Microbacterium paulum sp. nov., isolated from microfiltered milk

Abstract

A novel Gram-stain-positive, strictly aerobic, short rod-shaped bacterium, designated 2CT, was isolated from freshly packaged microfiltered milk. This strain was able to grow within the NaCl concentration range of 0–5 % (w/v), temperature range of 8–37 °C (optimally at 30 °C) and at pH 6.0–10.0. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain 2CT was closely related to species of the genus Microbacterium , with the highest sequence similarity (99.2 %) to Microbacterium lacticum DSM 20427T as well as Microbacterium flavum DSM 18909T (=YM18-098T). The phylogenetic tree based on 16S rRNA genes showed that strain 2CT clustered with M. flavum DSM 18909T. However, the phylogenetic tree based on concatenated 16S rRNA and four housekeeping genes showed that strain 2CT clustered with M. lacticum DSM 20427T. Furthermore, the phylogenomic tree showed that strain 2CT clustered with M. lacticum DSM 20427T and M. flavum DSM 18909T. The major respiratory quinones were MK-10, MK-11 and MK-12. The predominant cellular fatty acids were anteiso-C15 : 0, iso-C16 : 0 and anteiso-C17 : 0. The polar lipid composition of strain 2CT consisted of diphosphatidylglycerol, phosphatidylglycerol, three unidentified glycolipids and two unidentified lipids. The cell-wall peptidoglycan type was a variant of B1α {Gly} [l-Lys] d-Glu-l-Lys, with the amino acids lysine, glycine, alanine and glutamic acid. The whole-cell sugars consisted of galactose, glucose, ribose and minor amounts of rhamnose. In addition, strain 2CT showed a glycolyl-type cell wall. The genomic DNA G+C content was 69.8mol%, while the average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values with the closely related Microbacterium species were below the recognized thresholds of 95–96 % ANI and 70 % DDH for species definition. Based on the phenotypic and genotypic data, strain 2CT (=LMG 32277T=CECT 30329T) is considered to represent a new species, for which the name Microbacterium paulum sp. nov. is proposed.

Four new Microbacterium species isolated from seaweeds and reclassification of five Microbacterium species with a proposal of Paramicrobacterium gen. nov. under a genome-based framework of the genus Microbacterium

“…In the core genome-based phylogenetic tree ( Figure 2 ), all the Microbacterium strains were resolved into nine major clusters (clusters I–IX including ≥three type strains), and eight minor clusters (clusters X–XVII with two type strains) were defined mostly at GSI of 92 and mean intra-cluster OrthoANIu of >80.00% ( Table 2 ). The topologies of the nine major clusters were maintained irrespective of the addition/deletion of the related sequences and also supported in the core genome analyses reported previously (Dong et al, 2020 ; Bellassi et al, 2021 ; Tian et al, 2021 ). The origin of isolation for each type strain included in the clusters is given in Supplementary Table S3 .…”

“…Cluster II ( Microbacterium lacticum clade) contains seven species, namely, M. aurum (Yokota et al, 1993a ), M. dextranolyticum (Yokota et al, 1993a ), Microbacterium flavum (Kageyama et al, 2007a ), Microbacterium hominis (Takeuchi and Hatano, 1998a ) , Microbacterium laevaniformans (Dias and Bhat, 1962 ; Collins et al, 1983 ), M. lacticum (Orla-Jensen, 1919 ; Approved Lists, 1980), and Microbacterium paulum (Bellassi et al, 2021 ), and revealed intra-cluster OrthoANIu values≥79.20% (mean, 81.04%) and DNA G+C contents of 69.4–71.0% ( Table 2 ). Most of the type strains were also found as a single cluster in a previous core genome analysis (Bellassi et al, 2021 ), albeit that many of the strains formed well-separated sublines in the 16S rRNA gene trees (Dong et al, 2020 ; Lee and Kim, 2023 ; Figure 1 ).…”

“…Subcluster Va contains the type strains of M. algeriense, M. liquefaciens, M. maritypicum , and M. oxydans together with three study strains, KSW4-10 T , KSW4-16, and SSW1-7, having intra-subcluster OrthoANIu values of 81.40–87.46% (mean, 84.87%) and DNA G+C contents of 68.0–69.6%; subcluster Vb is composed of the type strain of M. paraoxydans and three study strains, KSW4-4, SSW1-47 T , and SSW1-51, with intra-subcluster OrthoANIu values of 82.67–84.51% (mean, 84.02%) and DNA G+C contents of 70.0–71.1%; subcluster Vc encompasses the five type strains of M. luteolum, M. saperdae, M. foliorum , and M. phyllosphaerae (Behrendt et al, 2001 ), and M. hydrocarbonoxydans (Schippers et al, 2005 ), and one study strain, SSW1-49 T , sharing intra-subcluster OrthoANIu values of 81.18–85.21% (mean, 82.23%) and DNA G+C contents of 68.0–68.9%; and subcluster Vd includes only the study strains, KSW2-24 T , KSW4-6, and SSW1-36, with intra-subcluster OrthoANIu values of 97.92–98.42% (mean, 98.09%) and DNA G+C contents of 69.2–69.4% ( Table 2 ). Among them, the six or seven type strains were associated together in the previous analyses of core genomes (Bellassi et al, 2021 ; Tian et al, 2021 ; Xie et al, 2021 ). In addition, most of the type strains were closely associated with one another in the 16S rRNA gene trees, while three species ( M. foliorum and M. phyllosphaerae , and M. hydrocarbonoxydans ) formed well-separated sublines from the other type strains (Dong et al, 2020 ; Lee and Kim, 2023 ; Figure 1 ).…”

“…Members of the genus contain different types of peptidoglycan with D-ornithine, L-lysine, or diaminobutyric acid as the diamino acid of the cell wall peptidoglycan (Suzuki and Hamada, 2012 ; Fidalgo et al, 2016 ) and can be readily differentiated from other genera of the family Microbacteriaceae using a combination of morphological and chemotaxonomic features (Evtushenko, 2012 ) as well as genome-based phylogeny (Nouioui et al, 2018 ). The intra-genus phylogenomic relationships have been partly studied, for < 50 species, through the descriptions of new species (Dong et al, 2020 ; Bellassi et al, 2021 ; Tian et al, 2021 ; Xie et al, 2021 ) or in the whole-genome-based taxonomic classification of the phylum Actinobacteria (Nouioui et al, 2018 ). In the recent descriptions of two new species from larvae of insects (Lee and Kim, 2023 ), the genomes of 81 Microbacterium strains were included in a core genome-based phylogeny.…”

Microbacterium abyssi sp. nov. and Microbacterium limosum sp. nov., two new species of the genus Microbacterium, isolated from deep-sea sediment samples

“…MK, menaquinone; PG, phosphatidylglycerol; DPG, diphosphatidylglycerol; PE, phosphatidylethanolamine; GL, unidentified glycolipids; PL, unidentified phospholipid; L, unidentified lipid. Data were from this study, except for cell morphology, menaquinones, polar lipids and DNA G+C contents of the reference strains[3,8,[35][36][37][38].…”