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
Authors: Lu Ling-Fei , Yang Yang , Zheng Lei , Zhang Rui , Liu Guang-Qian , Tu Ting-Yao , Xu Tao , Luo Xue , Ran Mao-Fang , Zhang Li-Qiang , Wang Song-Tao , Shen Cai-Hong , Zhang Ying-Gang 
Published: December 2021

A novel Gram-stain-positive, strictly anaerobic, elliptical, non-motile and non-flagellated bacterium, designed LZLJ-2T, was isolated from the mud in a fermentation cellar used for the production of Chinese Luzhou-flavour Baijiu. Growth occurred at 28–45 °C (optimum, 37 °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-2T belonged to the genus

Thermophilibacter

and was most closely related to

Thermophilibacter mediterraneus

Marseille-P3256T (similarity 96.9 %),

Olsenella gallinarum

ClaCZ62T (similarity 96.6 %) and

Thermophilibacter provencensis

Marseille-P2912T (similarity 96.4 %). In addition, strain LZLJ-2T had high similarity to the genus

Olsenella

, including

Olsenella profusa

DSM 13989T (similarity 94.9 %),

Olsenella umbonata

DSM 22620T (similarity 94.9 %),

Olsenella uli

ATCC 49627T (similarity 94.22 %),

Tractidigestivibacter scatoligenes

DSM 28304T (similarity 93.9 %) and

Paratractidigestivibacter faecalis

KCTC 15699T (similarity 93.25 %). Comparative genome analysis showed that orthoANI values between strain LZLJ-2T and

Thermophilibacter mediterraneus

Marseille-P3256T,

Olsenella gallinarum

ClaCZ62T,

Thermophilibacter provencensis

Marseille-P2912T,

Olsenella profusa

DSM 13989T,

Olsenella umbonata

DSM 22620T,

Olsenella uli

ATCC 49627T,

Tractidigestivibacter scatoligenes

DSM 28304T and

Paratractidigestivibacter faecalis

KCTC 15699T 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-2T was 65.21 mol%. The predominant cellular fatty acids (>10 %) of strain LZLJ-2T were C18 : 1
cis 9 (33.7 %), C14 : 0 (22.0 %) and C18 : 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-2T 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, α-glucosidase and β-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-2T 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-2T (=KCTC 25162T=JCM 34224T).