*2.3. Chemotaxonomic Analysis*

The freeze-dried cells used for chemotaxonomic analysis were obtained from cultures grown in GY medium on a rotary shaker for seven days at 28 ◦C. Cells were acquired and washed twice with sterile distilled water and freeze-dried. The isomer of diaminopimelic acid (DAP) in the cell wall hydrolysates was derivatized and analyzed by HPLC (Agilent TC-C18 Column, 250 × 4.6 mm, i.d. 5 μm) with a mobile phase consisting of acetonitrile/phosphate buffer (0.05 mol·L−1, pH 7.2, 15:85, *v*/*v*), and a flow rate of 0.5 mL·min−<sup>1</sup> at a column temperature of 28 ◦C [24]. An Agilent G1321A fluorescence detector was used to detect the peak with a 365 nm excitation and 455 nm longpass emission filters. The whole-cell sugars were analyzed according to Lechevalier [25]. The polar lipids were extracted and examined by two-dimensional TLC (thin-layer chromatography, Qingdao Marine Chemical Inc., Qingdao, China) and identified according to the method of Minnikin et al. [26]. Menaquinones were extracted and purified from freeze-dried biomass following the methods of Collins [27]. The extracts were analyzed by HPLC-UV (Agilent Extend-C18 Column, 150 <sup>×</sup> 4.6 mm, i.d. 5 <sup>μ</sup>m, 1.0 mL·min−<sup>1</sup> acetonitrile: iso-propyl alcohol = 60:40) at 270 nm [28]. Fatty acid methyl esters were performed by GC-MS according to the method of Xiang et al. [29] and identified with the NIST 14 database.

#### *2.4. Phylogenetic Analysis*

Strain NEAU-H2<sup>T</sup> was grown on ISP 3 agar plates for one week at 28 ◦C. Then, it was inoculated into 250-mL baffle Erlenmeyer flasks containing 50 mL of GY broth and cultivated for two days at 28 ◦C with shaking at 250 rpm. After that, the total DNA was extracted according to the lysozyme-sodium dodecyl sulfate-phenol/chloroform method [30]. The primers and procedure for PCR amplification were carried out as described by Yi et al. [31]. The PCR product was purified and cloned into the vector pMD19-T (Takara, Shiga, Japan) and sequenced using an Applied Biosystems DNA sequencer (model 3730XL, Applied Biosystems Inc., Foster City, CA, USA). Almost full-length 16S rRNA gene sequence (1519 bp) was multiply aligned in MEGA (Molecular Evolutionary Genetics Analysis) using the Clustal W algorithm and trimmed manually if necessary. Phylogenetic trees were constructed with neighbor-joining [32] and maximum likelihood [33] algorithms using MEGA software version 7.0 (Kumar S, Philly, PA, USA) [34]. The stability of the topology of the phylogenetic tree was assessed using the bootstrap method with 1000 repetitions [35]. A distance matrix was calculated using Kimura's two-parameter model [36]. All positions containing gaps and missing data were eliminated from the dataset (complete deletion option). The calculation of 16S rRNA gene sequence similarities between strains was carried out on the basis of pairwise alignment using the EzBioCloud server (https://www.ezbiocloud.net/) [37]. Phylogenetic relationships of strain NEAU-H2T were also confirmed using sequences for five individual housekeeping genes (*atpD*, *gyrB*, *recA*, *rpoB*, and *trpB*). These sequences of housekeeping genes of strain NEAU-H2T were obtained from the Whole Genome sequences. The sequences of each locus were aligned using the software package MEGA version 7.0 and trimmed manually at the same position before being used for further analysis. Phylogenetic analysis was performed as described above.
