Does virulence assessment of Vibrio anguillarum using sea bass (Dicentrarchus labrax) larvae correspond with genotypic and phenotypic characterization?

Background: Vibriosis is one of the most ubiquitous fish diseases caused by bacteria belonging to the genus Vibrio such as Vibrio (Listonella) anguillarum. Despite a lot of research efforts, the virulence factors and mechanism of V. anguillarum are still insufficiently known, in part because of the lack of standardized virulence assays. Methodology/Principal Findings: We investigated and compared the virulence of 15 V. anguillarum strains obtained from different hosts or non-host niches using a standardized gnotobiotic bioassay with European sea bass (Dicentrarchus labrax L.) larvae as model hosts. In addition, to assess potential relationships between virulence and genotypic and phenotypic characteristics, the strains were characterized by random amplified polymorphic DNA (RAPD) and repetitive extragenic palindromic PCR (rep-PCR) analyses, as well as by phenotypic analyses using Biolog's Phenotype MicroArray (TM) technology and some virulence factor assays. Conclusions/Significance: Virulence testing revealed ten virulent and five avirulent strains. While some relation could be established between serotype, genotype and phenotype, no relation was found between virulence and genotypic or phenotypic characteristics, illustrating the complexity of V. anguillarum virulence. Moreover, the standardized gnotobiotic system used in this study has proven its strength as a model to assess and compare the virulence of different V. anguillarum strains in vivo. In this way, the bioassay contributes to the study of mechanisms underlying virulence in V. anguillarum

Assessing genetic and phenotypic diversity of Brettanomyces yeast

Brettanomyces yeast species, withBrettanomyces (Dekkera) bruxellensis being the most important, are generally reported to be spoilage yeasts in the beer and wine industry due to the production of phenolic off-flavors. The aromas imparted, which can be described as ‘medicinal and rsquo;, are colloquially known as rdquo; character and are generally considered negative for beer and wine quality. However, the same compounds are regarded positively when produced during certain fermentation processes, such as the production of some styles of beer (e.g. lambic and gueuze). Despite its economic importance, surprisingly little is known about the biology, physiology and ecology of Brettanomyces yeasts. Herein, several aspects of Brettanomyces yeast biology and ecology were studied, with particular emphasis on B. Bruxellensis, thus contributing to a better understanding of the biology and ecology of these important influencers of flavor profile. In the first chapter (Chapter I), we give a comprehensive literature overview of the state-of-the-art of Brettanomyces research, emphasizing areas that were particularly well explored at the start of this PhD study, including aroma-associated aspects and methods for detection and identification. We also focused on recent genetic and genomic studies providing novel insights into the biology and evolution of B. bruxellensis. In Chapter II, we assessed the genetic relationships between 50 Brettanomyces strains belonging to all species presently identified within the genus and isolated from different food products and beverages using established DNA fingerprinting methods. These methods included ribosomal RNA (rRNA) gene sequencing, random amplified polymorphic DNA (RAPD) PCR, arbitrarily primed (AP) PCR and repetitive element PCR fingerprinting (rep-PCR). Our results support earlier findings that Brettanomyces yeasts form a genetically diverse clade, even within a species, and are represented by several subgroupings. Further our results revealed an intriguing correlation between B. bruxellensis genotype groups and the respective source of isolation, suggesting niche adaptation. To further explore this relationship, we first sequenced a (beneficial) beer isolate of B. bruxellensis (VIB X9085; ST05.12/22) and compared its genome sequence with the genome sequences of two wine spoilage strains (AWRI 1499 and CBS 2499) (Chapter III). In addition to strain-specific single nucleotide polymorphisms (SNPs) and insertions/deletions (InDels), structural genome variation was found between our strain and both wine strains, with some genomic regions specifically deleted in the beer strain. These included, but were not limited to, a region harboring the B. bruxellensis nitrate assimilation cluster and a region representing a cluster of genes mainly involved in carbon metabolism. Next, in Chapter IV, metabolic differences in carbon and nitrogen assimilation between different B. bruxellensis strains from different beverages (beer, wine and soft drink) were thoroughly assessed using Biolog Phenotype Microarrays. While some similarities of physiology were noted, many traits were variable among strains. Interestingly, some phenotypes were found that could be linked to strain origin, especially for the assimilation of particularalpha;- and β-glycosides as well as nbsp;and β-substituted monosaccharides. Based upon gene presence or absence, an nbsp;and β-glucosidase were found explaining the observed phenotypes. Further, using a PCR screen on a large number of B. bruxellensis isolates we have been able to specifically link a genomic deletion (e.g. harboring a β-glucosidase gene) to the beer strains, suggesting that this region may have a fitness cost for B. bruxellensis in certain fermentation systems such as brewing. Additionally, our work indicates that most beer strains are diploid, whereas the vast majority of wine strains are known to be triploid, suggesting that the additional set of chromosomes may confer a selective advantage in environments such as wineries. Finally, in Chapter V a series of fermentation tests were performed, in which different B. bruxellensis strains were inoculated in different media representative of different ecological niches of the yeast (i.e. beer, wine and soft drink). Utilisation of different sugars was quantified and production of (off-) flavors was monitored after one month of fermentation. Our results revealed that not only the medium (which may contain different (off-) flavor precursors), but also the yeast strain mediates the formation of the typical Brettanomyces (off-) flavors. Moreover, a (moderate) correlation was found between the origin of the strains and their impact on the volatile composition of the media. Only strains originally isolated from wine produced typical Brettanomyces off-flavors when inoculated in wine, whereas strains originally isolated from beer or soft drink did not, for example. Vice versa, in strong golden pale ale (Duvel), beer strains behaved differently from the other strains. Altogether, these results suggest that the interaction between medium and strain affects the outcome of potential (off-) flavor production. Together, the components of this PhD study provide tools to discriminate Brettanomyces strains and reveal a first glimpse into the genetic diversity and genomic and phenotypic plasticity of B. bruxellensis. Our findings are relevant for the wine industry as well as for the beer industry. After all, deeper understanding of the ecology of B. bruxellensis not only provides novel insights into the evolution of this intriguing yeast species, it will also facilitate avoidance of wine spoilage and improvement of B. bruxellensis strains for brewing.status: publishe

Current and emerging trends in techniques for plant pathogen detection

Plant pathogenic microorganisms cause substantial yield losses in several economically important crops, resulting in economic and social adversity. The spread of such plant pathogens and the emergence of new diseases is facilitated by human practices such as monoculture farming and global trade. Therefore, the early detection and identification of pathogens is of utmost importance to reduce the associated agricultural losses. In this review, techniques that are currently available to detect plant pathogens are discussed, including culture-based, PCR-based, sequencing-based, and immunology-based techniques. Their working principles are explained, followed by an overview of the main advantages and disadvantages, and examples of their use in plant pathogen detection. In addition to the more conventional and commonly used techniques, we also point to some recent evolutions in the field of plant pathogen detection. The potential use of point-of-care devices, including biosensors, have gained in popularity. These devices can provide fast analysis, are easy to use, and most importantly can be used for on-site diagnosis, allowing the farmers to take rapid disease management decisions

Full genomes of 15 Vibrio anguillarum strains

<p>Enclosed are the assembly (scaffold level) and annotations (amino acid level) of 15 Vibrio anguillarum strains as described in "Comparative genome sequencing to asses the genetic diversity and virulence attributes of 15 <em>Vibrio anguillarum</em> strains" (Busschaert <em>et al</em>, 2014)</p

Full Genome Brettanomyces bruxellensis ST05.12/22

<p>Enclosed are the assembly (scaffold level) and annotations (CDS and protein level) of a beneficial <em>Brettanomyces bruxellensis</em> strain (ST05.12/22) isolated from a Belgian beer brewery.</p

Sauerbier: die Renaissance alter Traditionen

Alte Traditionen – neue Chancen | Sauerbier kann auf sehr vielfältige Weise hergestellt werden. Dazu zählen moderne Kesselsäuerungsverfahren bis hin zu traditionellen, gemischten Gärungen mit langen Reifungszeiten. In diesem Artikel werden die wichtigsten Säuerungsverfahren und Chancen zur Diversifizierung des Sauerbiersortiments beschrieben.status: publishe

The power of sour - A review: Old traditions, new opportunities

Currently, there is a strong interest in sour beers, with more breweries producing sour styles and sales continuing to increase. There are many different ways for producing sour beer, offering breweries a variety of opportunities to pursue new beverages and diversify their portfolio. These methods range from modern kettle souring and short term mixed fermentation to traditional long term mixed fermentation and maturation in wooden barrels. While the first methods enable quick souring, these beers generally lack the flavour complexity that can be obtained through more traditional methods. Here, we discuss the microbiology of sour beers and review the most common production methods as well as the most important beer styles that are produced using these methods. Further, we report a number of novel methods that have the potential to produce the next generation of flavour-rich sour beers. Increased knowledge on the microbiology and flavour development in traditional sours will help developing such next generation of sour beers.status: publishe

Phenotypic characterization of Brettanomyces bruxellensis strains for the tolerance to stresses encountered during second generation bioethanol production

Brettanomyces (teleomorph Dekkera) bruxellensis has been generally considered a spoilage yeast in fuel ethanol production plants. However, due to its peculiar carbon- and nitrogen metabolism, the yeast is also believed to hold great potential for bioethanol production in continuous reactors. Nevertheless, before actually being useful in the production of second generation bioethanol there are still some challenges to overcome. For example, the use of lignocellulosic biomass results in the need for an extensive pretreatment process during which often several inhibiting compounds are released, leading to less efficient or stuck fermentations. The objective of this study was to phenotypically characterize B. bruxellensis strains for tolerance to stresses typically encountered during second generation bioethanol fermentation and to develop a screening plate for the evaluation of yeast strains against stresses relevant for the second generation bioethanol production. To this end, a plate was developed containing a dose range of different inhibitors (i.e. vanillin, catechol, levulinic acid, formic acid, furfural, ethanol, low pH, and high osmotic pressure). Further the plate included a negative and positive control well. Subsequently, several B. bruxellensis strains from different ecological niches were screened using the developed plate. Plates were incubated for eight days at 25 °C and analyzed by the OmniLog incubator/reader (Biolog, Hayward, CA, USA). All analysis were performed in duplicate. The different B. bruxellensis strains were ranked by calculating the average well colour development (AWCD). Additionally, each strain was scored for its tolerance to the tested inhibitory conditions. This resulted in a huge variation among strains, demonstrating the need for screening a large collection of strains to identify superior yeast strains. Highly ranked B. bruxellensis strains can then be further tested for tolerance against a mixture of inhibitors and real hydrolyzed biomass fermentation broths. Additionally, features such as ethanol yield and performance in pilot plants should be evaluated to truly see its potential for industrial second generation bioethanol production.status: publishe

Microbial counts of mealworm larvae (Tenebrio molitor) and crickets (Acheta domesticus and Gryllodes sigillatus) from different rearing companies and different production batches

The rising interest in insects for human consumption and the changing regulations in Europe require a profound insight into the food safety of insects reared and sold in Western society. The microbial quality of edible insects has only been studied occasionally. This study aimed at generating an overview of intrinsic parameters (pH, water activity and moisture content) and microbial quality of fresh mealworm larvae and crickets for several rearing companies and for several batches per rearer. In total, 21 batches obtained from 7 rearing companies were subjected to analysis of intrinsic parameters, a range of plate counts and presence-absence tests for Salmonella spp. and Listeria monocytogenes. The microbial counts of the fresh insects were generally high. Different rearing batches from a single rearing company showed differences in microbial counts which could not be explained by variations in intrinsic properties. The largest variations were found in numbers of bacterial endospores, psychrotrophs and fungi. Salmonella spp. and L. monocytogenes were not detected in any of the samples. Altogether, our study shows that large variations were found between batches from individual rearers. As a consequence, no overall differences between rearers could be observed.publisher: Elsevier articletitle: Microbial counts of mealworm larvae (Tenebrio molitor) and crickets (Acheta domesticus and Gryllodes sigillatus) from different rearing companies and different production batches journaltitle: International Journal of Food Microbiology articlelink: http://dx.doi.org/10.1016/j.ijfoodmicro.2016.11.007 content_type: article copyright: © 2016 Elsevier B.V. All rights reserved.status: publishe

The science of sour: Old traditions reborn

TRADITIONS AND OPPORTUNITIES | There are many different ways for producing sour beer, ranging from modern kettle souring to traditional long term mixed fermentation. This article describes the most important souring methods, emphasizing opportunities to diversify the portfolio of sour beer styles.status: publishe