Pseudomonas kielensis sp. nov. and Pseudomonas baltica sp. nov., isolated from raw milk in Germany

Abstract

In this study, nine Gram-negative, motile and rod-shaped bacteria were isolated during a Germany-wide investigation of raw milk microbiota. The strains could be differentiated from their closest relatives by phenotypic and chemotaxonomic characterization and average nucleotide identity (ANIb) values calculated from draft genome assemblies. Strains MBT-1T, MBT-8, MBT-9, MBT-10, MBT-11 and MBT-12 were related to the Pseudomonas chlororaphis subgroup. Isolates MBT-2T, MBT-13 and MBT-14 were closely related to Pseudomonas rhizosphaerae DSM 16299T with an ANIb of 88.2 % and a genome-to-genome distance result of 36.0 %. The G+C content of the DNA of strains MBT-1T and MBT-2T was 60.84 and 62.48 mol%, respectively. The major fatty acids were C16 : 1 ω7c (summed feature 3), C16 : 0 and C18 : 1 ω7c (summed feature 8). The strains were catalase-positive, while production of urease, β-galactosidase and indole were negative. Growth occurred at 4–30 °C and at pH values of pH 6.0–8.0. Based on these results, we conclude that the strains belong to two novel species, for which the names Pseudomonas kielensis sp. nov. and Pseudomonas baltica sp. nov. are proposed. The type strains are MBT-1T (=DSM 111668 T= LMG 31954T) and MBT-2T (=DSM 111761 T=LMG 31955T).

Pseudomonas rustica sp. nov., isolated from bulk tank raw milk at a German dairy farm

“…Genomic DNA was isolated using the ZR Fungal/ Bacterial DNA MiniPrep Kit (Zymo Research) according to the manufacturer's instructions. Library preparation was performed with the Illumina TruSeq DNA Nano Kit and DNA was sequenced on an Illumina MiSeq platform as described previously [13]. The draft genomes were assembled using Unicycler version 0.4.8 [14] and annotation was done by the NCBI Prokaryotic Genome Annotation Pipeline version 4.11 [15].…”

“…Due to the heat treatment used in dairy plants, Pseudomonas species are eliminated, but their exoenzymes, such as lipases and proteinases, are very heat-resistant and can therefore lead to quality loss in terms of extended shelf life [6]. Within a Germany-wide investigation of microbiological raw milk quality [13], approximately 815 strains, primary belonging to the genus Pseudomonas , were isolated and identified by the use of matrix-assisted laser desorption ionization/time-of-flight mass spectrometry (MALDI-TOF MS).…”

“…Milk-associated isolates were identified by MALDI-TOF MS analyses based on their species-specific protein fingerprints in the mass range of 2–20 kDa and in the positive mode. For this purpose, single colonies were extracted by the formic acid method as previously described [13]. Subsequently, 1 µl extract was transferred in duplicate to a steel target and, after drying, coated with 1 µl HCCA matrix containing 10 mg l −1 α-cyano-4-hydroxycinnamic acid dissolved in acetonitrile (50%), water (47.5%) and trifluoroacetic acid (2.5%).…”

Genome analysis reveals that the correct name of type strain Adlercreutzia caecicola DSM 22242T is Parvibacter caecicola Clavel et al. 2013

“…shows a phylogenetic tree inferred with FastME 2.1.6.1 [17] using the Genome blast Distance phylogeny (GBDP) approach calculated from genome sequences supplied by TYGS. For reasons of better visualisation, the tree was slightly modified using Inkscape 0.92.4.…”

“… Adlercreutzia caecicola DSM 22242 T is not clearly distinguishable on the basis of these phenotypic and biochemical results ( and S4). Therefore, MALDI-TOF MS analysis of all type strains of members of the genus Adlercreutzia was conducted as follows: Single colonies were prepared according to the formic acid extraction method recommended by Bruker Daltonik and 1 µl of the extract was spotted in duplicate onto a steel target [17]. After drying, the samples were coated with 1 µl HCCA matrix containing 10 mg l −1 α-cyano-4-hydroxycinnamic acid dissolved in acetonitrile (50%), water (47.5%) and trifluoroacetic acid (2.5%).…”