Actinoplanes aureus sp. nov., a novel protease-producing actinobacterium isolated from soil

Abstract

A novel protease-producing actinobacterium, designated strain NEAU-A11^T, was isolated from soil collected from Aohan banner, Chifeng, Inner Mongolia Autonomous Region, China, and characterised using a polyphasic approach. The hydrolytic enzymes, such as proteases, played critical roles in destruction of fungi by degrading the protein linkages to disrupt integrity in the cell wall. This suggested that the isolate could be a good biocontrol candidate against pathogens to control fungal diseases. On the basis of 16S rRNA gene sequence analysis, strain NEAU-A11^T was indicated to belong to the genus Actinoplanes and was most closely related to Actinoplanes rectilineatus JCM 3194 ^T (98.9%). Cell walls contained meso-diaminopimelic acid as the diagnostic diamino acid and the whole-cell sugars were arabinose, xylose and glucose. The phospholipid profile contained diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylinositol and two phosphatidylinositol mannosides. The predominant menaquinones were MK-9(H₄), MK-9(H₆) and MK-9(H₈). The major fatty acids were C_18:0, C_16:0, C_18:1 ω9c, C_17:0 and C_15:0. Genome sequencing revealed a genome size of 10,742,096 bp, a G + C content of 70.5% and 9,514 protein-coding genes (CDS), including 102 genes coding for protease. Moreover, Genome analysis showed that strain NEAU-A11^T contained 255 glycoside hydrolases (GHs), 152 glycosyl transferases (GTs), 40 carbohydrate esterases (CEs), 26 polysaccharide lyases (PLs), and 12 auxiliary activities (AAs) genes. Genome mining analysis using antiSMASH 5.0 led to the identification of 20 putative gene clusters responsible for the production of diverse secondary metabolites. Phylogenetic analysis using the 16S rRNA gene sequences showed that the strain formed a stable clade with A. rectilineatus JCM 3194 ^T in the genus Actinoplanes. Whole-genome phylogeny showed strain NEAU-A11^T formed a stable phyletic line with Actinoplanes lutulentus DSM 45883 ^T (97.6%). However, whole-genome average nucleotide identity value between strain NEAU-A11^T and its reference strains A. rectilineatus JCM 3194 ^T and A. lutulentus DSM 45883 ^T were found to be 81.1% and 81.6%, respectively. The levels of digital DNA-DNA hybridization between them were 24.6% (22.2-27.0%) and 24.8% (22.5-27.3%), respectively. The values were well below the criteria for species delineation of 70% for dDDH and 95-96% for ANI, suggesting that the isolate differed genetically from its closely related type strain. The content of G + C in genomic DNA was 70.5%, within the range of 67-76%. In addition, evidences from phenotypic, chemotaxonomic and genotypic studies indicated that strain NEAU-A11^T represents a novel species of the genus Actinoplanes, for which the name Actinoplanes aureus sp. nov. is proposed, with NEAU-A11^T (= CCTCC AA 2019063 ^T = JCM 33971 ^T) as the type strain.

Keywords: 16S rRNA gene; Actinoplanes aureus sp. nov; Genome; Polyphasic analysis.