@article{jensen_deichmann_ma_vilandt_schiesaro_rojek_lengger_eliasson_vento_durmusoglu_et al._2022, title={Engineered cell differentiation and sexual reproduction in probiotic and mating yeasts}, volume={13}, ISSN={["2041-1723"]}, DOI={10.1038/s41467-022-33961-y}, abstractNote={G protein-coupled receptors (GPCRs) enable cells to sense environmental cues and are indispensable for coordinating vital processes including quorum sensing, proliferation, and sexual reproduction. GPCRs comprise the largest class of cell surface receptors in eukaryotes, and for more than three decades the pheromone-induced mating pathway in baker's yeast Saccharomyces cerevisiae has served as a model for studying heterologous GPCRs (hGPCRs). Here we report transcriptome profiles following mating pathway activation in native and hGPCR-signaling yeast and use a model-guided approach to correlate gene expression to morphological changes. From this we demonstrate mating between haploid cells armed with hGPCRs and endogenous biosynthesis of their cognate ligands. Furthermore, we devise a ligand-free screening strategy for hGPCR compatibility with the yeast mating pathway and enable hGPCR-signaling in the probiotic yeast Saccharomyces boulardii. Combined, our findings enable new means to study mating, hGPCR-signaling, and cell-cell communication in a model eukaryote and yeast probiotics.}, number={1}, journal={NATURE COMMUNICATIONS}, author={Jensen, Emil D. and Deichmann, Marcus and Ma, Xin and Vilandt, Rikke U. and Schiesaro, Giovanni and Rojek, Marie B. and Lengger, Bettina and Eliasson, Line and Vento, Justin M. and Durmusoglu, Deniz and et al.}, year={2022}, month={Oct} }
 @article{siedler_rau_bidstrup_vento_aunsbjerg_bosma_mcnair_beisel_neves_2020, title={Competitive Exclusion Is a Major Bioprotective Mechanism of Lactobacilli against Fungal Spoilage in Fermented Milk Products}, volume={86}, ISSN={["1098-5336"]}, DOI={10.1128/AEM.02312-19}, abstractNote={In societies that have food choices, conscious consumers demand natural solutions to keep their food healthy and fresh during storage, simultaneously reducing food waste. The use of “good bacteria” to protect food against spoilage organisms has a long, successful history, even though the molecular mechanisms are not fully understood. In this study, we show that the depletion of free manganese is a major bioprotective mechanism of lactobacilli in dairy products. High manganese uptake and intracellular storage provide a link to the distinct, nonenzymatic, manganese-catalyzed oxidative stress defense mechanism, previously described for certain lactobacilli. The evaluation of representative Lactobacillus species in our study identifies multiple relevant species groups for fungal growth inhibition via manganese depletion. Hence, through the natural mechanism of nutrient depletion, the use of dedicated bioprotective lactobacilli constitutes an attractive alternative to artificial preservation. ABSTRACT A prominent feature of lactic acid bacteria (LAB) is their ability to inhibit growth of spoilage organisms in food, but hitherto research efforts to establish the mechanisms underlying bioactivity focused on the production of antimicrobial compounds by LAB. We show, in this study, that competitive exclusion, i.e., competition for a limited resource by different organisms, is a major mechanism of fungal growth inhibition by lactobacilli in fermented dairy products. The depletion of the essential trace element manganese by two Lactobacillus species was uncovered as the main mechanism for growth inhibition of dairy spoilage yeast and molds. A manganese transporter (MntH1), representing one of the highest expressed gene products in both lactobacilli, facilitates the exhaustive manganese scavenging. Expression of the mntH1 gene was found to be strain dependent, affected by species coculturing and the growth phase. Further, deletion of the mntH1 gene in one of the strains resulted in a loss of bioactivity, proving this gene to be important for manganese depletion. The presence of an mntH gene displayed a distinct phylogenetic pattern within the Lactobacillus genus. Moreover, assaying the bioprotective ability in fermented milk of selected lactobacilli from 10 major phylogenetic groups identified a correlation between the presence of mntH and bioprotective activity. Thus, manganese scavenging emerges as a common trait within the Lactobacillus genus, but differences in expression result in some strains showing more bioprotective effect than others. In summary, competitive exclusion through ion depletion is herein reported as a novel mechanism in LAB to delay the growth of spoilage contaminants in dairy products. IMPORTANCE In societies that have food choices, conscious consumers demand natural solutions to keep their food healthy and fresh during storage, simultaneously reducing food waste. The use of “good bacteria” to protect food against spoilage organisms has a long, successful history, even though the molecular mechanisms are not fully understood. In this study, we show that the depletion of free manganese is a major bioprotective mechanism of lactobacilli in dairy products. High manganese uptake and intracellular storage provide a link to the distinct, nonenzymatic, manganese-catalyzed oxidative stress defense mechanism, previously described for certain lactobacilli. The evaluation of representative Lactobacillus species in our study identifies multiple relevant species groups for fungal growth inhibition via manganese depletion. Hence, through the natural mechanism of nutrient depletion, the use of dedicated bioprotective lactobacilli constitutes an attractive alternative to artificial preservation.}, number={7}, journal={APPLIED AND ENVIRONMENTAL MICROBIOLOGY}, author={Siedler, Solvej and Rau, Martin Holm and Bidstrup, Susanne and Vento, Justin M. and Aunsbjerg, Stina Dissing and Bosma, Elleke F. and McNair, Laura M. and Beisel, Chase L. and Neves, Ana Rute}, year={2020}, month={Apr} }
 @misc{vento_crook_beisel_2019, title={Barriers to genome editing with CRISPR in bacteria}, volume={46}, ISSN={["1476-5535"]}, url={https://doi.org/10.1007/s10295-019-02195-1}, DOI={10.1007/s10295-019-02195-1}, abstractNote={Abstract}, number={9-10}, journal={JOURNAL OF INDUSTRIAL MICROBIOLOGY & BIOTECHNOLOGY}, publisher={Springer Science and Business Media LLC}, author={Vento, Justin M. and Crook, Nathan and Beisel, Chase L.}, year={2019}, month={Oct}, pages={1327–1341} }
 @article{leenay_vento_shah_martino_leulier_beisel_2019, title={Genome Editing with CRISPR-Cas9 in Lactobacillus plantarum Revealed That Editing Outcomes Can Vary Across Strains and Between Methods}, volume={14}, ISSN={["1860-7314"]}, DOI={10.1002/biot.201700583}, abstractNote={Lactic-acid bacteria such as Lactobacillus plantarum are commonly used for fermenting foods and as probiotics, where increasingly sophisticated genome-editing tools are employed to elucidate and enhance these microbes' beneficial properties. The most advanced tools to date utilize an oligonucleotide or double-stranded DNA donor for recombineering and Cas9 for targeted DNA cleavage. As the associated methods are often developed in isolation for one strain, it remains unclear how different Cas9-based editing methods compare across strains. Here, this work directly compares two methods in different strains of L. plantarum: one utilizing a plasmid-encoded recombineering template and another utilizing an oligonucleotide donor and an inducible DNA recombinase. This comparison reveals one instance in which only the recombineering-template method generates desired edits and another instance in which only the oligo method generates desired edits. It is further found that both methods exhibit highly variable success editing the same site across multiple L. plantarum strains. Finally, failure modes are identified for the recombineering-template method, including a consistent genomic deletion and reversion of a point mutation in the recombineering template. This study therefore highlights surprising differences for Cas9-mediated genome editing between methods and related strains, arguing for the need for multiple, distinct methods when performing CRISPR-based editing in bacteria.}, number={3}, journal={BIOTECHNOLOGY JOURNAL}, author={Leenay, Ryan T. and Vento, Justin M. and Shah, Malay and Martino, Maria Elena and Leulier, Francois and Beisel, Chase L.}, year={2019}, month={Mar} }