Biology subjects

Pfeilmeier, S.

Publications and source records attributed to Pfeilmeier, S..

6 recordsLinked to original sources

bioRxiv2024

Four cell wall-degrading enzymes of Xanthomonas campestris pv. campestris determine bacterial escape from hydathodes to the leaf vasculature

To colonize plants, pathogenic bacteria modulate the biology of the host employing different bacterial secretion systems. For example, the type II secretion system (T2SS) releases toxins, proteases, lipases and carbohydrate-degrading enzymes into the extracellular environment to promote tissue softening and soft rot. In this way, the T2SS promotes virulence of phytopathogenic Gram-negative bacteria. However, the role of the T2SS and its substrates for vascular disease remains enigmatic. Here, we show that the Xps-T2SS allows Xanthomonas campestris pv. campestris (Xcc) to breach the tissue barrier between hydathodes- the initial bacterial entry point - and xylem thereby gaining access to the leaf vasculature. Yet, Xps-T2SS was dispensable for bacterial multiplication in the leaf apoplast or inside the hydathode cavity suggesting a role beyond plant defense suppression or nutrient acquisition. Using comparative genomics, four plant cell wall-degrading enzymes (CWDEs) were found to be associated with vascular pathogenesis. Testing gene knockout combinations of those enzymes revealed that virulence of only the quadruple CWDE mutant was down to the level of the xps-T2SS mutant. Our results thus demonstrate that the Xps-T2SS and a set of CWDEs that is likely secreted by this system allow Xcc to break this tissue barrier enabling long-distance mobility of Xcc inside the host plant. We thus expand our understanding on how certain bacterial pathogens have specialized towards vascular pathogens.

plant biology↗

bioRxiv2024

Non-invasive, bioluminescence-based visualization and quantification of bacterial infections in Arabidopsis over time

Plant pathogenic bacteria use various entry strategies to colonize their host, like entering through natural openings and wounds in leaves and roots. The vascular pathogen Xanthomonas campestris pv. campestris (Xcc) enters through hydathodes, organs at the leaf margin involved in guttation. Subsequently, Xcc breaks out from infected hydathodes, progressing into the xylem vessels and causing systemic disease. To elucidate the mechanisms that underpin the different stages of Xcc pathogenesis, a need exists to image Xcc progression in planta in a non-invasive manner. Here, we describe a phenotyping setup and Python image analysis pipeline capturing the Xcc infection in 16 Arabidopsis thaliana plants in parallel over time. The setup used both an RGB to capture disease symptoms and an ultra-sensitive CCD camera to monitor bacterial progression inside the leaves using bioluminescence. We demonstrate that the image analysis pipeline reliably quantifies bacterial growth in planta for two bacterial species, that is vascular Xcc and the mesophyll pathogen Pseudomonas syringae pv. tomato. The resolution of the camera allowed early detection of Xcc in the hydathodes, yielding valuable information on this early stage of the Xcc infection process. The data obtained through the automated image analysis pipeline was robust and validated findings from other bioluminescence imaging methods, while requiring fewer samples. We can thus quantify the resistance level of a large number of Arabidopsis thaliana accessions and mutant lines to different bacterial strains in a non-invasive manner for phenotypic screenings.

plant biology↗

bioRxiv2024

Hydathode immunity against the vascular pathogen Xanthomonas campestris pv. campestris by the Arabidopsis CNL-type receptor SUT1

Bacterial plant pathogens exploit natural openings, such as pores or wounds, to enter the plant interior and cause disease. Plants guard these openings through defense mechanisms. However, bacteria from the genus Xanthomonas have specialized in that they enter their host via a special entry point, the hydathode--an organ at the leaf margin involved in xylem sap guttation. Hydathodes can mount an immune response against bacteria, including non-adapted and adapted pathogens like X. campestris pv. campestris (Xcc) that cause vascular disease. Previously, it was shown that the RKS1/ZAR1 immune complex confers vascular resistance against Xcc by recognizing XopAC activity, a type III effector (T3E). However, in absence of XopAC recognition, Arabidopsis Col-0 hydathodes still display resistance against Xcc. Here we mapped the causal gene using an inoculation method that promotes Xcc hydathode entry. Using a population of Recombinant Inbred Lines (RILs) of a cross between a susceptible (Oy-0) and resistant accession (Col-0), a major QTL for Xcc resistance was found on the right arm of Chromosome 5 in Col-0. Combining this result with a genome-wide association analysis yielded a single candidate gene encoding a coiled-coil nucleotide-binding leucine-rich repeat (CNL-type) immune receptor protein called SUPPRESSOR OF TOPP4 1 (SUT1). Expression of SUT1 was confirmed in hydathodes. We reveal that RKS1/ZAR1 and SUT1 confer different levels of Xcc resistance in different tissue types. Both RKS1/ZAR1 and SUT1 are alone sufficient for Xcc resistance in Col-0 hydathodes. However, RKS1/ZAR1 resistance is also effective in tissue types that represent late infection stages, i.e. xylem and mesophyll. In contrast, SUT1 resistance is not effective in the xylem, while weakly additive to RKS1/ZAR1 in the mesophyll. We thus identify a novel R gene, SUT1, that confers Xcc resistance primarily early in the infection during hydathode colonization. Author summaryBlack rot disease, caused by the bacterial pathogen Xanthomonas campestris pv. campestris (Xcc), is an economically important disease of cabbage crops. Xcc is unique in that it enters the plant interior through specialized organs at the leaf margin. These structures called hydathodes contain water pores and are involved in root pressure regulation. Although we know that hydathodes can mount an immune response against these bacteria, specific immune receptors still need to be discovered. Here we use the model plant Arabidopsis thaliana to map an hydathode-effective resistance gene. By screening two different Arabidopsis populations, we could map a single gene, SUT1, that is involved in this resistance. SUT1 restricts the early hydathode colonization by Xcc thus suppressing disease progression. Interestingly, SUT1 resistance was not effective in the plant vascular system, which the bacteria subsequently colonize. Therefore, this study provides a new insight into the role of hydathodes in anti-bacterial resistance in plants and opens the door for research on tissue- and organ-specific resistance mechanisms.

plant biology↗

bioRxiv2023

Dysbiosis of a leaf microbiome is caused by enzyme secretion of opportunistic Xanthomonas strains

Dysbiosis is characterized by a perturbed microbiota associated with host disease. In both plants and animals, the innate immune system contributes to maintain microbiota homeostasis in healthy organisms, with NADPH oxidases playing a crucial role. In Arabidopsis thaliana, the absence of NADPH oxidase RBOHD can lead to an altered leaf microbiota, including an enrichment of opportunistic Xanthomonas pathogens. It is currently unclear whether the microbiota change occurs independently of the opportunistic pathogens or is caused by the latter, and which virulence factors of Xanthomonas are essential for its opportunistic lifestyle. Here, we found that the opportunistic Xanthomonas strains secrete a cocktail of cell wall degrading enzymes via the type-2 secretion system (T2SS) that degrade leaf tissue and promote Xanthomonas growth during plant infection. Both disease severity and leaf degradation activity were increased in rbohD compared to Col-0 plants, attesting to the opportunistic behaviour of the Xanthomonas strains on immune compromised plants. Using gnotobiotic plant experiments with a synthetic bacterial community of more than 100 commensal strains and drop-in of Xanthomonas wildtype or mutant strains revealed that T2SS-dependent virulence is required for plant disease and for the shift in microbiota composition. Overall, our data indicate that a single opportunistic pathogen can drive community shifts, here caused by tissue damage in leaves, creating an environment in which specific commensal bacteria can thrive.

plant biology↗

bioRxiv2021

Pseudomonas syringae addresses distinct environmental challenges during plant infection through the coordinated deployment of polysaccharides

Prior to infection, phytopathogenic bacteria face a challenging environment on the plant surface, where they are exposed to nutrient starvation and abiotic stresses. Pathways enabling surface adhesion, stress tolerance and epiphytic survival are important for successful plant pathogenesis. Understanding the roles and regulation of these pathways is therefore crucial to fully understand bacterial plant infections. The phytopathogen Pseudomonas syringae pv. tomato (Pst) encodes multiple polysaccharides that are implicated in biofilm formation, stress survival and virulence in other microbes. To examine how these polysaccharides impact Pst epiphytic survival and pathogenesis, we analysed mutants in multiple polysaccharide loci to determine their intersecting contributions to epiphytic survival and infection. In parallel, we used qRT-PCR to analyse the regulation of each pathway. Pst polysaccharides are tightly coordinated by multiple environmental signals. Nutrient availability, temperature and surface association strongly affect the expression of different polysaccharides under the control of the signalling proteins ladS and cbrB and the second messenger cyclic-di-GMP. Furthermore, functionally redundant, combinatorial phenotypes were observed for several polysaccharides. Exopolysaccharides and WapQ-mediated lipopolysaccharide production are important for leaf adhesion, while -glucan and alginate together confer desiccation tolerance. Our results suggest that polysaccharides play important roles in overcoming environmental challenges to Pst during plant infection. HighlightPseudomonas syringae uses the coordinated deployment of polysaccharides to address environmental challenges during plant colonization. Functional redundancy renders individual polysaccharides dispensable during plant infection, but their combined loss impedes pathogenicity.

microbiology↗

bioRxiv2017

Heterologous expression of the immune receptor EFR in Medicago truncatula reduces pathogenic infection, but not rhizobial symbiosis

Interfamily transfer of plant pattern recognition receptors (PRRs) represents a promising biotechnological approach to engineer broad-spectrum, and potentially durable, disease resistance in crops. It is however unclear whether new recognition specificities to given pathogen-associated molecular patterns (PAMPs) affect the interaction of the recipient plant with beneficial microbes. To test this in a direct reductionist approach, we transferred the Brassicaceae-specific PRR ELONGATION FACTOR-THERMO UNSTABLE RECEPTOR (EFR) from Arabidopsis thaliana to the legume Medicago truncatula, conferring recognition of the bacterial EF-Tu protein. Constitutive EFR expression led to EFR accumulation and activation of immune responses upon treatment with the EF-Tu-derived elf18 peptide in leaves and roots. The interaction of M. truncatula with the bacterial symbiont Sinorhizobium meliloti is characterized by the formation of root nodules that fix atmospheric nitrogen. Although nodule numbers were slightly reduced at an early stage of the infection in EFR-Medicago when compared to control lines, nodulation was similar in all lines at later stages. Furthermore, nodule colonization by rhizobia, and nitrogen fixation were not compromised by EFR expression. Importantly, the M. truncatula lines expressing EFR were substantially more resistant to the root bacterial pathogen Ralstonia solanacearum. Our data suggest that the transfer of EFR to M. truncatula does not impede root nodule symbiosis, but has a positive impact on disease resistance against a bacterial pathogen. In addition, our results indicate that Rhizobium can either avoid PAMP recognition during the infection process, or is able to actively suppress immune signaling.\n\nSignificance StatementCrop engineering helps reducing the economic and environmental costs of plant disease. The genetic transfer of immune receptors across plant species is a promising biotechnological approach to increase disease resistance. Surface-localized pattern-recognition receptors (PRRs), which detect conserved characteristic microbial features, are functional in heterologous taxonomically-diverse plant species, and confer broad-spectrum disease resistance. It was unclear whether PRR transfer negatively impacts the association of the recipient plants with symbiotic microbes. Here, we show that a legume engineered with a novel PRR recognizing a conserved bacterial protein becomes more resistant to an important bacterial pathogen without significant impact on nitrogen-fixing symbiosis with rhizobia. This finding is of particular relevance as attempts to transfer this important symbiosis into non-legume plants are ongoing.

plant biology↗