RESUMEN

UNLABELLED: Spontaneous fermented sourdoughs prepared from amaranth flour were investigated for the presence of autochthonous lactic acid bacteria (LAB) predominating microbiota. The doughs were fermented with daily backslopping on a laboratory scale at 30°C for 10 days. LAB counts ranged from 2·60 to 8·54 log CFU g(-1) with a pH declined from 6·2 to 3·8 throughout fermentation. The combined use of randomly amplified polymorphic DNA (RAPD)-PCR analysis and sequence analysis of 16S rRNA was applied for LAB intraspecies differentiation and taxonomic identification, respectively. Enterococcus, Pediococcus and Lactobacillus species were present in amaranth sourdoughs (AS). After the first refreshment step, Lactobacillus plantarum dominated AS until the end of fermentation. In coincidence, when DGGE analysis was performed, the occurrence of a progressive change in bacterial communities allowed the selection of Lact. plantarum as a dominant species. Moreover, technological, functional and safety characteristics of representative RAPD-biotypes were investigated. Lact. plantarum CRL1898 was selected as a potential candidate for gluten-free amaranth sourdough starter. SIGNIFICANCE AND IMPACT OF THE STUDY: Nowadays, there is an increasing interest in ancient noncereal gluten-free (GF) crops such as amaranth, due to their reported nutritional and health benefits. However, the use of these grains is still limited to traditional foods and bread making processes that are not yet well standardized. Results on the dynamics of autochthonous lactic acid bacteria (LAB) microbiota during laboratory spontaneous amaranth sourdoughs (AS) fermentation will contribute to overcome challenges for GF-fermented products development. In addition, knowledge about LAB diversity involving Enterococcus, Pediococcus and Lactobacillus species, with Lactobacillus plantarum predominating during AS fermentation, and their technological and functional properties provides the basis for the selection of autochthonous strains as starters cultures for novel gluten-free bakery products with enhanced nutritional, sensory and/or safety quality.

Asunto(s)

Amaranthus/microbiología , Enterococcus/clasificación , Harina/microbiología , Lactobacillus plantarum/clasificación , Pediococcus/clasificación , Técnicas de Tipificación Bacteriana , Biodiversidad , Reactores Biológicos/clasificación , Reactores Biológicos/microbiología , Pan/microbiología , Dieta Sin Gluten , Enterococcus/aislamiento & purificación , Enterococcus/metabolismo , Fermentación , Microbiología de Alimentos , Ácido Láctico/metabolismo , Lactobacillus plantarum/aislamiento & purificación , Lactobacillus plantarum/metabolismo , Microbiota/genética , Pediococcus/aislamiento & purificación , Pediococcus/metabolismo , ARN Ribosómico 16S/genética , Técnica del ADN Polimorfo Amplificado Aleatorio

RESUMEN

A lactic-acid producing bacterium was isolated from the rumen of lambs with rumen acidosis. The cells were gram-positive, nonmotile, nonsporing, catalase negative spherical, 1.5-2.0 µm in diameter, and occur in pairs and tetrads. Analysis of 16S ribosomal RNA indicated that the rumen bacterium was a strain of Pediococcus acidilactici with 99% of nucleotide homology. This bacterium was sensible to monensin and lasalocid at the unique dose tested of 300 ppm. The concentration of lactic acid and DM degradation decreased (P<0.05) when monensin or lasalocid were added to the culture media after 24, 48 and 72 h of incubation. In contrast, total VFA concentration and pH were higher (P<0.05) in the culture media added with the ionophores. Up to now S. bovis is considered the main ruminal bacterium related with rumen acidosis, but the importance of P. acidilactici should be also reconsidered in experimental studies focused on the control rumen acidosis.

Asunto(s)

Acidosis/veterinaria , Lasalocido/farmacología , Monensina/farmacología , Pediococcus/aislamiento & purificación , Rumen/microbiología , Enfermedades de las Ovejas/inducido químicamente , Alimentación Animal/análisis , Alimentación Animal/toxicidad , Animales , Antibacterianos/farmacología , Dieta/veterinaria , Carbohidratos de la Dieta/análisis , Carbohidratos de la Dieta/toxicidad , Resistencia a Medicamentos , Historia del Siglo XVI , Concentración de Iones de Hidrógeno , Ionóforos/farmacología , Masculino , Pediococcus/clasificación , Pediococcus/efectos de los fármacos , Pediococcus/genética , Filogenia , ARN Ribosómico 16S/genética , Ovinos

RESUMEN

We established a Chelex 100-Microwave method for the purification of bacterial genomic DNA (gDNA) in less than 20 min with high yield and good quality, useful for multiple purposes. It combines Chelex 100, proteinase K, RNase A and heating in a microwave oven. The resulting gDNA was used directly to identify bacterial species of the Order Lactobacillales by means of PCR amplification of their 16S rDNA gene, isolated from sediments on the Yucatan Peninsula, Mexico. This method produced gDNA free of phenolic and protein residual contaminants from 100 of these isolated bacteria. 16S rDNA amplification and sequencing showed Pediococcus acidilactici to prevail in inland lagoons, and Pediococcus pentosaceus, Lactobacillus plantarum, Lactobacillus sp., and Lactobacillus fermentum to be most abundant in the soils of livestock farms. The combination of Chelex 100, enzymes and microwave heating used in the Chelex 100-Microwave method produced large amounts of highly pure gDNA from Gram-positive and Gram-negative bacteria, in less than 20 min.

Asunto(s)

ADN Bacteriano/aislamiento & purificación , Sedimentos Geológicos/microbiología , Lactobacillus/genética , Lactobacillus/aislamiento & purificación , Pediococcus/genética , Pediococcus/aislamiento & purificación , Microbiología del Suelo , ADN Bacteriano/análisis , ADN Ribosómico/genética , Endopeptidasa K , Genoma Bacteriano , Ácido Láctico/metabolismo , Lactobacillus/clasificación , México , Microondas , Pediococcus/clasificación , Reacción en Cadena de la Polimerasa , ARN Ribosómico 16S/genética , Resinas Sintéticas , Ribonucleasa Pancreática , Análisis de Secuencia de ADN , Humedales

RESUMEN

Mezcal is an alcoholic beverage obtained from the distillation of fermented juices of cooked Agave spp. plant stalks (agave must), and each region in Mexico with denomination of origin uses defined Agave species to prepare mezcal with unique organoleptic characteristics. During fermentation to produce mezcal in the state of Tamaulipas, not only alcohol-producing yeasts are involved, but also a lactic acid bacterial community that has not been characterized yet. In order to address this lack of knowledge on this traditional Mexican beverage, we performed a DGGE-16S rRNA analysis of the lactic acid bacterial diversity and metabolite accumulation during the fermentation of a typical agave must that is rustically produced in San Carlos County (Tamaulipas, Mexico). The analysis of metabolite production indicated a short but important malolactic fermentation stage not previously described for mezcal. The denaturing gradient gel electrophoresis (DGGE) analysis of the 16S rRNA genes showed a distinctive lactic acid bacterial community composed mainly of Pediococcus parvulus, Lactobacillus brevis, Lactobacillus composti, Lactobacillus parabuchneri, and Lactobacillus plantarum. Some atypical genera such as Weissella and Bacillus were also found in the residual must. Our results suggest that the lactic acid bacteria could strongly be implicated in the organoleptic attributes of this traditional Mexican distilled beverage.

Asunto(s)

Agave/microbiología , Bebidas Alcohólicas/microbiología , Bacillus/aislamiento & purificación , Ácido Láctico/metabolismo , Lactobacillales/aislamiento & purificación , Bacillus/clasificación , Bacillus/genética , Bacillus/metabolismo , Secuencia de Bases , Dermatoglifia del ADN , ADN Ribosómico/análisis , Electroforesis en Gel de Gradiente Desnaturalizante , Fermentación , Microbiología de Alimentos , Lactobacillales/clasificación , Lactobacillales/genética , Lactobacillales/metabolismo , Lactobacillus/clasificación , Lactobacillus/genética , Lactobacillus/aislamiento & purificación , Lactobacillus/metabolismo , Levilactobacillus brevis/clasificación , Levilactobacillus brevis/genética , Levilactobacillus brevis/aislamiento & purificación , Levilactobacillus brevis/metabolismo , Lactobacillus plantarum/clasificación , Lactobacillus plantarum/genética , Lactobacillus plantarum/aislamiento & purificación , Lactobacillus plantarum/metabolismo , México , Pediococcus/clasificación , Pediococcus/genética , Pediococcus/aislamiento & purificación , Pediococcus/metabolismo , Filogenia , Reacción en Cadena de la Polimerasa , ARN Bacteriano/genética , ARN Ribosómico 16S/análisis , Weissella/clasificación , Weissella/genética , Weissella/aislamiento & purificación , Weissella/metabolismo

RESUMEN

A Gram-positive, small coccus-shaped lactic acid bacterium, strain LMG 23999(T), was isolated from Argentinean wheat flour. 16S rRNA gene sequence analysis revealed that the phylogenetic position of the novel strain was within the genus Pediococcus, with Pediococcus stilesii, Pediococcus pentosaceus and Pediococcus acidilactici as its closest relatives (97.7, 97.3 and 96.9 % gene sequence similarity, respectively). Fluorescent amplified fragment length polymorphism fingerprinting of whole genomes and whole-cell protein electrophoresis confirmed the unique taxonomic status of the novel strain. DNA-DNA hybridizations, DNA G+C content determination, comparative sequence analysis of the pheS, rpoA and atpA genes and physiological and biochemical characterization demonstrated that strain LMG 23999(T) (=CCUG 54535(T)=CRL 776(T)) represents a novel species for which the name Pediococcus argentinicus sp. nov. is proposed. Multi-locus sequence analysis based on pheS, rpoA and atpA genes was found to be a suitable method for the identification of species of the genus Pediococcus.

Asunto(s)

Harina/microbiología , Genes Bacterianos/genética , Pediococcus/clasificación , Pediococcus/genética , Argentina , Fermentación , Datos de Secuencia Molecular , Pediococcus/fisiología , Filogenia , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADN , Especificidad de la Especie , Triticum/microbiología

RESUMEN

Seventy-two strains of pediococci isolated from human clinical sources were characterized by conventional physiological tests, chromogenic enzymatic tests, analysis of whole-cell protein profiles (WCPP) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and analysis of chromosomal DNA restriction profiles by pulsed-field gel electrophoresis (PFGE). Conventional tests allowed identification of 67 isolates: 52 strains were identified as Pediococcus acidilactici, 15 strains were identified as Pediococcus pentosaceus, and 5 strains were not identified because of atypical reactions. Analysis of WCPP identified all isolates since each species had a unique WCPP. By the WCPP method, the atypical strains were identified as P. acidilactici (two strains) and P. pentosaceus (three strains). The chromogenic substrate test with o-nitrophenyl-beta-D-glucopyranoside differentiated all 54 strains of P. acidilactici (negative reactions) and 13 (72%) of 18 strains of P. pentosaceus (positive reactions). Isolates of both species were shown to be nonclonal as revealed by the genetic diversity when chromosomal DNA was analyzed by PFGE. Using WCPP as the definitive identification procedure, P. acidilactici (28 of 54 strains; 51.8%) was more likely than P. pentosaceus (4 of 18 strains; 22.3%) to be isolated from blood cultures.

Asunto(s)

Técnicas de Tipificación Bacteriana , Infecciones por Bacterias Grampositivas/microbiología , Pediococcus/clasificación , Pediococcus/genética , Compuestos Cromogénicos/metabolismo , ADN Bacteriano/análisis , Electroforesis en Gel de Campo Pulsado , Electroforesis en Gel de Poliacrilamida , Genotipo , Humanos , Pediococcus/aislamiento & purificación , Pediococcus/fisiología , Fenotipo

RESUMEN

Lactic acid bacteria and more particularly lactobacilli and Leuconostoc, are widely found in a wide variety of traditional fermented foods of tropical countries, made with cereals, tubers, meat or fish. These products represent a source of bacterial diversity that cannot be accurately analysed using classical phenotypic and biochemical tests. In the present work, the identification and the molecular diversity of lactic acid bacteria isolated from cassava sour starch fermentation were assessed by using a combination of complementary molecular methods: Randomly Amplified Polymorphic DNA fingerprinting (RAPD), plasmid profiling, hybridization using rRNA phylogenetic probes and partial 16S rDNA sequencing. The results revealed a large diversity of bacterial species (Lb. manihotivorans, Lb. plantarum, Lb. casei, Lb. hilgardii, Lb. buchneri, Lb. fermentum, Ln. mesenteroides and Pediococcus sp.). However, the most frequently isolated species were Lb. plantarum and Lb. manihotivorans. The RAPD analysis revealed a large molecular diversity between Lb. manihotivorans or Lb. plantarum strains. These results, observed on a rather limited number of samples, reveal that significant bacterial diversity is generated in traditional cassava sour starch fermentations. We propose that the presence of the amylolytic Lb. manihotivorans strains could have a role in sour starch processing.

Asunto(s)

Microbiología de Alimentos , Ácido Láctico , Lactobacillus/genética , Manihot/microbiología , Pediococcus/genética , ADN Ribosómico , Fermentación , Variación Genética , Lactobacillus/clasificación , Datos de Secuencia Molecular , Pediococcus/clasificación , Filogenia , Plásmidos/genética , ARN Ribosómico 16S , Técnica del ADN Polimorfo Amplificado Aleatorio , Almidón