Isolation and identification of lactic acid bacteria from sediments of a coastal marsh using a differential selective medium

Abstract

Aims: To identify lactic acid bacteria (LAB) colonies isolated from sediments of a coastal marsh by the reduction of 2,3,5‐triphenyltetrazolium chloride (TTC) in MRS medium. Methods and Results: Single colonies isolated from sediments of a coastal marsh by enrichment in MRS broth were selected from MRS‐TTC plates and classified according to colony phenotype based on TTC reduction. A total of 37 colonies grouped in seven different phenotypes were identified by analysis of its 16S ribosomal gene sequence. Most isolates belonged to the Firmicutes phylum, mainly to orders Bacillales and Lactobacillales. LAB were represented by 20 isolates, 15 of which belong to the genus Weissella. Conclusions: Enrichment in MRS was highly selective for the isolation of bacteria belonging to phylum Firmicutes. Several different phenotypes were developed by LAB and must be considered during LAB isolation based on TTC reduction. Significance and Impact of the Study: To our knowledge, this is the first study aimed at determining a relationship between colony phenotype from TTC reduction and a partial identification of isolates based on 16S ribosomal gene sequence similarities. Besides, this is the first report of isolation of W. cibaria from environmental samples.

Diversity and Antimicrobial Properties of Lactic Acid Bacteria Isolated from Rhizosphere of Olive Trees and Desert Truffles of Tunisia

“…Moreover, the isolation of bacteria belonging to the genus Weissella was already reported either from soil [19] or plants [17]. With regard to others studies on LAB recovery from soil [19, 35, 47, 48] a higher number of species diversity is recorded in olive tree and truffle rhizospheric samples such as Lb. sakei, Pc.…”

Genotypic Diversity, Antibiotic Resistance and Bacteriocin Production of Enterococci Isolated from Rhizospheres

“…Although the major part of the country is characterized by a sub-arid climate, the northern regions differ mainly by darker soil and more abundant rainfall than in the southern parts. Enterococci were isolated by the enrichment method according to the procedure of Zamudio-Maya et al (47), with some modifications. Briefly, 1 g sample was resuspended in 5 mL MRS broth and incubated at 30°C for 2–3 days.…”

Purification of bacterial genomic DNA in less than 20 min using chelex-100 microwave: examples from strains of lactic acid bacteria isolated from soil samples

“…After TCC reduction, these strains were identified based on their LAB indicative phenotype (i.e., colors): pale pink; deep pink; pink with red center; and clear with a red or white center (Zamudio-Maya et al 2008). …”

“…All sediment bacterial strains used in the study were provided by the Jalisco State Center for Research and Technological and Design Assistance (Centro de Investigación y Asistencia en Tecnología y Diseño del Estado de Jalisco-CIATEJ, Unidad Sureste) (Table 1), and had been previously isolated using TCC reagent (Zamudio-Maya et al 2008). The LAB strains were grown in Man, Rogosa and Sharpe medium (MRS, Difco) (De Vuyst and Leroy 2007) for 16 h at 37°C under anaerobic conditions.…”

“…Identification is made more selective by adding 2,3,5-triphenyltetrazolium chloride (TTC) to the culture medium, because when it is reduced by LAB it generates pink and red colors that differ from undesired white colonies (Zamudio-Maya et al 2008). Other methods for LAB presence determination and species identification-without involving bacterial population isolation-is metagenome extraction and specific genes-identification, e.g., 16S ribosomal DNA (16S rDNA) (Luna et al 2006;Mohania et al 2008;Nocker et al 2007).…”