In Planta Study Localizes an Effector Candidate from Austropuccinia psidii Strain MF-1 to the Nucleus and Demonstrates In Vitro Cuticular Wax-Dependent Differential Expression.

Austropuccinia psidii is a biotrophic fungus that causes myrtle rust. First described in Brazil, it has since spread to become a globally important pathogen that infects more than 480 myrtaceous species. One of the most important commercial crops affected by A. psidii is eucalypt, a widely grown forestry tree. The A. psidii-Eucalyptus spp. interaction is poorly understood, but pathogenesis is likely driven by pathogen-secreted effector molecules. Here, we identified and characterized a total of 255 virulence effector candidates using a genome assembly of A. psidii strain MF-1, which was recovered from Eucalyptus grandis in Brazil. We show that the expression of seven effector candidate genes is modulated by cell wax from leaves sourced from resistant and susceptible hosts. Two effector candidates with different subcellular localization predictions, and with specific gene expression profiles, were transiently expressed with GFP-fusions in Nicotiana benthamiana leaves. Interestingly, we observed the accumulation of an effector candidate, Ap28303, which was upregulated under cell wax from rust susceptible E. grandis and described as a peptidase inhibitor I9 domain-containing protein in the nucleus. This was in accordance with in silico analyses. Few studies have characterized nuclear effectors. Our findings open new perspectives on the study of A. psidii-Eucalyptus interactions by providing a potential entry point to understand how the pathogen manipulates its hosts in modulating physiology, structure, or function with effector proteins.

Meta-omics integration approach reveals the effect of soil native microbiome diversity in the performance of inoculant Azospirillum brasilense.

Plant growth promoting bacteria (PGPB) have been used as integrative inputs to minimize the use of chemical fertilizers. However, a holistic comprehension about PGPB-plant-microbiome interactions is still incipient. Furthermore, the interaction among PGPB and the holobiont (host-microbiome association) represent a new frontier to plant breeding programs. We aimed to characterize maize bulk soil and rhizosphere microbiomes in irradiated soil (IS) and a native soil (NS) microbial community gradient (dilution-to-extinction) with Azospirillum brasilense Ab-V5, a PGPB commercial inoculant. Our hypothesis was that plant growth promotion efficiency is a result of PGPB niche occupation and persistence according to the holobiont conditions. The effects of Ab-V5 and NS microbial communities were evaluated in microcosms by a combined approach of microbiomics (species-specific qPCR, 16S rRNA metataxonomics and metagenomics) and plant phenomics (conventional and high-throughput methods). Our results revealed a weak maize growth promoting effect of Ab-V5 inoculation in undiluted NS, contrasting the positive effects of NS dilutions 10-3, 10-6, 10-9 and IS with Ab-V5. Alpha diversity in NS + Ab-V5 soil samples was higher than in all other treatments in a time course of 25 days after sowing (DAS). At 15 DAS, alpha diversity indexes were different between NS and IS, but similar in all NS dilutions in rhizospheric samples. These differences were not persistent at 25 DAS, demonstrating a stabilization process in the rhizobiomes. In NS 10-3 +Ab-V5 and NS 10-6 Ab-V5, Ab-V5 persisted in the maize rhizosphere until 15 DAS in higher abundances compared to NS. In NS + Ab-V5, abundance of six taxa were positively correlated with response to (a)biotic stresses in plant-soil interface. Genes involved in bacterial metabolism of riboses and amino acids, and cresol degradation were abundant on NS 10-3 + Ab-V5, indicating that these pathways can contribute to plant growth promotion and might be a result of Ab-V5 performance as a microbial recruiter of beneficial functions to the plant. Our results demonstrated the effects of holobiont on Ab-V5 performance. The meta-omics integration supported by plant phenomics opens new perspectives to better understanding of inoculants-holobiont interaction and for developing better strategies for optimization in the use of microbial products.

Revealing the high variability on nonconserved core and mobile elements of Austropuccinia psidii and other rust mitochondrial genomes.

Mitochondrial genomes are highly conserved in many fungal groups, and they can help characterize the phylogenetic relationships and evolutionary biology of plant pathogenic fungi. Rust fungi are among the most devastating diseases for economically important crops around the world. Here, we report the complete sequence and annotation of the mitochondrial genome of Austropuccinia psidii (syn. Puccinia psidii), the causal agent of myrtle rust. We performed a phylogenomic analysis including the complete mitochondrial sequences from other rust fungi. The genome composed of 93.299 bp has 73 predicted genes, 33 of which encoded nonconserved proteins (ncORFs), representing almost 45% of all predicted genes. A. psidii mtDNA is one of the largest rust mtDNA sequenced to date, most likely due to the abundance of ncORFs. Among them, 33% were within intronic regions of diverse intron groups. Mobile genetic elements invading intron sequences may have played significant roles in size but not shaping of the rust mitochondrial genome structure. The mtDNAs from rust fungi are highly syntenic. Phylogenetic inferences with 14 concatenated mitochondrial proteins encoded by the core genes placed A. psidii according to phylogenetic analysis based on 18S rDNA. Interestingly, cox1, the gene with the greatest number of introns, provided phylogenies not congruent with the core set. For the first time, we identified the proteins encoded by three A. psidii ncORFs using proteomics analyses. Also, the orf208 encoded a transmembrane protein repressed during in vitro morphogenesis. To the best of our knowledge, we presented the first report of a complete mtDNA sequence of a member of the family Sphaerophragmiacea.

YTOX: a rapid toxicity test based on the dehydrogenase activity of Saccharomyces cerevisiae for detection of contaminants in water samples.

A simple generic toxicity method (test) is proposed using baker's yeast to mediate the reduction of the colourless triphenyltetrazolium chloride (TTC) to red, 1,3,5-triphenyl formazan, which can be extracted by dimethyl sulfoxide (DMSO), enabling the identification of reducible toxic compounds (e.g. cadmium, fipronil) in water for consumption.