Assessment of bacterial diversity in selected Philippine fermented food products through PCR-DGGE

2011 ◽  
Vol 2 (4) ◽  
pp. 273-281 ◽  
Author(s):  
L.M. Dalmacio ◽  
A.K. Angeles ◽  
L.L. Larcia ◽  
M. Balolong ◽  
R. Estacio
Keyword(s):  
Bacterial Diversity ◽  
16S Rdna ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Acetic Acid Bacteria ◽  
Food Samples ◽  
Fermented Food ◽  
Probiotic Bacteria ◽  
Rrna Gene ◽  
Fermented Foods ◽  
16S Rdna Pcr

The bacterial population in several Philippine fermented food preparations was assessed by PCR-denaturing gradient gel electrophoresis (PCR-DGGE) of the 16S rRNA gene (16S rDNA). Genomic DNA was isolated directly from alamang (fermented shrimp paste), burong isda (fermented fish and rice), burong hipon (fermented shrimp and rice), burong mustasa (fermented mustard leaves), tuba (sugar cane wine), suka (vinegar) and sinamak (spiced vinegar) using one of two protocols, namely – MoBio DNA Extraction Kit procedure and a cetyltrimethylammonium bromide-based method. Samples recalcitrant to both methods underwent enrichment in three culture broths prior to DNA isolation. Isolated DNA was amplified using nested primer pairs targeting the bacterial 16S rDNA. PCR products were subjected to DGGE to elucidate the bacterial diversity in each fermented food. 16S rDNA sequence analyses revealed that lactic acid bacteria (LAB) and acetic acid bacteria (AAB) were dominant in the food samples. The LAB identified were Lactobacillus fermentum, Lactobacillus plantarum, Lactobacillus panis, Lactobacillus pontis and Weissella cibaria. Identified AAB were Acetobacter pomorum, Acetobacter ghanensis, Acetobacter orientalis, and Acetobacter pasteurianus. Among these, L. fermentum, L. plantarum and W. cibaria are established probiotic bacteria, while L. panis and L. pontis are potential probiotic bacteria. This finding would increase the appeal and significance of local fermented foods to consumers. Furthermore, the majority of the identified bacteria in the study have not been reported before in culture-dependent studies of similar food preparations. As such, some of the bacterial 16S rDNA obtained were cloned to have an initial partial bacterial 16S rDNA library for Philippine fermented foods.

Bacterial diversity in Philippine fermented mustard (burong mustasa) as revealed by 16S rRNA gene analysis

2011 ◽  
Vol 2 (4) ◽  
pp. 263-271 ◽  
Author(s):  
L.L. Larcia II ◽  
R. Estacio ◽  
L.M. Dalmacio
Keyword(s):  
Bacterial Diversity ◽  
16S Rdna ◽  
Nested Pcr ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Independent Study ◽  
Vector System ◽  
Rrna Gene ◽  
Bacterial Identification ◽  
16S Rrna Gene Analysis ◽  
Cloned Gene

Previous studies on the bacterial profile of burong mustasa, a traditional Philippine fermented food, had been conducted using culture-dependent techniques. Since these methods may underestimate the total microbiota of a sample, a culture-independent study was done to determine the bacterial diversity in burong mustasa through molecular biology techniques. Bacterial DNA was isolated from fermented mustard samples at different stages of fermentation. The isolated genomic DNA was amplified by PCR using specific primers for the 16S ribosomal RNA gene (16S rDNA). The 1.5 kb amplicons obtained were subjected to nested PCR using primers for the internal variable region of the 16S rDNA. The 585 bp nested PCR amplicons were then subjected to denaturing gradient gel electrophoresis (DGGE) to separate the different bacteria present in each sample. Distinct and unique bands in the DGGE profile were excised, reamplified, purified and sequenced for bacterial identification. Molecular cloning of the 1.5 kb 16S rDNA was also performed using the pGEM-T Easy Vector System. The cloned gene was sequenced for bacterial identification. The identified microbiota in burong mustasa at different stages of fermentation include lactic acid bacteria and several uncultured bacteria (initial up to the final stages); acetic acid bacteria (middle stage); and Streptobacillus and Fusobacterium species (initial stage). The potential probiotic bacteria found in burong mustasa are Weissella and Lactobacillus.

scholarly journals Molecular Diversity Assessment of Plant Growth Promoting Rhizobacteria Using Denaturing Gradient Gel Electrophoresis (DGGE) of 16s rRNA Gene

2017 ◽  
Vol 5 (1) ◽  
pp. 72-80
Author(s):  
Umesh Prasad Shrivastava
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
16S Rdna ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Uttar Pradesh ◽  
Plant Growth Promoting Rhizobacteria ◽  
Melting Behavior ◽  
Rrna Gene ◽  
Dgge Profile ◽  
Diversity Assessment

The rhizobacteria were isolated from rhizosphere of rice plant of different fields of 4 districts of Nepal and 5 districts of Bihar and Uttar Pradesh, adjoining states of India with Nepal. The DGGE analysis was performed for diversity analysis. For the construction of dendrogram, 16S rRNA gene was amplified by two different sets of primers. The DGGE ladder consisting of PCR amplified products of nine pure bacterial cultures were obtained. The first DGGE ladder was prepared by 400 bp fragment of 16S rDNA with GC clamp and the second DGGE ladder was prepared with 200 bp fragment of 16S rDNA with GC clamp. The perpendicular DGGE of these amplicons based on their melting behavior clearly demonstrated separation of different isolates. The 16S rDNA fragment amplified with primer set of V2-V3 regions with GC clamp showed separation between 40-60% of denaturant. The DGGE profile based on primer set F352T and 519r for various bacteria present in soil samples of 5 districts of India and 4 districts of Nepal revealed that the number of bands which might be specific for diazotrophic isolates varied from 2 to 11. The dendrogram constructed based on DGGE profile of various samples of 5 districts of India and 4 districts of Nepal showed that all the samples could be clustered in nine groups with 58-96% similarity to each other. Among all these 37 samples, only Var-4 and Var-5 showed 100% similarity, no other samples from any site showed 100% similarity. Int. J. Appl. Sci. Biotechnol. Vol 5(1): 72-80

scholarly journals Changes in Bacterial Diversity Associated with Epithelial Tissue in the Beef Cow Rumen during the Transition to a High-Grain Diet

2011 ◽  
Vol 77 (16) ◽  
pp. 5770-5781 ◽  
Author(s):  
Yanhong Chen ◽  
Gregory B. Penner ◽  
Meiju Li ◽  
Masahito Oba ◽  
Le Luo Guan
Keyword(s):  
Bacterial Diversity ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Pcr Analysis ◽  
Control Group ◽  
Epithelial Tissue ◽  
Rrna Gene ◽  
Content Type ◽  
Beef Cow ◽  
Dietary Transition ◽  
Gradient Gel Electrophoresis

ABSTRACTOur understanding of the ruminal epithelial tissue-associated bacterial (defined as epimural bacteria in this study) community is limited. In this study, we aimed to determine whether diet influences the diversity of the epimural bacterial community in the bovine rumen. Twenty-four beef heifers were randomly assigned to either a rapid grain adaptation (RGA) treatment (n= 18) in which the heifers were allowed to adapt from a diet containing 97% hay to a diet containing 8% hay over 29 days or to the control group (n= 6), which was fed 97% hay. Rumen papillae were collected when the heifers were fed 97%, 25%, and 8% hay diets. PCR-denaturing gradient gel electrophoresis (DGGE) and quantitative real-time PCR analysis were used to characterize rumen epimural bacterial diversity and to estimate the total epimural bacterial population (copy numbers of the 16S rRNA gene). The epimural bacterial diversity from RGA heifers changed (P= 0.01) in response to the rapid dietary transition, whereas it was not affected in control heifers. A total of 88 PCR-DGGE bands were detected, and 44 were identified from phyla includingFirmicutes,Bacteroidetes, andProteobacteria. The bacteriaTreponemasp.,Ruminobactersp., andLachnospiraceaesp. were detected only when heifers were fed 25% and 8% hay diets, suggesting the presence of these bacteria is the result of adaptation to the high-grain diets. In addition, the total estimated population of rumen epimural bacteria was positively correlated with molar proportions of acetate, isobutyrate, and isovalerate, suggesting that they may play a role in volatile fatty acid metabolism in the rumen.

PCR-DGGE Analysis of Bacterial Diversity of Intestinal System in Hyperlipidemia Rats

2013 ◽  
Vol 726-731 ◽  
pp. 898-901
Author(s):  
Ri Na Wu ◽  
Xiao Meng Pang ◽  
Xi Yan Wang ◽  
Jun Rui Wu
Keyword(s):  
Bacterial Diversity ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Intestinal Flora ◽  
Intestinal Bacteria ◽  
Control Group ◽  
Rrna Gene ◽  
Dgge Analysis ◽  
Gradient Gel Electrophoresis ◽  
The Relationship ◽  
Insight Into

Denaturing gradient gel electrophoresis (DGGE) analysis of 16S rRNA gene has been regarded as one of powerful tools for gaining insight into the bacterial diversity of intestinal system. In the present study, hyperlipidemia model was constructed in rat according to the tests of blood lipids. Fecal samples of rats were collected after 60d feeding, and DGGE was used to investigate the diversities of intestinal bacteria in the artificially-induced hyperlipidemia rats and normal rats. The results showed that two patterns had similarities, but there were also some different bacteria communities. Moreover, control group had much more bands than model group on gel, showing species in intestinal of model rats might be deduced by hyperlipidemia. It will be helpful to explore the relationship between hyperlipidemia and intestinal flora.

scholarly journals Dynamics and Biodiversity of Populations of Lactic Acid Bacteria and Acetic Acid Bacteria Involved in Spontaneous Heap Fermentation of Cocoa Beans in Ghana

2007 ◽  
Vol 73 (6) ◽  
pp. 1809-1824 ◽  
Author(s):  
Nicholas Camu ◽  
Tom De Winter ◽  
Kristof Verbrugghe ◽  
Ilse Cleenwerck ◽  
Peter Vandamme ◽  
...  
Keyword(s):  
Acetic Acid ◽  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Fermentation Process ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Acetic Acid Bacteria ◽  
Cocoa Bean ◽  
Rrna Gene ◽  
Cocoa Bean Fermentation ◽  
Leuconostoc Pseudomesenteroides

ABSTRACT The Ghanaian cocoa bean heap fermentation process was studied through a multiphasic approach, encompassing both microbiological and metabolite target analyses. A culture-dependent (plating and incubation, followed by repetitive-sequence-based PCR analyses of picked-up colonies) and culture-independent (denaturing gradient gel electrophoresis [DGGE] of 16S rRNA gene amplicons, PCR-DGGE) approach revealed a limited biodiversity and targeted population dynamics of both lactic acid bacteria (LAB) and acetic acid bacteria (AAB) during fermentation. Four main clusters were identified among the LAB isolated: Lactobacillus plantarum, Lactobacillus fermentum, Leuconostoc pseudomesenteroides, and Enterococcus casseliflavus. Other taxa encompassed, for instance, Weissella. Only four clusters were found among the AAB identified: Acetobacter pasteurianus, Acetobacter syzygii-like bacteria, and two small clusters of Acetobacter tropicalis-like bacteria. Particular strains of L. plantarum, L. fermentum, and A. pasteurianus, originating from the environment, were well adapted to the environmental conditions prevailing during Ghanaian cocoa bean heap fermentation and apparently played a significant role in the cocoa bean fermentation process. Yeasts produced ethanol from sugars, and LAB produced lactic acid, acetic acid, ethanol, and mannitol from sugars and/or citrate. Whereas L. plantarum strains were abundant in the beginning of the fermentation, L. fermentum strains converted fructose into mannitol upon prolonged fermentation. A. pasteurianus grew on ethanol, mannitol, and lactate and converted ethanol into acetic acid. A newly proposed Weissella sp., referred to as “Weissella ghanaensis,” was detected through PCR-DGGE analysis in some of the fermentations and was only occasionally picked up through culture-based isolation. Two new species of Acetobacter were found as well, namely, the species tentatively named“ Acetobacter senegalensis” (A. tropicalis-like) and “Acetobacter ghanaensis” (A. syzygii-like).

scholarly journals Polyphasic Study of the Spatial Distribution of Microorganisms in Mexican Pozol, a Fermented Maize Dough, Demonstrates the Need for Cultivation-Independent Methods To Investigate Traditional Fermentations

1999 ◽  
Vol 65 (12) ◽  
pp. 5464-5473 ◽  
Author(s):  
Frédéric Ampe ◽  
Nabil ben Omar ◽  
Claire Moizan ◽  
Carmen Wacher ◽  
Jean-Pierre Guyot
Keyword(s):  
Lactic Acid ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Fermented Food ◽  
Fermented Foods ◽  
Product Analysis ◽  
Culture Independent ◽  
Gradient Gel Electrophoresis ◽  
Traditional Fermented Foods

ABSTRACT The distribution of microorganisms in pozol balls, a fermented maize dough, was investigated by a polyphasic approach in which we used both culture-dependent and culture-independent methods, including microbial enumeration, fermentation product analysis, quantification of microbial taxa with 16S rRNA-targeted oligonucleotide probes, determination of microbial fingerprints by denaturing gradient gel electrophoresis (DGGE), and 16S ribosomal DNA gene sequencing. Our results demonstrate that DGGE fingerprinting and rRNA quantification should allow workers to precisely and rapidly characterize the microbial assemblage in a spontaneous lactic acid fermented food. Lactic acid bacteria (LAB) accounted for 90 to 97% of the total active microflora; no streptococci were isolated, although members of the genus Streptococcus accounted for 25 to 50% of the microflora. Lactobacillus plantarum and Lactobacillus fermentum, together with members of the generaLeuconostoc and Weissella, were the other dominant organisms. The overall activity was more important at the periphery of a ball, where eucaryotes, enterobacteria, and bacterial exopolysacharide producers developed. Our results also showed that the metabolism of heterofermentative LAB was influenced in situ by the distribution of the LAB in the pozol ball, whereas homolactic fermentation was controlled primarily by sugar limitation. We propose that starch is first degraded by amylases from LAB and that the resulting sugars, together with the lactate produced, allow a secondary flora to develop in the presence of oxygen. Our results strongly suggest that cultivation-independent methods should be used to study traditional fermented foods.

scholarly journals Potential of Bacteria from Alternative Fermented Foods as Starter Cultures for the Production of Wheat Sourdoughs

2020 ◽  
Vol 8 (10) ◽  
pp. 1534
Author(s):  
Andrea Comasio ◽  
Simon Van Kerrebroeck ◽  
Henning Harth ◽  
Fabienne Verté ◽  
Luc De Vuyst
Keyword(s):  
Starter Culture ◽  
Gluconobacter Oxydans ◽  
Acetic Acid Bacteria ◽  
Fermented Food ◽  
Starter Cultures ◽  
Fermented Foods ◽  
Water Mixture ◽  
Coagulase Negative Staphylococci ◽  
Fermented Sausage ◽  
Different Strains

Microbial strains for starter culture-initiated sourdough productions are commonly isolated from a fermenting flour–water mixture. Yet, starter culture strains isolated from matrices other than sourdoughs could provide the dough with interesting metabolic properties and hence change the organoleptic properties of the concomitant breads. Furthermore, the selection of sourdough starter cultures does not need to be limited to lactic acid bacteria (LAB), as other food-grade microorganisms are sometimes found in sourdoughs. Therefore, different strains belonging to LAB, acetic acid bacteria (AAB), and coagulase-negative staphylococci (CNS) that originated from different fermented food matrices (fermenting cocoa pulp-bean mass, fermented sausage, and water kefir), were examined as to their prevalence in a wheat sourdough ecosystem during 72-h fermentations. Limosilactobacillus fermentum IMDO 222 (fermented cocoa pulp-bean mass isolate) and Latilactobacillus sakei CTC 494 (fermented sausage isolate) seemed to be promising candidates as sourdough starter culture strains, as were the AAB strains Acetobacter pasteurianus IMDO 386B and Gluconobacter oxydans IMDO A845 (both isolated from fermented cocoa pulp-bean mass), due to their competitiveness in the wheat flour-water mixtures. Wheat breads made with G. oxydans IMDO A845 sourdoughs were significantly darker than reference wheat breads.

Bacterial diversity associated with subalpine fir (Abies lasiocarpa) ectomycorrhizae following wildfire and salvage-logging in central British Columbia

2002 ◽  
Vol 48 (7) ◽  
pp. 611-625 ◽  
Author(s):  
Madhukar B Khetmalas ◽  
Keith N Egger ◽  
Hugues B Massicotte ◽  
Linda E Tackaberry ◽  
M Jill Clapperton
Keyword(s):  
Bacterial Diversity ◽  
16S Rdna ◽  
Bacterial Communities ◽  
Restriction Analysis ◽  
Rrna Gene ◽  
Abies Lasiocarpa ◽  
Gram Positive ◽  
Salvage Logging ◽  
Gram Positive Bacteria ◽  
Significant Difference

To assess the effect of fire and salvage logging on the diversity of mycorrhizal–bacterial communities, bacteria associated with Cenococcum, Thelephora, Tomentella, Russulaceae, and E-strain ectomycorrhizae (ECM) of Abies lasiocarpa seedlings were characterized using two approaches. First, bacteria were isolated and characterized by Biolog©, gas chromatography fatty acid methyl ester (GC-FAME), and amplified 16S rDNA restriction analysis (ARDRA). The bacterial communities retrieved from ECM from both sites were dominated by Proteobacteria (groups gamma and beta). Pseudomonas was the most common genus isolated, followed by Variovorax, Burkholderia, and Xanthomonas. Gram-positive isolates (mostly high-G+C Gram-positive bacteria) were more frequently retrieved on the burned-salvaged site, many commonly associated with the two ascomycete ECM, Cenococcum and E-strain. Pseudomonas species were retrieved more frequently from Thelephora. Although actinomycetes were isolated from all sites, almost no actinomycetes or other Gram-positive bacteria were isolated from either Thelephora or Tomentella. Second, amplified 16S rRNA gene sequences were amplified directly from root tips and then cloned into the plasmid vector pAMP1, followed by restriction analysis. This technique distinguished more genotypes than isolates retrieved by culturing methods, but generally, results were similar in that the largest proportion of the bacteria were putatively Gram-negative; putative Gram-positive bacteria were fewer and most were from the burned–salvaged site. Direct cloning resulted in many patterns that did not match any identified isolates, suggesting that a large proportion of clones were unique or not culturable by the methods used. Analysis for both protocols showed no significant difference in bacterial diversity between the burned–salvaged and unburned sites. Key words: rhizosphere bacteria, ARDRA, 16S rDNA, Biolog©, GC-FAME.

Bacterial communities inside and in the vicinity of the Chinese Great Wall Station, King George Island, Antarctica

2007 ◽  
Vol 19 (1) ◽  
pp. 11-16 ◽  
Author(s):  
Xiang Xiao ◽  
Mingguang Li ◽  
Ziyong You ◽  
Fengping Wang
Keyword(s):  
Bacterial Diversity ◽  
Denaturing Gradient Gel Electrophoresis ◽  
Heterotrophic Bacteria ◽  
Soil Samples ◽  
Rrna Gene ◽  
Adjacent Area ◽  
Molecular Approaches ◽  
Cell Counts ◽  
Two Samples ◽  
Great Wall

Both bacterial culture and molecular approaches were used to investigate the bacterial diversity inside the Chinese Antarctic Great Wall Station and in its adjacent area. Heterotrophic bacteria were isolated from the samples using a direct plating method. γ-Proteobacteria, Actinobacteria, Flavobacteria and Firmicutes were isolated from these samples. In the three water samples, Pseudomonas species were dominant. In soil samples, Flavobacterium, Bacillus or Arthrobacter species dominated. Escherichia coli strains were isolated only in two samples from inside the station. Total cell counts in the six soil samples were semi-quantified by Quantitative Competitive-PCR of the 16S rRNA gene copies. The soil samples contained 105 to 109 cells g−1. Denaturing gradient gel electrophoresis (DGGE) was used further to investigate the bacterial diversity in the soil samples. A wider range of bacterial diversity including α-Proteobacteria, β-Proteobacteria, δ-Proteobacteria, γ-Proteobacteria, Flavobacteria, Actinobacteria and unclassified bacteria was discovered.

Sign in / Sign up

Export Citation Format

Share Document