Diverse Eukaryotes Research Articles

Ecological roles of nano-picoplankton in stratified waters of an embayment in the southern Benguela.

Nano-picoplankton are the dominant primary producers during the post-upwelling period in St Helena Bay, South Africa. Their dynamics on short time scales are not well understood and neither are the community composition, structure, and potential functionality of the surrounding microbiome. Samples were collected over five consecutive days in March 2018 from three depths (1 m, 25 m, 50 m) at a single sampling station in St Helena Bay. There was clear depth-differentiation between the surface and depth in both diversity and function throughout the sampling period for the archaea, bacteria and eukaryotes. Daily difference in eukaryote diversity, was more pronounced at 1 m and 25 m with increased abundances of Syndiniales and Bacillariophyta. Surface waters were dominated by photosynthetic and photoheterotrophic microorganisms, while samples at depth were linked to nitrogen cycling processes, with high abundances of nitrifiers and denitrifiers. Strong depth gradients found in the nutrient transporters for ammonia were good indicators of measured uptake rates. This study showed that nano-picoplankton dynamics were driven by light availability, nutrient concentrations, carbon biomass and oxygenation. The nano-picoplankton help sustain ecosystem functioning in St Helena Bay through their ecological roles, which emphasizes the need to monitor this size fraction of the plankton.

Interactions between SDBS and Hydrilla verticillata − epiphytic biofilm in wetland receiving STPs effluents: Nutrients removal and epiphytic microbial assembly

The fate and effects of sodium dodecyl benzene sulfonate (SDBS) in sewage treatment plants (STPs) effluents on nutrients and submerged macrophytes are far from clear in wetlands. This study conducted a 24-day experiment to investigate changes in nutrients and epiphytic biofilm of Hydrilla verticillata in wetlands receiving effluents with 0.5, 2 and 5 mg L−1 SDBS. The decrease of SDBS in overlying water followed pseudo-first-order kinetic equation, with over 80 % of SDBS removal achieved. 2 and 5 mg L−1 SDBS decreased nutrient removal efficiency, induced oxidative stress response and damaged cells of H. verticillata. SDBS altered bacterial and eukaryotic community diversity. 0.5 mg L−1 SDBS can promote carbon fixation and methane oxidation of microorganisms. Network analysis revealed that 0.5 mg L−1 SDBS decreased the stability of epiphytic ecosystems. Mantel tests indicated significant influences of SDBS, temperature, and total nitrogen on epiphytic microbial communities.

LukProt: A Database of Eukaryotic Predicted Proteins Designed for Investigations of Animal Origins.

The origins and early evolution of animals are subjects with many outstanding questions. One problem faced by researchers trying to answer them is the absence of a comprehensive database with sequences from nonbilaterians. Publicly available data are plentiful but scattered and often not associated with proper metadata. A new database presented in this paper, LukProt, is an attempt at solving this issue. The database contains protein sequences obtained mostly from genomic, transcriptomic, and metagenomic studies and is an extension of EukProt (Richter DJ, Berney C, Strassert JFH, Poh Y-P, Herman EK, Muñoz-Gómez SA, Wideman JG, Burki F, de Vargas C. EukProt: a database of genome-scale predicted proteins across the diversity of eukaryotes. Peer Community J. 2022:2:e56. https://doi.org/10.24072/pcjournal.173). LukProt adopts the EukProt naming conventions and includes data from 216 additional animals. The database is associated with a taxonomic grouping (taxogroup) scheme suitable for studying early animal evolution. Minor updates to the database will contain species additions or metadata corrections, whereas major updates will synchronize LukProt to each new version of EukProt, and releases are permanently stored on Zenodo (https://doi.org/10.5281/zenodo.7089120). A BLAST server to search the database is available at: https://lukprot.hirszfeld.pl/. Users are invited to participate in maintaining and correcting LukProt. As it can be searched without downloading locally, the database aims to be a convenient resource not only for evolutionary biologists, but for the broader scientific community as well.

Evolutionary origins of the lysosome-related organelle sorting machinery reveal ancient homology in post-endosome trafficking pathways

The major organelles of the endomembrane system were in place by the time of the last eukaryotic common ancestor (LECA) (~1.5 billion years ago). Their acquisitions were defining milestones during eukaryogenesis. Comparative cell biology and evolutionary analyses show multiple instances of homology in the protein machinery controlling distinct interorganelle trafficking routes. Resolving these homologous relationships allows us to explore processes underlying the emergence of additional, distinct cellular compartments, infer ancestral states predating LECA, and explore the process of eukaryogenesis itself. Here, we undertake a molecular evolutionary analysis (including providing a transcriptome of the jakobid flagellate Reclinomonas americana), exploring the origins of the machinery responsible for the biogenesis of lysosome-related organelles (LROs), the Biogenesis of LRO Complexes (BLOCs 1,2, and 3). This pathway has been studied only in animals and is not considered a feature of the basic eukaryotic cell plan. We show that this machinery is present across the eukaryotic tree of life and was likely in place prior to LECA, making it an underappreciated facet of eukaryotic cellular organisation. Moreover, we resolve multiple points of ancient homology between all three BLOCs and other post-endosomal retrograde trafficking machinery (BORC, CCZ1 and MON1 proteins, and an unexpected relationship with the "homotypic fusion and vacuole protein sorting" (HOPS) and "Class C core vacuole/endosomal tethering" (CORVET) complexes), offering a mechanistic and evolutionary unification of these trafficking pathways. Overall, this study provides a comprehensive account of the rise of the LROs biogenesis machinery from before the LECA to current eukaryotic diversity, integrating it into the larger mechanistic framework describing endomembrane evolution.

DNA-metabarcoding of cyanobacteria and microalgae in chernozem soils of temperate continental climate of the forest-steppe zone of Eurasia under different degrees of agrotechnology intensification.

Chernozem soil is a valuable resource and contains a great diversity of microorganisms that play a global role in the process of soil formation, the species diversity of which has changed over the last five years under the influence of different agrotechnologies. For the first time, under the conditions of the Central Chernozem region, grain and fallow crop rotation, studies using the DNA-metabarcoding method were carried out to study the taxonomic structure of bacteria, fungi, cyanobacteria, and microalgae communities in the arable horizon of typical medium loamy chernozem under winter wheat cultivation. A comparative analysis of the composition of the genotypes showed significant differences in the presented level of mineral nutrition of the soil NPK (60) and NPK (100) compared with the control variant. After processing the 16S and 18S rRNA datasets, a similar trend of decreasing numbers of pro- and eukaryotic species was found from 6296 (control without MF) to 5310 with NPK (60) and to 4643 with NPK (100), respectively. The Chao1 index indicated that the expected diversity within the prokaryotic group was higher in the control without MF at 211, but decreased to 182 and 193 with NPK (60) and NPK (100) fertilizers, respectively. Analysis of the eukaryotic group revealed a 2.6- and 2.9-fold decrease in diversity by class and genus, respectively, depending on the nutritional levels in agrotechnologies, owing to the use of MF. In the prokaryotic community, Alphaproteobacteria microorganisms predominated at an amount of 14.20-14.46%, with Cyanophyceae accounting for 5.2-9.9%. The diversity of eukaryotes was smaller than the number of classes of prokaryotes; the main dominant were Zygnematophyceae 19.5-41%, Chlorophyceae occupied 10.4-15.8%. On the other hand, the doses of fertilizers used contributed to the emergence of dominant species adapted to high doses of mineral nutrients for plants.

Metal tolerance of Río Tinto fungi.

Southwest Spain's Río Tinto is a stressful acidic microbial habitat with a noticeably high concentration of toxic heavy metals. Nevertheless, it has an unexpected degree of eukaryotic diversity in its basin, with a high diversity of fungal saprotrophs. Although some studies on the eukaryotic diversity in Rio Tinto have been published, none of them used molecular methodologies to describe the fungal diversity and taxonomic affiliations that emerge along the river in different seasons. The aim of the present study was to isolate and describe the seasonal diversity of the fungal community in the Río Tinto basin and its correlation with the physicochemical parameters existing along the river's course. The taxonomic affiliation of 359 fungal isolates, based on the complete internal transcribed spacer DNA sequences, revealed a high degree of diversity, identifying species belonging primarily to the phylum Ascomycota, but representatives of the Basidiomycota and Mucoromycota phyla were also present. In total, 40 representative isolates along the river were evaluated for their tolerance to toxic heavy metals. Some of the isolates were able to grow in the presence of 1000 mM of Cu2+, 750 mM of As5+ and Cd2+, and 100 mM of Co2+, Ni2+, and Pb2+.

Rethinking large scale phylogenomics with PhyloToL 6, a flexible toolkit to enable phylogeny-informed data curation and analysis.

The bulk of eukaryotic diversity is microbial, with macroscopic lineages such as plant, animals and fungi nesting among a plethora of diverse lineages that include amoebae, flagellates, ciliates, and many types of algae. Our understanding of the evolutionary relationships and genome properties of microbial eukaryotes is rapidly advancing through analyses of omics (transcriptomic, genomic) data. However, phylogenomic analyses are challenging for microeukaryotes, and particularly uncultivable lineages, as single-cell approaches generate a mixture of sequence data from hosts, associated microbiomes, and contaminants. Current practices include resampling of hand-curated gene sets that can be difficult for other researchers to replicate. To address these challenges, we present PhyloToL version 6.0, a modular, user-friendly pipeline that enables effective data curation that includes phylogeny-informed contamination removal, estimation of homologous gene families, and generation of both multisequence alignments and gene trees. We provide several databases that will be of use for those interested in eukaryotic evolution: a Hook Database of curated reference sequences for 15,000 gene families; a database of transcriptome and genomes from 1,000 taxa with GFs assigned; and a highly-curated set of MSA and gene trees for 500 GFs in these taxa. We also demonstrate a suite of stand-alone utilities that provide basic statistics on sequences, analyze compositional/codon patterns, and enable exploration of trees (e.g. clade-grabbing and efficient tip labeling). We exemplify the power of PhyloToL 6.0 in estimating eukaryotic phylogeny using the 500 conserved GFs, and set standards for curation of omics data for future research in the field.

Putative MutS2 Homologs in Algae: More Goods in Shopping Bag?

MutS2 proteins are presumably involved in either control of recombination or translation quality control in bacteria. MutS2 homologs have been found in plants and some algae; however, their actual diversity in eukaryotes remains unknown. We found putative MutS2 homologs in various species of photosynthetic eukaryotes and performed a detailed analysis of the revealed amino acid sequences. Three groups of homologs were distinguished depending on their domain composition: MutS2 homologs with full set of specific domains, MutS2-like sequences without endonuclease Smr domain, and MutS2-like homologs lacking Smr and clamp in domain IV, the extreme form of which are proteins with only a complete ATPase domain. We clarified the information about amino acid composition and set of specific motifs in the conserved domains in MutS2 and MutS2-like sequences. The models of the predicted tertiary structure were obtained for each group of homologs. The phylogenetic analysis demonstrated that all eukaryotic sequences split into two large groups. The first group included homologs belonging to species of Archaeplastida and a subset of haptophyte homologs, while the second-sequences of organisms from CASH groups (cryptophytes, alveolates, stramenopiles, haptophytes) and chlorarachniophytes. The cyanobacterial MutS2 clustered together with the first group, and proteins belonging to Deltaproteobacteria (orders Myxococcales and Bradymonadales) showed phylogenetic affinity to the CASH-including group with strong support. The observed tree pattern did not support a clear differentiation of eukaryotes into lineages with red and green algae-derived plastids. The results are discussed in the context of current conceptions of serial endosymbioses and genetic mosaicism in algae with complex plastids.

Chlormequat inhibits Vallisneria natans growth and shapes the epiphytic biofilm microbial community.

Submerged macrophytes can overgrow and negatively affect freshwater ecosystems. This study aimed to investigate the use of chlormequat (CQ) to regulate submerged Vallisneria natans growth as well as its impact on the microbial community of epiphytic biofilms. V.natans height under CQ dosages of 20, 100, and 200 mg/L decreased within 21 days by 12.57%, 30.07%, and 44.62%, respectively, while chlorophyll content increased by 1.94%, 20.39%, and 38.83%. At 100 mg/L, CQ reduced the diversity of bacteria in the biofilm attached to V.natans leaves but increased the diversity of the eukaryotic microbial community. CQ strongly inhibited Cyanobacteria; compared with the control group, the treatment group experienced a significant reduction from 36.54% to 2.61%. Treatment significantly inhibited Gastrotricha and Rotifera, two dominant phyla of eukaryotes in the leaf biofilm, reducing their relative abundances by 17.41% and 6.48%, respectively. CQ significantly changed the leaf biofilm microbial community correlation network. The treatment group exhibited lower modularity (2.012) compared with the control group (2.249); however, the central network of the treated group contained a higher number of microbial genera (13) than the control group (4), highlighting the significance of eukaryotic genera in the network. The results obtained from this study provide invaluable scientific context and technical understanding pertinent to the restoration of submerged macrophytes within aquatic ecosystems. PRACTITIONER POINTS: Chlormequat reduced the plant height but increased leaf chlorophyll content. Chlormequat reduced biofilm bacterial diversity but increased eukaryotic diversity. Chlormequat affected the bacterial-fungal association networks in biofilms.

Distinct Communities of Bacteria and Unicellular Eukaryotes in the Different Water Masses of Cretan Passage Water Column (Eastern Mediterranean Sea).

Elucidating marine microbiota diversity and dynamics holds significant importance due to their role in maintaining vital ecosystem functions and services including climate regulation. This work aims to contribute in the understanding of microbial ecology and networking in one of the world's most understudied marine regions, the Eastern Mediterranean Sea. High-throughput 16S and 18S rRNA gene sequencing analysis was applied to study the diversity of bacteria and unicellular eukaryotes in the different water masses of the Cretan Passage during two seasonally-different sampling expeditions. We assumed that microbial associations differ between the surface and deepwater masses and created co-occurrence networks to evaluate this hypothesis. Our results unveiled vertical variations in both bacterial and unicellular eukaryotic diversity with species fluctuations indicative of seasonality being recorded in the surface water mass. Heterotrophic taxa and grazers related to organic matter degradation and nutrient cycling were enriched in the deepest water layers. Moreover, surface waters presented a higher number of microbial associations indicating abundant ecological niches compared to the deepest layer, possibly related to the lack of bottom-up resources in the oligotrophic deep ocean. Overall, our data provide insight in a heavily stressed, yet underexplored, marine area that requires further research to unravel the ecological roles of marine microbes. To our knowledge, this is the first study that combines molecular biology tools to provide data on both planktic prokaryotes and unicellular eukaryotes across the different water masses in this marine region of the Eastern Mediterranean basin.

Revisiting biochemical pathways for lead and cadmium tolerance by domain bacteria, eukarya, and their joint action in bioremediation.

With the advent rise is in urbanization and industrialization, heavy metals (HMs) such as lead (Pb) and cadmium (Cd) contamination have increased considerably. It is among the most recalcitrant pollutants majorly affecting the biotic and abiotic components of the ecosystem like human well-being, animals, soil health, crop productivity, and diversity of prokaryotes (bacteria) and eukaryotes (plants, fungi, and algae). At higher concentrations, these metals are toxic for their growth and pose a significant environmental threat, necessitating innovative and sustainable remediation strategies. Bacteria exhibit diverse mechanisms to cope with HM exposure, including biosorption, chelation, and efflux mechanism, while fungi contribute through mycorrhizal associations and hyphal networks. Algae, especially microalgae, demonstrate effective biosorption and bioaccumulation capacities. Plants, as phytoremediators, hyperaccumulate metals, providing a nature-based approach for soil reclamation. Integration of these biological agents in combination presents opportunities for enhanced remediation efficiency. This comprehensive review aims to provide insights into joint action of prokaryotic and eukaryotic interactions in the management of HM stress in the environment.

Novel findings on the mitochondria in ciliates, with description of mitochondrial genomes of six representatives

AbstractDetermining and comparing mitochondrial genomes (mitogenomes) are essential for assessing the diversity and evolution of mitochondria. Ciliates are ancient and diverse unicellular eukaryotes, and thus are ideal models for elucidating the early evolution of mitochondria. Here, we report on six new mitogenomes of spirotrichs, a dominant ciliate group, and perform comparative analyses on 12 representative species. We show that: (1) the mitogenomes of spirotrichs are linear structures with high A+T contents (61.12–81.16%), bidirectional transcription, and extensive synteny (except for the nad5, ccmf and cob genes in Euplotia); (2) the non-split of NADH dehydrogenase subunit 2 gene (nad2) is a plesiomorphy of ciliates, whereas it has evolved into a split gene in Spirotrichea (apart from Euplotes taxa), Oligohymenophorea, and Armophorea; (3) the number of small subunit ribosomal proteins (rps) encoded in mitogenomes increases in the later branching classes of ciliates, whereas rps8 shows a loss trend during the evolution of Euplotes taxa; (4) the mitogenomes of spirotrichs exhibit A/T codon bias at the third position, and the codon bias is mainly due to DNA mutation in oligotrichs, hypotrichs and Diophrys appendiculata; (5) the phylogenetic position of D. appendiculata is unstable and controversial based on both phylogenetic analyses and mitogenome evidence. In summary, we investigated the mitogenome diversity of spirotrichs and broadened our understanding of the evolution of mitochondria in ciliates.

Tetracycline and quinolone contamination mediate microbial and antibiotic resistant gene composition in epiphytic biofilms of mesocosmic wetlands

The fate and ecological impact of antibiotics on aquatic ecosystems have not been properly elucidated in mesocosm wetlands scale. This study explored how tetracyclines (TCs, including tetracycline TC and oxytetracycline) and fluoroquinolones (QNs, including ciprofloxacin CIP and levofloxacin) affect mesocosm wetlands vegetated by V. spiralis, focusing on their impact on epiphytic biofilm microbial communities and antibiotic resistance genes (ARGs). Results showed that submerged plants absorbed more antibiotics than sediment. Both TCs and QNs disrupted microbial communities in different ways and increased eukaryotic community diversity in a concentration-dependent manner (2-4 mg/L for CIP, 4-8 mg/L for TC). TCs mainly inhibited epiphytic bacteria, while CIP increased bacterial phyla abundance. TC reduced Cyanobacteriota, Acidobacteriota, and Patescibacteria but increased Bacillota, Bacteroidota, and Armatimonadota. In contrast, CIP reduced Bacteroidota, Cyanobacteriota, and Gemmatimonadota but increased Bacillota, Planctomycetota, and Acidobacteriota. Significant differences in ARG profiles were observed between QNs and TCs, with TCs having a more substantial effect on ARGs due to their stronger impact on bacterial communities. Both antibiotics raised ARG levels with higher concentrations, particularly for multidrug resistance, tetracyclines, trimethoprim, sulfonamides, aminoglycosides, and fosfomycin, emphasizing their role in antimicrobial resistance. The study suggests that antibiotics can either stimulate or inhibit ARGs depending on their effects on bacterial communities. This study provides key evidence on the ecological mechanisms underlying the impact of TCs and QNs on epiphytic microbes of mesocosm wetlands.

The gut microbiota-immune-brain axis in a wild vertebrate: dynamic interactions and health impacts.

The gut microbiota-immune-brain axis is a feedback network which influences diverse physiological processes and plays a pivotal role in overall health and wellbeing. Although research in humans and laboratory mice has shed light into the associations and mechanisms governing this communication network, evidence of such interactions in wild, especially in young animals, is lacking. We therefore investigated these interactions during early development in a population of common buzzards (Buteo buteo) and their effects on individual condition. In a longitudinal study, we used a multi-marker approach to establish potential links between the bacterial and eukaryotic gut microbiota, a panel of immune assays and feather corticosterone measurements as a proxy for long-term stress. Using Bayesian structural equation modeling, we found no support for feedback between gut microbial diversity and immune or stress parameters. However, we did find strong relationships in the feedback network. Immunity was negatively correlated with corticosterone levels, and microbial diversity was positively associated with nestling body condition. Furthermore, corticosterone levels and eukaryotic microbiota diversity decreased with age while immune activity increased. The absence of conclusive support for the microbiota-immune-brain axis in common buzzard nestlings, coupled with the evidence for stress mediated immunosuppression, suggests a dominating role of stress-dominated maturation of the immune system during early development. Confounding factors inherent to wild systems and developing animals might override associations known from adult laboratory model subjects. The positive association between microbial diversity and body condition indicates the potential health benefits of possessing a diverse and stable microbiota.

Exploring the impact of urban pollution on ciliate diversity along the Sapucaí River (Minas Gerais, Brazil) via DNA metabarcoding.

Protists are diverse single-celled eukaryotes found in various habitats. They exhibit a wide range of forms and functions, representing a significant portion of the eukaryotic tree of life, which also includes animals, plants, and fungi. Due to their high sensitivity to environmental changes, these organisms are widely used as biological indicators of organic pollution. We investigated the molecular diversity of ciliate protists at seven strategic points along the Sapucaí River (Itajubá, Minas Gerais State, Brazil), to assess the impact of urban pollution on the richness, abundance, and diversity indexes of these communities. For each sampling point, values of physicochemical parameters were also recorded. DNA sequences were obtained by high-throughput sequencing (HTS) and analyzed using the V4 18S-rRNA molecular marker, employing the DNA metabarcoding method. We recorded 125 ciliate taxonomic units (OTUs), with nearly 80% corresponding to the classes Spirotrichea, Oligohymenophorea, and Litostomatea. At the genus level, 54 OTUs (43.2%) were identified, spanning 28 genera. The composition of ciliates varied significantly along the river's course, from upstream to downstream of Itajubá city. Samples collected from the urban area displayed the lowest richness and diversity, corroborating the influence of the pollution gradient on these communities. The physicochemical parameters showed little variation among the samples and were not linked to the observed changes in ciliate communities, revealing that these organisms are strongly affected by environmental changes and respond more sensitively to these disturbances than physicochemical parameters, emphasizing their potential as bioindicators.

A hybrid biosynthetic-catabolic pathway for norspermidine production.

The only known pathway for biosynthesis of the polyamine norspermidine starts from aspartate β-semialdehyde to form the diamine 1,3-diaminopropane, which is then converted to norspermidine via a carboxynorspermidine intermediate. This pathway is found primarily in the Vibrionales order of the γ-Proteobacteria. However, norspermidine is also found in other species of bacteria and archaea, and in diverse single-celled eukaryotes, chlorophyte algae and plants that do not encode the known norspermidine biosynthetic pathway. We reasoned that products of polyamine catabolism could be an alternative route to norspermidine production. 1,3-diaminopropane is formed from terminal catabolism of spermine and spermidine, and norspermidine can be formed from catabolism of thermospermine. We found that the single-celled chlorophyte alga Chlamydomonas reinhardtii thermospermine synthase (CrACL5) did not aminopropylate exogenously-derived 1,3-diaminopropane efficiently when expressed in Escherichia coli. In contrast, it completely converted all E. coli native spermidine to thermospermine. Co-expression in E. coli of the polyamine oxidase 5 from lycophyte plant Selaginella lepidophylla (SelPAO5), together with the CrACL5 thermospermine synthase, converted almost all thermospermine to norspermidine. Although CrACL5 was efficient at aminopropylating norspermidine to form tetraamine norspermine, SelPAO5 oxidizes norspermine back to norspermidine, with the balance of flux being inclined fully to norspermine oxidation. The steady-state polyamine content of E. coli co-expressing thermospermine synthase CrACL5 and polyamine oxidase SelPAO5 was an almost total replacement of spermidine by norspermidine. We have recapitulated a potential hybrid biosynthetic-catabolic pathway for norspermidine production in E. coli, which could explain norspermidine accumulation in species that do not encode the known aspartate β-semialdehyde-dependent pathway.

Addictive manipulation: a perspective on the role of reproductive parasitism in the evolution of bacteria-eukaryote symbioses.

Wolbachia bacteria encompass noteworthy reproductive manipulators of their arthropod hosts. which influence host reproduction to favour their own transmission, also exploiting toxin-antitoxin systems. Recently, multiple other bacterial symbionts of arthropods have been shown to display comparable manipulative capabilities. Here, we wonder whether such phenomena are truly restricted to arthropod hosts. We focused on protists, primary models for evolutionary investigations on eukaryotes due to their diversity and antiquity, but still overall under-investigated. After a thorough re-examination of the literature on bacterial-protist interactions with this question in mind, we conclude that such bacterial 'addictive manipulators' of protists do exist, are probably widespread, and have been overlooked until now as a consequence of the fact that investigations are commonly host-centred, thus ineffective to detect such behaviour. Additionally, we posit that toxin-antitoxin systems are crucial in these phenomena of addictive manipulation of protists, as a result of recurrent evolutionary repurposing. This indicates intriguing functional analogy and molecular homology with plasmid-bacterial interplays. Finally, we remark that multiple addictive manipulators are affiliated with specific bacterial lineages with ancient associations with diverse eukaryotes. This suggests a possible role of addictive manipulation of protists in paving the way to the evolution of bacteria associated with multicellular organisms.

V9 Hypervariable Region Metabarcoding Primers for Euglenozoa and Metamonada

ABSTRACTShort amplicon sequencing is a commonly used method to study the diversity of organisms in various habitats. The hypervariable regions of the small subunit rRNA gene (18S rDNA) are the most general barcodes for eukaryotes, which can provide detailed taxonomic information across a wide range of eukaryotic diversity. However, some organisms are often missed by universal primers, which have difficulty amplifying their barcodes. In this study, specific primers were designed for the amplification of the highly diverse 18S‐V9 region of the Euglenozoa and Metamonada groups. The performance of the newly designed primers—V9Eug and V9Meta—was compared with the universal V9 primer on cultured communities derived from a range of freshwater environments of the Soos Natural Reserve and the Slavkov Forest in the Czech Republic. The V9Eug primer was more specific with Euglenozoa representing 91.8% of reads and 57.0% of OTUs, while the V9Meta primer showed lower specificity with only 48.4% of reads and 19.7% of OTUs assigned to Metamonada. Both the Euglenozoa and Metamonada primer pairs significantly improved recovery of their target groups compared to the universal V9 primer pair, detecting 2.7 and 1.8 times more OTUs, respectively. These results provide a more sensitive protocol for studying the diversity of these eukaryotic taxa.

Molecular diversity of green-colored microbial mats from hot springs of northern Japan.

We acquired and analyzed metagenome and 16S/18S rRNA gene amplicon data of green-colored microbial mats from two hot springs within the Onikobe geothermal region (Miyagi Prefecture, Japan). The two collection sites-Tamago and Warabi-were in proximity and had the same temperature (40°C), but the Tamago site was connected to a nearby stream, whereas the Warabi site was isolated. Both the amplicon and metagenome data suggest the bacterial, especially cyanobacterial, dominance of the mats; other abundant groups include Chloroflexota, Pseudomonadota, Bacteroidota/Chlorobiota, and Deinococcota. At finer resolution, however, the taxonomic composition entirely differed between the mats. A total of 5 and 21 abundant bacterial 16S rRNA gene OTUs were identified for Tamago and Warabi, respectively; of these, 12 are putative chlorophyll- or rhodopsin-based phototrophs. The presence of phylogenetically diverse microbial eukaryotes was noted, with ciliates and amoebozoans being the most abundant eukaryote groups for Tamago and Warabi, respectively. Fifteen metagenome-assembled genomes (MAGs) were obtained, represented by 13 bacteria, one ciliate (mitochondrion), and one giant virus. A total of 15 novel taxa, including a new deeply branching Chlorobiota species, is noted from the amplicon and MAG data, highlighting the importance of environmental sequencing in uncovering hidden microorganisms.

Unveiling the above-ground eukaryotic diversity supported by individual large old trees: the “Life on Trees” integrative protocol

Unveiling the above-ground eukaryotic diversity supported by individual large old trees: the “Life on Trees” integrative protocol