Methane-utilizing Bacteria Research Articles

State-of the-Art Constraint-Based Modeling of Microbial Metabolism: From Basics to Context-Specific Models with a Focus on Methanotrophs.

Methanotrophy is the ability of an organism to capture and utilize the greenhouse gas, methane, as a source of energy-rich carbon. Over the years, significant progress has been made in understanding of mechanisms for methane utilization, mostly in bacterial systems, including the key metabolic pathways, regulation and the impact of various factors (iron, copper, calcium, lanthanum, and tungsten) on cell growth and methane bioconversion. The implementation of -omics approaches provided vast amount of heterogeneous data that require the adaptation or development of computational tools for a system-wide interrogative analysis of methanotrophy. The genome-scale mathematical modeling of its metabolism has been envisioned as one of the most productive strategies for the integration of muti-scale data to better understand methane metabolism and enable its biotechnological implementation. Herein, we provide an overview of various computational strategies implemented for methanotrophic systems. We highlight functional capabilities as well as limitations of the most popular web resources for the reconstruction, modification and optimization of the genome-scale metabolic models for methane-utilizing bacteria.

Identification of a TonB-Dependent Receptor Involved in Lanthanide Switch by the Characterization of Laboratory-Adapted Methylosinus trichosporium OB3b.

Two methanol dehydrogenases (MDHs), MxaFI and XoxF, have been characterized in methylotrophic and methanotrophic bacteria. MxaFI contains a calcium ion in its active site, whereas XoxF contains a lanthanide ion. Importantly, the expression of MxaFI and XoxF is inversely regulated by lanthanide bioavailability, i.e., the "lanthanide switch." To reveal the genetic and environmental factors affecting the lanthanide switch, we focused on two Methylosinus trichosporium OB3b mutants isolated during routine cultivation. In these mutants, MxaF was constitutively expressed, but lanthanide-dependent XoxF1 was not, even in the presence of 25 μM cerium ions, which is sufficient for XoxF expression in the wild type. Genotyping showed that both mutants harbored a loss-of-function mutation in the CQW49_RS02145 gene, which encodes a TonB-dependent receptor. Gene disruption and complementation experiments demonstrated that CQW49_RS02145 was required for XoxF1 expression in the presence of 25 μM cerium ions. Phylogenetic analysis indicated that CQW49_RS02145 was homologous to the Methylorubrum extorquens AM1 lanthanide transporter gene (lutH). These findings suggest that CQW49_RS02145 is involved in lanthanide uptake across the outer membrane. Furthermore, we demonstrated that supplementation with cerium and glycerol caused severe growth arrest in the wild type. CQW49_RS02145 underwent adaptive laboratory evolution in the presence of cerium and glycerol ions, resulting in a mutation that partially mitigated the growth arrest. This finding implies that loss-of-function mutations in CQW49_RS02145 can be attributed to residual glycerol from the frozen stock. IMPORTANCE Lanthanides are widely used in many industrial applications, including catalysts, magnets, and polishing. Recently, lanthanide-dependent metabolism was characterized in methane-utilizing bacteria. Despite the global demand for lanthanides, few studies have investigated the mechanism of lanthanide uptake by these bacteria. In this study, we identify a lanthanide transporter in Methylosinus trichosporium OB3b and indicate the potential interaction between intracellular lanthanide and glycerol. Understanding the genetic and environmental factors affecting lanthanide uptake should not only help improve the use of lanthanides for the bioconversion of methane into valuable products like methanol but also be of value for developing biomining to extract lanthanides under neutral conditions.

Methane monooxygenases; physiology, biochemistry and structure

Methane monooxygenases (MMOs) are potential biocatalysts for industrial methanol production from methane. We summarize recent structural and biochemical studies of pMMO and discuss the future directions to develop efficient and robust biocatalysts.

Prospecting the significance of methane-utilizing bacteria in agriculture.

Microorganisms act as both the source and sink of methane, a potent greenhouse gas, thus making a significant contribution to the environment as an important driver of climate change. The rhizosphere and phyllosphere of plants growing in natural (mangroves) and artificial wetlands (flooded agricultural ecosystems) harbor methane-utilizing bacteria that oxidize methane at the source and reduce its net flux. For several decades, microorganisms have been used as biofertilizers to promote plant growth. However, now their role in reducing net methane flux, especially from flooded agricultural ecosystems is gaining momentum globally. Research in this context has mainly focused on taxonomic aspects related to methanotrophy among diverse bacterial genera, and environmental factors that govern methane utilization in natural and artificial wetland ecosystems. In the last few decades, concerted efforts have been made to develop multifunctional microbial inoculants that can oxidize methane and alleviate greenhouse gas emissions, as well as promote plant growth. In this context, combinations of taxonomic groups commonly found in rice paddies and those used as biofertilizers are being explored. This review deals with methanotrophy among diverse bacterial domains, factors influencing methane-utilizing ability, and explores the potential of novel methane-utilizing microbial consortia with plant growth-promoting traits in flooded ecosystems.

Isolation, Screening, and Characterization of Methane-Utilizing Bacteria From the Sediment of Lowland Rice

Methane is a major greenhouse gas that contributes to climate change. Methanogen and methanotrophic group microbes play a role in methane emissions in lowland rice fields. Methane is produced by methanogenic bacteria that decompose organic matter in anaerobic conditions. These bacteria will be active if the soil is inundated for an extended period of time. Some of this methane will be oxidized by methanotrophic bacteria in the rhizosphere. The researchers aimed to isolate, screen, and characterize methane-utilizing bacteria in lowland rice sediment from several Indonesian provinces. 27 methane- utilizing bacteria were isolated from rice field sediments in Lampung, West Java, and East Nusa Tenggara Province. Six of them had the potential to reduce methane emissions by more than half. A pmoA-like gene could be found in all of the selected isolates. The bacterial isolates were identified as Mycobacterium senegalense, Bacillus marisflavi, Bacillus methylotrophicus, Flavobacterium tirrenicum, Providencia stuartii, and Rhizobium rhizoryzae after characterization and identification with the Biolog OmniLog® ID system. These were all capable of nitrogen fixation, phosphorus solubilization, and IAA production. These isolates have the potential to be used as biofertilizers and methane mitigation agents.
 Keywords: biofertilizers, greenhouse gas, lowland rice, methane-utilizing bacteria

Mitigation of yield-scaled greenhouse gas emissions from irrigated rice through Azolla, Blue-green algae, and plant growth-promoting bacteria.

Irrigated transplanted flooded rice is a major source of methane (CH4) emission. We carried out experiments for 2 years in irrigated flooded rice to study if interventions like methane-utilizing bacteria, Blue-green algae (BGA), and Azolla could mitigate the emission of CH4 and nitrous oxide (N2O) and lower the yield-scaled global warming potential (GWP). The experiment included nine treatments: T1 (120 kg N ha-1 urea), T2 (90 kg N ha-1 urea + 30 kg N ha-1 fresh Azolla), T3 (90 kg N ha-1 urea + 30 kg N ha-1 Blue-green algae (BGA), T4 (60 kg N ha-1 urea + 30 kg N ha-1 BGA + 30 kg N ha-1 Azolla, T5 (120 kg N ha-1 urea + Hyphomicrobium facile MaAL69), T6 (120 kg N ha-1 by urea + Burkholderia vietnamiensis AAAr40), T7 (120 kg N ha-1 by urea + Methylobacteruim oryzae MNL7), T8 (120 kg N ha-1 urea + combination of Burkholderia AAAr40, Hyphomicrobium facile MaAL69, Methylobacteruim oryzae MNL7), and T9 (no N fertilizer). Maximum decrease in cumulative CH4 emission was observed with the application of Methylobacteruim oryzae MNL7 in T7 (19.9%), followed by Azolla + BGA in T4 (13.2%) as compared to T1 control. N2O emissions were not significantly affected by the application of CH4-oxidizing bacteria. However, significantly lower (P<0.01) cumulative N2O emissions was observed in T4 (40.7%) among the fertilized treatments. Highest yields were observed in Azolla treatment T2 with 25% less urea N application. The reduction in yield-scaled GWP was at par in T4 (Azolla and BGA) and T7 (Methylobacteruim oryzae MNL7) treatments and reduced by 27.4% and 15.2% in T4 and T7, respectively, as compared to the T1 (control). K-means clustering analysis showed that the application of Methylobacteruim oryzae MNL7, Azolla, and Azolla + BGA can be an effective mitigation option to reduce the global warming potential while increasing the yield.

Preferential Binding of Lanthanides to Methanol Dehydrogenase Evaluated with Density Functional Theory.

Methanol dehydrogenase (MDH) is an enzyme used by certain bacteria for the oxidation of methanol to formaldehyde, which is a necessary metabolic reaction. The discovery of a lanthanide-dependent MDH reveals that lanthanide ions (Ln3+) have a role in biology. Two types of MDH exist in methane-utilizing bacteria: one that is Ca2+-dependent (MxaF) and another that is Ln3+-dependent. Given that the triply charged Ln3+ are strongly hydrated, it is not clear how preference for Ln3+ is manifested and if the Ca2+-dependent MxaF protein can also bind Ln3+ ions. A computational approach was used to estimate the Gibbs energy differences between the binding of Ln3+ and Ca2+ to MDH using density functional theory. The results show that both proteins bind La3+ with higher affinity than Ca2+, albeit with a more pronounced difference in the case of Ln3+-dependent MDH. Interestingly, the binding of heavier lanthanides is preferred over the binding of La3+, with Gd3+ showing the highest affinity for both proteins of all Ln3+ ions that were tested (La3+, Sm3+, Gd3+, Dy3+, and Lu3+). Energy decomposition analysis reveals that the higher affinity of La3+ than Ca2+ to MDH is due to stronger contributions of electrostatics and polarization, which overcome the high cost of desolvating the ion.

Methane utilizing plant growth-promoting microbial diversity analysis of flooded paddy ecosystem of India.

Methane utilizing bacteria (MUB) are known to inhabit the flooded paddy ecosystem where they play an important role in regulating net methane (CH4) emission. We hypothesize that efficient MUB having plant growth-promoting (PGP) attributes can be used for developing novel bio-inoculant for flooded paddy ecosystem which might not only reduce methane emission but also assist in improving the plant growth parameters. Hence, soil and plant samples were collected from the phyllosphere, rhizosphere, and non-rhizosphere of five rice-growing regions of India at the tillering stage and investigated for efficient methane-oxidizing and PGP bacteria. Based on the monooxygenase activity and percent methane utilization on NMS medium with methane as the sole C source, 123 isolates were identified and grouped phylogenetically into 13 bacteria and 2 yeast genera. Among different regions, a significantly higher number of isolates were obtained from lowland flooded paddy ecosystems of Aduthurai (33.33%) followed by Ernakulum (20.33%) and Brahmaputra valley (19.51%) as compared to upland irrigated regions of Gaya (17.07%) and Varanasi (8.94%). Among sub-samples, a significantly higher number of isolates were found inhabiting the phyllosphere (58.54%) followed by non-rhizosphere (25.20%) and rhizosphere (15.45%). Significantly higher utilization of methane and PGP attributes were observed in 30 isolates belonging to genera Hyphomicrobium, Burkholderia, Methylobacterium, Paenibacillus,Pseudomonas,Rahnella, andMeyerozyma.M. oryzaeMNL7 showed significantly better growth with 74.33% of CH4 utilization at the rate of 302.9 ± 5.58 and exhibited half-maximal growth rate, Ksof 1.92 ± 0.092mg CH4L-1. Besides the ability to utilize CH4,P. polymyxaMaAL70 possessed PGP attributes such as solubilization of P, K, and Zn, fixation of atmospheric N and production of indole acetic acid (IAA). Both these promising isolates can be explored in the future for developing novel biofertilizers for flooded paddies.

Impact of methane-utilizing bacteria on rice yield, inorganic fertilizers efficiency and methane emissions

Methane (CH4) is one of the greenhouse gases that contribute to climate change. Lowland rice cultivation is one of the main sources of methane emissions, accounting for around 5 to 19% of total global CH4. One of the efforts to reduce CH4 gas emissions that is environmentally friendly is through methane-utilizing bacteria. This study aimed to determine the effect of methane-utilizing bacteria on rice yield and methane emissions. The bacterial strains used were Mycobacterium senegalense LM1, Providencia stuartii LM18, Rhizobium rhizoryzae BMU, and Bacillus methylotrophicus N2P4. The research was conducted in the experimental field of The Indonesian Center for Rice Research, Bogor, Indonesia. The experiment was carried out by using a factorial randomized block design with two factors and three replications. The first factor was the dose of NPK inorganic fertilizer (50%, 75%, 100%). The second factor was bacterial consortium formulas (without bacteria, bacterial consortium 1, bacterial consortium 2). The application of a bacterial consortium containing all strains increased the efficiency of inorganic fertilizers by 25%, increased rice production by 33.55%, and reduced CH4 emissions by 37.26%. It seems that the consortium of methane-utilizing bacteria has the prospect of biofertilizer and mitigation agents to mitigate the impact of global warming.

How Lanthanide Ions Affect the Addition-Elimination Step of Methanol Dehydrogenases.

The recently discovered methanol dehydrogenase, XoxF, is a widespread enzyme used by methylotrophic bacteria to oxidize methanol for carbon and energy, and requires lanthanide ions for its activity. This enzyme represents an essential component of methanol utilization by both methanol- and methane-utilizing bacteria. The present investigation looks on the electronic, energetic and geometrical behavior of the methanol dehydrogenase from Methylacidiphilum fumariolicum SolV, which is strictly dependent on early lanthanide metals with +3 oxidation states, by examining enzyme-substrate complexes of all the lanthanides. We focus on the catalytic reaction mechanism of two methanol dehydrogenases having as cofactor europium and ytterbium belonging to the mid- and later- series of lanthanides, in comparison with the methanol dehydrogenase containing the cerium, one early lanthanide. Our results provide evidence for the influence of the lanthanide contraction effect in all the elementary steps of the catalytic reaction mechanism. This indication may prove useful for developing new catalytic machineries of enzymes that adopt new-to-nature transformations.

メタノール資化性酵母と細菌及びメタン利用細菌の保存(研究報告,第26回凍結及び乾燥研究会特別講演及び研究報告)

メタノール資化性酵母と細菌及びメタン利用細菌の保存(研究報告,第26回凍結及び乾燥研究会特別講演及び研究報告)

Genome-scale metabolic reconstructions and theoretical investigation of methane conversion in Methylomicrobium buryatense strain 5G(B1).

BackgroundMethane-utilizing bacteria (methanotrophs) are capable of growth on methane and are attractive systems for bio-catalysis. However, the application of natural methanotrophic strains to large-scale production of value-added chemicals/biofuels requires a number of physiological and genetic alterations. An accurate metabolic model coupled with flux balance analysis can provide a solid interpretative framework for experimental data analyses and integration.ResultsA stoichiometric flux balance model of Methylomicrobium buryatense strain 5G(B1) was constructed and used for evaluating metabolic engineering strategies for biofuels and chemical production with a methanotrophic bacterium as the catalytic platform. The initial metabolic reconstruction was based on whole-genome predictions. Each metabolic step was manually verified, gapfilled, and modified in accordance with genome-wide expression data. The final model incorporates a total of 841 reactions (in 167 metabolic pathways). Of these, up to 400 reactions were recruited to produce 118 intracellular metabolites. The flux balance simulations suggest that only the transfer of electrons from methanol oxidation to methane oxidation steps can support measured growth and methane/oxygen consumption parameters, while the scenario employing NADH as a possible source of electrons for particulate methane monooxygenase cannot. Direct coupling between methane oxidation and methanol oxidation accounts for most of the membrane-associated methane monooxygenase activity. However the best fit to experimental results is achieved only after assuming that the efficiency of direct coupling depends on growth conditions and additional NADH input (about 0.1–0.2 mol of incremental NADH per one mol of methane oxidized). The additional input is proposed to cover loss of electrons through inefficiency and to sustain methane oxidation at perturbations or support uphill electron transfer. Finally, the model was used for testing the carbon conversion efficiency of different pathways for C1-utilization, including different variants of the ribulose monophosphate pathway and the serine cycle.ConclusionWe demonstrate that the metabolic model can provide an effective tool for predicting metabolic parameters for different nutrients and genetic perturbations, and as such, should be valuable for metabolic engineering of the central metabolism of M. buryatense strains.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-015-0377-3) contains supplementary material, which is available to authorized users.

Gas chromatography–mass spectrometry with chemometric analysis for determining 12C and 13C labeled contributions in metabolomics and 13C flux analysis

A novel method for the analysis of nearly co-eluting 12C and 13C isotopically labeled metabolites has been developed and evaluated for gas chromatography coupled to mass spectrometry (GC–MS) data. The method utilizes parallel factor analysis (PARAFAC) with two-dimensional GC–MS data when sample replicates are aligned and stacked in series to create a three-dimensional data cube for mathematical peak deconvolution and 12C and 13C contribution isolation, with the intent of increasing the accuracy and precision of quantitative metabolomics and 13C flux analysis. The platform is demonstrated with 13C-labeled metabolite extracts, generated via biosynthesis, added as an internal standard to unlabeled 12C metabolites extracted from the methanol-utilizing bacterium Methylobacterium extorquens AM1. Eleven representative metabolites that are common targets for flux analysis were chosen for validation. Good quantitative accuracy and precision were acquired for a 5.00μM known metabolite concentration (for the 11 metabolites), with an average predicted concentration of 5.07μM, and a RSD range of 1.2–13.0%. This study demonstrates the ability to reliably deconvolute 12C-unlabeled and 13C-labeled contributions for a given metabolite. Additionally, using this chemical analysis platform, a dynamic flux experiment is presented in which the incorporation of 13C-labeled cell extract can be detected in the methane-utilizing bacterium Methylosinus trichosporium OB3b and measured temporally.

Methylobacterium populi sp. nov., a novel aerobic, pink-pigmented, facultatively methylotrophic, methane-utilizing bacterium isolated from poplar trees (Populus deltoides x nigra DN34).

A pink-pigmented, aerobic, facultatively methylotrophic bacterium, strain BJ001T, was isolated from internal poplar tissues (Populus deltoidesxnigra DN34) and identified as a member of the genus Methylobacterium. Phylogenetic analyses showed that strain BJ001T is related to Methylobacterium thiocyanatum, Methylobacterium extorquens, Methylobacterium zatmanii and Methylobacterium rhodesianum. However, strain BJ001T differed from these species in its carbon-source utilization pattern, particularly its use of methane as the sole source of carbon and energy, an ability that is shared with only one other member of the genus, Methylobacterium organophilum. In addition, strain BJ001T is the only member of the genus Methylobacterium to be described as an endophyte of poplar trees. On the basis of its physiological, genotypic and ecological properties, the isolate is proposed as a member of a novel species of the genus Methylobacterium, Methylobacterium populi sp. nov. (type strain, BJ001T=ATCC BAA-705T=NCIMB 13946T).

Methylomonas scandinavica sp.nov., a New Methanotrophic Psychrotrophic Bacterium isolated from Deep Igneous Rock Ground Water of Sweden

Methane-utilizing bacteria were enriched from deep igneous rock environments and affiliated by amplification of functional and phylogenetic gene probes. Type I methanotrophs belonging to the genera Methylomonas and Methylobacter dominated in enrichment cultures from depths below 400 m. A pure culture of an obligate methanotroph (strain SR5) was isolated and characterized. Pink-pigmented motile rods of the new isolate contained intracytoplasmic membranes as stacks of vesicles, assimilated methane via the ribulose monophosphate pathway and had an incomplete tricarboxylic acid cycle. Phosphatidyl glycerol, methylene ubiquinone and cytochrome c552 were prevailing. The DNA G+C content is 53.3 mol %. Strain SR5 grew at temperatures between 5 and 30 degrees C with optimum at 15 degrees C, close to its in situ temperature. Analyses of 16S rRNA gene, whole cell protein, enzymatic and physiological analyses of strain SR-5 revealed significant differences compared to the other representatives of Type I methanotrophs. Based on pheno- and genotypic characteristics we propose to refer the strain SR5 as to a new species, Methylomonas scandinavica.

Bioremediation of trichloroethylene and cis-1,2-dichloroethylene-contaminated groundwater by methane-utilizing bacteria.

Experimental studies on the bioremediation of groundwater contaminated with low concentration trichloroethylene (TCE) and cis1,2-dichloroethylene (DCE) were performed with two sets of bioreactors. Reactors No. 1 and No. 2 were operated without and with methane supplement, respectively. No inoculum was used. The concentrations of TCE and DCE in the effluent and the off gas from reactor No. 2 were much lower than those from reactor No. 1. When air and an H2O2 solution were supplied to reactor No. 2, concentrations of TCE and DCE in the effluent and the off gas were lower than the lowest detectable limit. The population of methane-utilizing bacteria in reactor No. 2 was 1,000 times higher than that in groundwater or in the effluent from reactor No. 1. These methane-utilizing bacteria were apparently attributable to the treatment of TCE.

Some characteristics of bacteria found in a bioreactor to treat trichloroethylene-contaminated groundwater.

A mixture of bacteria, having a methane-utilizing ability, was separated from a bioreactor supplied with air and methane gas. The bioreactor was operated to treat trichloroethylene (TCE)-contaminated groundwater. The mixture was composed of an obligate methane-utilizing bacterium and a heterotroph, identified as Methylomonas methanica and Pseudomonas sp., respectively. The mixed culture of these two strains removed TCE. In addition, it appeared that a cooperative metabolic interaction of these strains enabled Meth.methanica to maintain the TCE degradation ability.

Methanotrophic bacteria.

Methane-utilizing bacteria (methanotrophs) are a diverse group of gram-negative bacteria that are related to other members of the Proteobacteria. These bacteria are classified into three groups based on the pathways used for assimilation of formaldehyde, the major source of cell carbon, and other physiological and morphological features. The type I and type X methanotrophs are found within the gamma subdivision of the Proteobacteria and employ the ribulose monophosphate pathway for formaldehyde assimilation, whereas type II methanotrophs, which employ the serine pathway for formaldehyde assimilation, form a coherent cluster within the beta subdivision of the Proteobacteria. Methanotrophic bacteria are ubiquitous. The growth of type II bacteria appears to be favored in environments that contain relatively high levels of methane, low levels of dissolved oxygen, and limiting concentrations of combined nitrogen and/or copper. Type I methanotrophs appear to be dominant in environments in which methane is limiting and combined nitrogen and copper levels are relatively high. These bacteria serve as biofilters for the oxidation of methane produced in anaerobic environments, and when oxygen is present in soils, atmospheric methane is oxidized. Their activities in nature are greatly influenced by agricultural practices and other human activities. Recent evidence indicates that naturally occurring, uncultured methanotrophs represent new genera. Methanotrophs that are capable of oxidizing methane at atmospheric levels exhibit methane oxidation kinetics different from those of methanotrophs available in pure cultures. A limited number of methanotrophs have the genetic capacity to synthesize a soluble methane monooxygenase which catalyzes the rapid oxidation of environmental pollutants including trichloroethylene.

The mxaAKL genes of Methylobacter albus BG8

The facultative methanol utilizer Methylobacterium extorquens AM1 contains at least three genes (mxaA, K and L) that encode functions involved in providing calcium to the holoenzyme of methanol dehydrogenase, the enzyme that oxidizes methanol to formaldehyde in this strain. Methane-utilizing bacteria (methanotrophs) also contain methanol dehydrogenase, and evidence suggests that similar methanol oxidation (Mox) functions may be present in some of these strains. DNA fragments from Methylobacterium extorquens AM1 specific to mxaA, mxaK and mxaL were isolated for use as hybridization probes against genomic digests of a variety of methanotrophic bacteria. Only the mxaL probe showed substantial hybridization, and it was used to identify and isolate an 8.5 kb HindIII fragment from Methylobacter albus BG8 (a Type I methanotroph). Hybridization of restriction digests of this fragment to individual probes for Methylobacterium extorquens AM1 mxaA, K and L indicated that the relative mxa gene order in Methylobacter albus BG8 is A-K-L. A T7 dual promoter/polymerase protein expression system indicated that five polypeptides are expressed from a 4.5 kb region of Methylobacter albus BG8 DNA in Escherichia coli, all transcribed in the same direction, and they apparently correspond to mxaACKDL. The functions of mxaC and mxaD are currently not known, but the order of mxaDL is reversed in Methylobacter albus BG8 compared to Methylobacterium extorquens AM1. When subclones of the Methylobacter albus BG8 fragment containing these genes were used as hybridization probes to genomic digests of methanotrophic bacteria, specific bands were detected that suggested a similar gene order in most cases. These data indicate that the mxaAKL region is relatively highly conserved in methanotrophs, and that in most cases the mxaAKL genes are grouped together in the same order as in the facultative methanol utilizer Methylobacterium extorquens AM1.

Aerobic transformation of linear alkylbenzenesulphonates by mixed methane-utilizing bacteria

Biodegradation of commercial linear alkylbenzenesulphonate (LAS) was performed in shake flasks in the presence and absence of methane as a primary growth substrate for methanotrophs and as an inducer for catabolic enzymes. Methane-utilizing culture MM1 was dominated by type II methanotrophs and originated from a groundwater aquifer. Methane, carbon dioxide, and oxygen concentrations were measured by using gas chromatography. Disappearance of LAS, a mixture of C10-C13 homologues, was monitored by reversed-phase high-performance liquid chromatography (RP-HPLC). Methane reduced LAS transformation rate in a concentration dependent manner, suggesting competitive inhibition between natural substrate, methane, and fortuitous substrate, LAS. The fastest LAS transformation was determined in the absence of methane. Simultaneous methane oxidation and LAS degradation, and the inhibition of both transformation processes by acetylene, indicated the involvement of a methane-monooxygenase enzyme system in LAS transformation. RP-HPLC analysis showed that culture MM1 exhibited preferential capability to transform LAS with longer alkyl side-chain. Consequently, the highest transformation rate was observed with the homologue of C13 LAS.