Genomic Insights into ANI-dDDH Relationships in Nocardiopsis and the Novel Species Nocardiopsis camelliae sp. nov

Average nucleotide identity (ANI) values of 95–96% and digital DNA–DNA hybridization (dDDH) values of 70% are currently regarded as the gold standards for bacterial species delineation. However, the accuracy and applicability of ANI thresholds may vary among genera and depend on genome quality. In this study, three genome quality standards were applied to re-evaluate ANI classification thresholds for the genus Nocardiopsis: >90% completeness and 90% completeness and 50% completeness and <10% contamination (40 strains, 780 genome pairs). Based on these analyses, we preliminarily estimated the species delineation thresholds for the genus Nocardiopsis as 96.68% for ANIm and 96.15% for ANIb, using the currently available genome datasets. Strain HUAS JQ3T was isolated from leaves of Camellia oleifera Abel collected in Taoyuan County, northwestern Hunan Province, China. Phylogenetic, genomic, morphological, cultural, and physiological analyses were conducted to determine its taxonomic status. Full-length 16S rRNA gene sequence analysis showed that strain HUAS JQ3T belongs to the genus Nocardiopsis and shares the highest sequence similarities with N. dassonvillei subsp. crassaminis D1T (100.00%), N. alborubida NBRC 13392T (99.86%), N. synnemataformans DSM 44143T (99.73%), and N. akebiae HDS 12T (99.73%). Phylogenetic analysis based on 16S rRNA gene sequences clustered strain HUAS JQ3T with N. alborubida, N. synnemataformans, and N. dassonvillei subsp. crassaminis. In contrast, whole-genome phylogenetic analysis indicated that strain HUAS JQ3T is most closely related to N. akebiae HDS 12T. Nevertheless, the ANIm/ANIb and dDDH values between strain HUAS JQ3T and N. akebiae HDS 12T were below the proposed species delineation thresholds of 70% dDDH and 96.68%/96.15% ANIm/b for the genus Nocardiopsis. In addition, strain HUAS JQ3T exhibited distinct morphological, cell culture, physiological, and biochemical characteristics compared with N. akebiae HDS 12T. The combined genotypic and phenotypic evidence gathered in this study demonstrates that strain HUAS JQ3T (= MCCC 1K08696T = JCM 36305T) represents a novel species of the genus Nocardiopsis, for which the name Nocardiopsis camelliae sp. nov. is proposed.