scholarly journals Mixed-Culture Metagenomics of the Microbes Making Sour Beer

Fermentation ◽  
2021 ◽  
Vol 7 (3) ◽  
pp. 174
Author(s):  
Renan Eugênio Araujo Piraine ◽  
Fábio Pereira Leivas Leite ◽  
Matthew Bochman
Keyword(s):  
Genetic Material ◽  
Rrna Gene ◽  
Internal Transcribed Spacer Region ◽  
Brettanomyces Bruxellensis ◽  
Fungal Identification ◽  
Sample Composition ◽  
Fungal Dna ◽  
Mixed Microbial Cultures ◽  
The 16S Rrna Gene ◽  
Generation Sequencing

Mixed microbial cultures create sour beers but many brewers do not know which microbes comprise their cultures. The objective of this work was to use deep sequencing to identify microorganisms in sour beers brewed by spontaneous and non-spontaneous methods. Twenty samples were received from brewers, which were processed for microbiome analysis by next generation sequencing. For bacteria, primers were used to amplify the V3-V4 region of the 16S rRNA gene; fungal DNA detection was performed using primers to amplify the entire internal transcribed spacer region. The sequencing results were then used for taxonomy assignment, sample composition, and diversity analyses, as well as nucleotide BLAST searching. We identified 60 genera and 140 species of bacteria, of which the most prevalent were Lactobacillus acetotolerans, Pediococcus damnosus, and Ralstonia picketti/mannitolilytica. In fungal identification, 19 genera and 26 species were found, among which the most common yeasts were Brettanomyces bruxellensis and Saccharomyces cerevisiae. In some cases, genetic material from more than 60 microorganisms was found in a single sample. In conclusion, we were able to determine the microbiomes of various mixed cultures used to produce beer, providing useful information to better understand the sour beer fermentation process and brewing techniques.

scholarly journals Mixed culture metagenomics of the microbes making sour beer

2021 ◽  
Author(s):  
Renan Eugenio Araujo Piraine ◽  
Fabio Pereira Leivas Leite ◽  
Matthew L. Bochman
Keyword(s):  
Genetic Material ◽  
Rrna Gene ◽  
Internal Transcribed Spacer Region ◽  
Microbiome Analysis ◽  
Fungal Identification ◽  
Sample Composition ◽  
Fungal Dna ◽  
Mixed Microbial Cultures ◽  
The 16S Rrna Gene ◽  
Generation Sequencing

Mixed microbial cultures create sour beers, but many brewers do not know which microbes comprise their cultures. The objective of this work was to use deep sequencing to identify mi-croorganisms in sour beers brewed by spontaneous and non-spontaneous methods. Twenty samples were received from brewers, which were processed for microbiome analysis by next generation sequencing. For bacteria, primers were used to amplify the V3-V4 region of the 16S rRNA gene; fungal DNA detection was performed using primers to amplify the entire internal transcribed spacer region. The sequencing results were then used for taxonomy assignment, sample composition, and diversity analyses, as well as nucleotide BLAST searching. We identi-fied 60 genera and 140 species of bacteria, of which the most prevalent were Lactobacillus acetotol-erans, Pediococcus damnosus, and Ralstonia picketti/mannitolilytica. In fungal identification, 19 genera and 26 species were found, among which the most common yeasts were Brettanomyces bruxellen-sis and Saccharomyces cerevisiae. In some cases, genetic material from more than 60 microorgan-isms was found in a single sample. In conclusion, we were able to determine the microbiomes of various mixed cultures used to produce beer, providing useful information to better understand the sour beer fermentation process and brewing techniques.

A case of feline leprosy caused by Mycobacterium lepraemurium originating from the island of Kythira (Greece): diagnosis and treatment

2007 ◽  
Vol 9 (3) ◽  
pp. 238-241 ◽  
Author(s):  
Francois Courtin ◽  
Michel Huerre ◽  
Janet Fyfe ◽  
Paul Dumas ◽  
Maria L. Boschiroli
Keyword(s):  
Rrna Gene ◽  
Internal Transcribed Spacer Region ◽  
Leukaemia Virus ◽  
Acid Fast Bacilli ◽  
Immunodeficiency Virus ◽  
Feline Leukaemia Virus ◽  
Polymerase Chain ◽  
Mycobacterium Lepraemurium ◽  
Foamy Macrophages ◽  
The 16S Rrna Gene

A 2-year-old, 4 kg, healthy, domestic shorthair female cat presented with ulcerated subcutaneous nodules on the commissures of its mouth. The cat was negative for feline leukaemia virus and feline immunodeficiency virus. Skin mycobacteriosis was diagnosed after detection of numerous acid-fast bacilli in Ziehl Neelsen-stained smears from the ulcers. Feline leprosy was suspected following preliminary polymerase chain reaction results: positive for Mycobacterium genus but negative for Mycobacterium tuberculosis and Mycobacterium avium complexes. Mycobacterium lepraemurium was later identified following DNA sequence analysis of the 5′ end of the 16S rRNA gene and the 16S–23S internal transcribed spacer region. Microscopic lesions consisted of pyogranulomas containing mainly large foamy macrophages with 10–100 intra-cellular acid-fast bacilli per field. The cat was cured after surgery and a 14-week course of clofazimine (30 mg daily) and clarithromycin (50 mg twice daily).

scholarly journals Amplification of the V5 – V8 region of the 16S rRNA gene effectively speciates medically important genital tract Lactobacillus species in the upper female genital tract

2019 ◽  
Author(s):  
Jessica L. O’Callaghan ◽  
Dana Willner ◽  
Melissa Buttini ◽  
Flavia Huygens ◽  
Elise S. Pelzer
Keyword(s):  
Next Generation Sequencing ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Genital Tract ◽  
Rrna Gene ◽  
Next Generation ◽  
Sequencing Technology ◽  
Next Generation Sequencing Technology ◽  
The 16S Rrna Gene ◽  
Generation Sequencing

ABSTRACTBackgroundThe endometrial cavity is an upper genital tract site largely heralded as sterile, however, advances in culture-independent, next generation sequencing technology have revealed that this site harbours a rich microbial community which includes multiple Lactobacillus species. These bacteria are considered to be the most common non-pathogenic genital tract commensals. Next-generation sequencing of the female lower genital tract has revealed significant variation amongst microbial community composition with respect to Lactobacillus sp. in samples collected from healthy and diseased women. The aim of this study was to evaluate the ability of the 16S rRNA gene to characterize genital tract lactobacilli to species-level taxonomy.MethodsSamples were interrogated for the presence of microbial DNA using two-step next generation sequencing technology to exploit the V5–V8 regions of the 16S rRNA gene and compared to standard speciation using qPCR.ResultsThe V5-V8 region of the 16S rRNA gene has sufficient sequence variation within frequently encountered genital tract lactobacilli to allow accurate determination of relative abundance within the community, and speciation for several key community members without completing additional experimentation.ConclusionsNext-generation sequencing of clinical genital tract isolates is an effective method for high throughput identification to species-level of key Lactobacillus sp.IMPORTANCEHuman microbiome experiments, including the low biomass organs such as the upper genital tract, require the development of consensus protocols to ensure accurate comparison between such studies and our data forms an important foundation for future protocols.This paper provides evidence to support the selection of the V5-V8 regions of the 16S rRNA gene improved Lactobacillus speciation using next generation sequencing technology. The choice of variable region for broad-range amplification in microbiome studies is important due to preferential primer binding associated with some genera based on nucleotide sequence patterns. By utilising the V5-V8 region, multiple species of Lactobacillus can be characterised with relative confidence.

Sequence-based Identification and Classification of Fungi.

2021 ◽  
pp. 198-216
Author(s):  
Andrew M. Borman ◽  
Elizabeth M. Johnson
Keyword(s):  
Dna Sequences ◽  
Its Region ◽  
Pcr Primers ◽  
Taxonomic Resolution ◽  
Its2 Region ◽  
Rrna Gene ◽  
5.8S Rrna Gene ◽  
Fungal Identification ◽  
Identification Of Fungi ◽  
Fungal Dna

Abstract This book chapter describes the advantages and limitations of the ITS Region as a universal barcode for fungal identification. The ITS region offers several practical advantages as a universal fungal barcode region. The region encompasses segments that permit resolution at different taxonomic levels as it includes the highly conserved 5.8S rRNA gene, the moderately rapidly evolving ITS2 region and the rapidly evolving ITS1 region, flanked by the highly conserved SSU and LSU genes which permit design of PCR primers that are almost panfungal. Over the last two decades the sequence-based identification of fungi has certainly come of age. The ITS region is universally accepted as the primary fungal barcoding region owing to the high barcode gap with the locus for many groups of fungi. Since the species-resolution power of ITS is poor for certain groups of fungi, and higher-level taxonomic resolution is greater with proteincoding genes, the TEF1α locus has been proposed as the universal secondary barcode region. In addition, the historical problems surrounding the reliability of fungal DNA sequences in centralized repositories are slowly being resolved by the development of an increasing number of publicly accessible, curated databases.

scholarly journals A Deeper Look into the Biodiversity of the Extremely Acidic Copahue volcano-Río Agrio System in Neuquén, Argentina

2019 ◽  
Vol 8 (1) ◽  
pp. 58 ◽  
Author(s):  
Germán Lopez Bedogni ◽  
Francisco L. Massello ◽  
Alejandra Giaveno ◽  
Edgardo Rubén Donati ◽  
María Sofía Urbieta
Keyword(s):  
Extreme Environment ◽  
Low Ph ◽  
Rrna Gene ◽  
Prokaryotic Community ◽  
Operational Taxonomic Units ◽  
Acidic River ◽  
First Time ◽  
The 16S Rrna Gene ◽  
Generation Sequencing ◽  
Autochthonous Species

The Copahue volcano-Río Agrio system, on Patagonia Argentina, comprises the naturally acidic river Río Agrio, that runs from a few meters down the Copahue volcano crater to more than 40 km maintaining low pH waters, and the acidic lagoon that sporadically forms on the crater of the volcano, which is studied for the first time in this work. We used next-generation sequencing of the 16S rRNA gene of the entire prokaryotic community to study the biodiversity of this poorly explored extreme environment. The correlation of the operational taxonomic units (OTUs)s presence with physicochemical variables showed that the system contains three distinct environments: the crater lagoon, the Upper Río Agrio, and the Salto del Agrio waterfall, a point located approximately 12 km down the origin of the river, after it emerges from the Caviahue lake. The prokaryotic community of the Copahue Volcano-Río Agrio system is mainly formed by acidic bacteria and archaea, such as Acidithiobacillus, Ferroplasma, and Leptospirillum, which have been isolated from similar environments around the world. These results support the idea of a ubiquitous acidic biodiversity; however, this highly interesting extreme environment also has apparently autochthonous species such as Sulfuriferula, Acidianus copahuensis, and strains of Acidibacillus and Alicyclobacillus.

scholarly journals Limitations of 16S rRNA Gene Sequencing to Characterize Lactobacillus Species in the Upper Genital Tract

2021 ◽  
Vol 9 ◽  
Author(s):  
Jessica L. O’Callaghan ◽  
Dana Willner ◽  
Melissa Buttini ◽  
Flavia Huygens ◽  
Elise S. Pelzer
Keyword(s):  
Microbial Community ◽  
Next Generation Sequencing ◽  
16S Rrna ◽  
16S Rrna Gene ◽  
Genital Tract ◽  
Rrna Gene ◽  
Next Generation ◽  
Variable Regions ◽  
The 16S Rrna Gene ◽  
Generation Sequencing

The endometrial cavity is an upper genital tract site previously thought as sterile, however, advances in culture-independent, next-generation sequencing technology have revealed that this low-biomass site harbors a rich microbial community which includes multiple Lactobacillus species. These bacteria are considered to be the most abundant non-pathogenic genital tract commensals. Next-generation sequencing of the female lower genital tract has revealed significant variation amongst microbial community composition with respect to Lactobacillus sp. in samples collected from healthy women and women with urogenital conditions. The aim of this study was to evaluate our ability to characterize members of the genital tract microbial community to species-level taxonomy using variable regions of the 16S rRNA gene. Samples were interrogated for the presence of microbial DNA using next-generation sequencing technology that targets the V5–V8 regions of the 16S rRNA gene and compared to speciation using qPCR. We also performed re-analysis of published data using alternate variable regions of the 16S rRNA gene. In this analysis, we explore next-generation sequencing of clinical genital tract isolates as a method for high throughput identification to species-level of key Lactobacillus sp. Data revealed that characterization of genital tract taxa is hindered by a lack of a consensus protocol and 16S rRNA gene region target allowing comparison between studies.

scholarly journals Evaluation of Compatibility of 16S rRNA V3V4 and V4 Amplicon Libraries for Clinical Microbiome Profiling

2020 ◽  
Author(s):  
Po-Yu Liu ◽  
Wei-Kai Wu ◽  
Chieh-Chang Chen ◽  
Suraphan Panyod ◽  
Lee-Yan Sheen ◽  
...  
Keyword(s):  
16S Rrna ◽  
16S Rrna Gene ◽  
In Silico ◽  
Rrna Gene ◽  
Systematic Bias ◽  
Fecal Microbiome ◽  
Hypervariable Regions ◽  
Microbiome Profiling ◽  
The 16S Rrna Gene ◽  
Generation Sequencing

ABSTRACTSequencing of the 16S rRNA gene by Illumina next-generation sequencing is broadly used in microbiome studies. Different hypervariable regions of the 16S rRNA gene, V3V4 (amplified with primers 341F–805R) or V4 (V4O; primers 515F–806R), are selected, depending on the targeted resolution. However, in population-based clinical studies, combining V3V4 and V4 data from different studies for a meta-analysis is challenging. Reads generated by short-read (150-bp) high-throughput sequencing platforms do not fully recover the V4 region read-length. Here, we evaluated the compatibility of 16S rRNA V3V4 and V4 amplicons for microbiome profiling. We compared taxonomic compositions obtained by the analysis of V3V4 and V4 amplicons, and V4 fragments trimmed from V3V4 amplicons. We also evaluated an alternative V4 region (V4N; primers 519F–798R) designed for efficient stitching with 150-bp paired-end sequencing. First, we simulated a global investigation of environmental prokaryotes in silico. This revealed that V4O primers recovered the highest proportion of fragments (81.7%) and most phyla, including archaea. Empirical sequencing of standard (mock) and human fecal samples revealed biased patterns of each primer that were similar to the ones determined by in silico simulation. Further, for human fecal microbiome profiling, the between-sample variance was greater than the systematic bias of each primer. The use of trimmed V4 fragments and single-end amplicons resulted in the same systematic bias. In conclusion, paired-end V4O sequencing yielded the most accurate data for both, simulation and mock community sequencing; the V4O amplicons were compatible with trimmed V4 sequences for microbiome profiling.IMPORTANCENext-generation sequencing of the 16S rRNA gene is a commonly used approach for clinical microbiome studies. Different amplicons of the 16S rRNA hypervariable regions are used in different studies, which creates incompatible sequence features when comparing and integrating data among studies by using 16S denoising pipelines. Here we compared the type of data and coverage obtained when different 16S rRNA amplicons were analyzed. In silico and empirical analyses of the human fecal microbiome revealed that the V3V4 amplicons are compatible with V4 amplicons after trimming up to the same region. These observations demonstrate that reconciling the compatibility of clinical microbiome data from different studies improve not only the sample size but also the confidence of the hypothesis tested.

scholarly journals The Milk Microbiota of the Spanish Churra Sheep Breed: New Insights into the Complexity of the Milk Microbiome of Dairy Species

Animals ◽  
2020 ◽  
Vol 10 (9) ◽  
pp. 1463
Author(s):  
Cristina Esteban-Blanco ◽  
Beatriz Gutiérrez-Gil ◽  
Héctor Marina ◽  
Rocío Pelayo ◽  
Aroa Suárez-Vega ◽  
...  
Keyword(s):  
Animal Species ◽  
Massively Parallel Sequencing ◽  
Massively Parallel ◽  
Rrna Gene ◽  
Microbiota Composition ◽  
Complex Issue ◽  
The Core ◽  
Sheep Milk ◽  
The 16S Rrna Gene ◽  
Generation Sequencing

Milk from healthy animals has classically been considered a sterile fluid. With the development of massively parallel sequencing and its application to the study of the microbiome of different body fluids, milk microbiota has been documented in several animal species. In this study, the main objective of this work was to access bacterial profiles of healthy milk samples using the next-generation sequencing of amplicons from the 16S rRNA gene to characterise the milk microbiome of the Churra breed. A total of 212 samples were collected from two Churra dairy farms with a different management system. The core milk microbiota in Churra ewes includes lesser genera (only two taxa: Staphylococcus and Escherichia/Shigella) than studies reported in other dairy species or even in a previous study in Assaf sheep milk. We found that diversity values in the two flocks of Churra breed were lower than the diversity of the milk microbiota in Assaf. The non-metric multidimensional scaling (NMDS) ordination using Bray-Curtis distance separates samples based on their microbiota composition. The information reported here might be used to understand the complex issue of milk microbiota composition.

scholarly journals The combination of mare's milk and grape polyphenol extract for treatment of dysbiosis induced by dextran sulfate sodium

2020 ◽  
Vol 21 (5) ◽  
Author(s):  
Samat Kozhakhmetov ◽  
DMITRIY BABENKO ◽  
MADIYAR NURGAZIYEV ◽  
ALTYNAY TUYAKOVA ◽  
AYAULYM NURGOZHINA ◽  
...  
Keyword(s):  
Ulcerative Colitis ◽  
Intestinal Microflora ◽  
Structural Changes ◽  
Biological Diversity ◽  
Dextran Sulfate ◽  
Dextran Sulfate Sodium ◽  
Rrna Gene ◽  
Biological Product ◽  
The 16S Rrna Gene ◽  
Generation Sequencing

Abstract. Kozhakhmetov S, Babenko D, Nurgaziyev M, Tuyakova A, Nurgozhina A, Muhanbetganov N, Chulenbayeva L, Sergazy S, Gulyayev A, Saliev T, Kushugulova A. 2020. The combination of mare's milk and grape polyphenol extract for treatment of dysbiosis induced by dextran sulfate sodium. Biodiversitas 21: 2275-2280. This study showed the potential of a biological product based on mare's milk and a complex of grape polyphenols to modulate intestinal microflora after dextran sulfate sodium  (DSS)-induced dysbiosis. Rat ulcerative colitis has been developed using intra-gastric administration of 10% DSS solution. To track changes in the structure of the microbiome at all stages of the study, the next-generation sequencing of the 16S rRNA gene section and LotuS conveyor were used. The results of sequencing demonstrated a decrease in biological diversity of microbiota after the induction of colitis, and recovery after 7 days of use of the  (MMGPE). The product induced the structural changes of the microbiome damaged by DSS. Representatives of SCFA producing bacteria increased concentrations of  Prevotella, Alloprevotella, Lactobacillus, Ruminococcaceae, and Blautia.

scholarly journals Influence of Traditional vs Alternative Dietary Carbohydrates Sources on the Large Intestinal Microbiota in Post-Weaning Piglets

Animals ◽  
2019 ◽  
Vol 9 (8) ◽  
pp. 516 ◽  
Author(s):  
Tretola ◽  
Luciano ◽  
Ottoboni ◽  
Baldi ◽  
Pinotti
Keyword(s):  
Growth Performance ◽  
Rrna Gene ◽  
Microbiota Composition ◽  
Gut Health ◽  
Faecal Microbiota ◽  
Alternative Ingredients ◽  
Weaning Piglets ◽  
The 16S Rrna Gene ◽  
Generation Sequencing ◽  
Gut Microbiota Composition

In this study, common cereal grains were partially replaced by former foodstuffs products (FFPs) in post-weaning piglets’ diets, to investigate how these alternative ingredients influence the faecal microbiota in the post-weaning period. Twelve post-weaning piglets were housed for 16 days in individual pens and were then fed two diets: a standard wheat-barley-corn meal diet and a diet containing 30% FFPs, thus partially substituting conventional cereals. The growth performance was monitored and faecal microbiota was characterized by the next generation sequencing of the 16S rRNA gene. The results showed no detrimental effects on growth performance when FFPs were used. However, the FFP diet decreased the bacterial richness and evenness in the large intestine, while minor differences were observed in the taxa composition. The core microbiota composition was only slightly affected, and no differences between the two groups in the gut microbiota composition at the phylum level over time were observed. Thus, although these results should be interpreted with caution, as they are case-specific, FFPs can be potentially used as alternative carbohydrate sources in post-weaning piglets, but further investigations are necessary to clarify their impact on gut health when used for a longer period.

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