Analysis of electrophoretic whole-cell protein profiles as a tool for characterization ofEnterococcus species

Merquior, Vânia Lúcia Carreira; Peralta, José Mauro; Facklam, Richard R.; Teixeira, Lúcia Martins

doi:10.1007/bf01571056

Cited by 52 publications

(46 citation statements)

References 27 publications

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“…The classification of the genus Enterococcus has undergone considerable changes as a consequence of the increase in the number of novel species and also improvements in the methods used to discriminate separate species (Baele et al, 2000;Merquior et al, 1994;Naser et al, 2005). At the time of writing, 34 Enterococcus species names have been validly published (Euzéby, 1997; last full update May 14, 2008).…”

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confidence: 99%

Enterococcus thailandicus sp. nov., isolated from fermented sausage ('mum') in Thailand

Tanasupawat

Sukontasing

Lee

2008

INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLO

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A Gram-positive and catalase-negative coccus that formed chains, designated strain FP48-3 T , isolated from fermented sausage ('mum'), was studied systematically. Strain FP48-3 T was facultatively anaerobic and produced L-lactic acid from glucose. Straight-chain fatty acids C 18 : 1 and C 16 : 0 were the dominant components. The DNA G+C content of strain FP48-3 T was 37.9 mol%. On the basis of 16S rRNA and RNA polymerase a-subunit (rpoA) gene sequence analysis, strain FP48-3 T was closely related to Enterococcus hirae LMG 6399 T , Enterococcus durans LMG 10746 T and Enterococcus faecium LMG 11423 T , with 99.3-99.6 and 95.1-96.9 % sequence similarities, respectively. Strain FP48-3 T could be clearly distinguished from E. hirae LMG 6399 T , E. durans LMG 10746 T and E. faecium LMG 11423 T by low DNA-DNA relatedness (¡14 %) and phenotypic characteristics. Therefore, this strain represents a novel species of the genus Enterococcus, for which the name Enterococcus thailandicus sp. nov. is proposed. The type strain is FP48-3 T (5KCTC 13134 T 5NBRC 101867 T 5NRIC 0107 T 5TISTR 933 T 5PCU 282 T ).Enterococci have been isolated from plants, soil, animals, humans and raw and prepared food including meat, poultry, fish, egg, milk, cheese and vegetables and traditional fermented foods (Knudtson & Hartman, 1992; Sukontasing et al., 2007). The classification of the genus Enterococcus has undergone considerable changes as a consequence of the increase in the number of novel species and also improvements in the methods used to discriminate separate species (Baele et al., 2000;Merquior et al., 1994;Naser et al., 2005). At the time of writing, 34 Enterococcus species names have been validly published (Euzéby, 1997; last full update May 14, 2008). Several species groups, Enterococcus faecium, E. avium, E. gallinarum, E. cecorum, E. faecalis and other species groups, including E. saccharolyticus, E. sulfureus and E. dispar, were revealed based on comparative 16S rRNA gene sequence analysis (Hardie & Whiley, 1997; Švec et al., 2006). Here we describe a novel bacterium, strain FP48-3 T , isolated from fermented sausage ('mum'), and propose that it represents a novel species, based on phenotypic and chemotaxonomic characteristics, DNA-DNA relatedness and 16S rRNA and RNA polymerase a-subunit (rpoA) gene sequence analysis.Samples of fermented sausage were collected from Khonkaen province, north-eastern Thailand. Cocci arranged in chains were isolated from the samples using GYP-CaCO 3 agar (Tanasupawat et al., 1992). Cultures grown using GYP-sodium acetate-mineral salt broth (Tanasupawat et al., 1992), adjusted to pH 7.2, were used for working cultures. All tests were performed by incubating the cultures at 30 u C. Cell shape, size, arrangement and colony appearance were examined using cells grown on GYP agar for 3 days. Gram staining was done as described by Hucker & Conn (1923). Spore formation was examined using the Gram-stained specimen. Results of the oxidation-fermentation test and motility were examined in soft agar (Whittenbur...

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confidence: 99%

Enterococcus thailandicus sp. nov., isolated from fermented sausage ('mum') in Thailand

Tanasupawat

Sukontasing

Lee

2008

INTERNATIONAL JOURNAL OF SYSTEMATIC AND EVOLUTIONARY MICROBIOLO

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“…Extract preparation and whole-cell protein profile analysis using one-dimensional SDS-PAGE were performed as described previously (Merquior et al, 1994). Dice indexes were determined for each isolate by using the Molecular Analyst Fingerprint Plus Software Package Version 1.6 (Bio-Rad) and a dendrogram was constructed by using the UPGMA algorithm.…”

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confidence: 99%

Vagococcus carniphilus sp. nov., isolated from ground beef

Shewmaker

Steigerwalt

Morey

et al. 2004

International Journal of Systematic and Evolutionary Microbiology

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Nine enterococcus-like strains were referred to the Streptococcus Laboratory at the Centers for Disease Control and Prevention (CDC) for further identification from the National Antimicrobial Resistance Monitoring System Laboratory at the CDC. The cultures were isolated from ground beef purchased from retail in Oregon in 2000. Conventional biochemical testing and analysis of whole-cell protein electrophoretic profiles distinguished these strains from known species of enterococci and vagococci. Comparative 16S rRNA gene sequencing studies revealed that these strains were most closely related to the species Vagococcus fluvialis. DNA–DNA reassociation studies confirmed that these nine strains represented a new taxon. The relative binding ratio was 87 % or greater at the optimal temperature, and the divergence was less than 1 % for strains hybridized against the isolate designated the type strain. DNA–DNA relatedness was 25 % to V. fluvialis and 9 % or less to the other three species of Vagococcus. DNA–DNA relatedness was 33 % or less to the 25 currently described species of Enterococcus. On the basis of this evidence, it is proposed that these strains be classified as Vagococcus carniphilus sp. nov. The type strain of V. carniphilus is 1843-02T (=ATCC BAA-640T=CCUG 46823T). The clinical significance (if any) of these strains is yet to be determined.

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“…Analysis of purified 221 preparations was carried out by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) [12]. After electrophoresis, gels were either silver stained [13] or bioassayed by a modification of the method of [14].…”

Section: Sds-page and Bioassaymentioning

confidence: 99%

Puriﬁcation and Characterization of 221, the Cationic Peptide that Inhibits Staphylococcus aureus

Mahrous¹,

Burnie²,

Tagg³

et al. 2017

JMEN

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Submit Manuscript | http://medcraveonline.com treat staphylococcal infections has risen to an alarming level [1,2]. A contributing factor has been the widespread use of antibiotics to treat non-life-threatening infections. In recent years, there has been much focus on a promising new class of bacteriocins known as lantibiotics [3,4].Bacteriocins are proteinaceous substances produced by certain bacteria, which display growth-inhibitory activity against a range of other bacteria belonging to the same or closely-related species. Bacteriocins of gram-negative bacteria (colisins) have been extensively characterized, resulting in comprehensive knowledge of the genetic basis, domain structure, mode of formation, and killing action of these molecules [5,6]. On the other hand, the bacteriocins of gram-positive bacteria have been classified into numerous subtypes [7][8][9]. The first class of bacteriocins (lantibiotics) is distinguishable from other antibiotics because of their polypeptide nature and bioactive properties. We here investigate the administration of the novel cationic peptide antibiotic (221) as a future opportunity for treating staphylococcal infections. Material and Methods Bacterial strainsS. epidermidis 221 was (The University of Manchester, Clinical Microbiology Department Library Strains, Strain no. 221) maintained on Columbia blood agar but was propagated in TSAYca (30g of Oxoid Trypticase soy powder per liter, 20g of Bacto yeast extract per liter, 5g of Bacto Calcium carbonate per liter, 10g of Oxoid Biological agar) at 37 o C, then 221 inhibitor was extracted by freeze-thaw method on which 221 strain was grown as a lawn under appropriate incubation conditions. After incubation, the plates were frozen at -70 o C for 3-4 hr, and then thawed at room temperature. The frozen agar exudes liquid as it thaws. This was collected, clarified by centrifugation at 15,764g for 5 min and tested for inhibitory activity by the spot or well diffusion assay.Staphylococcal core indicator strains (Table 1) were used in subsequent assays to test for S. aureus susceptibility to activity of 221 strains and used as representative of sensitive, moderate and highly resistant species to synthetic antibiotics. These strains were clinical isolates selected from a wide range of S. aureus library with the exception of an M. leutus strain, which was a reference isolate used commonly as a sensitive biological indicator of BLIS [10]. Bacteriocin assayBacteriocin production was determined as described previously by either agar well diffusion assay or spot-on-lawn assay [11]. purificationAn overnight culture of S. epidermidis 221 was used to inoculate TSAYca media. The culture was incubated at 37°C, and the cells were then removed using a sterile spatula. The free bacterial cells plate culture was then frozen at -70 o C for 3-4h, and then thawed at room temperature. The liquor was then collected and centrifuged at 15,764g for 15min. The liquor was then filter-sterilized by passage through a 0.45-mm-pore-size membrane filter.Solid (NH 4...

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Analysis of electrophoretic whole-cell protein profiles as a tool for characterization ofEnterococcus species

Cited by 52 publications

References 27 publications

Enterococcus thailandicus sp. nov., isolated from fermented sausage ('mum') in Thailand

Enterococcus thailandicus sp. nov., isolated from fermented sausage ('mum') in Thailand

Vagococcus carniphilus sp. nov., isolated from ground beef

Puriﬁcation and Characterization of 221, the Cationic Peptide that Inhibits Staphylococcus aureus

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