We analyzed the large subunit ribosomal RNA (rRNA) gene [LSU ribosomal DNA (rDNA)] as a phylogenetic marker for arbuscular mycorrhizal (AM) fungal taxonomy. Partial LSU rDNA sequences were obtained from ten AM fungal isolates, comprising seven species, with two new primers designed for Glomeromycota LSU rDNA. The sequences, together with 58 sequences available from the databases, represented 31 AM fungal species. Neighbor joining and parsimony analyses were performed with the aim of evaluating the potential of the LSU rDNA for phylogenetic resolution. The resulting trees indicated that Archaeosporaceae are a basal group in Glomeromycota, Acaulosporaceae and Gigasporaceae belong to the same clade, while Glomeraceae are polyphyletic. The results support data obtained with the small subunit (SSU) rRNA gene, demonstrating that the LSU rRNA gene is a useful molecular marker for clarifying taxonomic and phylogenetic relationships in Glomeromycota.

The identification of filamentous fungi through culture characterization may be hampered by phenotypic variability. Information obtained from the identification of microorganisms are important for investigation of sources of contamination of a product or process. The aim of this study was to identify filamentous fungal strains (n = 50) isolated from a pharmaceutical facility by using Matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS), as well as D2 domain of the large-subunit (LSU) ribosomal RNA gene and internal transcribed spacer regions (ITS) sequencing. MALDI-TOF MS system only identified five strains at the species level, while 45 were not identified. The analysis through GenBank allowed the identification of up to 19 strains at the species level, while MycoBank allowed the identification of up to nine strains at the species level. The databases identified up to 11 genera: Penicillium, Aspergillus, Cladosporium, Chaetomium, Coniochaeta, Curvularia, Diaporthe, Fusarium, Trichoderma, Rhizopus and Microdochium. MALDI-TOF MS showed an insufficient database to identify the species of fungi. DNA sequencing was the best methodology to identify to the genus level but was unable to differentiate between closely related species. Therefore further methods for the identification of filamentous fungi from pharmaceutical areas at species level need to be developed. (Copyright © 2023 Elsevier B.V. All rights reserved.)

Members of the plant-specific LSU (RESPONSE TO LOW SULFUR) family are strongly induced during sulfur starvation. The molecular functions of these proteins are unknown; however, they were identified as important stress-related hubs in several studies. In Arabidopsis thaliana, there are four members of the LSU family (LSU1–4). These proteins are small (approximately 100 amino acids), with coiled-coil structures. In this work, we investigated interactions between different monomers of LSU1–4. Differences in homo- and heterodimer formation were observed. Our structural models of LSU1–4 homo- and heterodimers were in agreement with our experimental observations and may help understand their binding properties. LSU proteins are involved in multiple protein–protein interactions, with the literature suggesting they can integrate abiotic and biotic stress responses. Previously, LSU partners were identified using the yeast two hybrid approach, therefore we sought to determine proteins co-purifying with LSU family members using protein extracts isolated from plants ectopically expressing TAP-tagged LSU1–4 constructs. These experiments revealed 46 new candidates for LSU partners. We tested four of them (and two other proteins, CAT2 and NBR1) for interaction with LSU1–4 by other methods. Binding of all six proteins with LSU1–4 was confirmed by Bimolecular Fluorescence Complementation, while only three of them were interacting with LSUs in yeast-two-hybrid. Additionally, we conducted network analysis of LSU interactome and revealed novel clues for the possible cellular function of these proteins.

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