Lacticaseibacillus parahuelsenbergensis sp. nov., Lacticaseibacillus styriensis sp. nov. and Lacticaseibacillus zeae subsp. silagei subsp. nov., isolated from different grass and corn silage

Four rod- shaped, non- motile, non- spore- forming, facultative anaerobic, Gram- stain- positive lactic acid bacteria, designated as EB0058T, SCR0080, LD0937T and SCR0063T, were isolated from different corn and grass silage samples. The isolated strains were characterized using a polyphasic approach and EB0058T and SCR0080 were identified as Lacticaseibacillus zeae by 16S rRNA gene sequence analysis. Based on whole- genome sequence- based characterization, EB0058T and SCR0080 were separated into a distinct clade from Lacticaseibacillus zeae DSM 20178T, together with CECT9104 and UD2202, whose genomic sequences are available from NCBI GenBank. The average nucleotide identity (ANI) values within the new subgroup are 99.9 % and the digital DNA-DNA hybridization (dDDH) values are 99.3-99.9 %, respectively. In contrast, comparison of the new subgroup with publicly available genomic sequences of L. zeae strains, including the type strain DSM 20178T, revealed dDDH values of 70.2-72.5 % and ANI values of 96.2-96.6 %. Based on their chemotaxonomic, phenotypic and phylogenetic characteristics, EB0058T and SCR0080 represent a new subspecies of L. zeae. The name Lacticaseibacillus zeae subsp. silagei subsp. nov. is proposed with the type strain EB0058T (=DSM 116376T=NCIMB 15474T). According to the results of 16S rRNA gene sequencing, LD0937T and SCR0063T are members of the Lacticaseibacillus group. The dDDH value between the isolates LD0937T and SCR0063T was 67.6 %, which is below the species threshold of 70 %, clearly showing that these two isolates belong to different species. For both strains, whole genome- sequencing revealed that the closest relatives within the Lacticaseibacillus group were Lacticaseibacillus huelsenbergensis DSM 115425 (dDDH 66.5 and 65.9%) and Lacticaseibacillus casei DSM 20011T (dDDH 64.1 and 64.9 %). Based on the genomic, chemotaxonomic and morphological data obtained in this study, two novel species, Lacticaseibacillus parahuelsenbergensis sp. nov. and Lacticaseibacillus styriensis sp. nov. are proposed and the type strains are LD0937T (=DSM 116105T=NCIMB 15471T) and SCR0063T (=DSM 116297T=NCIMB 15473T), respectively.