Characterization of Slimes Produced by Pseudomonas Phaseolicola in Vitro and in Planta, and the Demonstration of the Slime Polysaccharide Alginate in Seven Pseudomonas Syringae Pathovars

AbstractThe purpose of this study was to determine whether zinc phosphate treatments of tomato plants (Solanum lycopersicum L.) can attenuate bacterial speck disease severity through reduction of Pseudomonas syringae pv. tomato (Pst) growth in planta and induce morphological and biochemical plant defence responses. Tomato plants were treated with 10 ppm (25.90 µM) zinc phosphate and then spray inoculated with strain DAPP-PG 215, race 0 of Pst. Disease symptoms were recorded as chlorosis and/or necrosis per leaf (%) and as numbers of necrotic spots. Soil treatments with zinc phosphate protected susceptible tomato plants against Pst, with reductions in both disease severity and pathogen growth in planta. The reduction of Pst growth in planta combined with significantly higher zinc levels in zinc-phosphate-treated plants indicated direct antimicrobial toxicity of this microelement, as also confirmed by in vitro assays. Morphological (i.e. callose apposition) and biochemical (i.e., expression of salicylic-acid-dependent pathogenesis-related protein PR1b1 gene) defence responses were induced by the zinc phosphate treatment, as demonstrated by histochemical and qPCR analyses, respectively. In conclusion, soil treatments with zinc phosphate can protect tomato plants against Pst attacks through direct antimicrobial activity and induction of morphological and biochemical plant defence responses.

Download Full-text

HopA1 Effector from Pseudomonas syringae pv syringae Strain 61 Affects NMD Processes and Elicits Effector-Triggered Immunity

International Journal of Molecular Sciences ◽

10.3390/ijms22147440 ◽

2021 ◽

Vol 22 (14) ◽

pp. 7440

Author(s):

Shraddha K. Dahale ◽

Daipayan Ghosh ◽

Kishor D. Ingole ◽

Anup Chugani ◽

Sang Hee Kim ◽

...

Keyword(s):

Pseudomonas Syringae ◽

Disease Susceptibility ◽

Null Mutant ◽

In Planta ◽

Host Range Expansion ◽

Unique System ◽

Effector Triggered Immunity ◽

Transcriptomic Changes ◽

Immune Detection

Pseudomonas syringae-secreted HopA1 effectors are important determinants in host range expansion and increased pathogenicity. Their recent acquisitions via horizontal gene transfer in several non-pathogenic Pseudomonas strains worldwide have caused alarming increase in their virulence capabilities. In Arabidopsis thaliana, RESISTANCE TO PSEUDOMONAS SYRINGAE 6 (RPS6) gene confers effector-triggered immunity (ETI) against HopA1pss derived from P. syringae pv. syringae strain 61. Surprisingly, a closely related HopA1pst from the tomato pathovar evades immune detection. These responsive differences in planta between the two HopA1s represents a unique system to study pathogen adaptation skills and host-jumps. However, molecular understanding of HopA1′s contribution to overall virulence remain undeciphered. Here, we show that immune-suppressive functions of HopA1pst are more potent than HopA1pss. In the resistance-compromised ENHANCED DISEASE SUSCEPTIBILITY 1 (EDS1) null-mutant, transcriptomic changes associated with HopA1pss-elicited ETI are still induced and carry resemblance to PAMP-triggered immunity (PTI) signatures. Enrichment of HopA1pss interactome identifies proteins with regulatory roles in post-transcriptional and translational processes. With our demonstration here that both HopA1 suppress reporter-gene translations in vitro imply that the above effector-associations with plant target carry inhibitory consequences. Overall, with our results here we unravel possible virulence role(s) of HopA1 in suppressing PTI and provide newer insights into its detection in resistant plants.

Download Full-text

The algT Gene of Pseudomonas syringae pv. glycinea andNew Insights into the Transcriptional Organization of thealgT-muc Gene Cluster

Journal of Bacteriology ◽

10.1128/jb.01160-06 ◽

2006 ◽

Vol 188 (23) ◽

pp. 8013-8021 ◽

Cited By ~ 14

Author(s):

Alexander Schenk ◽

Michael Berger ◽

Lisa M. Keith ◽

Carol L. Bender ◽

Georgi Muskhelishvili ◽

...

Keyword(s):

Gene Cluster ◽

Pseudomonas Syringae ◽

Azotobacter Vinelandii ◽

Regulatory Gene ◽

In Planta ◽

Phytopathogenic Bacterium ◽

Reverse Transcription Pcr ◽

Alginate Production ◽

Alginate Biosynthesis

ABSTRACT The phytopathogenic bacterium Pseudomonas syringae pv. glycinea infects soybean plants and causes bacterial blight. In addition to P. syringae, the human pathogen Pseudomonas aeruginosa and the soil bacterium Azotobacter vinelandii produce the exopolysaccharide alginate, a copolymer of d-mannuronic and l-guluronic acids. Alginate production in P. syringae has been associated with increased fitness and virulence in planta. Alginate biosynthesis is tightly controlled by proteins encoded by the algT-muc regulatory gene cluster in P. aeruginosa and A. vinelandii. These genes encode the alternative sigma factor AlgT (σ22), its anti-sigma factors MucA and MucB, MucC, a protein with a controversial function that is absent in P. syringae, and MucD, a periplasmic serine protease and homolog of HtrA in Escherichia coli. We compared an alginate-deficient algT mutant of P. syringae pv. glycinea with an alginate-producing derivative in which algT is intact. The alginate-producing derivative grew significantly slower in vitro growth but showed increased epiphytic fitness and better symptom development in planta. Evaluation of expression levels for algT, mucA, mucB, mucD, and algD, which encodes an alginate biosynthesis gene, showed that mucD transcription is not dependent on AlgT in P. syringae in vitro. Promoter mapping using primer extension experiments confirmed this finding. Results of reverse transcription-PCR demonstrated that algT, mucA, and mucB are cotranscribed as an operon in P. syringae. Northern blot analysis revealed that mucD was expressed as a 1.75-kb monocistronic mRNA in P. syringae.

Download Full-text

Discovery and Characterization of Low-Molecular Weight Inhibitors of Erwinia tracheiphila

Phytopathology ◽

10.1094/phyto-11-19-0440-r ◽

2020 ◽

Vol 110 (5) ◽

pp. 989-998

Author(s):

Cláudio M. Vrisman ◽

Loïc Deblais ◽

Yosra A. Helmy ◽

Reed Johnson ◽

Gireesh Rajashekara ◽

...

Keyword(s):

Pseudomonas Syringae ◽

High Throughput Screening ◽

Pathogenic Bacteria ◽

Bacterial Species ◽

Bacillus Species ◽

Erwinia Tracheiphila ◽

Static Activity ◽

Low Toxicity

Plant pathogenic bacteria in the genus Erwinia cause economically important diseases, including bacterial wilt of cucurbits caused by Erwinia tracheiphila. Conventional bactericides are insufficient to control this disease. Using high-throughput screening, 464 small molecules (SMs) with either cidal or static activity at 100 µM against a cucumber strain of E. tracheiphila were identified. Among them, 20 SMs (SM1 to SM20), composed of nine distinct chemical moiety structures, were cidal to multiple E. tracheiphila strains at 100 µM. These lead SMs had low toxicity to human cells and honey bees at 100 µM. No phytotoxicity was observed on melon plants at 100 µM, except when SM12 was either mixed with Silwet L-77 and foliar sprayed or when delivered through the roots. Lead SMs did not inhibit the growth of beneficial Pseudomonas and Enterobacter species but inhibited the growth of Bacillus species. Nineteen SMs were cidal to Xanthomonas cucurbitae and showed >50% growth inhibition against Pseudomonas syringae pv. lachrymans. In addition, 19 SMs were cidal or static against Erwinia amylovora in vitro. Five SMs demonstrated potential to suppress E. tracheiphila when foliar sprayed on melon plants at 2× the minimum bactericidal concentration. Thirteen SMs reduced Et load in melon plants when delivered via roots. Temperature and light did not affect the activity of SMs. In vitro cidal activity was observed after 3 to 10 h of exposure to these five SMs. Here, we report 19 SMs that provide chemical scaffolds for future development of bactericides against plant pathogenic bacterial species.

Download Full-text

Immunogold Labeling of Hrp Pili of Pseudomonas syringae pv. tomato Assembled in Minimal Medium and In Planta

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi.2001.14.2.234 ◽

2001 ◽

Vol 14 (2) ◽

pp. 234-241 ◽

Cited By ~ 25

Author(s):

Wenqi Hu ◽

Jing Yuan ◽

Qiao-Ling Jin ◽

Patrick Hart ◽

Sheng Yang He

Keyword(s):

Arabidopsis Thaliana ◽

Pseudomonas Syringae ◽

Minimal Medium ◽

Leaf Tissue ◽

Hypersensitive Reaction ◽

Immunogold Labeling ◽

Structural Protein ◽

In Planta ◽

Hrp Genes

Hypersensitive reaction and pathogenicity (hrp) genes are required for Pseudomonas syringae pv. tomato (Pst) DC3000 to cause disease in susceptible tomato and Arabidopsis thaliana plants and to elicit the hypersensitive response in resistant plants. The hrp genes encode a type III protein secretion system known as the Hrp system, which in Pst DC3000 secretes HrpA, HrpZ, HrpW, and AvrPto and assembles a surface appendage, named the Hrp pilus, in hrp-gene-inducing minimal medium. HrpA has been suggested to be the Hrp pilus structural protein on the basis of copurification and mutational analyses. In this study, we show that an antibody against HrpA efficiently labeled Hrp pili, whereas antibodies against HrpW and HrpZ did not. Immunogold labeling of bacteria-infected Arabidopsis thaliana leaf tissue with an Hrp pilus antibody revealed a characteristic lineup of gold particles around bacteria and/or at the bacterium-plant contact site. These results confirm that HrpA is the major structural protein of the Hrp pilus and provide evidence that Hrp pili are assembled in vitro and in planta.

Download Full-text

Identification of a putative DNA-binding protein in Arabidopsis that acts as a susceptibility hub and interacts with multiple Pseudomonas syringae effectors

Molecular Plant-Microbe Interactions ◽

10.1094/mpmi-10-20-0291-r ◽

2020 ◽

Author(s):

Karl Schreiber ◽

Jennifer D Lewis

Keyword(s):

Pseudomonas Syringae ◽

Transcriptional Profiling ◽

Effector Proteins ◽

Ribosomal Protein Genes ◽

Loss Of Function ◽

Wild Type ◽

In Planta ◽

Arabidopsis Protein ◽

Wide Range

Phytopathogens use secreted effector proteins to suppress host immunity and promote pathogen virulence, and there is increasing evidence that the host-pathogen interactome comprises a complex network. In an effort to identify novel interactors of the Pseudomonas syringae effector HopZ1a, we performed a yeast two-hybrid screen that identified a previously uncharacterized Arabidopsis protein that we designate HopZ1a Interactor 1 (ZIN1). Additional analyses in yeast and in planta revealed that ZIN1 also interacts with several other P. syringae effectors. We show that an Arabidopsis loss-of-function zin1 mutant is less susceptible to infection by certain strains of P. syringae, while overexpression of ZIN1 results in enhanced susceptibility. Functionally, ZIN1 exhibits topoisomerase-like activity in vitro. Transcriptional profiling of wild-type and zin1 Arabidopsis plants inoculated with P. syringae indicated that while ZIN1 regulates a wide range of pathogen-responsive biological processes, the list of genes more highly expressed in zin1 versus wild-type plants was particularly enriched for ribosomal protein genes. Altogether, these data illuminate ZIN1 as a potential susceptibility hub that interacts with multiple effectors to influence the outcome of plant-microbe interactions.

Download Full-text

Molecular characterization of differences between the tomato immune receptors Fls3 and Fls2

10.1101/2020.02.04.934133 ◽

2020 ◽

Author(s):

Robyn Roberts ◽

Alexander E. Liu ◽

Lingwei Wan ◽

Annie M. Geiger ◽

Sarah R. Hind ◽

...

Keyword(s):

Pseudomonas Syringae ◽

Molecular Mechanisms ◽

Bacterial Pathogen ◽

Transcript Abundance ◽

Defense Responses ◽

Structural Features ◽

Host Responses ◽

Pathogen Invasion

AbstractPlants mount defense responses by recognizing indications of pathogen invasion, including microbe-associated molecular patterns (MAMPs). Flagellin from the bacterial pathogen Pseudomonas syringae pv. tomato (Pst) contains two MAMPs, flg22 and flgII-28, that are recognized by tomato receptors Flagellin sensing 2 (Fls2) and Flagellin sensing 3 (Fls3), respectively. It is unknown to what degree each receptor contributes to immunity and if they promote immune responses using the same molecular mechanisms. Characterization of CRISPR/Cas9-generated Fls2 and Fls3 tomato mutants revealed that the two receptors contribute equally to disease resistance both on the leaf surface and in the apoplast. However, striking differences were observed in certain host responses mediated by the two receptors. Compared to Fls2, Fls3 mediated a more sustained production of reactive oxygen species (ROS) and an increase in transcript abundance of 44 tomato genes, with two genes serving as reporters for Fls3. Fls3 had greater in vitro kinase activity and interacted differently with the Pst effector AvrPtoB as compared to Fls2. Using chimeric Fls2/Fls3 proteins, we found that no receptor domain was solely responsible for the Fls3 sustained ROS, suggesting involvement of multiple structural features. This work reveals differences in the immunity outputs between Fls2 and Fls3, suggesting they use distinct molecular mechanisms to activate pattern-triggered immunity in response to flagellin-derived MAMPs.

Download Full-text

The ecological genetics ofPseudomonas syringaefrom kiwifruit leaves

10.1101/235853 ◽

2017 ◽

Cited By ~ 1

Author(s):

Christina Straub ◽

Elena Colombi ◽

Li Li ◽

Hongwen Huang ◽

Matthew D. Templeton ◽

...

Keyword(s):

Population Structure ◽

Pseudomonas Syringae ◽

Bacterial Pathogen ◽

Ecological Factors ◽

Ecological Genetics ◽

Plant Interactions ◽

In Planta ◽

Canker Disease ◽

Pathogen Populations

SUMMARYInteractions between commensal microbes and invading pathogens are understudied, despite their likely effects on pathogen population structure and infection processes. We describe the population structure and genetic diversity of a broad range of co-occurringPseudomonas syringaeisolated from infected and uninfected kiwifruit during an outbreak of bleeding canker disease caused byP. syringaepv.actinidiae(Psa) in New Zealand. Overall population structure was clonal and affected by ecological factors including infection status and cultivar. Most isolates are members of a new clade in phylogroup 3 (PG3a), also present on kiwifruit leaves in China and Japan. Stability of the polymorphism between pathogenicPsaand commensalP. syringaePG3a isolated from the same leaf was tested using reciprocal invasion from rare assaysin vitroand in planta.P. syringaeG33C (PG3a) inhibitedPsaNZ54, while the presence ofPsaNZ54 enhanced the growth ofP. syringaeG33C. This effect could not be attributed to virulence activity encoded by the Type 3 secretion system ofPsa. Together our data contribute toward the development of an ecological perspective on the genetic structure of pathogen populations.ORIGINALITY-SIGNIFICANT STATEMENTBacterial pathogen populations are often studied with little consideration of co-occurring microbes and yet interactions between pathogens and commensals can affect both population structure and disease progression. A fine-scale sampling of commensals present on kiwifruit leaves during an outbreak of bleeding canker disease caused byP. syringaepv.actinidiaereveals a clonal population structure. A new clade of non-pathogenicP. syringae(PG3a) appears to be associated with kiwifruit on a global scale. The presence of PG3a on kiwifruit has significant effects on the outcome of infection byP. syringaepv.actinidiae. This emphasises the value of studying the effect of co-occurring bacteria on pathogen-plant interactions.

Download Full-text