Methylocapsa acidiphila gen. nov., sp. nov., a novel methane-oxidizing and dinitrogen-fixing acidophilic bacterium from Sphagnum bog

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A novel species, Methylocella tundrae, is proposed for three methanotrophic strains (T4 T , TCh1 and TY1) isolated from acidic Sphagnum tundra peatlands. These strains are aerobic, Gram-negative, non-motile, dinitrogen-fixing rods that possess a soluble methane monooxygenase and utilize the serine pathway for carbon assimilation. Strains T4

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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.

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The terminal oxidases of the respiratory chain of seven strains of gram-negative bacteria were shown to be involved in the reduction of tellurite. The rate of tellurite reduction correlated with the intensity of respiration. The inhibitors of terminal oxidases, carbon monoxide and cyanide, inhibited the reduction of tellurite. In Pseudomonas aeruginosa PAO ML4262 and P. aeruginosa PAO ML4262 (pBS 10), the respiratory chain was found to contain three types of cytochrome c, one of which (the carbon monoxide-binding cytochrome c) was involved in the reduction of tellurite. Agrobacterium tumefaciens VKM B-1219, P. aeruginosa IBPM B-13, and Escherichia coli G0-102bd++ cells contained oxidases aa3, bb3, and bd, respectively. The respiratory chain of other strains contained two oxidases: E. coli DH5alpha of bb3- and bd-type, and Erwinia carotovora VKM B-567 of bo3- and bd-type. All the strains under study reduced tellurite with the formation of tellurium crystallites. Depending on the position of the active center of terminal oxidases in the plasma membrane, the crystallites appeared either in the periplasmic space [P. aeruginosa PAO ML4262 and P. aeruginosa PAO ML4262 (pBS10)], or on the outer surface of the membrane (A. tumefaciens VKM B-1219 and P. aeruginosa IBPM B-13), its inner surface (E. coli G0-102bd++), or on both surfaces (E. coli DHaalpha and E. carotovora VKM B-567).

Bryobacter aggregatus gen. nov., sp. nov., a peat-inhabiting, aerobic chemo-organotroph from subdivision 3 of the Acidobacteria MPL1011 and MOB76 were acidotolerant, mesophilic organisms capable of growth at pH 4.5-7.2 and between 4 and 33 6C (optimum growth at pH 5.5-6.5 and 22-28 6C). The preferred growth substrates were sugars, some heteropolysaccharides and galacturonic and glucuronic acids, which are released during decomposition of Sphagnum moss. The major fatty acids were iso-C 15 : 0 , C 16 : 0 and summed feature 3 (iso-C 15 : 0 2-OH and/or C 16 : 1 v7c); the major quinones were MK-9 and MK-10. The DNA G+C content was 55.5-56.5 mol%. Strains MPL3 T , MPL1011 and MOB76 possessed nearly identical 16S rRNA gene sequences and belonged to the phylum Acidobacteria. They represent the first taxonomically characterized members of acidobacterial subdivision 3 and display only 81.7-86.7 % 16S rRNA gene sequence similarity to other members of the Acidobacteria with validly published names. Therefore, strains MPL3 T , MPL1011 and MOB76 are classified as representatives of a novel species in a new genus, for which the name Bryobacter aggregatus gen. nov., sp. nov. is proposed; strain MPL3 T (5ATCCis the type strain of Bryobacter aggregatus.The Acidobacteria is one of the globally distributed but little understood phylogenetic groups of the domain Bacteria Recently, we showed that members of the Acidobacteria are a characteristic component of the microbial community in Sphagnum-dominated acidic wetlands (Dedysh et al., 2006;Pankratov et al., 2008). Using low-nutrient media and fluorescent in situ hybridization-mediated monitoring of the isolation procedure, several peat-inhabiting acidobacteria were obtained in pure culture. Three of these strains, MPL3T , MPL1011 and MOB76, belonged to subdivision 3 of the Acidobacteria, for which no representatives with validly published names are known. Here, the characteristics of our novel isolates are described and a novel species in a new genus is proposed to accommodate these bacteria.The GenBank/EMBL/DDBJ accession numbers for the 16S rRNA gene sequences of Bryobacter aggregatus strains MPL3 T , MPL1011 and MOB76 are AM162405, AM887761 and AM887762, respectively.