scholarly journals Prediction of Genetic Groups within Brettanomyces bruxellensis through Cell Morphology Using a Deep Learning Tool

2021 ◽  
Vol 7 (8) ◽  
pp. 581
Author(s):  
Manon Lebleux ◽  
Emmanuel Denimal ◽  
Déborah De Oliveira ◽  
Ambroise Marin ◽  
Nicolas Desroche ◽  
...  
Keyword(s):  
Deep Learning ◽  
Random Amplified Polymorphic Dna ◽  
Microsatellite Analysis ◽  
Intraspecific Diversity ◽  
Brettanomyces Bruxellensis ◽  
Genetic Characteristics ◽  
Genetic Groups ◽  
Spoilage Yeast ◽  
Microbial Ecosystems ◽  
Pcr Method

Brettanomyces bruxellensis is described as a wine spoilage yeast with many mainly strain-dependent genetic characteristics, bestowing tolerance against environmental stresses and persistence during the winemaking process. Thus, it is essential to discriminate B. bruxellensis isolates at the strain level in order to predict their stress resistance capacities. Few predictive tools are available to reveal intraspecific diversity within B. bruxellensis species; also, they require expertise and can be expensive. In this study, a Random Amplified Polymorphic DNA (RAPD) adapted PCR method was used with three different primers to discriminate 74 different B. bruxellensis isolates. High correlation between the results of this method using the primer OPA-09 and those of a previous microsatellite analysis was obtained, allowing us to cluster the isolates among four genetic groups more quickly and cheaply than microsatellite analysis. To make analysis even faster, we further investigated the correlation suggested in a previous study between genetic groups and cell polymorphism using the analysis of optical microscopy images via deep learning. A Convolutional Neural Network (CNN) was trained to predict the genetic group of B. bruxellensis isolates with 96.6% accuracy. These methods make intraspecific discrimination among B. bruxellensis species faster, simpler and less costly. These results open up very promising new perspectives in oenology for the study of microbial ecosystems.

scholarly journals Real-Time PCR Assay for Detection and Enumeration of Dekkera bruxellensis in Wine

2003 ◽  
Vol 69 (12) ◽  
pp. 7430-7434 ◽  
Author(s):  
Trevor G. Phister ◽  
David A. Mills
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Pcr Primers ◽  
Rrna Gene ◽  
Dekkera Bruxellensis ◽  
Pcr Assay ◽  
Specific Pcr ◽  
Spoilage Yeast ◽  
Pcr Method ◽  
26S Rrna

ABSTRACT Traditional methods to detect the spoilage yeast Dekkera bruxellensis from wine involve lengthy enrichments. To overcome this difficulty, we developed a quantitative real-time PCR method to directly detect and enumerate D. bruxellensis in wine. Specific PCR primers to D. bruxellensis were designed to the 26S rRNA gene, and nontarget yeast and bacteria common to the winery environment were not amplified. The assay was linear over a range of cell concentrations (6 log units) and could detect as little as 1 cell per ml in wine. The addition of large amounts of nontarget yeasts did not impact the efficiency of the assay. This method will be helpful to identify possible routes of D. bruxellensis infection in winery environments. Moreover, the time involved in performing the assay (3 h) should enable winemakers to more quickly make wine processing decisions in order to reduce the threat of spoilage by D. bruxellensis.

Genomics Perspectives on Metabolism, Survival Strategies, and Biotechnological Applications of Brettanomyces bruxellensis LAMAP2480

2017 ◽  
Vol 27 (3) ◽  
pp. 147-158 ◽  
Author(s):  
Liliana Godoy ◽  
Evelyn Silva-Moreno ◽  
Wladimir Mardones ◽  
Darwin Guzman ◽  
Francisco A. Cubillos ◽  
...  
Keyword(s):  
Ethanol Production ◽  
Survival Strategies ◽  
Whole Genome ◽  
Biotechnological Applications ◽  
Genome Sequences ◽  
Wine Production ◽  
Brettanomyces Bruxellensis ◽  
Biotechnological Potential ◽  
Spoilage Yeast ◽  
Genome Comparisons

Wine production is an important commercial issue for the liquor industry. The global production was estimated at 275.7 million hectoliters in 2015. The loss of wine production due to <i>Brettanomyces bruxellensis </i>contamination is currently a problem. This yeast causes a “horse sweat” flavor in wine, which is an undesired organoleptic attribute. To date, 6 <i>B. bruxellensis </i>annotated genome sequences are available (LAMAP2480, AWRI1499, AWRI1608, AWRI1613, ST05.12/22, and CBS2499), and whole genome comparisons between strains are limited. In this article, we reassembled and reannotated the genome of <i>B. bruxellensis</i> LAMAP2480, obtaining a 27-Mb assembly with 5.5 kb of N50. In addition, the genome of <i>B. bruxellensis</i> LAMAP2480 was analyzed in the context of spoilage yeast and potential as a biotechnological tool. In addition, we carried out an exploratory transcriptomic analysis of this strain grown in synthetic wine. Several genes related to stress tolerance, micronutrient acquisition, ethanol production, and lignocellulose assimilation were found. In conclusion, the analysis of the genome of <i>B. bruxellensis</i> LAMAP2480 reaffirms the biotechnological potential of this strain. This research represents an interesting platform for the study of the spoilage yeast <i>B. bruxellensis</i>.

scholarly journals PMKT2, a new killer toxin from Pichia membranifaciens, and its promising biotechnological properties for control of the spoilage yeast Brettanomyces bruxellensis

Microbiology ◽  
2009 ◽  
Vol 155 (2) ◽  
pp. 624-634 ◽  
Author(s):  
A. Santos ◽  
M. San Mauro ◽  
E. Bravo ◽  
D. Marquina
Keyword(s):  
Biochemical Characterization ◽  
Gel Filtration ◽  
Chemical Properties ◽  
Killer Toxin ◽  
Small Scale ◽  
Brettanomyces Bruxellensis ◽  
Killer Activity ◽  
Pichia Membranifaciens ◽  
Spoilage Yeast ◽  
Spoilage Yeasts

Pichia membranifaciens CYC 1086 secretes a killer toxin (PMKT2) that is inhibitory to a variety of spoilage yeasts and fungi of agronomical interest. The killer toxin in the culture supernatant was concentrated by ultrafiltration and purified to homogeneity by two successive steps, including native electrophoresis and HPLC gel filtration. Biochemical characterization of the toxin showed it to be a protein with an apparent molecular mass of 30 kDa and an isoelectric point of 3.7. At pH 4.5, optimal killer activity was observed at temperatures up to 20 °C. Above approximately this pH, activity decreased sharply and was barely noticeable at pH 6. The toxin concentrations present in the supernatant during optimal production conditions exerted a fungicidal effect on a variety of fungal and yeast strains. The results obtained suggest that PMKT2 has different physico-chemical properties from PMKT as well as different potential uses in the biocontrol of spoilage yeasts. PMKT2 was able to inhibit Brettanomyces bruxellensis while Saccharomyces cerevisiae was fully resistant, indicating that PMKT2 could be used in wine fermentations to avoid the development of the spoilage yeast without deleterious effects on the fermentative strain. In small-scale fermentations, PMKT2, as well as P. membranifaciens CYC 1086, was able to inhibit B. bruxellensis, verifying the biocontrol activity of PMKT2 in simulated winemaking conditions.

scholarly journals Morphological, Chemical, and Genetic Characteristics of Korean Native Thyme Bak-Ri-Hyang (Thymus quinquecostatus Celak.)

Antibiotics ◽  
2020 ◽  
Vol 9 (6) ◽  
pp. 289
Author(s):  
Minju Kim ◽  
Jun-Cheol Moon ◽  
Songmun Kim ◽  
Kandhasamy Sowndhararajan
Keyword(s):  
Essential Oil ◽  
Rapd Markers ◽  
Rapd Analysis ◽  
Random Amplified Polymorphic Dna ◽  
Essential Oil Composition ◽  
Similarity Coefficient ◽  
Gas Chromatography Mass Spectrometry ◽  
Aromatic Plant ◽  
Oil Composition ◽  
Genetic Characteristics

Bak-ri-hyang (Thymus quinquecostatus Celak.) is an important medicinal and aromatic plant in Korea. T. quinquecostatus population and is always mixed with other thyme cultivars during cultivation and marketing. Hence, this study aimed to determine the genetic variability and the essential oil composition of three Korean native thyme, T. quinquecostatus cultivars collected from the Wolchul, Jiri, and Odae mountains, in comparison with six commercial thyme cultivars (T. vulgaris), to distinguish Bak-ri-hyang from other thyme cultivars. The composition of essential oils obtained from nine individuals was analyzed by gas chromatography–mass spectrometry (GC–MS). The random amplified polymorphic DNA (RAPD) analysis was accomplished using 16 different primers. The GC–MS analysis revealed that Wolchul, creeping, golden, and orange cultivars belong to the geraniol chemotype. Whereas the Odae, lemon, and silver cultivars belong to the thymol chemotype. Further, linalool was the most abundant component in carpet and Jiri cultivars. The RAPD analysis demonstrated that all thyme cultivars showed characteristic RAPD patterns that allowed their identification. In total, 133 bands were obtained using 16 primers, and 124 bands were polymorphic, corresponding to 93.2% polymorphism. Cluster analysis of RAPD markers established the presence of clear separation from nine thyme cultivars. The highest dissimilarity and similarity coefficient of the RAPD markers were 0.58 and 0.98, respectively. According to the RAPD patterns, the nine thyme cultivars could be divided into two major clusters. Among three Korean cultivars, the Wolchul and Odae cultivars were placed into the same cluster, but they did not show identical clustering with their essential oil compositions. The findings of the present study suggest that RAPD analysis can be a useful tool for marker-assisted identification of T. quinquecostatus from other Thymus species.

scholarly journals Presence and Persistence of Legionella spp. in Groundwater

2005 ◽  
Vol 71 (2) ◽  
pp. 663-671 ◽  
Author(s):  
Joana Costa ◽  
Igor Tiago ◽  
Milton S. da Costa ◽  
António Veríssimo
Keyword(s):  
Legionella Pneumophila ◽  
Random Amplified Polymorphic Dna ◽  
Acid Methyl Ester ◽  
Gene Exchange ◽  
Rapd Profile ◽  
Genetic Groups ◽  
Acid Methyl ◽  
Groundwater Samples ◽  
Rapd Pcr ◽  
Reference Strains

ABSTRACT Groundwater samples (111) from six different boreholes located in two geographical areas were examined for the presence of legionellae over a 7-year period. The number of Legionella isolates detected was generally low. The colonization of the aquifers was not uniform, and the persistence of Legionella was independent of the hydraulic pumps and the plumbing system present in the borehole. A total of 374 isolates identified by fatty acid methyl ester analysis belonged to Legionella pneumophila, L. oakridgensis, L. sainthelensi, and L. londiniensis. In area 1, L. oakridgensis constituted the major population detected, exhibiting only one random amplified polymorphic DNA (RAPD)-PCR profile. L. sainthelensi strains were less frequently isolated and also displayed a single RAPD profile, while L. pneumophila was only sporadically detected. In contrast, L. pneumophila comprised the vast majority of the isolates in area 2 and exhibited six distinct RAPD patterns, indicating the presence of different genetic groups; three L. londiniensis RAPD types were also detected. Two of the L. pneumophila and one of the L. londiniensis RAPD types were persistent in this environment for at least 12 years. The genetic structure of L. pneumophila groundwater populations, inferred from rpoB and dotA gene sequences, was peculiar, since the majority of the isolates were allied in a discrete group different from the lineages containing most of the type and reference strains of the three subspecies of L. pneumophila. Furthermore, gene exchange events related to the dotA allele could be envisioned.

scholarly journals Occurrence of Natural Bacillus thuringiensis Contaminants and Residues of Bacillus thuringiensis-Based Insecticides on Fresh Fruits and Vegetables

2006 ◽  
Vol 72 (5) ◽  
pp. 3435-3440 ◽  
Author(s):  
Kristine Frederiksen ◽  
Hanne Rosenquist ◽  
Kirsten J�rgensen ◽  
Andrea Wilcks
Keyword(s):  
Bacillus Thuringiensis ◽  
Fruits And Vegetables ◽  
Dna Analysis ◽  
Random Amplified Polymorphic Dna ◽  
The Other ◽  
Crystal Proteins ◽  
Enterotoxin Genes ◽  
Pcr Method ◽  
Plasmid Profiling ◽  
Hemolysin Bl

ABSTRACT A total of 128 Bacillus cereus-like strains isolated from fresh fruits and vegetables for sale in retail shops in Denmark were characterized. Of these strains, 39% (50/128) were classified as Bacillus thuringiensis on the basis of their content of cry genes determined by PCR or crystal proteins visualized by microscopy. Random amplified polymorphic DNA analysis and plasmid profiling indicated that 23 of the 50 B. thuringiensis strains were of the same subtype as B. thuringiensis strains used as commercial bioinsecticides. Fourteen isolates were indistinguishable from B. thuringiensis subsp. kurstaki HD1 present in the products Dipel, Biobit, and Foray, and nine isolates grouped with B. thuringiensis subsp. aizawai present in Turex. The commercial strains were primarily isolated from samples of tomatoes, cucumbers, and peppers. A multiplex PCR method was developed to simultaneously detect all three genes in the enterotoxin hemolysin BL (HBL) and the nonhemolytic enterotoxin (NHE), respectively. This revealed that the frequency of these enterotoxin genes was higher among the strains indistinguishable from the commercial strains than among the other B. thuringiensis and B. cereus-like strains isolated from fruits and vegetables. The same was seen for a third enterotoxin, CytK. In conclusion, the present study strongly indicates that residues of B. thuringiensis-based insecticides can be found on fresh fruits and vegetables and that these are potentially enterotoxigenic.

scholarly journals Diversity in Spoilage Yeast Dekkera/Brettanomyces bruxellensis Isolated from French Red Wine. Assessment During Fermentation of Synthetic Wine Medium

2008 ◽  
Vol 114 (1) ◽  
pp. 69-75 ◽  
Author(s):  
Pascal Barbin ◽  
Jean-Luc Cheval ◽  
Jean-François Gilis ◽  
Pierre Strehaiano ◽  
Patricia Taillandier
Keyword(s):  
Red Wine ◽  
Brettanomyces Bruxellensis ◽  
Spoilage Yeast

Morphological and molecular data to determine the origin and taxonomic status of Prosopis chilensis var. riojana (Fabaceae, Mimosoideae)

2003 ◽  
Vol 81 (9) ◽  
pp. 905-917 ◽  
Author(s):  
Soledad Vázquez-Garcidueñas ◽  
Ramón A Palacios ◽  
Jenny Segovia-Quiroz ◽  
Juan T Frías-Hernández ◽  
Victor Olalde-Portugal ◽  
...  
Keyword(s):  
Random Amplified Polymorphic Dna ◽  
Taxonomic Status ◽  
Molecular Data ◽  
Genetic Distances ◽  
Group Method ◽  
Sympatric Populations ◽  
Arithmetic Means ◽  
Genetic Characteristics ◽  
Genetic Traits ◽  
Prosopis Chilensis

This study analyzes the morphological and genetic characteristics of three sympatric populations of Prosopis from Argentina. Although morphological and geographical data suggest that Prosopis chilensis var. riojana is an interspecific hybrid of Prosopis chilensis var. chilensis and Prosopis flexuosa var. flexuosa, no correlation was found between morphological traits and genetic distances generated by random amplified polymorphic DNA - polymerase chain reaction (RAPD-PCR). Genetic similarity is greater among P. chilensis var. chilensis and P. flexuosa var. flexuosa than between either of these two taxa with P. chilensis var. riojana. Also, P. chilensis var. riojana has unique genetic markers that are absent from its putative parents. Additionally, dendrograms generated by unweighted pair group method with arithmetic means (UPGMA) and Neighbor-Joining clustering criteria group P. chilensis var. chilensis and P. flexuosa var. flexuosa apart from P. chilensis var. riojana. Possibilities for the lack of congruence between morphology and RAPD markers are discussed. The results obtained are insufficient to conclusively establish the origin of P. chilensis var. riojana; however, the morphological and genetic traits observed suggest this taxon is well differentiated from P. chilensis var. chilensis and P. flexuosa var. flexuosa.Key words: genetic distances, hybridization, morphology, Prosopis, RAPD, rare taxon.

scholarly journals Rapid Identification and Enumeration of Saccharomyces cerevisiae Cells in Wine by Real-Time PCR

2005 ◽  
Vol 71 (11) ◽  
pp. 6823-6830 ◽  
Author(s):  
P. Martorell ◽  
A. Querol ◽  
M. T. Fernández-Espinar
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Real Time ◽  
Real Time Pcr ◽  
Yeast Species ◽  
Random Amplified Polymorphic Dna ◽  
Rapid Identification ◽  
Sequence Information ◽  
Spoilage Yeast ◽  
Saccharomyces Pastorianus ◽  
Wine Spoilage

ABSTRACT Despite the beneficial role of Saccharomyces cerevisiae in the food industry for food and beverage production, it is able to cause spoilage in wines. We have developed a real-time PCR method to directly detect and quantify this yeast species in wine samples to provide winemakers with a rapid and sensitive method to detect and prevent wine spoilage. Specific primers were designed for S. cerevisiae using the sequence information obtained from a cloned random amplified polymorphic DNA band that differentiated S. cerevisiae from its sibling species Saccharomyces bayanus, Saccharomyces pastorianus, and Saccharomyces paradoxus. The specificity of the primers was demonstrated for typical wine spoilage yeast species. The method was useful for estimating the level of S. cerevisiae directly in sweet wines and red wines without preenrichment when yeast is present in concentrations as low as 3.8 and 5 CFU per ml. This detection limit is in the same order as that obtained from glucose-peptone-yeast growth medium (GPY). Moreover, it was possible to quantify S. cerevisiae in artificially contaminated samples accurately. Limits for accurate quantification in wine were established, from 3.8 × 105 to 3.8 CFU/ml in sweet wine and from 5 × 106 to 50 CFU/ml in red wine.

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