Browsing by Person "Pfannstiel, Jens"

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Characterization of Bacillus velezensis UTB96, demonstrating improved lipopeptide production compared to the strain B. velezensis FZB42
(2022) Vahidinasab, Maliheh; Adiek, Isabel; Hosseini, Behnoush; Akintayo, Stephen Olusanmi; Abrishamchi, Bahar; Pfannstiel, Jens; Henkel, Marius; Lilge, Lars; Vögele, Ralf ; Hausmann, Rudolf
Bacillus strains can produce various lipopeptides, known for their antifungal properties. This makes them attractive metabolites for applications in agriculture. Therefore, identification of productive wild-type strains is essential for the development of biopesticides. Bacillus velezensis FZB42 is a well-established strain for biocontrol of plant pathogens in agriculture. Here, we characterized an alternative strain, B. velezensis UTB96, that can produce higher amounts of all three major lipopeptide families, namely surfactin, fengycin, and iturin. UTB96 produces iturin A. Furthermore, UTB96 showed superior antifungal activity towards the soybean fungal pathogen Diaporthe longicolla compared to FZB42. Moreover, the additional provision of different amino acids for lipopeptide production in UTB96 was investigated. Lysine and alanine had stimulatory effects on the production of all three lipopeptide families, while supplementation of leucine, valine and isoleucine decreased the lipopeptide bioproduction. Using a 45-litre bioreactor system for upscaling in batch culture, lipopeptide titers of about 140 mg/L surfactin, 620 mg/L iturin A, and 45 mg/L fengycin were achieved. In conclusion, it becomes clear that B. velezensis UTB96 is a promising strain for further research application in the field of agricultural biological controls of fungal diseases.
Characterization ofantifungal properties of lipopeptide-producing Bacillus velezensis strains and their proteome-based response to the phytopathogens, Diaporthe spp
(2023) Akintayo, Stephen Olusanmi; Hosseini, Behnoush; Vahidinasab, Maliheh; Messmer, Marc; Pfannstiel, Jens; Bertsche, Ute; Hubel, Philipp; Henkel, Marius; Hausmann, Rudolf; Vögele, Ralf; Lilge, Lars
Introduction: B. velezensis strains are of interest in agricultural applications due to their beneficial interactions with plants, notable through their antimicrobial activity. The biocontrol ability of two new lipopeptides-producing B. velezensis strains ES1-02 and EFSO2-04, against fungal phytopathogens of Diaporthe spp., was evaluated and compared with reference strains QST713 and FZB42. All strains were found to be effective against the plant pathogens, with the new strains showing comparable antifungal activity to QST713 and slightly lower activity than FZB42. Methods: Lipopeptides and their isoforms were identified by high-performance thin-layer chromatography (HPTLC) and mass spectrometric measurements. The associated antifungal influences were determined in direct in vitro antagonistic dual culture assays, and the inhibitory growth effects on Diaporthe spp. as representatives of phytopathogenic fungi were determined. The effects on bacterial physiology of selected B. velezensis strains were analyzed by mass spectrometric proteomic analyses using nano-LC-MS/MS. Results and Discussion: Lipopeptide production analysis revealed that all strains produced surfactin, and one lipopeptide of the iturin family, including bacillomycin L by ES1-02 and EFSO2-04, while QST713 and FZB42 produced iturin A and bacillomycin D, respectively. Fengycin production was however only detected in the reference strains. As a result of co-incubation of strain ES1-02 with the antagonistic phytopathogen D. longicolla, an increase in surfactin production of up to 10-fold was observed, making stress induction due to competitors an attractive strategy for surfactin bioproduction. An associated global proteome analysis showed a more detailed overview about the adaptation and response mechanisms of B. velezensis, including an increased abundance of proteins associated with the biosynthesis of antimicrobial compounds. Furthermore, higher abundance was determined for proteins associated with oxidative, nitrosative, and general stress response. In contrast, proteins involved in phosphate uptake, amino acid transport, and translation were decreased in abundance. Altogether, this study provides new insights into the physiological adaptation of lipopeptide-producing B. velezensis strains, which show the potential for use as biocontrol agents with respect to phytopathogenic fungi.
High-resolution proteomics reveals differences in the proteome of spelt and bread wheat flour representing targets for research on wheat sensitivities
(2020) Afzal, Muhammad; Pfannstiel, Jens; Zimmermann, Julia; Bischoff, Stephan C.; Würschum, Tobias; Longin, C. Friedrich H.
Wheat consumption can trigger celiac disease, allergic reactions and non-celiac wheat sensitivity (NCWS) in humans. Some people with NCWS symptoms claim a better tolerability of spelt compared to bread wheat products. We therefore investigated potential differences in the proteomes of spelt and bread wheat flour using nano LC–ESI–MS/MS on a set of 15 representative varieties for each of the two species. Based on the bread wheat reference, we detected 3,050 proteins in total and for most of them the expression was mainly affected by the environment. By contrast, 274 and 409 proteins in spelt and bread wheat, respectively, had a heritability ≥ 0.4 highlighting the potential to influence their expression level by varietal choice. We found 84 and 193 unique proteins for spelt and bread wheat, respectively, and 396 joint proteins, which expression differed significantly (p ≤ 0.05) when comparing both species. Thus, about one third of proteins differed significantly between spelt and bread wheat. Of them, we identified 81 proteins with high heritability, which therefore might be interesting candidates for future research on wheat hypersensitivities.
Mineral and phytic acid content as well as phytase activity in flours and breads made from different wheat species
(2023) Longin, C. Friedrich. H.; Afzal, Muhammad; Pfannstiel, Jens; Bertsche, Ute; Melzer, Tanja; Ruf, Andrea; Heger, Christoph; Pfaff, Tobias; Schollenberger, Margit; Rodehutscord, Markus
Wheat is of high importance for a healthy and sustainable diet for the growing world population, partly due to its high mineral content. However, several minerals are bound in a phytate complex in the grain and unavailable to humans. We performed a series of trials to compare the contents of minerals and phytic acid as well as phytase activity in several varieties from alternative wheat species spelt, emmer and einkorn with common wheat. Additionally, we investigated the potential of recent popular bread making recipes in German bakeries to reduce phytic acid content, and thus increase mineral bioavailability in bread. For all studied ingredients, we found considerable variance both between varieties within a species and across wheat species. For example, whole grain flours, particularly from emmer and einkorn, appear to have higher mineral content than common wheat, but also a higher phytic acid content with similar phytase activity. Bread making recipes had a greater effect on phytic acid content in the final bread than the choice of species for whole grain flour production. Recipes with long yeast proofing or sourdough and the use of whole grain rye flour in a mixed wheat bread minimized the phytic acid content in the bread. Consequently, optimizing food to better nourish a growing world requires close collaboration between research organizations and practical stakeholders ensuring a streamlined sustainable process from farm to fork.
New approaches to manage Asian soybean rust (Phakopsora pachyrhizi) using Trichoderma spp. or their antifungal secondary metabolites
(2022) El-Hasan, Abbas; Walker, Frank; Klaiber, Iris; Schöne, Jochen; Pfannstiel, Jens; Voegele, Ralf T.
Attempts have been made to determine the in vitro and in planta suppressive potential of particular Trichoderma strains (T16 and T23) and their secondary metabolites (SMs) against Asian soybean rust (ASR) incited by Phakopsora pachyrhizi. Aside from the previously identified SMs 6-pentyl-α-pyrone (6PAP) and viridiofungin A (VFA), the chemical structures of harzianic acid (HA), iso-harzianic acid (iso-HA), and harzianolide (HZL) were characterized in this study. Our results indicate that exposure of urediospores to 200 ppm 6PAP completely inhibits germination. A slightly higher dosage (250 ppm) of HZL and VFA reduces germination by 53.7% and 44%, respectively. Germ tube elongation seems more sensitive to 6PAP than urediospore germination. On detached leaves, application of conidia of T16 and T23 results in 81.4% and 74.3% protection, respectively. Likewise, 200 ppm 6PAP recorded the highest ASR suppression (98%), followed by HZL (78%) and HA (69%). Treatment of undetached leaves with 6PAP, HA, or HZL reduces ASR severity by 84.2%, 65.8%, and 50.4%, respectively. Disease reduction on the next, untreated trifoliate by T23 (53%), T16 (41%), HZL (42%), and 6PAP (32%) suggests a translocation or systemic activity of the SMs and their producers. To our knowledge, this study provides the first proof for controlling ASR using antifungal SMs of Trichoderma. Our findings strongly recommend the integration of these innovative metabolites, particularly 6PAP and/or their producers in ASR management strategies.
Surfactin shows relatively low antimicrobial activity against Bacillus subtilis and other bacterial model organisms in the absence of synergistic metabolites
(2022) Lilge, Lars; Ersig, Nadine; Hubel, Philipp; Aschern, Moritz; Pillai, Evelina; Klausmann, Peter; Pfannstiel, Jens; Henkel, Marius; Morabbi Heravi, Kambiz; Hausmann, Rudolf
Surfactin is described as a powerful biosurfactant and is natively produced by Bacillus subtilis in notable quantities. Among other industrially relevant characteristics, antimicrobial properties have been attributed to surfactin-producing Bacillus isolates. To investigate this property, stress approaches were carried out with biotechnologically established strains of Corynebacterium glutamicum, Bacillus subtilis, Escherichia coli and Pseudomonas putida with the highest possible amounts of surfactin. Contrary to the popular opinion, the highest growth-reducing effects were detectable in B. subtilis and E. coli after surfactin treatment of 100 g/L with 35 and 33%, respectively, while P. putida showed no growth-specific response. In contrast, other antimicrobial biosurfactants, like rhamnolipids and sophorolipids, showed significantly stronger effects on bacterial growth. Since the addition of high amounts of surfactin in defined mineral salt medium reduced the cell growth of B. subtilis by about 40%, the initial stress response at the protein level was analyzed by mass spectrometry, showing induction of stress proteins under control of alternative sigma factors σB and σW as well as the activation of LiaRS two-component system. Overall, although surfactin is associated with antimicrobial properties, relatively low growth-reducing effects could be demonstrated after the surfactin addition, challenging the general claim of the antimicrobial properties of surfactin.
Toward effects of hydrophobicity on biosurfactant production by Bacillus subtilis isolates from crude-oil-exposed environments
(2024) Hashemi, Seyedeh Zahra; Fooladi, Jamshid; Vahidinasab, Maliheh; Hubel, Philipp; Pfannstiel, Jens; Pillai, Evelina; Hrenn, Holger; Hausmann, Rudolf; Lilge, Lars
Background: Due to their structural features, biosurfactants reveal promising physicochemical properties, making them interesting for various applications in different fields, such as the food, cosmetics, agriculture, and bioremediation sectors. In particular, the bioproduction of surfactin, one of the most potent microbially synthesized biosurfactant molecules, is of great interest. However, since the wild-type productivities are comparably low, stimulatory environmental conditions have to be identified for improved bioproduction This study aims to find a correlation between the hydrophobicity and production of the biosurfactant surfactin by B. subtilis isolates from crude-oil-contaminated soil and water. Methods: The surfactin production yield was characterized in adapted batch cultivations using high-performance thin-layer liquid chromatography (HPTLC). Defined hydrophobic environmental conditions were achieved by supplementation with hexadecane or polystyrene beads, and the effects on biosurfactant production were measured. Adaptations at the protein level were analyzed using mass spectrometry measurements. Results: The correlation between hydrophobicity and surfactin production was characterized using Bacillus subtilis strains ZH1 and P7 isolated from crude-oil-contaminated soil and water. Since these isolates show the biodegradation of crude oil and hexadecane as hydrophobic substrates, respectively, a first-time approach, using polystyrene beads, was applied to provide a hydrophobic environment. Interestingly, contrary to popular opinion, reduced biosurfactant production was determined. Using mass spectrometric approaches, the physiological effects of co-cultivation and the cellular response at the protein level were investigated, resulting in altered quantities of stress proteins and proteins involved in the carbon metabolism counter to polystyrene beads. Conclusions: Contrary to common opinion, increasing hydrophobicity does not have a stimulating effect, and even reduces the effect on the bioproduction of surfactin as the main biosurfactant using selected B. subtilis strains.