Sphingomonas difficilis sp. nov., a difficultly cultivable bacterium that grows on solid but not in liquid medium, isolated from an abandoned lead–zinc mine

A difficult to cultivate bacterial strain, designated 1PNM-26 T , isolated from a lead-zinc mine, was investigated using a polyphasic taxonomic approach. The strain was able to grow on solid medium but not in liquid medium. Cells were Gram-reaction-negative, aerobic, non-spore-forming, non-motile and rod-shaped. It showed positive reactions for catalase and oxidase and hydrolysis of aesculin. The results of phylogenetic analyses based on 16S rRNA gene sequences indicated that strain 1PNM-26 T represents a member of the genus Sphingomonas and forms a stable cluster with Sphingomonas morindae KCTC 42183 T , Sphingomonas polyaromaticivorans JCM 16711 T and Sphingomonas oligoaromativorans NBRC 105508 T . The major fatty acids were summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c), summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c) and C16 : 0. C14 : 0 2-OH was present as the major hydroxyl fatty acid. The major polyamine was sym-homospermidine, and ubiquinone 10 (Q-10) was the predominant respiratory quinone. The genomic DNA G+C content of strain 1PNM-26 T was determined to be 66.3±0.3 mol%, and the polar lipids consisted of sphingoglycolipid, phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylcholine, phosphatidyldimethylethanolamine, phosphatidylmonomethylethanolamine, an unidentified glycolipid, three unidentified aminolipids and three unidentified lipids. The phenotypic, phylogenetic and chemotaxonomic results strongly supported the hypothesis that strain 1PNM-26 T represents a novel species of the genus Sphingomonas, for which the name Sphingomonasdifficilis sp. nov. is proposed. The type strain is 1PNM-26 T (=GDMCC 1.664 T =KCTC 42758 T =DSM 27573 T ).