Cytochalasin effects in plants and eukaryotic microbial systems.
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Cytochalasin B↗
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Crawling under the radar: Two novel Paulinella species expand knowledge about the ecology and evolution of a primary plastid-containing amoeba lineage.
The genus Paulinella represents a rare, independent case of primary endosymbiosis, providing a unique system to study the early stages of organelle evolution. Here, we expand current understanding of primary plastid endosymbiosis through the discovery and characterization of two novel photosynthetic amoebae, Paulinella marae sp. nov. and Paulinella murrayi sp. nov., isolated from a brackish water habitat in North Carolina, United States. Complete chromatophore genomes and mitochondrial data revealed conserved gene content but notable structural variation, including genome rearrangements and inversion events. Phylogenetic analyses uncovered significant discordance between nuclear and organelle datasets, likely driven by substitution saturation, limited taxon sampling, and differing evolutionary signals across loci. Ecological observations over multiple years indicate that both species are in low abundance but consistently present, and when coupled with hobbyist data, support the hypothesis that photosynthetic Paulinella species are globally distributed yet under-sampled. These results increase known species diversity within the clade from four to six and highlight the importance of integrating field-based observations with genomic approaches. Overall, this work advances Paulinella as a model for studying ongoing primary endosymbiosis, lineage divergence, and the ecological strategies of low-abundance microbial eukaryotes.
Paulinella↗
Yeast chromosomal DNA molecules have strands which are cross-linked at their termini.
The microbial eukaryote Saccharomyces cerevisiae has 18 chromosomes, each consisting of a DNA molecule of 1 x 15 x 10(8) daltons (150 to 2,300 kilobase pairs). Interstand cross-links have now been found in molecules of all sizes by examining the ability of high molecular weight DNA to snap back, i.e., to rapidly renature after denaturation. Experiments in which snap back was assessed for molecules broken by shearing indicate that there are probably two cross-links in each chromosome. Evidence that the cross-links occur at specific sites in the genome was obtained by treating total chromosomal DNA with the endonuclease EcoRI which cleaves the yeast genome into approximately 2,000 discrete fragments. Cross-link containing fragments were separated from fragments without cross-links. This purification resulted in enrichment for about 18 specific fragments. To determine whether the cross-links are terminal or at internal sites in chromosomal DNA, large shear-produced fragments were examined by electron microsopy. With complete denaturation few fragments exhibit the X-shaped single strand configuration expected for internal cross-links. When partially denatured fragments were examined some ends had single strand loops as expected for (AT-rich) cross-linked termini. We propose that a duplex chromosomal DNA molecules have cross-linked termini. We propose that a duplex chromosomal DNA molecule in this eukaryote consists of a continuous, single, self-complementary strand of DNA. This structure has implications for the mechanism of chromosome replication and may be the basis of telomere behavior.
Chromosomes↗
Unisexual reproduction in the global human fungal pathogen Cryptococcus neoformans.
The human fungal pathogen Cryptococcus species complex (encompassing Cryptococcus neoformans, Cryptococcus deneoformans, and the Cryptococcus gattii species complexes) exhibits diversity in sexual reproduction, including α-a mating, pseudosexual reproduction, as well as unisexual reproduction initiated from a single isolate or between isolates of the same mating type. A central conundrum is that while most Cryptococcus natural populations exhibit significant α mating-type bias, genetic and genomic analyses show recombination occurs in nature. The discovery of unisexual reproduction in C. deneoformans provided insight; however, thus far, unisexual reproduction has never been directly observed in the predominant global pathogenic species C. neoformans. Here, we provide evidence that mutating the RIC8 gene, which encodes a conserved guanine nucleotide exchange factor (GEF) involved in both chaperoning and activating Gα proteins, enables unisexual reproduction in C. neoformans. Additionally, we show that genetic variation in the natural population promotes unisexual reproduction, and unisexual reproduction in C. neoformans involves canonical meiotic recombination. Finally, we found that deletion of both GPA2 and GPA3 in the MATα background leads to self-filamentation without sporulation, suggesting that differential modulation of the Gα proteins, likely involving Ric8, could underlie the switch between different modes of sexual reproduction in Cryptococcus. Our study further highlights that the highly conserved Ric8 GEF can act as an important regulator of cellular development in response to environmental stimuli and could modulate sexual reproduction in nature. We hypothesize that unisexual reproduction occurs much more frequently in nature than currently appreciated, and possibly in other fungi and microbial eukaryotes as well.
Cryptococcus neoformans↗
Biocontrol potential and molecular basis of predation in a marine raptorial ciliate.
Predator-prey interactions are widespread across organisms and are key drivers of morphological and behavioral evolution. Despite this, predation remains poorly understood among microbial eukaryotes, mostly due to the absence of a tractable experimental system that allows quantitative, reproducible investigation. This study establishes the marine raptorial ciliate Chaenea vorax as a highly efficient predator, with Rosenzweig-MacArthur model simulations based on predation data showing that only a few dozen individuals can eliminate the vast majority of the facultatively pathogenic ciliate Uronema marinum within 1-2 days, providing a quantitative basis for developing predator-based biocontrol strategies in aquaculture. Genomic analysis shows that C. vorax possesses a highly fragmented macronuclear genome enriched with predation-related pathways, including calcium-mediated contractility, cellular proteolysis, toxin expulsion systems, among others. Transcriptomic profiling during predation events further demonstrates significant upregulation of genes involved in cytoskeletal remodeling, proteolytic activity, and cellular detoxification. Evolutionary analyses suggest that C. vorax has an extremely long evolutionary history, exceptionally high nucleotide diversity even among ciliates, and gene family expansions linked to predatory adaptation. Although the prey possesses certain defensive mechanisms (e.g. trichocysts), these are largely ineffective against short-term predation in closed aquatic environments. These findings provide fundamental insights into the molecular basis of predation in ciliates and suggest the potential utility of C. vorax in biocontrol applications targeting pathogenic ciliates.
Ciliophora↗
Unisexual reproduction in the global human fungal pathogen Cryptococcus neoformans.
The human fungal pathogen Cryptococcus species complex (encompassing C. neoformans, C. deneoformans, and the C. gattii species complexes) exhibit diversity in sexual reproduction, including α-a mating, pseudosexual reproduction, as well as unisexual reproduction initiated from a single isolate or between isolates of the same mating type. A central conundrum is that while most Cryptococcus natural populations exhibit significant α mating-type bias, genetic and genomic analyses show recombination occurs in nature. The discovery of unisexual reproduction in C. deneoformans provided insight; however, thus far unisexual reproduction has never been directly observed in the predominant global pathogenic species C. neoformans. Here, we provide evidence that mutating the RIC8 gene, which encodes a conserved guanine nucleotide exchange factor (GEF) involved in both chaperoning and activating Gα proteins, enables unisexual reproduction in C. neoformans. Additionally, we show that genetic variation in the natural population promotes unisexual reproduction, and unisexual reproduction in C. neoformans involves canonical meiotic recombination. Finally, we found that deletion of both GPA2 and GPA3 in the MATα background leads to self-filamentation without sporulation, suggesting that differential modulation of the Gα proteins, likely involving Ric8, could underlie the switch between different modes of sexual reproduction in Cryptococcus. Our study further highlights that the highly conserved Ric8 GEF can act as an important regulator of cellular development in response to environmental stimuli and could modulate sexual reproduction in nature. We hypothesize that unisexual reproduction occurs much more frequently in nature than currently appreciated, and possibly in other fungi and microbial eukaryotes as well.
G protein↗
Persistent gaps and errors in reference databases impede ecologically meaningful taxonomy assignments in 18S rRNA studies: a case study of terrestrial and marine nematodes.
In metabarcoding studies, Linnaean taxonomy assignments of Operational Taxonomic Units (OTUs) or Amplicon Sequence Variants (ASVs) underpin many downstream bioinformatics analyses and ecological interpretations of environmental DNA (eDNA) datasets. However, public molecular databases (i.e., SILVA, EUKARYOME, BOLD) for most microbial metazoan phyla (nematodes, tardigrades, kinorhynchs, etc.) are sparsely populated, negatively impacting our ability to assign ecologically meaningful taxonomy to these understudied groups. Additionally, the choice of bioinformatics parameters and computational algorithms can further impact the accuracy of eDNA taxonomy assignments. Here, we use two in-silico datasets to show that taxonomy assignments using the 18S rRNA gene can be dramatically improved by curating Linnaean taxonomy strings associated with each reference sequence and closing phylogenetic gaps by improving taxon sampling. Using free-living nematodes as a case study, we applied two commonly used taxonomy assignment algorithms (BLAST+ and the QIIME2 Naïve Bayes classifier) across six iterations of the SILVA 138 reference database to evaluate the precision and accuracy of taxonomy assignments. The BLAST+ top hit with a 90% sequence similarity cutoff often returned the highest percentage of correctly assigned taxonomy at the genus level, and the QIIME2 Naïve Bayes classifier performed similarly well when paired with a reference database containing corrected taxonomy strings. Our results highlight the urgent need for phylogenetically-informed expansions of public reference databases (encompassing both genomes and common gene markers), focused on poorly sampled lineages which are now robustly recovered via eDNA metabarcoding approaches. Additional taxonomy curation efforts should be applied to popular reference databases such as SILVA, and taxon sampling could be rapidly improved by more frequent incorporation of newly published GenBank sequences linked to genus and/or species level identifications.
18S rRNA metabarcoding↗
Biological activity of polyene antibiotics.
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Animals↗
Nonmutagenic action of cannabinoids in vitro.
Under the specific conditions reported for the separate tests delta9-tetrahydrocannabinol (THC) did not elicit a mutagenic response in microbial and eukaryotic in vitro test systems. THC treatment to histidine auxotrophs of Salmonella typhimurium strains TA 98 (susceptible to frame shift mutation) and TA 100 (susceptible to base pair substitution) were investigated. Analysis for possible revertance in the presence and absence of S9 microsomal activation system indicated an absence of induction of gene mutation. Cultured fibroblasts from healthy individuals and DNA repair deficient Xeroderma pigmentosum patients display similar survival activity upon exposure to THC. There was no observable increase in the number of chromosome breaks or chromatid exchanges following exposure to THC or THC plus S9 microsomal fraction. THC, 11-OHdelta9-THC, cannabinol, and cannabidiol did not induce unscheduled DNA repair synthesis in cultured human fibroblasts. Moreover, THC did not suppress UV-induced DNA repair synthesis.
Adult↗
Concurrent stimulation of diflufenican biodegradation and changes in the active microbiome in gravel revealed by Total RNA.
The use of slowly degraded pesticides poses a particular problem when these are applied to urban areas such as gravel paths. The urban gravel provides an environment very different from agricultural soils; i.e., it is both lower in carbon and microbial activity. We, therefore, endeavored to stimulate the degradation of the pesticide diflufenican added to urban gravel microcosms amended with dry alfalfa to increase microbial activity. In the present study, alfalfa addition significantly increased the formation of diflufenican's primary metabolite, 2-[3-(trifluoromethyl)phenoxy]nicotinic acid (AE-B), indicating stimulated biotransformation. The concurrent changes of the active microbial communities within the gravel were explored using shotgun metatranscriptomic sequencing of ribosomal RNA and messenger RNA. Although bacterial taxa remained dominant (87.0%-98.5% relative abundance), the alfalfa treatment led to a 4-5-fold increase in eukaryotic groups, including fungi and microbial grazers. Several microbial taxa potentially involved in the degradation of complex carbon compounds and aromatic pollutants-including Bacteroidetes, Verrucomicrobia, Sordariomycetes, Mortierellales, Tremellales, Sphingopyxis, and Phenylobacterium-increased in relative abundance following alfalfa amendment. Functional gene profiling revealed elevated expression of genes related to microbial activity and biomass production. Genes with potential roles in the breakdown of complex carbon structures (e.g., xylanases/chitin deacetylases) and in the transformation of aromatic compounds (e.g., ring-cleaving dioxygenases) were revealed. We conclude that complex carbon amendments can enhance the microbial activity, promoting the biotransformation of diflufenican in urban gravel environments. These findings provide new insights into the interactions between microbial community dynamics, gene expression profiles, and pesticide biotransformation in non-agricultural matrices.IMPORTANCEPesticides used on urban areas, e.g., gravel paths, are likely to have different effects and fates than when these are used on agricultural soils. Hence, studies into the degradation of pesticides applied to urban matrices are needed. We have previously shown that metabolites of the persistent pesticide diflufenican are even more persistent in urban soils, and it has also previously been shown that these metabolites leach from gravel surfaces. The reasons behind this are that the urban gravel provides an environment very different from agricultural soils; i.e., it is both lower in carbon and microbial activity. In the present study, we, therefore, endeavored to stimulate the degradation of the pesticide diflufenican added to urban gravel microcosms amended with dry alfalfa to increase microbial activity, concurrently studying the changes in the active microbiome by Total RNA-metatranscriptomics.
Biodegradation, Environmental↗
Protein methylation in behavioural control mechanisms and in signal transduction.
In both prokaryotes and eukaryotes methyl groups can be added to and removed from the carboxyl groups of proteins. Recent work has revealed that these reactions have a role in several behavioural phenomena. The nature of this role has been uncovered in one case--that of bacterial chemotaxis.
Adaptation, Physiological↗
Deep subsurface organic-rich shale supports abundant, diverse, and novel fungi.
As Earth's principal reservoir of organic carbon and microbial biomass, the deep subsurface hosts microorganisms capable of mobilizing this once-sequestered carbon. Contrary to standard assumptions of eukaryotic scarcity, this study documents abundant fungal communities, ranging from 4.2 × 103 to 6.8 × 103 fungal cells ml-1, across a methane-producing organic-rich shale 247-556 meters below the surface. Although fungal:bacterial cell ratios ranged from 1:7028 to 1:713, application of biomass conversion factors developed for oceanic systems yielded a median fungal:bacterial biomass ratio of 1:4.7. 16S ribosomal ribonucleic acid (rRNA) gene amplicons revealed bacterial and archaeal communities mirroring those found in well-characterized extremophilic, carbon-degrading environments, while sequencing of 18S rRNA gene and internal transcribed spacer rRNA spacer amplicons collectively identified a eukaryotic hotspot with 689 fungal operational taxonomic units across six phyla. The dominant fungal classes, Agaricomycetes and Dothideomycetes, are well-established degraders of recalcitrant carbon compounds at the surface, suggesting they may similarly contribute to organic matter degradation and ecosystem maintenance in the subsurface. Cultivation and isolation efforts yielded 205 fungal strains, including 13 candidate novel taxa, underscoring the deep subsurface as an underexplored eukaryotic habitat. Stable carbon isotopes indicate methane is predominantly generated via microbial conversion of the fossil carbon, while water isotopes suggest in situ geochemical conditions have been relatively stable since the Late Pleistocene, with subglacial recharge as a plausible mechanism for microbial introduction. Collectively, these findings suggest that fungi are underrecognized contributors to organic matter transformation and functional diversity in the deep biosphere, revealing a critical gap in our understanding of deep subsurface ecosystem processes.
Fungi↗
Large enrichments in fatty acid 2H/1H ratios distinguish respiration from aerobic fermentation in yeast Saccharomyces cerevisiae.
Shifts in the hydrogen stable isotopic composition (2H/1H ratio) of lipids relative to water (lipid/water 2H-fractionation) at natural abundances reflect different sources of the central cellular reductant, NADPH, in bacteria. Here, we demonstrate that lipid/water 2H-fractionation (2εfattyacid/water) can also constrain the relative importance of key NADPH pathways in eukaryotes. We used the metabolically flexible yeast Saccharomyces cerevisiae, a microbial model for respiratory and fermentative metabolism in industry and medicine, to investigate 2εfattyacid/water. In chemostats, fatty acids from glycerol-respiring cells were >550‰ 2H-enriched compared to those from cells aerobically fermenting sugars via overflow metabolism, a hallmark feature in cancer. Faster growth decreased 2H/1H ratios, particularly in glycerol-respiring cells by 200‰. Variations in the activities and kinetic isotope effects among NADP+-reducing enzymes indicate cytosolic NADPH supply as the primary control on 2εfattyacid/water. Contributions of cytosolic isocitrate dehydrogenase (cIDH) to NAPDH production drive large 2H-enrichments with substrate metabolism (cIDH is absent during fermentation but contributes up to 20 percent NAPDH during respiration) and slower growth on glycerol (11 percent more NADPH from cIDH). Shifts in NADPH demand associated with cellular lipid abundance explain smaller 2εfattyacid/water variations (<30‰) with growth rate during fermentation. Consistent with these results, tests of murine liver cells had 2H-enriched lipids from slower-growing, healthy respiring cells relative to fast-growing, fermenting hepatocellular carcinoma. Our findings point to the broad potential of lipid 2H/1H ratios as a passive natural tracker of eukaryotic metabolism with applications to distinguish health and disease, complementing studies that rely on complex isotope-tracer addition methods.
Saccharomyces cerevisiae↗
Oral and gut microbiota profiles in patients with locally advanced rectal cancer with varying responses to neoadjuvant chemoradiotherapy.
Recent research has focused on gut bacteria in colorectal cancer, but the influence of other microbiota, including oral and nonbacterial gut microbiota, on treatment efficacy remains insufficiently explored. This study aimed to investigate their relationship with the efficacy of neoadjuvant chemoradiotherapy (nCRT) in locally advanced rectal cancer (LARC). Saliva and fecal samples were collected from patients with LARC before treatment. Shotgun metagenomic sequencing was used to profile bacterial, archaeal, eukaryotic, and viral taxonomic groups and to examine oral and gut microbial functions. An artificial intelligence-based prediction model was developed by integrating oral and gut microbiome data with clinical information. Statistical analyses compared diversity and response-associated microbial features between responders and non-responders to nCRT. Response-associated differences were observed in bacterial and nonbacterial taxonomic profiles and in oral and gut microbial functional profiles. In the internal test subset, the integrated analysis yielded an observed AUC of 0.917. Given the small cohort and the exploratory comparison of candidate classifiers, this estimate requires confirmation in larger, independent cohorts. Baseline oral and gut microbiome profiles were associated with response to nCRT. Integrating microbiome and clinical features showed potential for response prediction, but the model remains exploratory and requires validation in larger, independent cohorts before clinical application. Retrospectively registered on 01/08/2026, NCT07346729.
Aged↗
Cell division from a genetic perspective.
A novel view of the eukaryotic cell cycle is taking form as genetic strategies borrowed from investigations of microbial gene regulation and bacteriophage morphogenesis are being applied to the process of cell division. It is a genetic construct in which mutational lesions identify the primary events, thermolabile gene products reveal temporal order, mutant phenotypes yield pathways of causality, and regulatory events are localized within sequences of gene controlled steps.
Animals↗
Ecological and methodological insights from genetic and coprological profiling of gastrointestinal communities in wild howler monkeys.
The gastrointestinal tract hosts a complex community of microorganisms and helminth parasites that collectively contribute to host health and fitness. Analysis of these communities provides insight into diverse aspects of host dietary ecology, immunity, nutrition, and host-parasite interactions. However, research methodologies, such as sample preservation and sequencing approach, can influence how we understand and characterize these features. Here, we profiled the gastrointestinal microbial and helminth communities in different groups of wild Costa Rican mantled howler monkeys (Alouatta palliata palliata). We compared samples stored in ethanol versus directly flash frozen, and contrasted conclusions drawn from 16S versus shotgun sequencing approaches. Bacterial, archaeal, and eukaryotic taxa associated with the digestion of plant material dominated the GI communities. Storage and sequencing methods influenced microbial profiles: ethanol-stored samples exhibited higher diversity than frozen samples, and 16S sequencing detected lower diversity than shotgun. Helminths were detected via coprological microscopy in 71% of individuals, whereas metagenomic detection was inconsistent. This study provides new data on the microorganisms and their putative digestive functions in the gut of a folivorous primate, and highlights the pros and cons of different methodological choices when profiling host-microbiome and host-parasite interactions.
Animals↗
Purification and properties of a soluble nicotinamide adenine dinucleotide-linked isocitrate dehydrogenase from Crithidia fasciculata.
A soluble NAD+-linked isocitrate dehydrogenase has been isolated from Crithidia fasciculata. The enzyme was purified 128-fold, almost to homogeneity, and was highly specific for NAD+ as the coenzyme. There is also a cytoplasmic NADP+-linked and a mitochondrial isocitrate dehydrogenase in the organism. Studies of the physical and kinetic properties of the soluble NAD+-isocitrate dehydrogenase from this organism showed that it resembled microbial NADP+-isocitrate dehydrogenases in general, all of which are cytoplasmic enzymes. The enzyme appeared not to be related to other NAD+-isocitrate dehydrogenases, which are found in the mitochondria of eukaryotic cells. The molecular weight of the soluble NAD+-isocitrate dehydrogenase was 105,000 which is within the range of the values for microbial NADP+-isocitrate dehydrogenases. Similar to the NADP+-isocitrate dehydrogenase in this organism, the enzyme was inhibited in a concerted manner by glyoxalate plus oxalacetate. Kinetic analysis revealed that Mn2+ was involved in the binding of isocitrate to the enzyme. Inhibition of the NAD+-linked isocitrate dehydrogenase by p-chloromercuribenzoate could be prevented by prior incubation of the enzyme with both Mn2+ and isocitrate; however, neither ion alone conferred protection. Free isocitrate, free Mn2+, and the Mn2+-isocitrate complex could all bind to the enzyme. Four different mechanisms with respect to the binding of isocitrate to the enzyme were tested. Of these, the formation of the active enzyme-Mn2+-isocitrate complex from (a) the random binding of Mn2+, isocitrate, and the Mn2+-isocitrate complex, or (b) the binding of Mn2+-isocitrate with free Mn2+ and isocitrate acting as dead-end competitors were both in agreement with these data.
Animals↗
Unveiling microbial communities and biogeochemical cycles in Antarctic colored snow.
Snow cover, the extensive terrestrial habitat in Antarctica, sometimes exhibits vivid coloration, yet the structure and function of its microbial communities remain poorly characterized. Using metagenomic sequencing of red snow (RS) and green snow (GS) from the Fildes Peninsula, we found that bacterial, eukaryotic, and archaeal relative abundances were 85.82%, 13.52% and 0.16%, respectively. β-Diversity differed significantly between RS and GS across these three domains (P < 0.05). Dominant bacterial phyla included Bacteroidota (RS: 62.61%; GS: 38.72%) and Pseudomonadota (RS: 32.80%; GS: 54.10%). Among eukaryotes, Chlorophyta (RS: 58.10%; GS: 52.98%) and Basidiomycota (RS: 14.80%; GS: 8.08%) were prevalent. Nanobdellota dominated archaea, with lower abundance in RS than GS. In the algal community, Sanguina, Gonium and Chloromonas were significantly enriched in red snow, while Chlorella and Micractinium were enriched in green snow (P < 0.05). Marker genes associated with carbon (C), nitrogen (N), phosphorus (P) and sulfur (S) cycles were identified in green and red snow. Aerobic respiration and phosphate regulation were significantly enriched in red snow, while CO oxidation, fermentation, and denitrification were significantly enriched in green snow. Key microbial genera associated with these functional pathways also varied. In the denitrification of red snow, Stutzerimonas was the most abundant genus, while Janthinobacterium was abundant in green snow. Nitrification-related genes were detected only in red snow based on the present metagenomic data. The network of the red snow microbial community was potentially more complex and resistant based on topology, which not only benefited its own long-term survival but might also have potentially influenced the positive feedback effect of snowmelt by maintaining a low-albedo snow surface. This provided an ecological implication under climate warming: the expansion of red snow patches showed the potential to the increase nitrate runoff export, which would affect nitrogen nutrient levels in coastal Antarctic waters. Overall, this study used metagenomics to compare the multidomain (bacteria, archaea and eukaryotes) composition and diversity between red snow and green snow, and directly linked key microbial taxa with functional genes of biogeochemical cycles. This study provided new insights into the biological characteristics and functional potential of Antarctic colored snow.
Snow↗