms (aerobic versus anaerobic, live versus dead)., Competing Interests: The authors have declared that no competing interests exist.

The objective of this work is to provide a mathematical analysis on how a Thoroughbred horse should regulate its speed over the course of a race to optimize performance. Because Thoroughbred horses are not capable of running the whole race at top speed, determining what pace to set and when to unleash the burst of speed is essential. Our model relies on mechanics, energetics (both aerobic and anaerobic) and motor control. It is a system of coupled ordinary differential equations on the velocity, the propulsive force and the anaerobic energy, that leads to an optimal control problem that we solve. In order to identify the parameters meaningful for Thoroughbred horses, we use velocity data on races in Chantilly (France) provided by France Galop, the French governing body of flat horse racing in France. Our numerical simulations of performance optimization then provide the optimal speed along the race, the oxygen uptake evolution in a race, as well as the energy or the propulsive force. It also predicts how the horse has to change its effort and velocity according to the topography (altitude and bending) of the track., Competing Interests: The authors have declared that no competing interests exist.