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The global interest in fatty acids is steadily rising due to their wealth of industrial potential ranging from cosmetics to biofuels. Unfortunately, certain fatty acids, such as monounsaturated C12, cannot be produced cost and energy-efficiently using conventional methods. Biosynthesis of fatty acids using microorganisms can overcome this drawback. However, rewiring a microbe’s metabolome for increased production remains challenging. To overcome this, sophisticated genome-wide metabolic network models have become available. These models predict the effect of genetic perturbations on the metabolism, thereby serving as a guide for metabolic pathways optimization. In this work, we used constraint-based modeling in combination with the algorithm Optknock to identify gene deletions in Escherichia coli that improve the C12 fatty acid production. Nine gene targets were identified that, when deleted, were predicted to increase C12 titers. Targets play a role in anaplerotic reactions, amino acid synthesis, carbon metabolism and cofactor-balancing. Subsequently, we constructed the corresponding (combinatorial) deletion mutants to validate the in silico predictions in vivo. Our highest producer (Δ maeB Δ ndk Δ pykA) reaches a titer of 6.7 mg/L, corresponding to a 7.5-fold increase in C12 fatty acid production. This study demonstrates that model-guided metabolic engineering is a useful tool to improve C12 fatty acid production.

Strain MDTJ8T is a chain-elongating thermophilic bacterium isolated from a thermophilic acidogenic anaerobic digestor treating human waste while producing the high commodity chemical n-caproate. The strain grows and produces formate, acetate, n-butyrate, n-caproate and lactate from mono-, di- and polymeric saccharides at 37–60 °C (optimum, 50–55 °C) and at pH 5.0–7.0 (optimum, pH 6.5). The organism is an obligate anaerobe, is motile and its cells form rods (0.3–0.5×1.0–3.0 µm) that stain Gram-positive and occur primarily as chains. Phylogenetic analysis of both the 16S rRNA gene and full genome sequence shows that strain MDTJ8T belongs to a group that consists of mesophylic chain-elongating bacteria within the family Oscillospiraceae , being nearest to Caproicibacter fermentans EA1T (94.8 %) and Caproiciproducens galactitolivorans BS-1T (93.7 %). Its genome (1.96 Mbp) with a G+C content of 49.6 mol% is remarkably smaller than those of other chain-elongating bacteria of the family Oscillospiraceae . Pairwise average nucleotide identity and DNA–DNA hybridization values between strain MDJT8T and its mesophilic family members are less than 70 and 35 %, respectively, while pairwise average amino acid identity values are less than 68 %. In addition, strain MDJT8T uses far less carbohydrate and non-carbohydrate substrates compared to its nearest family members. The predominant cellular fatty acids of strain MDTJ8T are C14 : 0, C14 : 0 DMA (dimethyl acetal) and C16 : 0, while its polar lipid profile shows three unidentified glycophospholipids, 11 glycolipids, 13 phospholipids and six unidentified lipids. No respiratory quinones and polyamines are detected. Based on its phylogenetic, genotypic, morphological, physiological, biochemical and chemotaxonomic characteristics, strain MDTJ8T represents a novel species and novel genus of the family Oscillospiraceae and Thermocaproicibacter melissae gen. nov., sp. nov. is proposed as its name. The type strain is MDTJ8T (=DSM 114174T=LMG 32615T=NCCB 100883T).

In vitro periodontal model systems are required for further elucidating phenomena behind periodontal pathogenesis and treatment applications. However, most oral biofilm models apply static conditions and are restricted to end-point measurements. This study established a multispecies periodontal model in the drip flow biofilm reactor in order to mimic shear flow in the oral cavity. The design is re-engineered to enable real-time characterization. A community of five bacteria, Streptococcus gordonii-GFPmut3*, Streptococcus oralis-GFPmut3*, Streptococcus sanguinis-pVMCherry, Fusobacterium nucleatum, and Porphyromonas gingivalis-SNAP26 is visualized using two distinct fluorescent proteins and the SNAP-tag. The biofilm in the reactor develops into a spatially uniform, dense and metabolically active biofilm with relative cell abundances similar to those in a healthy individual. Metabolic activity, structural features and bacterial composition of the biofilm remain stable from 3 up to 6 days. As a proof of concept for our periodontal model, the 3-day developed biofilm is exposed to a prebiotic treatment with L-arginine. Multifaceted effects of L-arginine on the oral biofilm were validated by this model setup. L-arginine showed to inhibit growth and co-aggregations of the pathogenic species, and to reduce biofilm thickness and volume. Additionally, L-arginine is metabolized by Streptococcus gordonii-GFPmut3* and Streptococcus sanguinis-pVMCherry, producing high levels of ornithine in the biofilm. In conclusion, our drip flow reactor setup is promising in studying spatiotemporal behavior of a multispecies periodontal community.

We explore the impact of nutrient scarcity on the interaction dynamics of two common human gut bacteria,Bacteroides thetaiotaomicron(BT) andRoseburia intestinalis(RI). Upon depletion of glucose, the viable cell numbers of BT decrease, whereas RI enters a slow growth mode. When mucin beads are supplemented, both species attach to the beads and the number of viable BT but not RI cells in liquid increases significantly. In co-culture, viable cell numbers decrease or increase significantly compared to monoculture, depending on the time point. A combination of targeted metabolomics and RNA-seq showed that RI’ s slow growth mode represents a diauxic shift towards acetate and lactate consumption, whereas BT survives glucose depletion by foraging mucin sugars. Most tested mucin monosaccharides inhibit the growth of RI but not BT on glucose. We encoded our findings in a kinetic model, which qualitatively reproduces the observed dynamics.

Objective Chlorhexidine mouthrinses are marketed in different formulations. This study aimed at investigating qualitative and quantitative changes in in-vitro multispecies oral biofilms, induced by different chlorhexidine-containing mouthrinses. Background data Earlier studies comparing chlorhexidine mouthrinses are either clinical studies or in-vitro studies assessing the antimicrobial efficacy of the mouthrinses. However, no clear investigations are available regarding ecological impact of different chlorhexidine formulations on in-vitro multispecies oral biofilms after rinsing with different chlorhexidine formulations. Methods Nine commercially available chlorhexidine mouthrinses were selected. Multispecies oral communities (14 species) were grown for 48 h in a Biostat-B Twin bioreactor. After that, they were used to develop biofilms on the surface of hydroxyapatite disks in 24-well pates for 48 h. Biofilms were then rinsed once or multiple times with the corresponding mouthrinse. Biofilms were collected before starting the rinsing experiment and every 24 h for 3 days and vitality quantitative PCR was performed. The experiment was repeated 3 independent times on 3 different days and the results were analyzed using a linear mixed model. Results The mouthrinses provoked different effects in terms of change in total viable bacterial load (VBL), ecology, and community structure of the multispecies biofilms. There was no relation between chlorhexidine concentrations, presence, or absence of cetylpyridinium chloride and/or alcohol, and the observed effects. Some tested chlorhexidine mouthrinses (MC, HG, HH, and HI) strongly lowered the total VBL (≈1007 Geq/ml), but disrupted biofilm symbiosis (≥40% of the biofilms communities are pathobionts). On the other hand, other tested chlorhexidine mouthrinses (MD, ME, and HF) had limited impact on total VBL (≥1010 Geq/ml), but improved the biofilm ecology and community structure (≤10% of the biofilms communities are pathobionts). Conclusion Not all chlorhexidine mouthrinses have the same effect on oral biofilms. Their effect seems to be strongly product dependent and vary according to their compositions and formulations.

The choice of an expression system for the metagenomic DNA of interest is of vital importance for the detection of any particular gene or gene cluster. Most of the screens to date have used the Gram-negative bacterium Escherichia coli as a host for metagenomic gene libraries. However, the use of E. coli introduces a potential host bias since only 40% of the enzymatic activities may be readily recovered by random cloning in E. coli. To recover some of the remaining 60%, alternative cloning hosts such as Streptomyces spp. have been used. Streptomycetes are high-GC Gram-positive bacteria belonging to the Actinomycetales and they have been studied extensively for more than 25 years as an alternative expression system. They are extremely well suited for the expression of DNA from other actinomycetes and genomes of high GC content. Furthermore, due to its high innate, extracellular secretion capacity, Streptomyces can be a better system than E. coli for the production of many extracellular proteins. In this article, an overview is given about the materials and methods for growth and successful expression and secretion of heterologous proteins from diverse origin using Streptomyces lividans as a host. More in detail, an overview is given about the protocols of transformation, type of plasmids used and of vectors useful for integration of DNA into the host chromosome, and accompanying cloning strategies. In addition, various control elements for gene expression including synthetic promoters are discussed, and methods to compare their strength are described. Stable and efficient marker-less integration of the gene of interest under the control of the promoter of choice into S. lividans chromosome via homologous recombination using pAMR23A-based system will be explained. Finally, a basic protocol for bench-top bioreactor experiments which can form the start in the production process optimization and up-scaling will be provided.

Operator training in the chemical industry is important because of the potentially hazardous nature of procedures and the way operators' mistakes can have serious consequences on process operation and safety. Currently, operator training is facing some challenges, such as high costs, safety limitations and time constraints. Also, there have been some indications of a lack of engagement of employees during mandatory training. Immersive technologies can provide solutions to these challenges. Specifically, virtual reality (VR) has the potential to improve the way chemical operators experience training sessions, increasing motivation, virtually exposing operators to unsafe situations, and reducing classroom training time. In this paper, we present research being conducted to develop a virtual reality training solution as part of the EU Horizon 2020 CHARMING Project, a project focusing on the education of current and future chemical industry stakeholders. This paper includes the design principles for a virtual reality training environment including the features that enhance the effectiveness of virtual reality training such as game-based learning elements, learning analytics, and assessment methods. This work can assist those interested in exploring the potential of virtual reality training environments in the chemical industry from a multidisciplinary perspective. (c) 2021 Institution of Chemical Engineers. Published by Elsevier B.V. All rights reserved.

Both in vitro and in vivo studies have shown that the probiotic Limosilactobacillus reuteri can improve oral health. Limosilactobacillus reuteri species are known to produce the antimicrobial "reuterin" from glycerol. In order to further increase its antimicrobial activity, this study evaluated the effect of the combined use of glycerol and Limosilactobacillus reuteri (ATCC PTA 5289) in view of using a synergistic synbiotic over a probiotic. An antagonistic agar growth and a multispecies biofilm model showed that the antimicrobial potential of the probiotic was significantly enhanced against periodontal pathobionts and anaerobic commensals when supplemented with glycerol. Synbiotic biofilms also showed a significant reduction in inflammatory expression of human oral keratinocytes (HOK-18A), but only when the keratinocytes were preincubated with the probiotic. Probiotic preincubation of keratinocytes or probiotic and synbiotic treatment of biofilms alone was insufficient to significantly reduce inflammatory expression. Overall, this study shows that combining glycerol with the probiotic L. reuteri into a synergistic synbiotic can greatly improve the effectiveness of the latter.

A bottleneck for microbial community experiments with many samples and/or replicates is the fast quantification of individual taxon abundances, which is commonly achieved through sequencing marker genes such as the 16S rRNA gene. Here, we propose a new approach for high-throughput and high-quality enumeration of human gut bacteria in a defined community, combining flow cytometry and supervised classification to identify and quantify species mixed in silico and in defined communities in vitro. We identified species in a 5-species in silico community with an F1 score of 71%. In addition, we demonstrate in vitro that our method performs equally well or better than 16S rRNA gene sequencing in two-species cocultures and agrees with 16S rRNA gene sequencing data on the most abundant species in a four-species community. We found that shape and size differences alone are insufficient to distinguish species, and that it is thus necessary to exploit the multivariate nature of flow cytometry data. Finally, we observed that variability of flow cytometry data across replicates differs between gut bacterial species. In conclusion, the performance of supervised classification of gut species in flow cytometry data is species-dependent, but is for some combinations accurate enough to serve as a faster alternative to 16S rRNA gene sequencing.

The increasing amount of available high-content data in genomics, proteomics, and metabolomics has significantly improved the predictive power and model accuracy of genome-scale metabolic network models in recent years. We review recent constraint-based modeling approaches that incorporate genomics and proteomics data to form resource allocation models. Different modeling approaches to build resource allocation models and the related enzyme-constrained genome-scale metabolic models are discussed and evaluated with respect to differences regarding model features. In addition, an overview of the data required to construct, simulate and validate models for the different approaches is given, together with a list of relevant databases.

Temperate bacterial viruses are commonly thought to favor vertical (lysogenic) transmission over horizontal (lytic) transmission when the virion-to-host-cell ratio is high and available host cells become scarce. In P22-infected Salmonella Typhimurium populations, however, we find that host subpopulations become lytically consumed despite high phage-to-host ratios that would normally favor lysogeny. These subpopulations originate from the proliferation of P22-free siblings that spawn off from P22-carrier cells from which they cytoplasmically inherit P22-borne superinfection exclusion factors (SEFs). In fact, we demonstrate that the gradual dilution of these SEFs in the growing subpopulation of P22-free siblings restricts the number of incoming phages, thereby imposing the perception of a low phage-to-host ratio that favors lytic development. Although their role has so far been neglected, our data indicate that phage-borne SEFs can spur complex infection dynamics and a history-dependent switch from vertical to horizontal transmission in the face of host-cell scarcity. ispartof: Cell Reports vol:39 issue:6 ispartof: location:United States status: published

A thermophilic chain elongating bacterium, strain MDTJ8, was isolated from a thermophilic acidogenic anaerobic digestor producing n-caproate from human waste, growing optimally at 50-55 °C and pH 6.5. 16S rRNA gene analysis suggests that MDTJ8 represents a new species/genus within a group currently composed of mesophilic chain elongators of the Oscillospiraceae family. Genome analysis showed that strain MDTJ8 contains homologues of genes encoding for chain elongation and energy conservation but also indicated n-caproate production from carbohydrates including polymeric substances. This was confirmed by culturing experiments in which MDTJ8 converted, at pH 6.5 and 55 °C, mono-, di- and polymeric carbohydrates (starch and hemicellulose) to n-caproate reaching concentrations up to 283 mg/L and accounting for up to 10 % of the measured fermentation products. MDTJ8 is the first axenic organism that thermophilically performs chain elongation, opening doors to understand and intensify thermophilic bioprocesses targeting anaerobic digestion towards the production of the value-added chemical n-caproate.

Practical laboratory courses are an essential part of chemistry education. However, they can be costly and time-consuming. They also require physical presence of the teacher and students and access to well-equipped laboratories, which can be hindered due to equipment cost or a pandemic lockdown. Virtual chemical laboratories are digital tools that become very useful in these situations, but what research has been done on these tools and what elements are important in terms of technology and instructional design? This systematic literature review presents an extensive overview of previous research and addresses the different types of technology and instructional design elements. Results of this study show that virtual labs can be more effective than passive teaching methods (e.g., lecture, text and video), but show equal or greater effectiveness compared to hands-on laboratory. Better results are shown when virtual labs and traditional methods are combined. Most of the included studies use 3D Desktop technology, while immersive VR technology is trending in the last few years. This review also identified instructional design elements used in context of virtual chemical labs, for example, inquiry-based learning, modality, instructional scaffolding. Virtual laboratories can be used as an effective complementary tool or temperate alternative to real hands-on laboratory, but future research should put more emphasis in investigating skill-based learning outcomes using immersive VR and NUI technologies and in considering instructional design in virtual chemical laboratories. keywords: Virtual laboratory, Chemistry, Distance education and online learning, Media in education, Virtual reality ispartof: Computers & Education Open vol:2 status: published

Virtual reality (VR) technologies are becoming more and more popular, not only as a gaming console, but also as a viable training tool. Especially for health and safety training programmes, VR can be very useful to train people in dangerous environments and situations without imposing real danger on them or others. Chemical laboratories are environments where risks of severe injury or even fatality are always present. For this, we developed a VR serious game, called VR LaboSafe Game, as a tool for laboratory safety training. However, designing a VR serious game is a challenging task. There are many factors to consider for an optimal game design. In this study, we discuss important design considerations and we present the game design of VR LaboSafe Game. Preliminary tests of an early version of the game show usability issues and minor discomforting symptoms for some participants. Nevertheless, participants do agree that VR LaboSafe Game is useful for learning laboratory safety; is more active and responsive in their learning process and makes safety training more engaging. ispartof: pages:23-33 ispartof: Games and Learning Alliance vol:1 pages:23-33 ispartof: 10th International Conference on Games and Learning Alliance location:Online date:1 Dec - 2 Dec 2021 status: published

Modulation of the commensal oral microbiota is a promising preventive or therapeutic strategy for oral health and can for instance be achieved by increasing the abundance and/or activity of certain species. This study evaluated whether 10 selected substrates could modulate in vitro multi-species oral biofilms towards a more health-associated state. These substrates were chosen based on the possibility that they could stimulate H2O2 production by certain commensal species and/or increase their abundance, as previously reported or as hypothesized based on known bacterial H2O2 pathways. Biofilms grown in presence of the substrates at a clinically relevant concentration of 1%(w/v) often showed increased abundances of commensal species and decreased abundances of periodontal pathogens. Furthermore, most biofilms also showed an altered metabolic profile. Effects on the expression of a selection of virulence genes were substrate-dependent, but often a decreased expression of certain genes could be observed. In conclusion, this study found that a selection of substrates chosen for their hypothesized beneficial effects on the commensal oral microbiota were able to modulate in vitro multi-species oral biofilms towards a more health-associated state. These modulatory effects were found to be substrate-dependent.

Previous research identified potential prebiotic substrates for oral health like the structural analogues N-acetyl-d-mannosamine (NADM) and N-acetyl-d-glucosamine (NADG). The main hypothesis of the current study was twofold. Firstly, it was hypothesized that the modulatory effects of NADM are not limited to changes in multi-species oral biofilm composition, but also include effects on metabolism, virulence, and inflammatory potential. Secondly, the presence and orientation of their N-acetyl group could play a role. Therefore, a comparison was made between the effects of NADM, NADG and d-(+)-mannose on multi-species oral biofilms. Besides a beneficial compositional shift, NADM-treated biofilms also showed an altered metabolism, a reduced virulence and a decreased inflammatory potential. At a substrate concentration of 1 M, these effects were pronounced for all biofilm aspects, whereas at ~ 0.05 M (1%(w/v)) only the effects on virulence were pronounced. When comparing between substrates, both the presence and orientation of the N-acetyl group played a role. However, this was generally only at 1 M and dependent on the biofilm aspect. Overall, NADM was found to have different effects at two concentrations that beneficially modulate in vitro multi-species oral biofilm composition, metabolism, virulence and inflammatory potential. The presence and orientation of the N-acetyl group influenced these effects.

This work highlights the issue of interference by growth media when measuring bacterial H2O2 production. H2O2 was shown to be stable in phosphate buffered saline (PBS) but not in growth media. The protocol used for evaluating the intrinsic capacity of oral streptococci to produce H2O2 was shown to be reliable. ispartof: JOURNAL OF MICROBIOLOGICAL METHODS vol:182 ispartof: location:Netherlands status: published

The development of viability quantitative PCR (v-qPCR) has allowed for a more accurate assessment of the viability of a microbial sample by limiting the amplification of DNA from dead cells. Although valuable, v-qPCR is not infallible. One of the most limiting factors for accurate live/dead distinction is the length of the qPCR amplicon used. However, no consensus or guidelines exist for selecting and designing amplicon lengths for optimal results. In this study, a wide range of incrementally increasing amplicon lengths (68 to 906 base pairs [bp]) was used on live and killed cells of nine bacterial species treated with a viability dye (propidium monoazide [PMA]). Increasing amplicon lengths up to approximately 200 bp resulted in increasing quantification cycle (Cq) differences between live and killed cells while maintaining a good qPCR efficiency. Longer amplicon lengths, up to approximately 400 bp, further increased the Cq difference but at the cost of qPCR efficiency. Above 400 bp, no valuable increase in Cq differences was observed. IMPORTANCE Viability quantitative PCR (v-qPCR) has evolved into a valuable, mainstream technique for determining the number of viable microorganisms in samples by qPCR. Amplicon length is known to be positively correlated with the ability to distinguish between live and dead bacteria but is negatively correlated with qPCR efficiency. This trade-off is often not taken into account and might have an impact on the accuracy of v-qPCR data. Currently, there is no consensus on the optimal amplicon length. This paper provides methods to determine the optimal amplicon length and suggests an amplicon length range for optimal v-qPCR, taking into consideration the trade-off between qPCR efficiency and live/dead distinction. ispartof: APPLIED AND ENVIRONMENTAL MICROBIOLOGY vol:87 issue:5 ispartof: location:United States status: published

This work highlights the issue of interference by growth media when measuring bacterial H

Background: Modulation of the commensal oral microbiota constitutes a promising preventive/therapeutic approach in oral healthcare. The use of prebiotics for maintaining/restoring the health-associated homeostasis of the oral microbiota has become an important research topic. Aims: This study hypothesised that in vitro 14-species oral biofilms can be modulated by (in)direct stimulation of beneficial/commensal bacteria with new potential prebiotic substrates tested at 1 M and 1%((w/v)), resulting in more host-compatible biofilms with fewer pathogens, decreased virulence and less inflammatory potential. Methods: Established biofilms were repeatedly rinsed with N-acetyl-D-glucosamine, alpha-D-lactose, D-(+)-trehalose or D-(+)-raffinose at 1 M or 1%((w/v)). Biofilm composition, metabolic profile, virulence and inflammatory potential were eventually determined. Results: Repeated rinsing caused a shift towards a more health-associated microbiological composition, an altered metabolic profile, often downregulated virulence gene expression and decreased the inflammatory potential on oral keratinocytes. At 1 M, the substrates had pronounced effects on all biofilm aspects, whereas at 1%((w/v)) they had a pronounced effect on virulence gene expression and a limited effect on inflammatory potential. Conclusion: Overall, this study identified four new potential prebiotic substrates that exhibit different modulatory effects at two different concentrations that cause in vitro multi-species oral biofilms to become more host-compatible.

In the past two decades, metabolomics has proved to be a valuable tool with many potential applications in different areas of science. However, there are still some challenges that need to be addressed, particularly for multicenter studies. These challenges are mainly attributed to various sources of fluctuation and unwanted variations that can be introduced at pre-analytical, analytical, and/or post-analytical steps of any metabolomics experiment. Thus, this study aimed at using Streptomyces lividans TK24 as the model organism in a cross-laboratory experiment in Manchester and Leuven to evaluate the reproducibility of a standard sample preparation method, and determine the optimal sample format (cell extract or quenched biomass) required to preserve the metabolic profile of the cells during cross-lab sample transportation and storage. Principal component analysis (PCA) scores plot of the gas chromatography-mass spectrometry (GC-MS) data from both laboratories displayed clear growth-dependent clustering patterns which was in agreement with the Procrustes analysis findings. In addition, the data generated in Manchester displayed tight clustering of cell pellets (quenched biomass) and metabolite extracts, confirming the stability of both sample formats during the transportation and storage period. ispartof: METABOLITES vol:10 issue:9 ispartof: location:Switzerland status: published

Oral biofilms form on all hard and soft surfaces of the oral cavity. When the microbial balance in this biofilm is disturbed, pathogens can take the overhand, and this can lead to periodontitis (i.e. a chronic and inflammatory disease of the gum and tooth supporting tissues). In this work, a dual species community model containing one commensal bacterium and the most common periopathogen, is presented. The commensal bacterium is Streptococcus gordonii, the periopathogen is Porphyromonas gingivalis. Existing Genome-Scale Metabolic Models (GSMNs) are curated and transformed into planktonic Dynamic Flux Balance Analysis (dFBA) models in DFBAlab (Gomez et al., 2014). In the planktonic model for S. gordonii, split ratio’s are used to ensure correct flux distributions between the glycolysis and the pentose phosphate pathway and around the pyruvate node. The split ratios are required to simulate the suboptimal growth behaviour of this bacterium. Simulation results for the planktonic S. gordonii model are compared to experiments for pure cultures. The planktonic P. gingivalis model gives feasible results for biomass growth and nutrient uptake. Finally, both planktonic models are transformed into a biofilm model in DFBAlab by introduction of nutrient gradients over the depth of the biofilm. This first oral biofilm model predicted the partitioning of both bacteria in the biofilm, similar to what has been described in literature.

For the production of edible microbial protein (MP), ammonia generated by the Haber-Bosch process or reclaimed ammonia from waste streams is typically considered as the nitrogen source. These processes for ammonia production are highly energy intensive. In this study, the potential for using nitrogen gas (N2) as a direct nitrogen source for MP production by hydrogen-oxidizing bacteria (HOB) was evaluated. The use of N2 versus ammonium as nitrogen source during the enrichment process resulted in differentiation of the bacterial community composition of the enrichments. A few previously unknown potential N2-fixing HOB taxa (i.e., representatives of the genus Azonexus and the family Comamonadaceae) dominated the enrichments. The biomass yield of a N2-fixing HOB enrichment was 30-50% lower than that of the ammonium-based HOB enrichment from the same inoculum source. The dried biomass of N2-fixing HOB had a high protein content (62.0 ± 6.3%) and an essential amino acid profile comparable to MP from ammonium-based HOB. MP from N2-fixing HOB could potentially be produced in situ without entailing the emissions caused by ammonia production and transportation by conventional means. It could be a promising substitute for N2-fixing protein-rich soybean because it has 70% higher protein content and double energy conversion efficiency from solar energy to biomass.

Background and Objective Little is known about the initiation of dysbiosis in oral biofilms, a topic of prime importance for understanding the etiology of, and preventing, periodontitis. The aim of this study was to evaluate the effect of different concentrations of crevicular and salivary peroxidase and catalase on dysbiosis in multispecies biofilms in vitro. Material and Methods The spotting technique was used to identify the effect of different concentrations of myeloperoxidase, lactoperoxidase, erythrocyte catalase, and horseradish peroxidase in salivary and crevicular fluid on the inhibitory effect of commensals on pathobiont growth. Vitality–quantitative real-time PCR was performed to quantify the dysbiotic effect of the peroxidases (adjusted to concentrations found in periodontal health, gingivitis, and periodontitis) on multispecies microbial communities. Results Agar plate and multispecies ecology experiments showed that production of hydrogen peroxide (H2O2) by commensal bacteria decreases pathobiont growth and colonization. Peroxidases at concentrations found in crevicular fluid and saliva neutralized this inhibitory effect. In multispecies communities, myeloperoxidase, at the crevicular fluid concentrations found in periodontitis, resulted in a 1-3 Log increase in pathobionts when compared with the crevicular fluid concentrations found in periodontal health. The effect of salivary lactoperoxidase and salivary myeloperoxidase concentrations was, in general, similar to the effect of crevicular myeloperoxidase concentrations. Conclusions Commensal species suppress pathobionts by producing H2O2. Catalase and peroxidases, at clinically relevant concentrations, can neutralize this effect and thereby can contribute to dysbiosis by allowing the outgrowth of pathobionts.