Lien Bosmans

Assessment of the genetic and phenotypic diversity among rhizogenic Agrobacterium biovar 1 strains infecting solanaceous and cucurbit crops.

Rhizogenic Agrobacterium biovar 1 strains have been found to cause extensive root proliferation on hydroponically grown Cucurbitaceae and Solanaceae crops, resulting in substantial economic losses. As these agrobacteria live under similar ecological conditions, infecting a limited number of crops, it may be hypothesized that genetic and phenotypic variation among such strains is relatively low. In this study we assessed the phenotypic diversity as well as the phylogenetic and evolutionary relationships of several rhizogenic Agrobacterium biovar 1 strains from cucurbit and solanaceous crops. A collection of 41 isolates was subjected to a number of phenotypic assays and characterized by MLSA targeting four housekeeping genes (16S rRNA gene, recA, rpoB and trpE) and two loci from the root-inducing Ri-plasmid (part of rolB and virD2). Besides phenotypic variation, remarkable genotypic diversity was observed, especially for some chromosomal loci such as trpE. In contrast, genetic diversity was lower for the plasmid-borne loci, indicating that the studied chromosomal housekeeping genes and Ri-plasmid-borne loci might not exhibit the same evolutionary history. Furthermore, phylogenetic and network analyses and several recombination tests suggested that recombination could be contributing in some extent to the evolutionary dynamics of rhizogenic Agrobacterium populations. Finally, a genomospecies-level identification analysis revealed that at least four genomospecies may occur on cucurbit and tomato crops (G1, G3, G8 and G9). Together, this study gives a first glimpse at the genetic and phenotypic diversity within this economically important plant pathogenic bacterium.

Fermentation assays reveal differences in sugar and (off-) flavor metabolism across different Brettanomyces bruxellensis strains

Abstract Brettanomyces (Dekkera) bruxellensis is an ascomycetous yeast of major importance in the food, beverage and biofuel industry. It has been isolated from various man‐made ecological niches that are typically characterized by harsh environmental conditions such as wine, beer, soft drink, etc. Recent comparative genomics studies revealed an immense intraspecific diversity, but it is still unclear whether this genetic diversity also leads to systematic differences in fermentation performance and (off‐)flavor production, and to what extent strains have evolved to match their ecological niche. Here, we present an evaluation of the fermentation properties of eight genetically diverse B. bruxellensis strains originating from beer, wine and soft drinks. We show that sugar consumption and aroma production during fermentation are determined by both the yeast strain and composition of the medium. Furthermore, our results indicate a strong niche adaptation of B. bruxellensis, most clearly for wine strains. For example, only strains originally isolated from wine were able to thrive well and produce the typical Brettanomyces‐related phenolic off‐flavors 4‐ethylguaiacol and 4‐ethylphenol when inoculated in red wine. Sulfite tolerance was found as a key factor explaining the observed differences in fermentation performance and off‐flavor production. Sequence analysis of genes related to phenolic off‐flavor production, however, revealed only marginal differences between the isolates tested, especially at the amino acid level. Altogether, our study provides novel insights in the Brettanomyces metabolism of flavor production, and is highly relevant for both the wine and beer industry.

Agar composition affects in vitro screening of biocontrol activity of antagonistic microorganisms.

Agar-based screening assays are the method of choice when evaluating antagonistic potential of bacterial biocontrol-candidates against pathogens. We showed that when using the same medium, but different agar compositions, the activity of a bacterial antagonist against Agrobacterium was strongly affected. Consequently, results from in vitro screenings should be interpreted cautiously.

Efficacy of hydrogen peroxide treatment for control of hairy root disease caused by rhizogenic agrobacteria.

Rhizogenic Agrobacterium strains are the causative agent of hairy root disease (HRD), an increasing problem in the hydroponic cultivation of tomato and cucumber in Europe. A previous study has demonstrated that different lineages of rhizogenic agrobacteria are able to form biofilms. Although hydrogen peroxide (H2O2) is a frequently used biocide in hydroponic systems, until now its effectiveness to remove rhizogenic agrobacteria has not been unequivocally demonstrated. Therefore, the main objective of this study was to assess the efficacy of H2O2 in controlling Agrobacterium‐containing biofilms.

Sweet scents: Nectar Specialist Yeasts Enhance Nectar Attraction of a Generalist Aphid Parasitoid without Affecting Survival

Floral nectar is commonly inhabited by microorganisms, mostly yeasts and bacteria, which can have a strong impact on nectar chemistry and scent. Yet, little is known about the effects of nectar microbes on the behavior and survival of insects belonging to the third trophic level such as parasitoids. Here, we used five nectar-inhabiting yeast species to test the hypothesis that yeast species that almost solely occur in nectar, and therefore substantially rely on floral visitors for dispersal, produce volatile compounds that enhance insect attraction without compromising insect life history parameters, such as survival. Experiments were performed using two nectar specialist yeasts (Metschnikowia gruessii and M. reukaufii) and three generalist species (Aureobasidium pullulans, Hanseniaspora uvarum, and Sporobolomyces roseus). Saccharomyces cerevisiae was included as a reference yeast. We compared olfactory responses of the generalist aphid parasitoid Aphidius ervi (Haliday) (Hymenoptera: Braconidae) when exposed to these microorganisms inoculated in synthetic nectar. Nectar-inhabiting yeasts had a significant impact on nectar chemistry and produced distinct volatile blends, some of which were attractive, while others were neutral or repellent. Among the different yeast species tested, the nectar specialists M. gruessii and M. reukaufii were the only species that produced a highly attractive nectar to parasitoid females, which simultaneously had no adverse effects on longevity and survival of adults. By contrast, parasitoids that fed on nectars fermented with the reference strain, A. pullulans, H. uvarum or S. roseus showed shortest longevity and lowest survival. Additionally, nectars fermented by A. pullulans or S. roseus were consumed significantly less, suggesting a lack of important nutrients or undesirable changes in the nectar chemical profiles. Altogether our results indicate that nectar-inhabiting yeasts play an important, but so far largely overlooked, role in plant-insect interactions by modulating the chemical composition of nectar, and may have important ecological consequences for plant pollination and biological control of herbivorous insects.

Potential for biocontrol of hairy root disease by a Paenibacillus clade

Rhizogenic Agrobacterium biovar 1 is the causative agent of hairy root disease (HRD) in the hydroponic cultivation of tomato and cucumber causing significant losses in marketable yield. In order to prevent and control the disease chemical disinfectants such as hydrogen peroxide or hypochlorite are generally applied to sanitize the hydroponic system and/or hydroponic solution. However, effective control of HRD sometimes requires high disinfectant doses that may have phytotoxic effects. Moreover, several of these chemicals may be converted to unwanted by-products with human health hazards. Here we explored the potential of beneficial bacteria as a sustainable means to control HRD. A large collection of diverse bacterial genera was screened for antagonistic activity against rhizogenic Agrobacterium biovar 1 using the agar overlay assay. Out of more than 150 strains tested, only closely related Paenibacillus strains belonging to a particular clade showed antagonistic activity, representing the species P. illinoisensis, P. pabuli, P. taichungensis, P. tundrae, P. tylopili, P. xylanexedens, and P. xylanilyticus. Assessment of the spectrum of activity revealed that some strains were able to inhibit the growth of all 35 rhizogenic agrobacteria strains tested, while others were only active against part of the collection, suggesting a different mode of action. Preliminary characterization of the compounds involved in the antagonistic activity of two closely related Paenibacillus strains, tentatively identified as P. xylanexedens, revealed that they are water-soluble and have low molecular weight. Application of a combination of these strains in greenhouse conditions resulted in a significant reduction of HRD, indicating the great potential of these strains to control HRD.

Rhizogenic agrobacteria in hydroponic crops: epidemics, diagnostics and control

&NA; Rhizogenic Agrobacterium biovar 1, harbouring an Ri‐plasmid (root‐inducing plasmid), is the causative agent of hairy root disease (HRD) in the hydroponic cultivation of tomato, cucumber and aubergine. The disease is characterized by extensive root proliferation leading to strong vegetative growth and, in severe cases, substantial losses in marketable yield. Agrobacterium biovar 1 is a heterogeneous group of agrobacteria and includes at least 10 genomospecies, among which at least four (G1, G3, G8 and G9) have been associated with HRD in hydroponically grown vegetables. This review has synthesized the current knowledge on rhizogenic Agrobacterium biovar 1, including infection process, current taxonomic status, genetic and phenotypic diversity, detection methods and strategies for disease control. With regard to the latter, symptom reduction and prevention of infection through cultivation methods and chemical disinfection (e.g. by the use of chlorine‐based disinfectants and hydrogen peroxide) are discussed and biocontrol strategies are elaborated on. Recent research has led to the identification of a phylogenetically related clade of Paenibacillus strains that have antagonistic activity against rhizogenic Agrobacterium biovar 1 strains, holding great potential for HRD control. Finally, possible directions for future research are proposed.