One Step Closer to Enigmatic USCα Methanotrophs: Isolation of a Methylocapsa-like Bacterium from a Subarctic Soil.

The scavenging of atmospheric trace gases has been recognized as one of the lifestyle-defining capabilities of microorganisms in terrestrial polar ecosystems. Several metagenome-assembled genomes of as-yet-uncultivated methanotrophic bacteria, which consume atmospheric CH4 in these ecosystems, have been retrieved in cultivation-independent studies. In this study, we isolated and characterized a representative of these methanotrophs, strain D3K7, from a subarctic soil of northern Russia. Strain D3K7 grows on methane and methanol in a wide range of temperatures, between 5 and 30 °C. Weak growth was also observed on acetate. The presence of acetate in the culture medium stimulated growth at low CH4 concentrations (~100 p.p.m.v.). The finished genome sequence of strain D3K7 is 4.15 Mb in size and contains about 3700 protein-encoding genes. According to the result of phylogenomic analysis, this bacterium forms a common clade with metagenome-assembled genomes obtained from the active layer of a permafrost thaw gradient in Stordalen Mire, Abisco, Sweden, and the mineral cryosol at Axel Heiberg Island in the Canadian High Arctic. This clade occupies a phylogenetic position in between characterized Methylocapsa methanotrophs and representatives of the as-yet-uncultivated upland soil cluster alpha (USCα). As shown by the global distribution analysis, D3K7-like methanotrophs are not restricted to polar habitats but inhabit peatlands and soils of various climatic zones.

Methylomonas rapida sp. nov., a novel species of fast-growing, carotenoid-producing obligate methanotrophs with high biotechnological potential.

The genus Methylomonas accommodates strictly aerobic, obligate methanotrophs, with their sole carbon and energy sources restricted to methane and methanol. These bacteria inhabit oxic-anoxic interfaces of various freshwater habitats and have attracted considerable attention as potential producers of a single-cell protein. Here, we characterize two fast-growing representatives of this genus, strains 12 and MP1T, which are phylogenetically distinct from the currently described Methylomonas species (94.0-97.3 % 16S rRNA gene sequence similarity). Strains 12 and MP1T were isolated from freshwater sediments collected in Moscow and Krasnodar regions, respectively. Cells of these strains are Gram-negative, red-pigmented, highly motile thick rods that contain a type I intracytoplasmic membrane system and possess a particulate methane monooxygenase (pMMO) enzyme. These bacteria grow between 8 and 45 °C (optimum 35 °C) in a relatively narrow pH range of 5.5-7.3 (optimum pH 6.6-7.2). Major carotenoids synthesized by these methanotrophs are 4,4'-diaplycopene-4,4'-dioic acid, 1,1'-dihydroxy-3,4-didehydrolycopene and 4,4'-diaplycopenoic acid. High biomass yield, of up to 3.26 g CDW/l, is obtained during continuous cultivation of MP1T on natural gas in a bioreactor at a dilution rate of 0.22 h-1. The complete genome sequence of strain MP1T is 4.59 Mb in size; the DNA G + C content is 52.8 mol%. The genome encodes four rRNA operons, one pMMO operon and 4,216 proteins. The genome sequence displays 82-85 % average nucleotide identity to those of earlier described Methylomonas species. We propose to classify these bacteria as representing a novel species of the genus Methylomonas, M. rapida sp. nov., with the type strain MP1T (=KCTC 92586T = VKM B-3663T).

Building a Cell House from Cellulose: The Case of the Soil Acidobacterium Acidisarcina polymorpha SBC82T.

Acidisarcina polymorpha SBC82T is a recently described representative of the phylum Acidobacteriota from lichen-covered tundra soil. Cells of this bacterium occur within unusual saccular chambers, with the chamber envelope formed by tightly packed fibrils. These extracellular structures were most pronounced in old cultures of strain SBC82T and were organized in cluster-like aggregates. The latter were efficiently destroyed by incubating cell suspensions with cellulase, thus suggesting that they were composed of cellulose. The diffraction pattern obtained for 45-day-old cultures of strain SBC82T by using small angle X-ray scattering was similar to those reported earlier for mature wood samples. The genome analysis revealed the presence of a cellulose biosynthesis locus bcs. Cellulose synthase key subunits A and B were encoded by the bcsAB gene whose close homologs are found in genomes of many members of the order Acidobacteriales. More distant homologs of the acidobacterial bcsAB occurred in representatives of the Proteobacteria. A unique feature of bcs locus in strain SBC82T was the non-orthologous displacement of the bcsZ gene, which encodes the GH8 family glycosidase with a GH5 family gene. Presumably, these cellulose-made extracellular structures produced by A. polymorpha have a protective function and ensure the survival of this acidobacterium in habitats with harsh environmental conditions.

Corrigendum: Hydrolytic Capabilities as a Key to Environmental Success: Chitinolytic and Cellulolytic Acidobacteria From Acidic Sub-arctic Soils and Boreal Peatlands.

[This corrects the article DOI: 10.3389/fmicb.2018.02775.].

Expanding Characterized Diversity and the Pool of Complete Genome Sequences of Methylococcus Species, the Bacteria of High Environmental and Biotechnological Relevance.

The bacterial genus Methylococcus, which comprises aerobic thermotolerant methanotrophic cocci, was described half-a-century ago. Over the years, a member of this genus, Methylococcus capsulatus Bath, has become a major model organism to study genomic and metabolic basis of obligate methanotrophy. High biotechnological potential of fast-growing Methylococcus species, mainly as a promising source of feed protein, has also been recognized. Despite this big research attention, the currently cultured Methylococcus diversity is represented by members of the two species, M. capsulatus and M. geothermalis, while finished genome sequences are available only for two strains of these methanotrophs. This study extends the pool of phenotypically characterized Methylococcus strains with good-quality genome sequences by contributing four novel isolates of these bacteria from activated sludge, landfill cover soil, and freshwater sediments. The determined genome sizes of novel isolates varied between 3.2 and 4.0Mb. As revealed by the phylogenomic analysis, strains IO1, BH, and KN2 affiliate with M. capsulatus, while strain Mc7 may potentially represent a novel species. Highest temperature optima (45-50°C) and highest growth rates in bioreactor cultures (up to 0.3h-1) were recorded for strains obtained from activated sludge. The comparative analysis of all complete genomes of Methylococcus species revealed 4,485 gene clusters. Of these, pan-genome core comprised 2,331 genes (on average 51.9% of each genome), with the accessory genome containing 846 and 1,308 genes in the shell and the cloud, respectively. Independently of the isolation source, all strains of M. capsulatus displayed surprisingly high genome synteny and a striking similarity in gene content. Strain Mc7 from a landfill cover soil differed from other isolates by the high content of mobile genetic elements in the genome and a number of genome-encoded features missing in M. capsulatus, such as sucrose biosynthesis and the ability to scavenge phosphorus and sulfur from the environment.

Methane-Oxidizing Communities in Lichen-Dominated Forested Tundra Are Composed Exclusively of High-Affinity USCα Methanotrophs.

Upland soils of tundra function as a constant sink for atmospheric CH4 but the identity of methane oxidizers in these soils remains poorly understood. Methane uptake rates of -0.4 to -0.6 mg CH4-C m-2 day-1 were determined by the static chamber method in a mildly acidic upland soil of the lichen-dominated forested tundra, North Siberia, Russia. The maximal CH4 oxidation activity was localized in an organic surface soil layer underlying the lichen cover. Molecular identification of methanotrophic bacteria based on retrieval of the pmoA gene revealed Upland Soil Cluster Alpha (USCα) as the only detectable methanotroph group. Quantification of these pmoA gene fragments by means of specific qPCR assay detected ~107pmoA gene copies g-1 dry soil. The pmoA diversity was represented by seven closely related phylotypes; the most abundant phylotype displayed 97.5% identity to pmoA of Candidatus Methyloaffinis lahnbergensis. Further analysis of prokaryote diversity in this soil did not reveal 16S rRNA gene fragments from well-studied methanotrophs of the order Methylococcales and the family Methylocystaceae. The largest group of reads (~4% of all bacterial 16S rRNA gene fragments) that could potentially belong to methanotrophs was classified as uncultivated Beijerinckiaceae bacteria. These reads displayed 96-100 and 95-98% sequence similarity to 16S rRNA gene of Candidatus Methyloaffinis lahnbergensis and "Methylocapsa gorgona" MG08, respectively, and were represented by eight species-level operational taxonomic units (OTUs), two of which were highly abundant. These identification results characterize subarctic upland soils, which are exposed to atmospheric methane concentrations only, as a unique habitat colonized mostly by USCα methanotrophs.

Methylotetracoccus oryzae Strain C50C1 Is a Novel Type Ib Gammaproteobacterial Methanotroph Adapted to Freshwater Environments.

Methane-oxidizing microorganisms perform an important role in reducing emissions of the greenhouse gas methane to the atmosphere. To date, known bacterial methanotrophs belong to the Proteobacteria, Verrucomicrobia, and NC10 phyla. Within the Proteobacteria phylum, they can be divided into type Ia, type Ib, and type II methanotrophs. Type Ia and type II are well represented by isolates. Contrastingly, the vast majority of type Ib methanotrophs have not been able to be cultivated so far. Here, we compared the distributions of type Ib lineages in different environments. Whereas the cultivated type Ib methanotrophs (Methylococcus and Methylocaldum) are found in landfill and upland soils, lineages that are not represented by isolates are mostly dominant in freshwater environments, such as paddy fields and lake sediments. Thus, we observed a clear niche differentiation within type Ib methanotrophs. Our subsequent isolation attempts resulted in obtaining a pure culture of a novel type Ib methanotroph, tentatively named "Methylotetracoccus oryzae" C50C1. Strain C50C1 was further characterized to be an obligate methanotroph, containing C16:1ω9c as the major membrane phospholipid fatty acid, which has not been found in other methanotrophs. Genome analysis of strain C50C1 showed the presence of two pmoCAB operon copies and XoxF5-type methanol dehydrogenase in addition to MxaFI. The genome also contained genes involved in nitrogen and sulfur cycling, but it remains to be demonstrated if and how these help this type Ib methanotroph to adapt to fluctuating environmental conditions in freshwater ecosystems.IMPORTANCE Most of the methane produced on our planet gets naturally oxidized by a group of methanotrophic microorganisms before it reaches the atmosphere. These microorganisms are able to oxidize methane, both aerobically and anaerobically, and use it as their sole energy source. Although methanotrophs have been studied for more than a century, there are still many unknown and uncultivated groups prevalent in various ecosystems. This study focused on the diversity and adaptation of aerobic methane-oxidizing bacteria in different environments by comparing their phenotypic and genotypic properties. We used lab-scale microcosms to create a countergradient of oxygen and methane for preenrichment, followed by classical isolation techniques to obtain methane-oxidizing bacteria from a freshwater environment. This resulted in the discovery and isolation of a novel methanotroph with interesting physiological and genomic properties that could possibly make this bacterium able to cope with fluctuating environmental conditions.

Fatty Acid and Hopanoid Adaption to Cold in the Methanotroph Methylovulum psychrotolerans.

Three strains of aerobic psychrotolerant methanotrophic bacteria Methylovulum psychrotolerans, isolated from geographically remote low-temperature environments in Northern Russia, were grown at three different growth temperatures, 20, 10 and 4°C and were found to be capable of oxidizing methane at all temperatures. The three M. psychrotolerans strains adapted their membranes to decreasing growth temperature by increasing the percent of unsaturated fatty acid (FAs), both for the bulk and intact polar lipid (IPL)-bound FAs. Furthermore, the ratio of ßOH-C16:0 to n-C16:0 increased as growth temperature decreased. The IPL head group composition did not change as an adaption to temperature. The most notable hopanoid temperature adaptation of M. psychrotolerans was an increase in unsaturated hopanols with decreasing temperature. As the growth temperature decreased from 20 to 4°C, the percent of unsaturated M. psychrotolerans bulk-FAs increased from 79 to 89 % while the total percent of unsaturated hopanoids increased from 27 to 49 %. While increased FA unsaturation in response to decreased temperature is a commonly observed response in order to maintain the liquid-crystalline character of bacterial membranes, hopanoid unsaturation upon cold exposition has not previously been described. In order to investigate the mechanisms of both FA and hopanoid cold-adaption in M. psychrotolerans we identified genes in the genome of M. psychrotolerans that potentially code for FA and hopanoid desaturases. The unsaturation of hopanoids represents a novel membrane adaption to maintain homeostasis upon cold adaptation.

Hydrolytic Capabilities as a Key to Environmental Success: Chitinolytic and Cellulolytic Acidobacteria From Acidic Sub-arctic Soils and Boreal Peatlands.

Members of the Acidobacteria are among the most efficient colonizers of acidic terrestrial habitats but the key traits underlying their environmental fitness remain to be understood. We analyzed indigenous assemblages of Acidobacteria in a lichen-covered acidic (pH 4.1) soil of forested tundra dominated by uncultivated members of subdivision 1. An isolate of these bacteria with cells occurring within saccular chambers, strain SBC82T, was obtained. The genome of strain SBC82T consists of a 7.11-Mb chromosome and four megaplasmids, and encodes a wide repertoire of enzymes involved in degradation of chitin, cellulose, and xylan. Among those, four secreted chitinases affiliated with the glycoside hydrolase family GH18 were identified. Strain SBC82T utilized amorphous chitin as a source of carbon and nitrogen; the respective enzyme activities were detected in tests with synthetic substrates. Chitinolytic capability was also confirmed for another phylogenetically related acidobacterium isolated from a Sphagnum peat bog, strain CCO287. As revealed by metatranscriptomic analysis of chitin-amended peat, 16S rRNA reads from these acidobacteria increased in response to chitin availability. Strains SBC82T and CCO287 were assigned to a novel genus and species, Acidisarcina polymorpha gen. nov., sp. nov. Members of this genus colonize acidic soils and peatlands and specialize in degrading complex polysaccharides.

Draft Genome Sequence of Methylovulum psychrotolerans Sph1T, an Obligate Methanotroph from Low-Temperature Environments.

Methylovulum psychrotolerans Sph1T is an aerobic, obligate methanotroph, which was isolated from cold methane seeps in West Siberia. This bacterium possesses only a particulate methane monooxygenase and is widely distributed in low-temperature environments. Strain Sph1T has the genomic potential for biosynthesis of hopanoids required for the maintenance of intracytoplasmic membranes.

Edaphobacter lichenicola sp. nov., a member of the family Acidobacteriaceae from lichen-dominated forested tundra.

An isolate of aerobic, Gram-stain-negative, rod-shaped, non-motile and light-pink pigmented bacteria, designated SBC68T, was obtained from slightly decomposed thalli of the lichen Cladonia sp. collected from the forested tundra of north-western Siberia. Cells of this isolate occurred singly, in pairs or in rosettes. These bacteria were acidophilic (optimum growth at pH 4.3-5.6) and mesophilic (optimum growth at 20-30 °C) but were also capable of growth at low temperatures, down to 7 °C. The preferred growth substrates were sugars, some organic acids and lichenan. The major fatty acids were iso-C15 : 0, C16 : 1ω7c, C16 : 0, C16 : 1ω7t, and 13,16-dimethyl octacosanedioic acid. The only quinone was MK-8, and the G+C content of the DNA was 54.7 mol%. SBC68T represented a member of the family Acidobactericeae; the closest taxonomically described relatives were Edaphobacter dinghuensis DHF9T and Granulicella aggregans TPB6028T (97.2 and 97.1 % 16S rRNA gene sequence similarity, respectively). In 16S rRNA gene-based trees, SBC68T clustered together with species of the genus Edaphobacter. However, this isolate differed from all previously described species of the genus Edaphobacter with respect to the pink pigmentation, formation of cell rosettes and substrate utilization pattern. On the basis of these data, strain SBC68T should be considered to represent a novel species of acidobacteria, for which the name Edaphobacter lichenicola sp. nov. is proposed. The type strain is SBC68T (=DSM 104462T=VKM B-3208T).

Methylovulum psychrotolerans sp. nov., a cold-adapted methanotroph from low-temperature terrestrial environments, and emended description of the genus Methylovulum.

Two isolates of aerobic methanotrophic bacteria, strains Sph1T and Sph2, were obtained from cold methane seeps in a floodplain of the river Mukhrinskaya, Irtysh basin, West Siberia. Another morphologically and phenotypically similar methanotroph, strain OZ2, was isolated from a sediment of a subarctic freshwater lake, Archangelsk region, northern Russia. Cells of these three strains were Gram-stain-negative, light-pink-pigmented, non-motile, encapsulated, large cocci that contained an intracytoplasmic membrane system typical of type I methanotrophs. They possessed a particulate methane monooxygenase enzyme and utilized only methane and methanol. Strains Sph1T, Sph2 and OZ2 were able to grow at a pH range of 4.0-8.9 (optimum at pH 6.0-7.0) and at temperatures between 2 and 36 °C. Although their temperature optimum was at 20-25 °C, these methanotrophs grew well at lower temperatures, down to 4 °C. The major cellular fatty acids were C16 : 1ω5c, C16 : 1ω6c, C16 : 1ω7c, C16 : 1ω8c, C16 : 0 and C14 : 0; the DNA G+C content was 51.4-51.9 mol%. Strains Sph1T, Sph2 and OZ2 displayed nearly identical (99.1-99.7 % similarity) 16S rRNA gene sequences and belonged to the family Methylococcaceae of the class Gammaproteobacteria. The most closely related organism was Methylovulum miyakonense HT12T (96.0-96.5 % 16S rRNA gene sequence similarity and 90 % pmoA sequence similarity). The novel isolates, however, differed from Methylovulum miyakonense HT12T by cell morphology, pigmentation, absence of soluble methane monooxygenase, more active growth at low temperatures, growth over a broader pH range and higher DNA G+C content. On the basis of these differences, we propose a novel species, Methylovulum psychrotolerans sp. nov., to accommodate these methanotrophs. Strain Sph1T (=LMG 29227T=VKM B-3018T) is the type strain.

Methylocapsa palsarum sp. nov., a methanotroph isolated from a subArctic discontinuous permafrost ecosystem.

An aerobic methanotrophic bacterium was isolated from a collapsed palsa soil in northern Norway and designated strain NE2T. Cells of this strain were Gram-stain-negative, non-motile, non-pigmented, slightly curved thick rods that multiplied by normal cell division. The cells possessed a particulate methane monooxygenase enzyme (pMMO) and utilized methane and methanol. Strain NE2T grew in a wide pH range of 4.1­8.0 (optimum pH 5.2­6.5) at temperatures between 6 and 32 °C (optimum 18­25 °C), and was capable of atmospheric nitrogen fixation under reduced oxygen tension. The major cellular fatty acids were C18 : 1ω7c, C16 : 0 and C16 : 1ω7c, and the DNA G+C content was 61.7 mol%. The isolate belonged to the family Beijerinckiaceae of the class Alphaproteobacteria and was most closely related to the facultative methanotroph Methylocapsa aurea KYGT (98.3 % 16S rRNA gene sequence similarity and 84 % PmoA sequence identity). However, strain NE2T differed from Methylocapsa aurea KYGT by cell morphology, the absence of pigmentation, inability to grow on acetate, broader pH growth range, and higher tolerance to NaCl. Therefore, strain NE2T represents a novel species of the genus Methylocapsa, for which we propose the name Methylocapsa palsarum sp. nov. The type strain is NE2T ( = LMG 28715T = VKM B-2945T).

Methylocystis bryophila sp. nov., a facultatively methanotrophic bacterium from acidic Sphagnum peat, and emended description of the genus Methylocystis (ex Whittenbury et al. 1970) Bowman et al. 1993.

A novel species is proposed for two facultatively methanotrophic representatives of the genus Methylocystis, strains H2s(T) and S284, which were isolated from an acidic (pH 4.3) Sphagnum peat-bog lake (Teufelssee, Germany) and an acidic (pH 3.8) peat bog (European North Russia), respectively. Cells of strains H2s(T) and S284 are aerobic, Gram-negative, non-motile, curved coccoids or short rods that contain an intracytoplasmic membrane system typical of type-II methanotrophs. They possess both a soluble and a particulate methane monooxygenase (MMO); the latter is represented by two isozymes, pMMO1 and pMMO2. The preferred growth substrates are methane and methanol. In the absence of C1 substrates, however, these methanotrophs are capable of slow growth on acetate. Atmospheric nitrogen is fixed by means of an aerotolerant nitrogenase. Strains H2s(T) and S284 grow between pH 4.2 and 7.6 (optimum pH 6.0-6.5) and at 8-37 °C (optimum 25-30 °C). The major fatty acids are C18 : 1ω8c, C18 : 1ω7c and C16 : 1ω7c; the major quinone is Q-8. The DNA G+C content is 62.0-62.3 mol%. Strains H2s(T) and S284 share identical 16S rRNA gene sequences, which displayed 96.6-97.3 % similarity to sequences of other taxonomically characterized members of the genus Methylocystis. Therefore, strains H2s(T) and S284 are classified as members of a novel species, for which the name Methylocystis bryophila sp. nov. is proposed; strain H2s(T) ( = DSM 21852(T)  = VKM B-2545(T)) is the type strain.

Acetate utilization as a survival strategy of peat-inhabiting Methylocystis spp.

Representatives of the genus Methylocystis are traditionally considered to be obligately methanotrophic bacteria, which are incapable of growth on multicarbon substrates. Here, we describe a novel member of this genus, strain H2s, which represents a numerically abundant and ecologically important methanotroph population in northern Sphagnum-dominated wetlands. This isolate demonstrates a clear preference for growth on methane but is able to grow slowly on acetate in the absence of methane. Strain H2s possesses both forms of methane monooxygenase (particulate and soluble MMO) and a well-developed system of intracytoplasmic membranes (ICM). In cells grown for several transfers on acetate, these ICM are maintained, although in a reduced form, and mRNA transcripts of particulate MMO are detectable. These cells resume their growth on methane faster than those kept for the same period of time without any substrate. Growth on acetate leads to a major shift in the phospholipid fatty acid composition. The re-examination of all type strains of the validly described Methylocystis species showed that Methylocystis heyeri H2(T) and Methylocystis echinoides IMET10491(T) are also capable of slow growth on acetate. This capability might represent an important part of the survival strategy of Methylocystis spp. in environments where methane availability is variable or limited.

Methylocapsa aurea sp. nov., a facultative methanotroph possessing a particulate methane monooxygenase, and emended description of the genus Methylocapsa.

An aerobic, methanotrophic bacterium, designated KYG(T), was isolated from a forest soil in Germany. Cells of strain KYG(T) were Gram-negative, non-motile, slightly curved rods that multiplied by binary fission and produced yellow colonies. The cells contained intracellular granules of poly-ß-hydroxybutyrate at each cell pole, a particulate methane monooxygenase (pMMO) and stacks of intracytoplasmic membranes (ICMs) packed in parallel along one side of the cell envelope. Strain KYG(T) grew at pH 5.2-7.2 and 2-33 °C and could fix atmospheric nitrogen under reduced oxygen tension. The major cellular fatty acid was C(18 : 1)ω7c (81.5 %) and the DNA G+C content was 61.4 mol%. Strain KYG(T) belonged to the family Beijerinckiaceae of the class Alphaproteobacteria and was most closely related to the obligate methanotroph Methylocapsa acidiphila B2(T) (98.1 % 16S rRNA gene sequence similarity and 84.7 % pmoA sequence similarity). Unlike Methylocapsa acidiphila B2(T), which grows only on methane and methanol, strain KYG(T) was able to grow facultatively on acetate. Facultative acetate utilization is a characteristic of the methanotrophs of the genus Methylocella, but the genus Methylocella does not produce pMMO or ICMs. Strain KYG(T) differed from Methylocapsa acidiphila B2(T) on the basis of substrate utilization pattern, pigmentation, pH range, cell ultrastructure and efficiency of dinitrogen fixation. Therefore, we propose a novel species, Methylocapsa aurea sp. nov., to accommodate this bacterium. The type strain is KYG(T) (=DSM 22158(T) =VKM B-2544(T)).

Acidisoma tundrae gen. nov., sp. nov. and Acidisoma sibiricum sp. nov., two acidophilic, psychrotolerant members of the Alphaproteobacteria from acidic northern wetlands.

Three obligately aerobic, heterotrophic bacteria, designated strains WM1T, TPB606T and TPB621, were isolated from acidic Sphagnum-dominated tundra and Siberian wetlands in Russia. Cells of these isolates were Gram-negative, non-motile coccobacilli that occurred singly, in pairs or in chains, and were covered by large capsules. The novel strains were moderately acidophilic and psychrotolerant organisms capable of growth at pH 3.0-7.6 and 2-30 degrees C. Cells contained numerous intracellular poly-beta-hydroxybutyrate granules (3-4 per cell). The major cellular fatty acid was cyclo C19:0omega8c and the predominant quinone was Q-10. Strains TPB606T and TPB621, isolated from Siberian wetland, possessed almost identical 16S rRNA gene sequences and shared 97.2% sequence similarity with tundra strain WM1T. The three strains were shown to belong to the Alphaproteobacteria, but were related only distantly to the type strains of acidophilic bacteria Acidisphaera rubrifaciens (93.4-94.3% 16S rRNA gene sequence similarity), Rhodopila globiformis (92.2-93.3%), and members of the genera Acidiphilium (91.3-93%) and Acidocella (91.8-92.4%). The DNA G+C contents of the novel strains were 60.5-61.9 mol%. The low levels of DNA-DNA relatedness (37%) and a number of phenotypic differences between the Siberian strains TPB606T and TPB621 and the tundra strain WM1T indicated that they represent two separate species. As the three isolates are clearly distinct from all recognized acidophilic members of the Alphaproteobacteria, they are considered to represent two novel species of a new genus, for which the names Acidisoma tundrae gen. nov., sp. nov. and Acidisoma sibiricum sp. nov. are proposed. The type strain of Acidisoma sibiricum is TPB606T (=DSM 21000T=VKM B-2487T) and the type strain of Acidisoma tundrae is WM1T (=DSM 19999T=VKM B-2488T).

Schlesneria paludicola gen. nov., sp. nov., the first acidophilic member of the order Planctomycetales, from Sphagnum-dominated boreal wetlands.

Three strains of budding, ellipsoid-shaped and rosette-forming bacteria were isolated from acidic Sphagnum-dominated boreal wetlands of northern Russia and were designated strains MPL7T, MOB77 and SB2. The presence of crateriform pits and numerous fibrillar appendages on the cell surface and an unusual spur-like projection on one pole of the cell indicated a planctomycete morphotype. These isolates are moderately acidophilic, mesophilic organisms capable of growth at pH values between 4.2 and 7.5 (with an optimum at pH 5.0-6.2) and at temperatures between 4 and 32 degrees C (optimum 15-26 degrees C). The major fatty acids are C16:0 and C16:1omega7c; the major quinone is MK-6. The G+C content of the DNA is 54.4-56.5 mol%. Strains MPL7T, MOB77 and SB2 possess nearly identical 16S rRNA gene sequences and belong to the planctomycete lineage defined by the genus Planctomyces, being most closely related to Planctomyces limnophilus DSM 3776T (86.9-87.1% sequence similarity). However, strain MPL7T showed only 28% DNA-DNA hybridization with P. limnophilus DSM 3776T. Compared with currently described members of the genus Planctomyces, the isolates from northern wetlands do not form long and distinctive stalks, have greater tolerance of acidic conditions and low temperatures, are more sensitive to NaCl, lack pigmentation and degrade a wider range of biopolymers. The data therefore suggest that strains MPL7T, MOB77 and SB2 represent a novel genus and species, for which the name Schlesneria paludicola gen. nov., sp. nov., is proposed. Strain MPL7T (=ATCC BAA-1393T=VKM B-2452T) is the type strain of Schlesneria paludicola.

Methylocystis heyeri sp. nov., a novel type II methanotrophic bacterium possessing 'signature' fatty acids of type I methanotrophs.

A novel species is proposed for two strains of methanotrophic bacteria (H2(T) and Sakb1) isolated from an acidic (pH 4.3) Sphagnum peat bog lake (Teufelssee, Germany) and an acidic (pH 4.2) tropical forest soil (Thailand), respectively. Cells of strains H2(T) and Sakb1 were aerobic, Gram-negative, non-motile, straight or curved rods that were covered by large polysaccharide capsules and contained an intracytoplasmic membrane system typical of type II methanotrophs. They possessed both a particulate and a soluble methane monooxygenase and utilized the serine pathway for carbon assimilation. They were moderately acidophilic organisms capable of growth between pH 4.4 and 7.5 (optimum 5.8-6.2). The most unique characteristic of these strains was the phospholipid fatty acid profile. In addition to the signature fatty acid of type II methanotrophs (18 : 1omega8c), the cells also contained large amounts of what was previously considered to be a signature fatty acid of type I methanotrophs, 16 : 1omega8c. The DNA G+C contents of strains H2(T) and Sakb1 were 61.5 and 62.1 mol%, respectively. The 16S rRNA gene sequences possessed 96-98 % similarity to sequences of other type II methanotrophs in the genera Methylosinus and Methylocystis. 16S rRNA gene sequence and pmoA phylogeny demonstrated that the strains form a novel lineage within the genus Methylocystis. DNA-DNA hybridization values of strain H2(T) with Methylocystis parvus OBBP(T) and Methylocystis echinoides IMET 10491(T) were 18 and 25 %, respectively. Thus, it is proposed that these two strains represent a novel species, Methylocystis heyeri sp. nov. Strain H2(T) (=DSM 16984(T)=VKM B-2426(T)) is the type strain.

Phylogenetic analysis and in situ identification of bacteria community composition in an acidic Sphagnum peat bog.

The Bacteria community composition in an acidic Sphagnum peat bog (pH 3.9 to 4.5) was characterized by a combination of 16S rRNA gene clone library analysis, rRNA-targeted fluorescence in situ hybridization (FISH), and cultivation. Among 84 environmental 16S rRNA gene clones, a set of only 16 cloned sequences was closely related (>or=95% similarity) to taxonomically described organisms. Main groups of clones were affiliated with the Acidobacteria (24 clones), Alphaproteobacteria (20), Verrucomicrobia (13), Actinobacteria (8), Deltaproteobacteria (4), Chloroflexi (3), and Planctomycetes (3). The proportion of cells that hybridized with oligonucleotide probes specific for members of the domains Bacteria (EUB338-mix) and Archaea (ARCH915 and ARC344) accounted for only 12 to 22% of the total cell counts. Up to 24% of the EUB338-positive cells could be assigned by FISH to specific bacterial phyla. Alphaproteobacteria and Planctomycetes were the most numerous bacterial groups (up to 1.3x10(7) and 1.1x10(7) cells g-1 peat, respectively). In contrast to conventional plating techniques, a novel biofilm-mediated enrichment approach allowed us to isolate some representatives of predominant Bacteria groups, such as Acidobacteria and Planctomycetes. This novel strategy has great potential to enable the isolation of a significant proportion of the peat bog bacterial diversity.