Description of two novel non-pathogenic tomato-associated Clavibacter species: Clavibacter seminis sp. nov. and Clavibacter quasicaliforniensis sp. nov.

ABSTRACT The Clavibacter genus is currently split into ten species, including C. michiganensis , C. nebraskensis , C. capsici , C. sepedonicus , C. tessellarius , C. insidiosus , C. californiensis , C. phaseoli , C. zhangzhiyongii , and the newly described C. lycopersici . These bacterial species have been isolated from different hosts, and some members are reported as non-pathogenic. In this study, we have sequenced and reported high-quality complete genomes of three new Clavibacter strains A6099 T , A4868, and A6308 isolated from tomato using Oxford nanopore MinION and Illumina NovaSeq sequencing platforms. In addition, the taxonomy position of other three Clavibacter strains from the NCBI GenBank database LMG 26808, CFBP 7493, and VKM Ac-22542 were assessed and properly defined. Application of different in silico approaches such as calculation of the average nucleotide identity (ANI), digital DNA-DNA hybridization (dDDH), alignment coverage (AP), multi-locus sequence analysis (MLSA) of eight housekeeping genes atpD , dnaA , dnaK , pgi , gyrB , ppk , recA and rpoB , and a phylogenomic analysis of 808 core genes of 97 Clavibacter strains, including the three sequenced novel Clavibacter strains, led us to differentiate two novel clades within the Clavibacter genus that are closely related to C. michiganensis and C. californiensis . The ANI and dDDH values were lower than the 96 and 70% cut-off values for species delineation, respectively; likewise, the phylogenies showed a clear differentiation of these two novel clades with respect to the other Clavibacter species. Therefore, we suggest establishing two new species to correctly classify these two novel clades, for which we propose the species names: Clavibacter seminis sp. nov. formed by the type strain A6099 T , LMG 26808 and CFBP 7493, and Clavibacter quasicaliforniensis sp. nov., composed of the type strain A4868 T , A6308, and CFBP 7493. The two new species were non-pathogenic on tomato seedlings, their isolation host, which correlated with the absence of the major virulence-associated genes determined in our comparative genomics analyses.