Reclassification of Olsenella gallinarum as Thermophilibacter gallinarum comb. nov. and description of Thermophilibacter immobilis sp. nov., isolated from the mud in a fermentation cellar used for the production of Chinese Luzhou-flavour Baijiu

A novel Gram-stain-positive, strictly anaerobic, elliptical, non-motile and non-flagellated bacterium, designed LZLJ-2(T), was isolated from the mud in a fermentation cellar used for the production of Chinese Luzhou-flavour Baijiu. Growth occurred at 28-45 degrees C (optimum, 37 degrees C), at pH 6.0-7.0 (optimum, pH 6.0) and with concentrations of NaCl up to 2 % (w/v; optimum, 0%). On the basis of 16S rRNA gene sequence similarity, strain LZLJ-2(T) belonged to the genus Thermophilibacter and was most closely related to Thermophilibacter mediterraneus Marseille-P3256(T) (similarity 96.9 %), Olsenella gallinarum ClaCZ62(T) (similarity 96.6%) and Thermophilibacter provencensis Marseille-P2912(T) (similarity 96.4%). In addition, strain LZLJ-2(T) had high similarity to the genus Olsenella, including Olsenella profusa DSM 13989(T) (similarity 94.9%), Olsenella umbonata DSM 22620(T) (similarity 94.9%), Olsenella uli ATCC 49627(T) (similarity 94.22%), Tractidigestivibacter scatoligenes DSM 28304(T) (similarity 93.9%) and Paratractidigestivibacter faecalis KCTC 15699(T) (similarity 93.25 %). Comparative genome analysis showed that orthoANI values between strain LZLJ-2(T) and Thermophilibacter mediterraneus Marseille-P3256(T), Olsenella gallinarum ClaCZ62(T), Thermophilibacter provencensis Marseille-P2912(T), Olsenella profusa DSM 13989T, Olsenella umbonata DSM 22620T, Olsenella uli ATCC 49627T, Tractidigestivibacter scatoligenes DSM 28304(T) and Paratractidigestivibacter faecalis KCTC 15699(T) were 78.68, 78.99, 78.29, 73.40, 74.00, 74.30, 75.08 and 77.23 %, and the genome-to-genome distance values were respectively 22.3, 22.5, 22.4, 19.6, 20.5, 19.7, 20.5 and 21.5%. The genomic DNA G+C content of strain LZLJ-2(T) was 65.21 mol%. The predominant cellular fatty acids (>10%) of strain LZLJ-2(T) were C(18:1)cis 9 (33.7%), C-14:0 (22.0%) and C(18:1)cis 9 DMA (13.5%). d-Glucose, sucrose, mannose, maltose, lactose (weak), salicin, glycerol (weak), cellobiose and trehalose (weak) could be used by strain LZLJ-2(T) as sole carbon sources. Enzyme activity results showed positive reactions with valine arylamidase, leucine arylamidase, crystine arylamidase, acid phosphatase, alkaline phosphatase, esterase (C4) (weakly positive), naphthol-AS-BI-phosphohydrolase, a-glucosidase and beta-glucosidase. The major end products of glucose fermentation were lactic acid and acetic acid. It produced skatole from indole acetic acid, and produced p-cresol from modified peptone-yeast extract medium with glucose. Based on the 16S rRNA gene trees as well as the genome core gene tree, it is suggested that Olsenella gallinarum are transferred to genus Thermophilibacter as Thermophilibacter gallinarum comb. nov. Based on phenotypic, genotypic and phylogenetic data, strain LZLJ-2(T) is considered to represent a novel species of the genus Thermophilibacter, for which the name Thermophilibacter immobilis sp. nov. is proposed. The type strain is LZLJ-2(T) (=KCTC 25162(T)=JCM 34224(T)).