The Ralstonia pseudosolanacearum Type III Effector RipL Delays Flowering and Promotes Susceptibility to Pseudomonas syringae in Arabidopsis thaliana.

The plant defense responses to microbial infection are tightly regulated and integrated with the developmental program for optimal resources allocation. Notably, the defense- associated hormone salicylic acid (SA) acts as a promoter of flowering while several plant pathogens actively target the flowering signaling pathway to promote their virulence or dissemination. Ralstonia pseudosolanacearum inject tens of effectors in the host cells that collectively promote bacterial proliferation in plant tissues. Here, we characterized the function of the broadly conserved R. pseudosolanacearum effector RipL, through heterologous expression in Arabidopsis thaliana . RipL-expressing transgenic lines presented a delayed flowering, which correlated with a low expression of flowering regulator genes. Delayed flowering was also observed in Nicotiana benthamiana plants transiently expressing RipL. In parallel, RipL promoted plant susceptibility to virulent strains of Pseudomonas syringae in the effector-expressing lines or when delivered by the type III secretion system. Unexpectedly, SA accumulation and SA-dependent immune signaling were not significantly affected by RipL expression. Rather, the RNA-seq analysis of infected RipL-expressing lines revealed that the overall amplitude of the transcriptional response was dampened, suggesting that RipL could promote plant susceptibility in an SA-independent manner. Further elucidation of the molecular mechanisms underpinning RipL effect on flowering and immunity may reveal novel effector functions in host cells.

Manipulation of the Host Endomembrane System by Bacterial Effectors.

The endomembrane system, extending from the nuclear envelope to the plasma membrane, is critical to the plant response to pathogen infection. Synthesis and transport of immunity-related proteins and antimicrobial compounds to and from the plasma membrane are supported by conventional and unconventional processes of secretion and internalization of vesicles, guided by the cytoskeleton networks. Although plant bacterial pathogens reside mostly in the apoplast, major structural and functional modifications of the endomembrane system in the host cell occur during bacterial infection. Here, we review the dynamics of these cellular compartments, briefly, for their essential contributions to the plant defense responses and, in parallel, for their emerging roles in bacterial pathogenicity. We further focus on Pseudomonas syringae, Xanthomonas spp., and Ralstonia solanacearum type III secreted effectors that one or both localize to and associate with components of the host endomembrane system or the cytoskeleton network to highlight the diversity of virulence strategies deployed by bacterial pathogens beyond the inhibition of the secretory pathway. [Formula: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY-NC-ND 4.0 International license.

Identification of RipAZ1 as an avirulence determinant of Ralstonia solanacearum in Solanum americanum.

Ralstonia solanacearum causes bacterial wilt disease in many plant species. Type III-secreted effectors (T3Es) play crucial roles in bacterial pathogenesis. However, some T3Es are recognized by corresponding disease resistance proteins and activate plant immunity. In this study, we identified the R. solanacearum T3E protein RipAZ1 (Ralstonia injected protein AZ1) as an avirulence determinant in the black nightshade species Solanum americanum. Based on the S. americanum accession-specific avirulence phenotype of R. solanacearum strain Pe_26, 12 candidate avirulence T3Es were selected for further analysis. Among these candidates, only RipAZ1 induced a cell death response when transiently expressed in a bacterial wilt-resistant S. americanum accession. Furthermore, loss of ripAZ1 in the avirulent R. solanacearum strain Pe_26 resulted in acquired virulence. Our analysis of the natural sequence and functional variation of RipAZ1 demonstrated that the naturally occurring C-terminal truncation results in loss of RipAZ1-triggered cell death. We also show that the 213 amino acid central region of RipAZ1 is sufficient to induce cell death in S. americanum. Finally, we show that RipAZ1 may activate defence in host cell cytoplasm. Taken together, our data indicate that the nucleocytoplasmic T3E RipAZ1 confers R. solanacearum avirulence in S. americanum. Few avirulence genes are known in vascular bacterial phytopathogens and ripAZ1 is the first one in R. solanacearum that is recognized in black nightshades. This work thus opens the way for the identification of disease resistance genes responsible for the specific recognition of RipAZ1, which can be a source of resistance against the devastating bacterial wilt disease.

Perception of unrelated microbe-associated molecular patterns triggers conserved yet variable physiological and transcriptional changes in Brassica rapa ssp. pekinensis.

Pattern-triggered immunity (PTI) includes the different transcriptional and physiological responses that enable plants to ward off microbial invasion. Surface-localized pattern-recognition receptors (PRRs) recognize conserved microbe-associated molecular patterns (MAMPs) and initiate a branched signaling cascade that culminate in an effective restriction of pathogen growth. In the model species Arabidopsis thaliana, early PTI events triggered by different PRRs are broadly conserved although their nature or intensity is dependent on the origin and features of the detected MAMP. In order to provide a functional basis for disease resistance in leafy vegetable crops, we surveyed the conservation of PTI events in Brassica rapa ssp. pekinensis. We identified the PRR homologs present in B. rapa genome and found that only one of the two copies of the bacterial Elongation factor-Tu receptor (EFR) might function. We also characterized the extent and unexpected specificity of the transcriptional changes occurring when B. rapa seedlings are treated with two unrelated MAMPs, the bacterial flagellin flg22 peptide and the fungal cell wall component chitin. Finally, using a MAMP-induced protection assay, we could show that bacterial and fungal MAMPs elicit a robust immunity in B. rapa, despite significant differences in the kinetic and amplitude of the early signaling events. Our data support the relevance of PTI for crop protection and highlight specific functional target for disease resistance breeding in Brassica crops.

Host adaptation and microbial competition drive Ralstonia solanacearum phylotype I evolution in the Republic of Korea.

Bacterial wilt caused by the Ralstonia solanacearum species complex (RSSC) threatens the cultivation of important crops worldwide. We sequenced 30 RSSC phylotype I (R. pseudosolanacearum) strains isolated from pepper (Capsicum annuum) and tomato (Solanum lycopersicum) across the Republic of Korea. These isolates span the diversity of phylotype I, have extensive effector repertoires and are subject to frequent recombination. Recombination hotspots among South Korean phylotype I isolates include multiple predicted contact-dependent inhibition loci, suggesting that microbial competition plays a significant role in Ralstonia evolution. Rapid diversification of secreted effectors presents challenges for the development of disease-resistant plant varieties. We identified potential targets for disease resistance breeding by testing for allele-specific host recognition of T3Es present among South Korean phyloype I isolates. The integration of pathogen population genomics and molecular plant pathology contributes to the development of location-specific disease control and development of plant cultivars with durable resistance to relevant threats.

Erratum: Ralstonia solanacearum Type III Effectors with Predicted Nuclear Localization Signal Localize to Various Cell Compartments and Modulate Immune Responses in Nicotiana spp.

[This corrects the article on p. 43 in vol. 36, PMID: 32089660.].

Ralstonia solanacearum Type III Effectors with Predicted Nuclear Localization Signal Localize to Various Cell Compartments and Modulate Immune Responses in Nicotiana spp.

Ralstonia solanacearum (Rso) is a causal agent of bacterial wilt in Solanaceae crops worldwide including Republic of Korea. Rso virulence predominantly relies on type III secreted effectors (T3Es). However, only a handful of Rso T3Es have been characterized. In this study, we investigated subcellular localization of and manipulation of plant immunity by 8 Rso T3Es predicted to harbor a nuclear localization signal (NLS). While 2 of these T3Es elicited cell death in both Nicotiana benthamiana and N. tabacum, only one was dependent on suppressor of G2 allele of skp1 (SGT1), a molecular chaperone of nucleotide-binding and leucine-rich repeat immune receptors. We also identified T3Es that differentially regulate flg22-induced reactive oxygen species production and gene expression. Interestingly, several of the NLS-containing T3Es translationally fused with yellow fluorescent protein accumulated in subcellular compartments other than the cell nucleus. Our findings bring new clues to decipher Rso T3E function in planta.