Phylogenomic and comparative genomic analyses of Leuconostocaceae species: identification of molecular signatures specific for the genera Leuconostoc, Fructobacillus and Oenococcus and proposal for a novel genus Periweissella gen. nov.

Abstract

The genera Convivina , Fructobacillus , Leuconostoc , Oenococcus and Weissella , which formed the family Leuconostocaceae , have recently been merged within the family Lactobacillaceae . Using genome sequences for 47 of the 52 named species from these genera, we report here comprehensive phylogenomic and comparative analyses on protein sequences from these species using multiple approaches. In a phylogenomic tree based on concatenated sequences of 498 core proteins from these five genera, and in a 16S rRNA gene tree, members of the genera Fructobacillus , Leuconostoc and Oenococcus formed distinct strongly supported clades. In contrast, Weissella species grouped into two distinct unrelated clades designated as the ‘ Weissella main clade’ and ‘ Weissella clade 2’. The presence of these clades is also seen in a matrix of pairwise average amino acid identity based on core protein sequences. In parallel, comparative genomic studies on protein sequences from Leuconostocaceae genomes have identified 46 conserved signature indels (CSIs) in diverse proteins that are unique characteristics of the different observed species clades. Of these identified CSIs, five, five and 13 CSIs are uniquely present in members of the genera Fructobacillus , Leuconostoc and Oenococcus , respectively. We also report here six and five CSIs that are exclusively present in the species from the Weissella main clade and Weissella clade 2, respectively, providing independent evidence supporting their distinctness from each other. The remaining 12 identified CSIs are commonly shared by some or all of the species from the genera Convivina , Fructobacillus and Leuconostoc , clarifying their interrelationships. The identified CSIs provide novel and reliable means for the identification/circumscription of members of the genera Fructobacillus , Leuconostoc and Oenococcus as well as the two Weissella species clades in molecular terms. Based on the strong phylogenetic and molecular evidence presented here, we propose that the genus Weissella be limited to only the species from the Weissell a main clade, whereas the species forming Weissella clade 2 should be transferred to a new genus Periweissella gen. nov.

Phylogenomic analysis of the genus Leuconostoc

“…The genomes were inspected for completeness and contamination with CheckM (Parks et al, 2015 ). Genomes were also checked for the presence of the molecular signatures characterizing the genus Leuconostoc (Bello et al, 2022 ). The genomes were annotated with Prokka (Seemann, 2014 ) to calculate the pangenome using Roary (Page et al, 2015 ), with the minimum percentage identity parameter set at 80%.…”

“…The present project aims to investigate the evolutionary relationships within the genus Leuconostoc through a phylogenomic approach and to verify the consistency of phylogenetic relationships with the current taxonomy. This genus holds a group of Lactic Acid Bacteria (LAB) belonging, together with the genera Convivina, Fructobacillus, Oenococcus, and Weissella, to the family of Leuconostocaceae (www.bacterio.net; Nieminen et al, 2014;Bello et al, 2022), in its turn included in the order Lactobacillales, based on the average amino acid identity of core proteins (cAAI) (Zheng et al, 2020). The genus Leuconostoc encompasses 17 species according to the List of Prokaryotic names with Standing in Nomenclature (https://lpsn.dsmz.de/genus/leuconostoc) (Parte et al, 2020).…”

Identification of Leuconostoc species based on novel marker genes identified using real-time PCR via computational pangenome analysis

“…This genus has undergone several reclassifications. While some species that were originally classified under the genus Leuconostoc were reclassified as Oenococcus, Fructobacillus, and Weissella, others have been considered heterotypic synonyms (Björkroth et al, 2014;Bello et al, 2022). Most recently, the previously reported Leu.…”

“…Whole-genome sequencing is widely used to determine the taxonomic position of microorganisms (Kumar et al, 2022). Recent comparative genomic studies have confirmed that the average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) might suffice to classify Leuconostoc species at the species or subspecies level (Wu and Gu, 2021;Bello et al, 2022). Also, more robust approaches (e.g., pan-and core-genome analysis) were used to account for strain diversity and classify closely related species or subspecies (Kim et al, 2021(Kim et al, , 2022b.…”

Metataxonomic insights in the distribution ofLactobacillaceaein foods and food environments

“…The most recent iteration of FoodMicrobionet uses SILVA v138.1 as a taxonomic reference database (https://www.arb-silva.de/documentation/release-1381/ last accessed in July 2022). This allows the taxonomic assignment of Amplicon Sequence Variants (ASV) to the genera proposed by Zheng et al (2020) but does not allow the taxonomic assignment for the new genera Convivina and Periweissella (Bello et al, 2022; Praet et al, 2015). FoodMicrobionet includes studies which have used a large variety of target regions within the 16S gene, although those using V1-V3, V3-V4 and V4 represent the vast majority.…”