Methylococcus Capsulatus Research Articles

Online monitoring of methane transfer rates unveils nitrogen fixation dynamics in Methylococcus capsulatus.

This study explores methane utilization by the methanotrophic microorganism Methylococcus capsulatus (Bath) for biomass production, presenting a promising approach to mitigate methane emissions and foster the development sustainable biomaterials. Traditional screening methods for gas cultivations involve either serum flasks without online monitoring or costly, low-throughput fermenters. To address these limitations, the Respiration Activity MOnitoring System was augmented with methane sensors for real-time methane transfer rate (MTR) monitoring in shake flasks. Utilizing online monitoring of the MTR in shake flasks results in enhanced throughput and cost-effectiveness for cultivating M. capsulatus. Simultaneous monitoring of transfer rates for oxygen, methane, and carbon dioxide was conducted in up to eight shake flasks, ensuring the success of the cultivation process. Alterations in methane-to-oxygen transfer rate ratios and carbon fixation rates reveal the impact of transfer limitations on microbial growth. Detection of gas transfer limitations, exploration of process parameter influences, and investigations of medium components were enabled by the introduced method. Optimal nitrogen concentrations could be determined to ensure optimal growth. This streamlined approach accelerates the screening process, offering efficient investigations into metabolic effects, limitations, and parameter influences in gas fermentations without the need for elaborate offline sampling, significantly reducing costs and enhanced reproducibility.

Overexpression of native carbonic anhydrases increases carbon conversion efficiency in the methanotrophic biocatalyst Methylococcus capsulatus Bath.

Methanotrophic bacteria play a vital role in the biogeochemical carbon cycle due to their unique ability to use CH4 as a carbon and energy source. Evidence suggests that some methanotrophs, including Methylococcus capsulatus, can also use CO2 as a carbon source, making these bacteria promising candidates for developing biotechnologies targeting greenhouse gas capture and mitigation. However, a deeper understanding of the dual CH4 and CO2 metabolism is needed to guide methanotroph strain improvements and realize their industrial utility. In this study, we show that M. capsulatus expresses five carbonic anhydrase (CA) isoforms, one α-CA, one γ-CA, and three β-CAs, that play a role in its inorganic carbon metabolism and CO2-dependent growth. The CA isoforms are differentially expressed, and transcription of all isoform genes is induced in response to CO2 limitation. CA null mutant strains exhibited markedly impaired growth compared to an isogenic wild-type control, suggesting that the CA isoforms have independent, non-redundant roles in M. capsulatus metabolism and physiology. Overexpression of some, but not all, CA isoforms improved bacterial growth kinetics and decreased CO2 evolution from CH4-consuming cultures. Notably, we developed an engineered methanotrophic biocatalyst overexpressing the native α-CA and β-CA with a 2.5-fold improvement in the conversion of CH4 to biomass. Given that product yield is a significant cost driver of methanotroph-based bioprocesses, the engineered strain developed here could improve the economics of CH4 biocatalysis, including the production of single-cell protein from natural gas or anaerobic digestion-derived biogas.IMPORTANCEMethanotrophs transform CH4 into CO2 and multi-carbon compounds, so they play a critical role in the global carbon cycle and are of interest for biotechnology applications. Some methanotrophs, including Methylococcus capsulatus, can also use CO2 as a carbon source, but this dual one-carbon metabolism is incompletely understood. In this study, we show that M. capsulatus carbonic anhydrases are critical for this bacterium to optimally utilize CO2. We developed an engineered strain with improved CO2 utilization capacity that increased the overall carbon conversion to cell biomass. The improvements to methanotroph-based product yields observed here are expected to reduce costs associated with CH4 conversion bioprocesses.

Comparative metagenomic analysis from Sundarbans ecosystems advances our understanding of microbial communities and their functional roles

The Sundarbans mangrove, located at the mouth of the Ganges and Brahmaputra Rivers, is the world’s largest tidal mangrove forest. These mangroves are also one of the most striking sources of microbial diversity, essential in productivity, conservation, nutrient cycling, and rehabilitation. Hence, the main objective of this study was to use metagenome analysis and provide detailed insight into microbial communities and their functional roles in the Sundarbans mangrove ecosystem. A comparative analysis was also done with a non-mangrove region of the Sundarbans ecosystem to assess the capability of the environmental parameters to explain the variation in microbial community composition. The study found several dominant bacteria, viz., Alphaproteobacteria, Actinomycetota, Bacilli, Clostridia, Desulfobacterota, Gammaproteobacteria, and Nitrospira, from the mangrove region. The mangrove sampling site reports several salt-tolerant bacteria like Alkalibacillus haloalkaliphilus, Halomonas anticariensis, and Salinivibrio socompensis. We found some probiotic species, viz., Bacillus clausii, Lactobacillus curvatus, Vibrio mediterranei and Vibrio fluvialis, from the Sundarbans mangrove. Nitrifying bacteria in Sundarbans soils were Nitrococcus mobilis, Nitrosococcus oceani, Nitrosomonas halophila, Nitrospirade fluvii, and others. Methanogenic archaea, viz., Methanoculleus marisnigri, Methanobrevibacter gottschalkii, and Methanolacinia petrolearia, were highly abundant in the mangroves as compared to the non-mangrove soils. The identified methanotrophic bacterial species, viz., Methylobacter tundripaludum, Methylococcus capsulatus, Methylophaga thiooxydans, and Methylosarcina lacus are expected to play a significant role in the degradation of methane in mangrove soil. Among the bioremediation bacterial species identified, Pseudomonas alcaligenes, Pseudomonas mendocina, Paracoccus denitrificans, and Shewanella putrefaciens play a significant role in the remediation of environmental pollution. Overall, our study shows for the first time that the Sundarbans, the largest mangrove ecosystem in the world, has a wide range of methanogenic archaea, methanotrophs, pathogenic, salt-tolerant, probiotic, nitrifying, and bioremediation bacteria.

Conserved C-Terminal Tail Is Responsible for Membrane Localization and Function of Pseudomonas aeruginosa Hemerythrin.

Many bacteria have hemerythrin (Hr) proteins that bind O2, including Pseudomonas aeruginosa, in which microoxia-induced Hr (Mhr) provide fitness advantages under microoxic conditions. Mhr has a 23 amino-acid extension at its C-terminus relative to a well-characterized Hr from Methylococcus capsulatus, and similar extensions are also found in Hrs from other bacteria. The last 11 amino acids of this extended, C-terminal tail are highly conserved in gammaproteobacteria and predicted to form a helix with positively charged and hydrophobic faces. In cellular fractionation assays, wild-type (WT) Mhr was found in both membrane and cytosolic fractions, while a MhrW143* variant lacking the last 11 residues was largely in the cytosol and did not complement Mhr function in competition assays. MhrL112Y, a variant that has a much longer-lived O2-bound form, was fully functional and had a similar localization pattern to that of WT Mhr. Both MhrW143* and MhrL112Y had secondary structures, stabilities, and O2-binding kinetics similar to those of WT Mhr. Fluorescence studies revealed that the C-terminal tail, and particularly the fragment corresponding to its last 11 residues, was sufficient and necessary for association with lipid vesicles. Molecular dynamics simulations and subsequent cellular analysis of Mhr variants have demonstrated that conserved, positively charged residues in the tail are important for Mhr interactions with negatively charged membranes and the contribution of this protein to competitive fitness. Together, these data suggest that peripheral interactions of Mhr with membranes are guided by the C-terminal tail and are independent of O2-binding.

Replacement of fishmeal with a microbial single-cell protein induced enteropathy and poor growth outcomes in barramundi (Lates calcarifer) fry.

Fish meal (FM) replacement is essential for the sustainable expansion of aquaculture. This study focussed on the feasibility of replacing FM with a single-cell protein (SCP) derived from methanotrophic bacteria (Methylococcus capsulatus, Bath) in barramundi fry (Lates calcarifer). Three isonitrogenous and isoenergetic diets were formulated with 0%, 6.4% and 12.9% inclusion of the SCP, replacing FM by 0%, 25% and 50%. Barramundi fry (initial body weight 2.5 ± 0.1 g) were fed experimental diets for 21 days to assess growth performance, gut microbiome composition and gut histopathology. Our findings revealed that both levels of SCP inclusion induced detrimental effects in barramundi fry, including impaired growth and reduced survival compared with the control group (66.7% and 71.7% survival in diets replacing FM with SCP by 25% and 50%, respectively; p < .05). Both dietary treatments presented mild necrotizing enteritis with subepithelial oedema and accumulation of PAS positive, diastase resistant droplets within hepatocytes (ceroid hepatopathy) and pancreatic atrophy. Microbiome analysis revealed a marked shift in the gut microbial community with the expansion of potential opportunistic bacteria in the genus Aeromonas. Reduced overall performance in the highest inclusion level (50% SCP) was primarily associated with reduced feed intake, likely related to palatability issues, albeit pathological changes observed in gut and liver may also play a role. Our study highlights the importance of meticulous optimization of SCP inclusion levels in aquafeed formulations, and the need for species and life-stage specific assessments to ensure the health and welfare of fish in sustainable aquaculture practices.

New Solutions in Single-Cell Protein Production from Methane: Construction of Glycogen-Deficient Mutants of Methylococcus capsulatus MIR

The biotechnology of converting methane to single-cell protein (SCP) implies using fast-growing thermotolerant aerobic methanotrophic bacteria. Among the latter, members of the genus Methylococcus received significant research attention and are used in operating commercial plants. Methylococcus capsulatus MIR is a recently discovered member of this genus with the potential to be used for the purpose of SCP production. Like other Methylococcus species, this bacterium stores carbon and energy in the form of glycogen, particularly when grown under nitrogen-limiting conditions. The genome of strain MIR encodes two glycogen synthases, GlgA1 and GlgA2, which are only moderately related to each other. To obtain glycogen-free cell biomass of this methanotroph, glycogen synthase mutants, ΔglgA1, ΔglgA2, and ΔglgA1ΔglgA2, were constructed. The mutant lacking both glycogen synthases exhibited a glycogen-deficient phenotype, whereas the intracellular glycogen content was not reduced in strains defective in either GlgA1 or GlgA2, thus suggesting functional redundancy of these enzymes. Inactivation of the glk gene encoding glucokinase also resulted in a sharp decrease in glycogen content and accumulation of free glucose in cells. Wild-type strain MIR and the mutant strain ΔglgA1ΔglgA2 were also grown in a bioreactor operated in batch and continuous modes. Cell biomass of ΔglgA1ΔglgA2 mutant obtained during batch cultivation displayed high protein content (71% of dry cell weight (DCW) compared to 54% DCW in wild-type strain) as well as a strong reduction in glycogen content (10.8 mg/g DCW compared to 187.5 mg/g DCW in wild-type strain). The difference in protein and glycogen contents in biomass of these strains produced during continuous cultivation was less pronounced, yet biomass characteristics relevant to SCP production were slightly better for ΔglgA1ΔglgA2 mutant. Genome analysis revealed the presence of glgA1-like genes in all methanotrophs of the Gammaproteobacteria and Verrucomicrobia, while only a very few methanotrophic representatives of the Alphaproteobacteria possessed these determinants of glycogen biosynthesis. The glgA2-like genes were present only in genomes of gammaproteobacterial methanotrophs with predominantly halo- and thermotolerant phenotypes. The role of glycogen in terms of energy reserve is discussed.

Evaluation of fish meal replacement by Methylcoccus capsulatus protein in diets for juvenile Chinese softshell turtle (Pelodiscus sinensis)

The present study was to investigate the effects of Methylococcus capsulatus protein (MCP) replacing fish meal on the growth performance, immunity, and intestinal health of Chinese soft-shelled turtle (Pelodiscus sinensis). Four isonitrogenous (47.00% crude protein) and isolipidic (6.00% crude lipid) diets were formulated for an 8-week feeding trial. The fish meal group (FM) contained 55% fish meal. MCP was used to replace 10%, 15%, and 20% of the fish meal in the FM group to create MCP10, MCP15, and MCP20 treatment groups. No significant differences were observed in the weight gain, feed conversion efficiency, physical indicators, or muscle amino acid composition among all diet treatments (P > 0.05). The MCP15 group showed the best growth performance. There were no significant differences in serum biochemistry among the groups (P > 0.05), except for aspartate aminotransferase, which exhibited a significant linear increase trend with the increase of MCP replacement level (P = 0.049). In the MCP20 group, the complement C3 content, immunoglobulin M content, and lysozyme activity in serum were significantly lower than those in the FM, MCP10, and MCP15 groups (P < 0.05), showing a quadratic relationship characterized by an initial increase followed by a decrease as the MCP level in the feed increased (P < 0.05). The intestinal morphology in all treatment groups appeared normal and the expression levels of intestinal inflammatory cytokines showed no significant differences (P > 0.05). The community structure of the intestinal microbiota, as well as the phylum and genus composition underwent significant alterations in response to the MCP supplementation. The recommended practice for Chinese softshell turtle diet involves substituting 15% MCP for fish meal, which enhances growth, boosts immune response, and does not adversely affect serum biochemistry, amino acid composition, or intestinal health.

Novel sterol binding domains in bacteria.

Sterol lipids are widely present in eukaryotes and play essential roles in signaling and modulating membrane fluidity. Although rare, some bacteria also produce sterols, but their function in bacteria is not known. Moreover, many more species, including pathogens and commensal microbes, acquire or modify sterols from eukaryotic hosts through poorly understood molecular mechanisms. The aerobic methanotroph Methylococcus capsulatus was the first bacterium shown to synthesize sterols, producing a mixture of C-4 methylated sterols that are distinct from those observed in eukaryotes. C-4 methylated sterols are synthesized in the cytosol and localized to the outer membrane, suggesting that a bacterial sterol transport machinery exists. Until now, the identity of such machinery remained a mystery. In this study, we identified three novel proteins that may be the first examples of transporters for bacterial sterol lipids. The proteins, which all belong to well-studied families of bacterial metabolite transporters, are predicted to reside in the inner membrane, periplasm, and outer membrane of M. capsulatus, and may work as a conduit to move modified sterols to the outer membrane. Quantitative analysis of ligand binding revealed their remarkable specificity for 4-methylsterols, and crystallographic structures coupled with docking and molecular dynamics simulations revealed the structural bases for substrate binding by two of the putative transporters. Their striking structural divergence from eukaryotic sterol transporters signals that they form a distinct sterol transport system within the bacterial domain. Finally, bioinformatics revealed the widespread presence of similar transporters in bacterial genomes, including in some pathogens that use host sterol lipids to construct their cell envelopes. The unique folds of these bacterial sterol binding proteins should now guide the discovery of other proteins that handle this essential metabolite.

Novel sterol binding domains in bacteria

Sterol lipids are widely present in eukaryotes and play essential roles in signaling and modulating membrane fluidity. Although rare, some bacteria also produce sterols, but their function in bacteria is not known. Moreover, many more species, including pathogens and commensal microbes, acquire or modify sterols from eukaryotic hosts through poorly understood molecular mechanisms. The aerobic methanotroph Methylococcus capsulatus was the first bacterium shown to synthesize sterols, producing a mixture of C-4 methylated sterols that are distinct from those observed in eukaryotes. C-4 methylated sterols are synthesized in the cytosol and localized to the outer membrane, suggesting that a bacterial sterol transport machinery exists. Until now, the identity of such machinery remained a mystery. In this study, we identified three novel proteins that may be the first examples of transporters for bacterial sterol lipids. The proteins, which all belong to well-studied families of bacterial metabolite transporters, are predicted to reside in the inner membrane, periplasm, and outer membrane of M. capsulatus, and may work as a conduit to move modified sterols to the outer membrane. Quantitative analysis of ligand binding revealed their remarkable specificity for 4-methylsterols, and crystallographic structures coupled with docking and molecular dynamics simulations revealed the structural bases for substrate binding by two of the putative transporters. Their striking structural divergence from eukaryotic sterol transporters signals that they form a distinct sterol transport system within the bacterial domain. Finally, bioinformatics revealed the widespread presence of similar transporters in bacterial genomes, including in some pathogens that use host sterol lipids to construct their cell envelopes. The unique folds of these bacterial sterol binding proteins should now guide the discovery of other proteins that handle this essential metabolite.

Complete genome sequences of Methylococcus capsulatus (Norfolk) and Methylocaldum szegediense (Norfolk) isolated from a landfill methane biofilter.

Here we report the complete genome sequence of two moderately thermophilic methanotrophs isolated from a landfill methane biofilter, Methylococcus capsulatus (Norfolk) and Methylocaldum szegediense (Norfolk).

Effect of the hps/phi Copy Number on the Growth of Methylococcus capsulatus MIR

Effect of the hps/phi Copy Number on the Growth of Methylococcus capsulatus MIR

Engineered Methylococcus capsulatus Bath for efficient methane conversion to isoprene

Isoprene has numerous industrial applications, including rubber polymer and potential biofuel. Microbial methane-based isoprene production could be a cost-effective and environmentally benign process, owing to a reduced carbon footprint and economical utilization of methane. In this study, Methylococcus capsulatus Bath was engineered to produce isoprene from methane by introducing the exogenous mevalonate (MVA) pathway. Overexpression of MVA pathway enzymes and isoprene synthase from Populus trichocarpa under the control of a phenol-inducible promoter substantially improved isoprene production. M. capsulatus Bath was further engineered using a CRISPR-base editor to disrupt the expression of soluble methane monooxygenase (sMMO), which oxidizes isoprene to cause toxicity. Additionally, optimization of the metabolic flux in the MVA pathway and culture conditions increased isoprene production to 228.1 mg/L, the highest known titer for methanotroph-based isoprene production. The developed methanotroph could facilitate the efficient conversion of methane to isoprene, resulting in the sustainable production of value-added chemicals.

Effect of different types of bacterial single cell protein on feed intake, digestibility, growth and body composition of Pacific white shrimp (Penaeus vannamei)

This study assessed the potential of four bacterial (Methylococcus capsulatus) single cell protein (SCP) products as alternative protein sources for Pacific white shrimp (Penaeus vannamei) diets. A growth trial and a digestibility trial were undertaken, during which the bacterial SCP products were compared with a high-quality fishmeal and a soy protein concentrate, regarding their impact on ingredient digestibility, growth, feed intake and whole-body composition of juvenile P. vannamei. Seven diets were formulated; one reference diet (REF) and six test diets. The test diets consisted of 85% of the REF diet and 15% of a test ingredient. Ingredients tested were four bacterial SCP products (SCP1–4), which differed in processing conditions, fishmeal (FM) and soy protein concentrate (SoyProt). Growth and feed utilization were similar for P. vannamei fed either the FM diet or one of the bacterial SCP diets, whilst lowest growth and feed utilization were observed for shrimp fed the SoyProt diet. Final whole-body protein content did not differ between shrimp fed the FM diet or one of the four bacterial SCP diets. However, shrimp fed the SCP diets had a significantly higher final phosphorus body content and a higher phosphorus retention than shrimp fed the FM or SoyProt diets. This indicates a higher phosphorus availability in the bacterial SCP products compared to FM and SoyProt. Protein digestibility of the SCP products was similar to FM, whilst amino acid (AA) digestibility was comparable to FM for three of the four SCP products (SCP1, SCP2 and SCP4). The SCP3 product showed the lowest digestibility for most AA, indicating a possible influence of processing conditions on AA availability of bacterial SCPs. Overall, this study highlights that bacterial SCP originating from M. capsulatus is a viable alternative protein source for Pacific white shrimp diets, but processing conditions should be taken into account.

Transcriptional and metabolomic responses of Methylococcus capsulatus Bath to nitrogen source and temperature downshift

Methanotrophs play a significant role in methane oxidation, because they are the only biological methane sink present in nature. The methane monooxygenase enzyme oxidizes methane or ammonia into methanol or hydroxylamine, respectively. While much is known about central carbon metabolism in methanotrophs, far less is known about nitrogen metabolism. In this study, we investigated how Methylococcus capsulatus Bath, a methane-oxidizing bacterium, responds to nitrogen source and temperature. Batch culture experiments were conducted using nitrate or ammonium as nitrogen sources at both 37°C and 42°C. While growth rates with nitrate and ammonium were comparable at 42°C, a significant growth advantage was observed with ammonium at 37°C. Utilization of nitrate was higher at 42°C than at 37°C, especially in the first 24 h. Use of ammonium remained constant between 42°C and 37°C; however, nitrite buildup and conversion to ammonia were found to be temperature-dependent processes. We performed RNA-seq to understand the underlying molecular mechanisms, and the results revealed complex transcriptional changes in response to varying conditions. Different gene expression patterns connected to respiration, nitrate and ammonia metabolism, methane oxidation, and amino acid biosynthesis were identified using gene ontology analysis. Notably, key pathways with variable expression profiles included oxidative phosphorylation and methane and methanol oxidation. Additionally, there were transcription levels that varied for genes related to nitrogen metabolism, particularly for ammonia oxidation, nitrate reduction, and transporters. Quantitative PCR was used to validate these transcriptional changes. Analyses of intracellular metabolites revealed changes in fatty acids, amino acids, central carbon intermediates, and nitrogen bases in response to various nitrogen sources and temperatures. Overall, our results offer improved understanding of the intricate interactions between nitrogen availability, temperature, and gene expression in M. capsulatus Bath. This study enhances our understanding of microbial adaptation strategies, offering potential applications in biotechnological and environmental contexts.

P15-01: Toxicological examination of a novel food protein, derived from Methylococcus capsulatus biomass

P15-01: Toxicological examination of a novel food protein, derived from Methylococcus capsulatus biomass

Production of Bacillus subtillis protein mass on the microbial mass of Methylococcus capsulatus

The growing demand for food of animal origin has contributed to the annual increase in the agricultural population of productive animals in the world, which in turn has led to a significant shortage of feeds and an increase in their cost. Therefore, the purpose of this research was to explore the qualitative composition of feed products obtained by culturing methanotrophic microorganisms Methylococcus capsulatus with probiotic bacteria Bacillus subtillis. Arbitrage methods for determining the quality of feeds, which are regulated by regulatory documents (ISO), were used in the research. The results of the research demonstrate that the protein product obtained by growing Methylococcus capsulatus culture has a high content of “crude” protein at the level of 60.9%, which corresponds to high-protein feeds of animal origin by its technical characteristics. The addition of probiotic microorganisms Bacillus subtillis to the medium for cultivation significantly improved the qualitative parameters of the synthesised protein by increasing by 1.5% the amount of essential amino acids, increasing by 3.8% fatty acids and increasing the concentration of calcium and phosphorus. The increase in the concentration of lysine, isoleucine, valine and asparagic acid, due to the cocultivation of two microorganisms allowed increasing their nutritional value for cattle since these amino acids are the most demanded in ruminant diets when using corn silage and other plant feeds. The addition of Bacillus subtillis culture allowed reducing the content of “crude” fibre and improving its biological characteristics by changing the microbiological composition of the obtained product. In the future, the biotechnological method of obtaining feed protein for animals will reduce the dependence of animal breeding on the cultivation of fodder crops and weather conditions

The Role of Serine-Glyoxylate Aminotransferase and Malyl-CoA Lyase in the Metabolism of Methylococcus capsulatus Bath.

The genome of aerobic methanotroph Methylococcus capsulatus Bath possesses genes of three biochemical pathways of C1-carbon assimilation: the ribulose monophosphate cycle, the Calvin-Benson-Bassham cycle, and the partial serine cycle. Numerous studies have demonstrated that during methanotrophic growth cells of Methylococcus capsulatus Bath express key enzymes of these routes. In this study, the role of the serine cycle key enzymes, serine-glyoxylate aminotransferase (Sga) and malyl-CoA lyase (Mcl) in metabolism of Methylococcus capsulatus Bath was investigated by gene inactivation. The Δsga mutant obtained by double homologous recombination showed a prolonged lag phase, and after the lag period, the growth rate became similar to that of the wild type strain. The elevated intracellular levels of glutamate, serine, glycine, alanine, methionine, leucine, and succinate suggested significant metabolic changes in the mutant cells. Deletion of the mcl gene resulted in very poor growth and glycine only partially improved growth of the mutant strain. Cells of Δmcl mutant possess lower content of histidine, but enhanced level of alanine, leucine, and lysine than those of the wild type strain. Our data imply the importance of the serine cycle enzymes in metabolism of the methanotroph as well as relationships of the three C1 assimilation pathways in the gammaproteobacterial methanotrophs.

Structures and Mechanisms of a Novel Bacterial Transport System for Fatty Acids.

Bacterial acquisition of metabolites is largely facilitated by transporters with unique substrate scopes. The tripartite ATP-independent periplasmic (TRAP) transporters comprise a large family of bacterial proteins that facilitate the uptake of a variety of small molecules. It has been reported that some TRAP systems encode a fourth protein, the T component. The T-component, or TatT, is predicted to be a periplasmic-facing lipoprotein that enables the uptake of metabolites from the outer membrane. However, no substrates were revealed for any TatT and their functional role(s) remained enigmatic. We recently identified a homolog in Methylococcus capsulatus that binds to sterols, and herein, we report two additional homologs that demonstrate a preference for long-chain fatty acids. Our bioinformatics, quantitative analyses of protein-ligand interactions, and high-resolution crystal structures suggest that TatTs might facilitate the trafficking of hydrophobic or lipophilic substrates and represent a new class of bacterial lipid and fatty acid transporters.

Methanotroph (Methylococcus capsulatus, Bath) bacteria meal alleviates soybean meal-induced enteritis in spotted seabass (Lateolabrax maculatus) by modulating immune responses and the intestinal flora

The effect of methanotroph (Methylococcus capsulatus, Bath) bacteria meal (FeedKind®, FK) supplementation in a high-soybean meal diet on growth, non-specific immunity, and gut health in spotted seabass (Lateolabrax maculatus) was investigated. Six experimental diets were formulated: control (containing 38% fish meal, FM, and 16.5% soybean meal, SBM), a high-soybean meal diet (containing 28% FM and 35% SBM), and four experimental diets each containing 28% FM and 35% SBM with the addition of FK at 2% (FK2), 4% (FK4), 6% (FK6) or 8% (FK8) in the diet to replace wheat gluten. All diets also contained 0.1% yttrium oxide to determine apparent digestibility coefficients (ADCs) of dry matter, crude protein and crude lipid. Following a 14-day acclimation period, fish were fed the diets for 56 days. Results showed that the FK2 and FK4 groups showed significantly higher weight gain (WG) compared to the SBM group (P < 0.05), whilst the FK2, FK6, and FK8 groups showed higher WG compared to the FM group (P < 0.05). The FK2, FK4, and FK8 had lower feed conversion rate (FCR) compared to the SBM group (P < 0.05). No differences were noted in overall survival. Intestinal lipase activity was significantly increased in the FK2 and FK4 groups and trypsin activity was significantly increased in the FK2 group compared to the SBM group (P < 0.05). The FK2 group showed significantly higher apparent digestibility coefficients of nutrients than the SBM group (P < 0.05). The expressions of anti-inflammatory genes (tgfβ, il-4, and il-10) in the intestine of the FK-supplemented group showed a significant increment (P < 0.05), whilst the expressions of pro-inflammatory genes (tnfα, il-1β, and il-8) significantly decreased (P < 0.05). The results of 16S rRNA gene sequencing showed that indexes of alpha diversity in the SBM group were decreased (P < 0.05) compared with all other diets. In addition, the FK2 group showed significantly higher serum acid phosphatase (ACP), alkaline phosphatase (AKP), complement 4 (C4), and total protein (TP), levels than the SBM group (P < 0.05). Inclusion of FeedKind in high SBM diets alleviated SBM-induced intestinal inflammation by modulating the expression of intestinal inflammatory factors and regulating intestinal flora. Overall, supplementation of SBM diets with 2%–4% FK improved growth, non-specific immunity, and ADC's in spotted seabass.

Adsorption and intracellular uptake of mercuric mercury and methylmercury by methanotrophs and methylating bacteria

The cell surface adsorption and intracellular uptake of mercuric mercury Hg(II) and methylmercury (MeHg) are important in determining the fate and transformation of Hg in the environment. However, current information is limited about their interactions with two important groups of microorganisms, i.e., methanotrophs and Hg(II)-methylating bacteria, in aquatic systems. This study investigated the adsorption and uptake dynamics of Hg(II) and MeHg by three strains of methanotrophs, Methylomonas sp. strain EFPC3, Methylosinus trichosporium OB3b, and Methylococcus capsulatus Bath, and two Hg(II)-methylating bacteria, Pseudodesulfovibrio mercurii ND132 and Geobacter sulfurreducens PCA. Distinctive behaviors of these microorganisms towards Hg(II) and MeHg adsorption and intracellular uptake were observed. The methanotrophs took up 55−80% of inorganic Hg(II) inside cells after 24 h incubation, lower than methylating bacteria (>90%). Approximately 80–95% of MeHg was rapidly taken up by all the tested methanotrophs within 24 h. In contrast, after the same time, G. sulfurreducens PCA adsorbed 70% but took up <20% of MeHg, while P. mercurii ND132 adsorbed <20% but took up negligible amounts of MeHg. These results suggest that microbial surface adsorption and intracellular uptake of Hg(II) and MeHg depend on the specific types of microbes and appear to be related to microbial physiology that requires further detailed investigation. Despite being incapable of methylating Hg(II), methanotrophs play important roles in immobilizing both Hg(II) and MeHg, potentially influencing their bioavailability and trophic transfer. Therefore, methanotrophs are not only important sinks for methane but also for Hg(II) and MeHg and can influence the global cycling of C and Hg.