Pseudomonas aeruginosa LasI-dependent plant growth promotion requires the host nitrate transceptor AtNRT1.1/CHL1 and the nitrate reductases NIA1 and NIA2.

MAIN CONCLUSION: In P. aeruginosa, mutation of the gene encoding N-acyl-L-homoserine lactone synthase LasI drives defense and plant growth promotion, and this latter trait requires adequate nitrate nutrition. Cross-kingdom communication with bacteria is crucial for plant growth and productivity. Here, we show a strong induction of genes for nitrate uptake and assimilation in Arabidopsis seedlings co-cultivated with P. aeruginosa WT (PAO1) or ΔlasI mutants defective on the synthesis of the quorum-sensing signaling molecule N-(3-oxododecanoyl)-L-homoserine lactone. Along with differential induction of defense-related genes, the change from plant growth repression to growth promotion upon bacterial QS disruption, correlated with upregulation of the dual-affinity nitrate transceptor CHL1/AtNRT1/NPF6.3 and the nitrate reductases NIA1 and NIA2. CHL1-GUS was induced in Arabidopsis primary root tips after transfer onto P. aeruginosa ΔlasI streaks at low and high N availability, whereas this bacterium required high concentrations of nitrogen to potentiate root and shoot biomass production and to improve root branching. Arabidopsis chl1-5 and chl1-12 mutants and double mutants in NIA1 and NIA2 nitrate reductases showed compromised growth under low nitrogen availability and failed to mount an effective growth promotion and root branching response even at high NH4NO3. WT P. aeruginosa PAO1 and P. aeruginosa ΔlasI mutant promoted the accumulation of nitric oxide (NO) in roots of both the WT and nia1nia2 double mutants, whereas NO donors SNP or SNAP did not improve growth or root branching in nia1nia2 double mutants with or without bacterial cocultivation. Thus, inoculation of Arabidopsis roots with P. aeruginosa drives gene expression for improved nitrogen acquisition and this macronutrient is critical for the plant growth-promoting effects upon disruption of the LasI quorum-sensing system.

Novel Freshwater Ascomycetes from Spain.

Freshwater ascomycetes are a group of fungi of great ecological importance because they are involved in decomposition processes and the recycling of organic matter in aquatic ecosystems. The taxonomy of these fungi is complex, with representatives in several orders of the phylum Ascomycota. In the present study, we collected ninety-two samples of plant debris submerged in freshwater in different locations in Spain. The plant specimens were placed in wet chambers and developed several fungi that were later isolated in pure culture. A main phylogenetic tree using the nucleotide sequences of D1-D2 domains of the 28S nrRNA gene (LSU) was built to show the taxonomic placement of all our fungal strains, and, later, individual phylogenies for the different families were built using single or concatenated nucleotide sequences of the most suitable molecular markers. As a result, we found a new species of Amniculicola that produces a coelomycetous asexual state, a new species of Elongatopedicellata that produces an asexual state, a new species of Neovaginatispora that forms both sexual and asexual states in vitro, and the sexual states of two species of Pyrenochaetopsis, none of which have been reported before for these genera. In addition, we describe a new species of Pilidium characterized by the production of copper-colored globose conidiomata, and of Pseudosigmoidea, which produces well-developed conidiophores.

New Dothideomycetes from Freshwater Habitats in Spain.

The Dothideomycetes are a class of cosmopolitan fungi that are present principally in terrestrial environments, but which have also been found in freshwater and marine habitats. In the present study, more than a hundred samples of plant debris were collected from various freshwater locations in Spain. Its incubation in wet chambers allowed us to detect and to isolate in pure culture numerous fungi producing asexual reproductive fruiting bodies (conidiomata). Thanks to a morphological comparison and to a phylogenetic analysis that combined the internal transcribed spacer (ITS) region of the nrDNA with fragments of the RNA polymerase II subunit 2 (rpb2), beta tubulin (tub2), and the translation elongation factor 1-alpha (tef-1) genes, six of those strains were identified as new species to science. Three belong to the family Didymellaceae: Didymella brevipilosa, Heterophoma polypusiformis and Paraboeremia clausa; and three belong to the family Phaeosphaeriaceae:Paraphoma aquatica, Phaeosphaeria fructigena and Xenophoma microspora. The finding of these new taxa significantly increases the number of the coelomycetous fungi that have been described from freshwater habitats.

New Coelomycetous Fungi from Freshwater in Spain.

Coelomycetous fungi are ubiquitous in soil, sewage, and sea- and freshwater environments. However, freshwater coelomycetous fungi have been very rarely reported in the literature. Knowledge of coelomycetous fungi in freshwater habitats in Spain is poor. The incubation of plant debris, from freshwater in various places in Spain into wet chambers, allowed us to detect and isolate in pure culture several pycnidia-producing fungi. Fungal strains were phenotypically characterized, and a phylogenetic study was carried out based on the analysis of concatenated nucleotide sequences of the D1-D2 domains of the 28S nrRNA gene (LSU), the internal transcribed spacer region (ITS) of the nrDNA, and fragments of the RNA polymerase II subunit 2 (rpb2) and beta tubulin (tub2) genes. As a result of these, we report the finding of two novel species of Neocucurbitaria, three of Neopyrenochaeta, and one of Pyrenochaetopsis. Based on the phylogenetic study, we also transferred Neocucurbitaria prunicola to the genus Allocucurbitaria. This work makes an important contribution to the knowledge of the mycobiota of plant debris in freshwater habitats.

New Taxa of the Family Amniculicolaceae (Pleosporales, Dothideomycetes, Ascomycota) from Freshwater Habitats in Spain.

With the exception of the so-called Ingoldian fungi, the diversity and distribution of the freshwater aero-aquatic or facultative fungi are not well known in Spain. In view of that, we collected and placed into wet chambers 105 samples of submerged and decomposing plant debris from various places in Spain, looking for individuals belonging to these latter two morpho-ecological groups of fungi. As a result, we found and isolated in pure culture several fungi, the morphology of some of them belonging to the family Amniculicolaceae (order Pleosporales, class Dothideomycetes). After a careful phenotypic characterization and a phylogenetic tree reconstruction using a concatenated sequence dataset of D1-D2 domains of the 28S nrRNA gene (LSU), the internal transcribed spacer region (ITS) of the nrDNA, and a fragment of the translation elongation factor 1-alpha (tef1) gene, we report the finding of three new species of the genus Murispora: Murispora navicularispora, which produces cinnamon-colored, broadly fusiform to navicular ascospores; Murispora fissilispora, which has as a remarkable characteristic the production of both sexual and asexual morphs in vitro; and Murispora asexualis, the unique species of the genus that lacks a sexual morph. As a consequence of the phylogenetic study, we introduce the new aero-aquatic genus Fouskomenomyces, with a new combination (Fouskomenomyces cupreorufescens, formerly Spirosphaera cupreorufescens as the type species of the genus) and a new species, Fouskomenomyces mimiticus; we propose the new combinations Murispora bromicola (formerly Pseudomassariosphaeria bromicola) and Murispora triseptata (formerly Pseudomassariosphaeria triseptata); and we resurrect Massariosphaeria grandispora, which is transferred to the family Lopiostomataceae.

Two new species of Gloniopsis (Hysteriales, Ascomycota) from clinical specimens: Morphological and molecular characterisation.

BACKGROUND: The coelomycetes comprise a wide range of fungal species distributed in at least three different classes of the phylum Ascomycota. These are morphologically characterised by producing their conidia inside of fruiting bodies called pycnidia or acervuli, and only a reduced number of species are able to cause human infections. However, their identification in the clinical laboratory is often difficult, due to their few morphological features or because they remain sterile. MATERIALS AND METHODS: In the present study, three isolates of coelomycetes of clinical origin were phenotypically and molecularly studied, by sequencing the D1-D2 fragment of the 28S nuclear ribosomal RNA (nrRNA) (LSU), the internal transcribed spacer region (ITS1-5.8S-ITS2) and a fragment of the translation elongation factor 1-alpha (tef1) genes. RESULTS AND CONCLUSIONS: As result of the molecular analysis, the isolates were identified as belonging to the genus Gloniopsis (order Hysteriales, Dothideomycetes) but without the characteristics of any of the species described so far. Therefore, we propose the new species Gloniopsis percutanea and Gloniopsis pneumoniae. Furthermore, this study revealed that some isolates from clinical specimens identified previously as Rhytidhysteron spp. were misidentified, and considering the few studies in the order Hysteriales and the scarce number of sequences of phylogenetic markers, future revisions of this order should be performed to clarify their taxonomy and obtain a better identification from isolates involved in human mycoses.

Non-ribosomal Peptide Synthases from Pseudomonas aeruginosa Play a Role in Cyclodipeptide Biosynthesis, Quorum-Sensing Regulation, and Root Development in a Plant Host.

Diverse molecules mediate cross-kingdom communication between bacteria and their eukaryotic partners and determine pathogenic or symbiotic relationships. N-acyl-L-homoserine lactone-dependent quorum-sensing signaling represses the biosynthesis of bacterial cyclodipeptides (CDPs) that act as auxin signal mimics in the host plant Arabidopsis thaliana. In this work, we performed bioinformatics, biochemical, and plant growth analyses to identify non-ribosomal peptide synthase (NRPS) proteins of Pseudomonas aeruginosa, which are involved in CDP synthesis. A reverse genetics strategy allowed the identification of the genes encoding putative multi-modular-NRPS (MM-NRPS). Mutations in these genes affected the synthesis of the CDPs cyclo(L-Pro-L-Val), cyclo(L-Pro-L-Leu), and cyclo(L-Pro-L-Tyr), while showing wild-type-like levels of virulence factors, such as violacein, elastase, and pyocyanin. When analyzing the bioactivity of purified, naturally produced CDPs, it was found that cyclo(L-Pro-L-Tyr) and cyclo(L-Pro-L-Val) were capable of antagonizing quorum-sensing-LasR (QS-LasR)-dependent signaling in a contrasting manner in the cell-free supernatants of the selected NRPS mutants, which showed QS induction. Using a bacteria-plant interaction system, we further show that the pvdJ, ambB, and pchE P. aeruginosa mutants failed to repress primary root growth, but improved root branching in A. thaliana seedlings. These results indicated that the CDP production in P. aeruginosa depended on the functional MM-NRPS, which influences quorum-sensing of bacteria and plays a role in root architecture remodeling.