RESUMEN
Four methane-oxidizing bacteria, designated as strains WSC-6T, WSC-7T, SURF-1T, and SURF-2T, were isolated from Saddle Mountain Creek in southwestern Oklahoma, USA, and the Sanford Underground Research Facility (SURF) in Lead, South Dakota, USA. The strains were Gram-negative, motile, short rods that possessed intracytoplasmic membranes characteristic of type I methanotrophs. All four strains were oxidase-negative and weakly catalase-positive. Colonies ranged from pale pink to orange in colour. Methane and methanol were the only compounds that could serve as carbon and energy sources for growth. Strains WSC-6T and WSC-7T grew optimally at lower temperatures (25 and 20 °C, respectively) compared to strains SURF-1T and SURF-2T (40 °C). Strains WSC-6T and SURF-2T were neutrophilic (optimal pH of 7.5 and 7.3, respectively), while strains WSC-7T and SURF-1T were slightly alkaliphilic, with an optimal pH of 8.8. The strains grew best in media amended with ≤0.5% NaCl. The major cellular fatty acids were C14â:â0, C16â:â1 ω8c, C16â:â1 ω7c, and C16â:â1 ω5c. The DNA G+C content ranged from 51.5 to 56.0 mol%. Phylogenetic analyses indicated that the strains belonged to the genus Methylomonas, with each exhibiting 98.6-99.6% 16S rRNA gene sequence similarity to closely related strains. Genome-wide estimates of relatedness (84.5-88.4% average nucleotide identity, 85.8-92.4% average amino acid identity and 27.4-35.0% digital DNA-DNA hybridization) fell below established thresholds for species delineation. Based on these combined results, we propose to classify these strains as representing novel species of the genus Methylomonas, for which the names Methylomonas rivi (type strain WSC-6T=ATCC TSD-251T=DSM 112293T), Methylomonas rosea (type strain WSC-7T=ATCC TSD-252T=DSM 112281T), Methylomonas aurea (type strain SURF-1T=ATCC TSD-253T=DSM 112282T), and Methylomonas subterranea (type strain SURF-2T=ATCC TSD-254T=DSM 112283T) are proposed.
Asunto(s)
Técnicas de Tipificación Bacteriana , Composición de Base , ADN Bacteriano , Ácidos Grasos , Metano , Methylomonas , Filogenia , ARN Ribosómico 16S , Análisis de Secuencia de ADN , ARN Ribosómico 16S/genética , Metano/metabolismo , ADN Bacteriano/genética , Methylomonas/genética , Methylomonas/clasificación , Methylomonas/aislamiento & purificación , Oklahoma , Hibridación de Ácido Nucleico , Agua Dulce/microbiología , Oxidación-Reducción , Microbiología del SueloRESUMEN
A bacterial strain, designated NLS-7T, was isolated through enrichment of landfill cover soil in methane-oxidizing conditions. Strain NLS-7T is a Gram-stain negative, non-motile rod, approximately 0.8 µm wide by 1.3 µm long. Phylogenetic analysis based on 16S rRNA gene sequencing places it within the genus Methylocystis, with its closest relatives being M. hirsuta, M. silviterrae and M. rosea, with 99.9, 99.7 and 99.6â% sequence similarity respectively. However, average nucleotide identity and average amino acid identity values below the 95â% threshold compared to all the close relatives and digital DNA-DNA hybridization values between 20.9 and 54.1â% demonstrate that strain NLS-7T represents a novel species. Genome sequencing generated 4.31 million reads and genome assembly resulted in the generation of 244 contigs with a total assembly length of 3â820â957 bp (N50, 37â735 bp; L50, 34). Genome completeness is 99.5â% with 3.98â% contamination. It is capable of growth on methane and methanol. It grows optimally at 30â°C between pH 6.5 and 7.0. Strain NLS-7T is capable of atmospheric dinitrogen fixation and can use ammonium (as NH4Cl), l-aspartate, l-arginine, yeast extract, nitrate, l-leucine, l-proline, l-methionine, l-lysine and l-alanine as nitrogen sources. The major fatty acids are C18:1 ω8c and C18:1 ω7c. Based upon this polyphasic taxonomic study, strain NLS-7T represents a novel species of the genus Methylocystis, for which the name Methylocystis suflitae sp. nov. is proposed. The type strain is NLS-7T (=ATCC TSD-256T=DSM 112294T). The 16S rRNA gene and genome sequences of strain NLS-7T have been deposited in GenBank under accession numbers ON715489 and GCA_024448135.1, respectively.