Virulence, Host-Selective Toxin Production, and Development of Three Cochliobolus Phytopathogens Lacking the Sfp-Type 4'-Phosphopantetheinyl Transferase Ppt1.

The Sfp-type 4'-phosphopantetheinyl transferase Ppt1 is required for activation of nonribosomal peptide synthetases, including α-aminoadipate reductase (AAR) for lysine biosynthesis and polyketide synthases, enzymes that biosynthesize peptide and polyketide secondary metabolites, respectively. Deletion of the PPT1 gene, from the maize pathogen Cochliobolus heterostrophus and the rice pathogen Cochliobolus miyabeanus, yielded strains that were significantly reduced in virulence to their hosts. In addition, ppt1 mutants of C. heterostrophus race T and Cochliobolus victoriae were unable to biosynthesize the host-selective toxins (HST) T-toxin and victorin, respectively, as judged by bioassays. Interestingly, ppt1 mutants of C. miyabeanus were shown to produce tenfold higher levels of the sesterterpene-type non-HST ophiobolin A, as compared with the wild-type strain. The ppt1 strains of all species were also reduced in tolerance to oxidative stress and iron depletion; both phenotypes are associated with inability to produce extracellular siderophores biosynthesized by the nonribosomal peptide synthetase Nps6. Colony surfaces were hydrophilic, a trait previously associated with absence of C. heterostrophus Nps4. Mutants were decreased in asexual sporulation and C. heterostrophus strains were female-sterile in sexual crosses; the latter phenotype was observed previously with mutants lacking Nps2, which produces an intracellular siderophore. As expected, mutants were albino, since they cannot produce the polyketide melanin and were auxotrophic for lysine because they lack an AAR.

Improvement of fitness and biocontrol properties of Pseudomonas putida via an extracellular heme peroxidase.

The extracellular 373-kDa PehA heme peroxidase of Pseudomonas putida KT2440 has two enzymatic domains which depend on heme cofactor for their peroxidase activity. A null pehA mutant was generated to examine the impact of PehA in rhizosphere colonization competence and the induction of plant systemic resistance (ISR). This mutant was not markedly hampered in colonization efficiency. However, increase in pehA dosage enhanced colonization fitness about 30 fold in the root and 900 fold in the root apex. In vitro assays with purified His-tagged enzymatic domains of PehA indicated that heme-dependent peroxidase activity was required for the enhancement of root tip colonization. Evaluation of live/dead cells confirmed that overexpression of pehA had a positive effect on bacterial cell viability. Following root colonization of rice plants by KT2440 strain, the incidence of rice blast caused by Magnaporthe oryzae was reduced by 65% and the severity of this disease was also diminished in comparison to non-treated plants. An increase in the pehA dosage was also beneficial for the control of rice blast as compared with gene inactivation. The results suggest that PehA helps P. putida to cope with the plant-imposed oxidative stress leading to enhanced colonization ability and concomitant ISR-elicitation.

Comparative chemical screening and genetic analysis reveal tentoxin as a new virulence factor in Cochliobolus miyabeanus, the causal agent of brown spot disease on rice.

Brown spot disease, caused by Cochliobolus miyabeanus, is currently considered to be one of the most important yield reducers of rice (Oryza sativa L.). Despite its agricultural importance, little is known about the virulence mechanisms deployed by the fungus. Therefore, we set out to identify novel virulence factors with a role in disease development. This article reports, for the first time, the production of tentoxin by C. miyabeanus as a virulence factor during brown spot disease and the identification of the non-ribosomal protein synthetase (NRPS) CmNps3, responsible for tentoxin biosynthesis. We compared the chemical compounds produced by C. miyabeanus strains differing in virulence ability using ultra-high-performance liquid chromatography (UHPLC) coupled to high-resolution Orbitrap mass spectrometry (HRMS). The production of tentoxin by a highly virulent strain was revealed by principal component analysis of the detected ions and confirmed by UHPLC coupled to tandem-quadrupole mass spectrometry (MS/MS). The corresponding NRPS was identified by in silico genome analysis and confirmed by gene deletion. Infection tests with wild-type and Cmnps3 mutants showed that tentoxin acts as a virulence factor and is correlated with chlorosis development during the second phase of infection. Although rice has previously been classified as a tentoxin-insensitive plant species, our data demonstrate that tentoxin production by C. miyabeanus affects symptom development.

Connecting growth and defense: the emerging roles of brassinosteroids and gibberellins in plant innate immunity.

Brassinosteroids (BRs) and gibberellins (GAs) are two groups of phytohormones that regulate many common developmental processes throughout the plant life cycle. Fueled by large-scale 'omics' technologies and the burgeoning field of plant computational biology, the past few years have witnessed paradigm-shifting advances in our understanding of how BRs and GA are perceived and their signals transduced. Accumulating evidence also implicates BR and GA in the coordination and integration of plant immune responses. Similarly to other growth regulators, BR and GA play ambiguous roles in molding pathological outcomes, the effects of which may depend not only on the pathogen's lifestyle and infection strategy, but also on specialized features of each interaction. Analysis of the underpinning molecular mechanisms points to a crucial role of GA-inhibiting DELLA proteins and the BR-regulated transcription factor BZR1. Acting at the interface of developmental and defense signaling, these proteins likely serve as central hubs for pathway crosstalk and signal integration, allowing appropriate modulation of plant growth and defense in response to various stimuli. In this review, we outline the latest discoveries dealing with BR and GA modulation of plant innate immunity and highlight interactions between BR and GA signaling, plant defense, and microbial virulence.