Microbiology and Plant Pathology

Climate change is altering precipitation regimes that control nitrogen (N) cycling in terrestrial ecosystems. In ecosystems exposed to frequent drought, N can accumulate in soils as they dry, stimulating the emission of both nitric oxide (NO; an air pollutant at high concentrations) and nitrous oxide (N2O; a powerful greenhouse gas) when the dry soils wet up. Because changes in both N availability and soil moisture can alter the capacity of nitrifying organisms such as ammonia-oxidizing bacteria (AOB) and archaea (AOA) to process N and emit N gases, predicting whether shifts in precipitation may alter NO and N2O emissions requires understanding how both AOA and AOB may respond. Thus, we ask: How does altering summer and winter precipitation affect nitrifier-derived N trace gas emissions in a dryland ecosystem? To answer this question, we manipulated summer and winter precipitation and measured AOA- and AOB-derived N trace gas emissions, AOA and AOB abundance, and soil N concentrations. We found that excluding summer precipitation increased AOB-derived NO emissions, consistent with the increase in soil N availability, and that increasing summer precipitation amount promoted AOB activity. Excluding precipitation in the winter (the most extreme water limitation we imposed) did not alter nitrifier-derived NO emissions despite N accumulating in soils. Instead, nitrate that accumulated under drought correlated with high N2O emission via denitrification upon wetting dry soils. Increases in the timing and intensity of precipitation that are forecasted under climate change may, therefore, influence the emission of N gases according to the magnitude and season during which the changes occur.

Climate change is altering precipitation regimes that control nitrogen (N) cycling in terrestrial ecosystems. In ecosystems exposed to frequent drought, N can accumulate in soils as they dry, stimulating the emission of both nitric oxide (NO; an air pollutant at high concentrations) and nitrous oxide (N₂O; a powerful greenhouse gas) when the dry soils wet up. Because changes in both N availability and soil moisture can alter the capacity of nitrifying organisms such as ammonia-oxidizing bacteria (AOB) and archaea (AOA) to process N and emit N gases, predicting whether shifts in precipitation may alter NO and N₂O emissions requires understanding how both AOA and AOB may respond. Thus, we ask: How does altering summer and winter precipitation affect nitrifier-derived N trace gas emissions in a dryland ecosystem? To answer this question, we manipulated summer and winter precipitation and measured AOA- and AOB-derived N trace gas emissions, AOA and AOB abundance, and soil N concentrations. We found that excluding summer precipitation increased AOB-derived NO emissions, consistent with the increase in soil N availability, and that increasing summer precipitation amount promoted AOB activity. Excluding precipitation in the winter (the most extreme water limitation we imposed) did not alter nitrifier-derived NO emissions despite N accumulating in soils. Instead, nitrate that accumulated under drought correlated with high N₂O emission via denitrification upon wetting dry soils. Increases in the timing and intensity of precipitation that are forecasted under climate change may, therefore, influence the emission of N gases according to the magnitude and season during which the changes occur.

The exploration of our solar system for microbial extraterrestrial life is the primary goal of several space agencies. Mars has attracted substantial attention owing to its Earth-like geological history and potential niches for microbial life. This study evaluated the suitability of the polyextremophilic fungal strain Rhinocladiella similis LaBioMMi 1217 as a model eukaryote for astrobiology. Comprehensive genomic analysis, including taxonomic and functional characterization, revealed several R. similis genes conferring resistance to Martian-like stressors, such as osmotic pressure and ultraviolet radiation. When cultured in a synthetic Martian regolith (MGS-1), R. similis exhibited altered morphology and produced unique metabolites, including oxylipins, indolic acid derivatives, and siderophores, which might be potential biosignatures. Notably, oxylipins were detected using laser desorption ionization mass spectrometry, a technique slated for its use in the upcoming European Space Agency ExoMars mission. Our findings enhance the understanding of extremophilic fungal metabolism under Martian-like conditions, supporting the potential of black yeasts as viable eukaryotic models in astrobiological studies. Further research is necessary to validate these biosignatures and explore the broader applicability of R. similis in other extraterrestrial environments.

The fungal genus Neonectria contains many phytopathogenic species currently impacting forests and fruit trees worldwide. Despite their importance, a majority of Neonectria spp. lack sufficient genomic resources to resolve suspected cryptic species. Here, we report draft genomes and assemblies for Neonectria magnoliae NRRL 64651 and Neonectria punicea NRRL 64653.

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Abstract: Colletotrichumspp. have a complicated history of association with land plants. Perhaps most well-known as plant pathogens for the devastating effect they can have on agricultural crops, someColletotrichumspp. have been reported as beneficial plant endophytes. However, there have been only a handful of reports ofColletotrichumspp. isolated from aquatic plant hosts and their ecological role in the marine ecosystem is underexplored. To address this, we present the draft genome and annotation ofColletotrichumsp. CLE4, previously isolated from rhizome tissue from the seagrassZostera marina. This genome (48.03 Mbp in length) is highly complete (BUSCO ascomycota: 98.8%) and encodes 12,015 genes, of which 5.7% are carbohydrate-active enzymes (CAZymes) and 12.6% are predicted secreted proteins. Phylogenetic placement putsColletotrichumsp. CLE4 within theC. acutatumcomplex, closely related toC. godetiae. We found a 8.69% smaller genome size, 21.90% smaller gene count, and the absence of 591 conserved gene families inColletotrichumsp. CLE4 relative to other members of theC. acutatumcomplex, suggesting a streamlined genome possibly linked to its specialized ecological niche in the marine ecosystem. Machine learning analyses using CATAStrophy on CAZyme domains predict this isolate to be a hemibiotroph, such that it has a biotrophic phase where the plant is kept alive during optimal environmental conditions followed by a necrotrophic phase where the fungi actively serves a pathogen. While future work is still needed to definitively tease apart the lifestyle strategy ofColletotrichumsp. CLE4, this study provides foundational insight and a high-quality genomic resource for starting to understand the evolutionary trajectory and ecological adaptations of marine-plant associated fungi.

Viroids occur in plants as swarms of sequence variants clustered around a dominant variant, leading to adoption of the term quasispecies to describe the viroid population in an individual host. The composition of the quasispecies can potentially change according to the age of the infection, the position of the leaf or branch in the canopy, and the host species. The primary aim of this study was to investigate the quasispecies concept for citrus viroid VII (CVd-VII), a recently discovered member of the family Pospiviroidae. Three experiments were conducted to determine factors affecting viroid variability (i) within different tissues of a lemon plant, (ii) among different plants of the same species (citron), and (iii) among different species and hybrids of citrus. Using two primer sets to produce amplicons for high-throughput sequencing, viroid population profiles were generated for each sample. The number of variants that were identified with both primer sets ranged from 2 to 13 per sample, and each sample comprised 1 to 4 major (> 10% sample) variants. The composition of variants differed in samples from different plants and among tissue types of a single plant. Single-nucleotide polymorphisms (SNPs), mostly in the form of substitutions, were the primary source of variation; in this study, SNPs were observed in approximately 10% of the viroid genome. The results of the three experiments indicate that CVd-VII follows the quasispecies model as reported for other viroids and that variability occurs in viroid populations in different tissue types and host species.

UNLABELLED: Heterotrimeric G protein signaling pathways control growth and development in eukaryotes. In the multicellular fungus Neurospora crassa, the guanine nucleotide exchange factor RIC8 regulates heterotrimeric Gα subunits. In this study, we used RNAseq and liquid chromatography-mass spectrometry (LC-MS) to profile the transcriptomes and metabolomes of N. crassa wild type, the Gα subunit mutants Δgna-1 and Δgna-3, and Δric8 strains. These strains exhibit defects in growth and asexual development (conidiation), with wild-type and Δgna-1 mutants producing hyphae in submerged cultures, while Δgna-3 and Δric8 mutants develop conidiophores, particularly in the Δric8 mutant. RNAseq analysis showed that the Δgna-1 mutant possesses 159 mis-regulated genes, while Δgna-3 and Δric8 strains have more than 1,000 each. Many of the mis-regulated genes are involved in energy homeostasis, conidiation, or metabolism. LC-MS revealed changes in levels of primary metabolites in the mutants, with several arginine metabolic intermediates impacted in Δric8 strains. The differences in metabolite levels could not be fully explained by the expression or activity of pathway enzymes. However, transcript levels for two predicted vacuolar arginine transporters were affected in Δric8 mutants. Analysis of arginine and ornithine levels in transporter mutants yielded support for altered compartmentation of arginine and ornithine between the cytosol and vacuole in Δric8 strains. Furthermore, we validated previous reports that arginine and ornithine levels are low in wild-type conidia. Our results suggest that RIC8 regulates asexual sporulation in N. crassa at least in part through altered expression of vacuolar transporter genes and the resultant mis-compartmentation of arginine and ornithine. IMPORTANCE: Resistance to inhibitors of cholinesterase-8 (RIC8) is an important regulator of heterotrimeric Gα proteins in eukaryotes. In the filamentous fungus Neurospora crassa, mutants lacking ric8 undergo inappropriate asexual development (macroconidiation) during submerged growth. Our work identifies a role for RIC8 in regulating expression of transporter genes that retain arginine and ornithine in the vacuole (equivalent of the animal lysosome) and relates this function to the developmental defect. Arginine is a critical cellular metabolite, both as an amino acid for protein synthesis and as a precursor for an array of compounds, including proline, ornithine, citrulline, polyamines, creatine phosphate, and nitric oxide. These results have broad relevance to human physiology and disease, as arginine modulates immune, vascular, hormonal, and other functions in humans.

ABSTRACTHistoplasmosis is an endemic mycosis in North America frequently reported along the Ohio and Mississippi River Valleys, although autochthonous cases occur in non-endemic areas. In the United States, the disease is provoked by two genetically distinct clades of Histoplasma capsulatum sensu lato, Histoplasma mississippiense (Nam1) and H. ohiense (Nam2). To bridge the molecular epidemiological gap, we genotyped 93 Histoplasma isolates (62 novel genomes) including clinical, environmental, and veterinarian samples from a broader geographical range by whole-genome sequencing, followed by evolutionary and species niche modelling analyses. We show that histoplasmosis is caused by two major lineages, H. ohiense and H. mississippiense; with sporadic cases caused by H. suramericanum in California and Texas. While H. ohiense is prevalent in eastern states, H. mississipiense was found to be prevalent in the central and western portions of the United States, but also geographically overlapping in some areas suggesting that these species might co-occur. Species Niche Modelling revealed that H. ohiense thrives in places with warmer and drier conditions, while H. mississippiense is endemic to areas with cooler temperatures and more precipitation. In addition, we predicted multiple areas of secondary contact zones where the two species co-occur, potentially facilitating gene exchange and hybridization. This study provides the most comprehensive understanding of the genomic epidemiology of histoplasmosis in the USA and lays a blueprint for the study of invasive fungal diseases.

UNLABELLED: Bacteroides species are successful colonizers of the human colon and can utilize a wide variety of complex polysaccharides and oligosaccharides that are indigestible by the host. To do this, they use enzymes encoded in polysaccharide utilization loci (PULs). While recent work has uncovered the PULs required for the use of some polysaccharides, how Bacteroides utilize smaller oligosaccharides is less well studied. Raffinose family oligosaccharides (RFOs) are abundant in plants, especially legumes, and consist of variable units of galactose linked by α-1,6 bonds to a sucrose (glucose α-1-β-2 fructose) moiety. Previous work showed that an α-galactosidase, BT1871, is required for RFO utilization in Bacteroides thetaiotaomicron. Here, we identify two different types of mutations that increase BT1871 mRNA levels and improve B. thetaiotaomicron growth on RFOs. First, a novel spontaneous duplication of BT1872 and BT1871 places these genes under the control of a ribosomal promoter, driving high BT1871 transcription. Second, nonsense mutations in a gene encoding the PUL24 anti-sigma factor likewise increase BT1871 transcription. We then show that hydrolases from PUL22 work together with BT1871 to break down the sucrose moiety of RFOs and determine that the master regulator of carbohydrate utilization (BT4338) plays a role in RFO utilization in B. thetaiotaomicron. Examining the genomes of other Bacteroides species, we found homologs of BT1871 in a subset and showed that representative strains of species with a BT1871 homolog grew better on melibiose than species that lack a BT1871 homolog. Altogether, our findings shed light on how an important gut commensal utilizes an abundant dietary oligosaccharide. IMPORTANCE: The gut microbiome is important in health and disease. The diverse and densely populated environment of the gut makes competition for resources fierce. Hence, it is important to study the strategies employed by microbes for resource usage. Raffinose family oligosaccharides are abundant in plants and are a major source of nutrition for the microbiota in the colon since they remain undigested by the host. Here, we study how the model commensal organism, Bacteroides thetaiotaomicron utilizes raffinose family oligosaccharides. This work highlights how an important member of the microbiota uses an abundant dietary resource.