Pediococcus lolii DSM 19927T and JCM 15055T are strains of Pediococcus acidilactici

Abstract

Strain NGRI 0510QT , isolated from ryegrass silage, was recently classified as a representative of a novel Pediococcus species, Pediococcus lolii Doi et al. 2009. It was deposited in the DSMZ and JCM culture collections as DSM 19927 T and JCM 15055 T , respectively. A polyphasic taxonomic study, including matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, pheS and 16S rRNA gene sequence analysis, fluorescent amplified fragment length polymorphism and DNA-DNA hybridization, was used to prove that both subcultures of the type, and only, strain of this species are strains of Pediococcus acidilactici.In 2009, Doi and colleagues reported on a novel strain belonging to the genus Pediococcus, NGRI 0510QT , for which they proposed the name Pediococcus lolii (Doi et al., 2009). The description was based on a single strain isolated from ryegrass silage that was deposited in the DSMZ and JCM culture collections as DSM 19927 T and JCM 15055 T , respectively. Their study revealed that the strain exhibited distinct phenotypic characteristics, divergent sequences of the 16S rRNA gene and the 16S-23S rRNA intergenic spacer region, and low rates of DNA-DNA hybridization in comparison with the type strains of Pediococcus acidilactici DSM 20284 T (5LMG 11384 T ) and Pediococcus pentosaceus DSM 20336 T (5LMG 11488 T ).The present study was initiated upon analysis of the P. lolii type strain accessioned from the JCM culture collection, JCM 15055 T (5LMG 25667 T ), by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), which failed to discriminate P. lolii LMG 25667T from P. acidilactici strains. Subsequently, DSM 19927 T (5LMG 27029 T ) was accessioned from the DSMZ culture collection and the polyphasic taxonomic study described below was performed.All strains were grown on MRS agar (Oxoid) at 28 uC in anaerobic atmosphere, except strain P. acidilactici LMG 11384T , which was cultured in an aerobic atmosphere. Prior to MALDI-TOF MS analysis, strains were subcultured twice. Five to ten milligrams of wet cells were suspended in MilliQ-water comprising 75 % pure ethanol. Subsequently, formic acid and acetonitrile were added in a 1 : 1 (v/v) ratio to the bacterial cell pellet. After shaking vigorously, 1 ml supernatant (5the cell extract) was spotted onto a MALDI-TOF MS stainless steel target plate. Spots were overlaid with 1 ml matrix, which consisted of 5 mg acyano-4-hydroxycinnamic acid dissolved in 1 ml acetonitrile/trifluoroacetic acid/MilliQ-solvent (50 : 2 : 48). Prior to analysis, the mass spectrometer was externally calibrated using a peptide mixture of adrenocorticotropic hormone (fragment 18-39) (Sigma-Aldrich), insulin (Sigma-Aldrich), ubiquitin (Sigma-Aldrich), cytochrome C (Sigma-Aldrich) and myoglobin (Sigma-Aldrich). A 4800 Plus MALDI TOF/TOF Analyser (Applied Biosystems) was used in linear mode and covered a mass range of 2-20 kDa. The mass spectrometer used a 200 Hz frequency tripled Nd : YAG laser, operating at a wavelength of 355 nm. Generated ...

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“…Cell extraction for MALDI-TOF MS analysis was performed as described by Wieme et al (2012). Bacterial fingerprints were acquired using the 4800 Plus MALDI TOF/TOF Analyzer (Applied Biosystems) in linear positive ion mode as described previously (Wieme et al, 2012). For each spot, 50 sub-spectra for each of 40 randomized positions within the spot were collected and presented as one main spectrum.…”

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