Eukaryotic 18S Research Articles

Exploring the Genetic and Morphological Diversity of Argulus Ectoparasite Infecting the Aquaculture and Ornamental Fish in Tripura

Background: The present study screened for isolation of Argulus spp. infecting fishes based on the report of causing substantial economic losses both in aquaculture ponds and ornamental shops across several locations in Tripura, India. Methods: Identification of Argulus spp. was performed based on morphological, and molecular techniques. The molecular identification of Argulus spp. was performed using universal eukaryotic 18S rRNA genetic markers. Prevalence analysis was performed to determine the infestation rate of Argulus spp. in different hosts. Result: Seven A. japonicus, one A. coregoni and one A. foliaceus isolates were identified based on molecular and morphological features. In the present study it was found that A. japonicus infected a wide range of host species, including Goldfish, Milky Koi carp, Mrigal, Rohu, Catla and Subunkin. It was the first instance of A. japonicus infecting milky koi carp both in India and globally. On the other hand, A. foliaceus and A. coregoni infected only tiger oscar and goldfish respectively, indicating a rather narrow range of host diversity.

National-scale distribution of protists associated with sorghum leaves and roots.

Protists, as integral constituents of the plant microbiome, are posited to confer substantial benefits to plant health and performance. Despite their significance, protists have received considerably less attention compared to other constituents of the plant microbiome, such as bacteria and fungi. To investigate the diversity and community structure of protists in sorghum leaves and roots, we employed amplicon sequencing of the eukaryotic 18S rRNA gene in 563 leaf and root samples collected from 57 locations across China. We found significant differences in the diversity and community structure of protists in sorghum leaves and roots. The leaf was taxonomically dominated by Evosea, Cercozoa and Ciliophora, while the root was dominated by Endomyxa, Cercozoa and Oomycota. The functional taxa of protists exhibited notable differences between leaves and roots, with the former being predominantly occupied by consumers and the latter by parasites. The community composition of protists in the leaf was predominantly influenced by mean annual precipitation, whereas soil pH played a more significant role in the root. The present study identified the most abundant and distributed protists in sorghum leaves and roots and elucidated the underlying factors that govern their community structure. The present study offers a novel perspective on the factors that shape plant-associated protist communities and their potential roles in enhancing the functionality of plant ecosystems.

The influence of biotic and abiotic factors on the bacterial microbiome of gentoo penguins (Pygoscelis papua) in their natural environment

The microbiome is a key factor in the health, well-being, and success of vertebrates, contributing to the adaptive capacity of the host. However, the impact of geographic and biotic factors that may affect the microbiome of wild birds in polar environments is not well defined. To address this, we determined the bacterial 16S rRNA gene sequence profiles in faecal samples from pygoscelid penguin populations in the Scotia Arc, focusing on gentoo penguins. This mesopredatory group breeds in defined colonies across a wide geographic range. Since diet could influence microbiome structure, we extracted dietary profiles from a eukaryotic 18S rRNA gene sequence profile. The bacterial microbiome profiles were considered in the context of a diverse set of environmental and ecological measures. Integrating wide geographic sampling with bacterial 16S and eukaryotic 18S rRNA gene sequencing of over 350 faecal samples identified associations between the microbiome profile and a suite of geographic and ecological factors. Microbiome profiles differed according to host species, colony identity, distance between colonies, and diet. Interestingly there was also a relationship between the proportion of host DNA (in relation to total 18S rRNA gene signal) and the microbiome, which may reflect gut passage time. Colony identity provided the strongest association with differences in microbiome profiles indicating that local factors play a key role in the microbiome structure of these polar seabirds. This may reflect the influence of local transfer of microbes either via faecal-oral routes, during chick feeding or other close contact events. Other factors including diet and host species also associate with variation in microbiome profile, and in at least some locations, the microbiome composition varies considerably between individuals. Given the variation in penguin microbiomes associated with diverse factors there is potential for disruption of microbiome associations at a local scale that could influence host health, productivity, and immunological competence. The microbiome represents a sensitive indicator of changing conditions, and the implications of any changes need to be considered in the wider context of environmental change and other stressors.

Distinct changes in tomato-associated multi-kingdom microbiomes during Meloidogyne incognita parasitism

BackgroundThe interplay between root-knot nematode (RKN) parasitism and the complex web of host-associated microbiota has been recognized as pivotal for effective management of the pest. However, studies assessing this relationship have focussed on the bacterial and fungal communities, neglecting the unicellular eukaryotic members. Here, we employed amplicon sequencing analysis of the bacterial 16S rRNA, fungal ITS and eukaryotic 18S rRNA genes, and comprehensively examined how the microbiome composition, diversity and networking developed with time in the rhizospheres and roots of RKN-inoculated and non-inoculated tomato plants.ResultsAs expected, infection with the RKN Meloidogyne incognita decreased plant growth. At individual timepoints, we found distinct bacterial, fungal and eukaryote community structures in the RKN-inoculated and non-inoculated rhizospheres and roots, and RKN inoculation affected several taxa in the root-associated microbiome differentially. Correlation analysis revealed several bacterial and fungal and few protist taxa that correlated negatively or positively with M. incognita. Moreover, network analysis using bacterial, fungal and eukaryotic data revealed more dynamic networks with higher robustness to disturbances in the RKN-inoculated than in the non-inoculated rhizospheres/roots. Hub taxa displayed a noticeable successional pattern that coincided with different phases of M. incognita parasitism. We found that fungal hubs had strong negative correlations with bacteria and eukaryotes, while positive correlations characterized hub members within individual kingdoms.ConclusionOur results reveal dynamic tomato-associated microbiomes that develop along different trajectories in plants suffering M. incognita infestation and non-infested plants. Overall, the results identify stronger associations between RKN and bacterial and fungal taxa than between eukaryotic taxa and RKN, suggesting that fungal and bacterial communities could play a larger role in the regulation of RKN. The study identifies several putative RKN-antagonistic bacterial and fungal taxa and confirms the antagonistic potential previously identified in other taxa.

Environmental DNA reveals fine‐scale spatial and temporal variation of marine mammals and their prey species in a Scottish marine protected area

AbstractMarine mammal foraging grounds are popular focal points for marine protected area (MPA) implementation, despite being temporally dynamic, requiring continuous monitoring to infer prey availability and abundance. Marine mammal distributions are assumed to be driven by their prey in foraging areas, but limited understanding of prey distributions often prevents us from exploring how shifting prey availability impacts both seasonal and long‐term marine mammal distributions. Environmental DNA (eDNA) metabarcoding could enhance the understanding of marine mammal habitat use in relation to their prey through simultaneous monitoring of both. However, eDNA applications focused on marine mammals or predator–prey dynamics have been limited to date. In this study, we assess spatiotemporal changes in the distribution and abundance of cetaceans, minke whales (Balaenoptera acutorostrata), bottlenose dolphins (Tursiops truncatus) and harbor porpoises (Phocoena phocoena) in relation to key prey species in a newly established MPA, employing eDNA metabarcoding. We recovered 105 molecular operational taxonomic units (OTUs) from marine vertebrates using two primer sets targeting 12S and 16S genes, along with 112 OTUs from a broader eukaryotic primer set targeting 18S rRNA. Overall, key forage fish prey species, sandeels and clupeids, were the most abundant teleost fishes detected, although their availability varied temporally and with distance from shore. We also found clear spatial partitioning between coastal bottlenose dolphins and the more pelagic minke whales and harbor porpoises, paralleling availability of their main prey species. Other species of conservation interest were also detected including the critically endangered European eel (Anguilla anguilla), Atlantic bluefin tuna (Thunnus thynnus), and the invasive pink salmon (Oncorhynchus gorbuscha). This study demonstrates the application of eDNA to detect spatiotemporal trends in the occurrence and abundance of cetacean predators and their prey, furthering our understanding of fine‐scale habitat use within MPAs. Future long‐term monitoring of predator–prey dynamics with eDNA could improve our ability to predict climate‐induced shifts in foraging grounds and enhance rapid responses with appropriate management actions.

Prevalence and Preferred Niche of Small Eukaryotes with Mixotrophic Potentials in the Global Ocean.

Unicellular eukaryotes that are capable of phago-mixotrophy in the ocean compete for inorganic nutrients and light with autotrophs, and for bacterial prey with heterotrophs. In this study, we ask what the overall prevalence of eukaryotic mixotrophs in the vast open ocean is, and how the availability of inorganic nutrients, light, and prey affects their relative success. We utilized the Tara Oceans eukaryotic 18S rRNA gene and environmental context variables dataset to conduct a large-scale field analysis. We also performed isolate-based culture experiments to verify growth and nutritional resource relationships for representative mixotrophic taxa. The field analysis suggested that the overall prevalence of mixotrophs were negatively correlated with nutrient concentrations and positively associated with light availability. Concentrations of heterotrophic bacteria as a single variable also presented a positive correlation with mixotrophic prevalence, but to a lesser extent. On the other hand, the culture experiments demonstrated a taxa-specific relationship between mixotrophic growth and nutrition resources, i.e., the growth of one group was significantly dependent on light availability, while the other group was less affected by light when they received sufficient prey. Both groups were capable of growing efficiently with low inorganic nutrients when receiving sufficient prey and light. Therefore, our field analysis and culture experiments both suggest that phago-mixotrophy for ocean eukaryotes is seemingly an efficient strategy to compensate for nutrient deficiency but unnecessary to compensate for light scarcity. This study collectively revealed a close relationship between abiotic and biotic nutritional resources and the prevalence of trophic strategies, shedding light on the importance of light and nutrients for determining the competitive success of mixotrophs versus autotrophic and heterotrophic eukaryotes in the ocean.

Foundation species stabilize an alternative eutrophic state in nutrient-disturbed ponds via selection on microbial community.

Eutrophication due to nutrient addition can result in major alterations in aquatic ecosystem productivity. Foundation species, individually and interactively, whether present as invasive species or as instruments of ecosystem management and restoration, can have unwanted effects like stabilizing turbid eutrophic states. In this study, we used whole-pond experimental manipulations to investigate the impacts of disturbance by nutrient additions in the presence and absence of two foundation species: Dreissena polymorpha (a freshwater mussel) and Myriophyllum spicatum (a macrophyte). We tracked how nutrient additions to ponds changed the prokaryotic and eukaryotic communities, using 16S, 18S, and COI amplicon sequencing. The nutrient disturbance and foundation species imposed strong selection on the prokaryotic communities, but not on the microbial eukaryotic communities. The prokaryotic communities changed increasingly over time as the nutrient disturbance intensified. Post-disturbance, the foundation species stabilized the prokaryotic communities as observed by the reduced rate of change in community composition. Our analysis suggests that prokaryotic community change contributed both directly and indirectly to major changes in ecosystem properties, including pH and dissolved oxygen. Our work shows that nutrient disturbance and foundation species strongly affect the prokaryotic community composition and stability, and that the presence of foundation species can, in some cases, promote the emergence and persistence of a turbid eutrophic ecosystem state.

Exploring the use of metabarcoding to reveal eukaryotic associations with mononchids nematodes.

Nematodes play a vital ecological role in soil and marine ecosystems, but there is limited information about their dietary diversity and feeding habits. Due to methodological challenges, the available information is based on inference rather than confirmed observations. The lack of correct dietary requirements also hampers rearing experiments. To achieve insight into the prey of mononchid nematodes, this study employed high-throughput Illumina paired-end sequencing using universal eukaryotic species 18S primers on 10 pooled mononchid nematode species, namely Mylonchulus brachyuris, M. brevicaudatus, Mylonchulus sp., Clarkus parvus, Prionchulus sp. M. hawaiiensis, M. sigmaturellus, M. vulvapapillatus, Anatonchus sp. and Miconchus sp. The results indicate that mononchids are associated with a remarkable diversity of eukaryotes, including fungi, algae, and protists. While the metabarcoding approach, first introduced here for mononchids, proved to be a simple and rapid method, it has several limitations and crucial methodological challenges that should be addressed in future studies. Ultimately, such methods should be able to evaluate the dietary complexity of nematodes and provide a valuable avenue for unraveling the dietary requirements of previously unculturable nematodes. This can contribute to the methodology of understanding their feeding habits and contributions to ecosystem dynamics.

First report of Paracapillaria philippinensis infection in flowerhorn cichlid in India.

The flowerhorn cichlid is a popular ornamental fish in many Asian countries. The present study reports the occurrence of Paracapillaria philippinensis (Chitwood et al., 1968), a parasitic nematode in flowerhorn cichlid (Cichlasoma sp.) from south India. The infected fish demonstrated clinical symptoms viz. dark coloration, poor appetite, lethargy, erratic swimming, and white stringy faeces. Microscopic observation of the intestinal content and faeces revealed the presence of adult worms, larvae, and unembryonated eggs. PCR amplification of eukaryotic 18S rRNA, P. philippinensis-specific SSU rRNA gene, and the subsequent sequence analysis confirmed the species identity as P. philippinensis. The generated sequences were submitted in the GenBank, NCBI, under the accession numbers, MK895507.1, MK895446.1, MW144993.1, and OR685675.1. This is the first scientific report of P. philippinensis in fish from India, and it confirms that the flowerhorn cichlid can act as a definitive host for P. philippinensis. This report alerts fish handlers and enthusiasts to undertake suitable precautionary measures while handling live flowerhorn cichlids to prevent possible transmission of P. Philippinensis, which has the potential to infect humans causing intestinal capillariasis.

Seasonal dynamics of gut microbiome: A study of multi-kingdom microbiota of earthworm gut in an urban park

Earthworms play an important role in maintaining terrestrial ecosystem function. In urban areas, earthworms mainly inhabit parks where they function through feeding and excretion of soil elements and nutrient turnover. Seasonal diet strongly alters the earthworm gut microbiome and thus affects this ecological process, but the seasonal variation of eukaryotes is still poorly described. In this study, we obtained earthworm samples from three seasons: summer, autumn and winter, assessing the gut microbiome of earthworms using bacterial 16S rRNA and eukaryotic 18S rRNA gene amplicon sequencing techniques. We identified biomarkers that significantly change with seasons and analyzed eukaryotic-bacterial interactions under seasonal variation through inter-kingdom networks. The results of this study showed that season significantly influenced the diversity of earthworm gut bacterial community, while the diversity of eukaryotic community was relatively stable. Inter-kingdom molecular ecological network analysis revealed high degree of positive or negative interactions between fungi and bacteria, protists and bacteria, and fungi and protists as keystone taxonomies divided the network into several modules. In summary, the gut eukaryotic community is more stable than the bacterial community throughout the seasons, and it has shown to maintain season-long homeostasis in the earthworm gut.

Riboproteome remodeling during quiescence exit in Saccharomyces cerevisiae

The quiescent state is the prevalent mode of cellular life in most cells. Saccharomyces cerevisiae is a useful model for studying the molecular basis of the cell cycle, quiescence, and aging. Previous studies indicate that heterogeneous ribosomes show a specialized translation function to adjust the cellular proteome upon a specific stimulus. Using nano LC-MS/MS, we identified 69 of the 79 ribosomal proteins (RPs) that constitute the eukaryotic 80S ribosome during quiescence. Our study shows that the riboproteome is composed of 444 accessory proteins comprising cellular functions such as translation, protein folding, amino acid and glucose metabolism, cellular responses to oxidative stress, and protein degradation. Furthermore, the stoichiometry of both RPs and accessory proteins on ribosome particles is different depending on growth conditions and among monosome and polysome fractions. Deficiency of different RPs resulted in defects of translational capacity, suggesting that ribosome composition can result in changes in translational activity during quiescence.

Effect of dietary phytase on water and fecal prokaryotic and eukaryotic microbiomes in a hybrid tilapia (Oreochromis aureus x O. niloticus) mixotrophic biofloc production system

In this study we evaluated feed and temporal effects on prokaryotic (16S) and eukaryotic (18S) microbiome dynamics in an outdoor mixotrophic biofloc system used to grow hybrid tilapia (Oreochromis aureus x O. niloticus) to market size. The dicalcium phosphate -free practical feed treated with 6-phytase (3000 FTU/kg; P-Free+Phytase) and positive (Control) and negative (P-Free) control practical feeds used were formulated to contain 27.7% digestible protein and 4% lipid. Sex reversed, all-male hybrid tilapia (28.9 g/fish) were stocked (29 fish/m3) in 9 outdoor rectangular tanks (16.6 m3) and grown for 154 days. The Control feed was fed to all fish during the 14-day acclimation period after which each test feed was fed to fish in three tanks/feed. Biofloc water and fecal samples were collected from all tanks at time 0 (T0, study day 15) and T7, T70, and T140; prokaryotic and eukaryotic abundance and diversity were assessed using high-throughput amplicon sequencing of the 16S and 18S rRNA genes, respectively. Fecal 16S microbiome in all treatments was dominated by Fusobacteria (namely Cetobacterium) with minority representation by Proteobacteria, Firmicutes, Bacteroidetes, and Planctomycetes. Opisthokonts (namely Capsaspora) were the dominant eukaryote detected overall in all fecal samples. Fecal 16S and 18S relative abundances varied significantly with time. Relative abundance of fecal 16S differed significantly between Control and P-Free+Phytase feed treatments only at T140 (harvest). Fecal 18S relative abundance differences among feed treatments were detected only at T0 and T7 and only at the supergroup taxonomic level. Prokaryotic communities in biofloc water among feed treatments and across times were highly diverse and dominated by Proteobacteria, Bacteroidetes, Fusobacteria, Chloroflexi, Planctomycetes, and Actinobacteria. High relative abundances of Capsaspora across time characterized eukaryotic communities. Relative abundances of 16S and 18S taxa in water varied significantly over time. Control and P-Free feed 16S relative abundances in water differed significantly at T0 and T7. Time was the factor that explained most of the variation observed for 16S and 18S beta diversity. Overall, results of the present experiment suggest minimal influence of phytase-treated or dicalcium phosphate-free feeds on hybrid tilapia 16S and 18S water and fecal microbiomes throughout the experiment, particularly because the base feed formulation was the same for all treatments. During this experiment and at harvest tilapia in all treatments appeared healthy and robust, and no known pathogenic bacteria or eukaryotes were detected in any sample, suggesting the observed microbiomes may represent system-specific “healthy” core microbiomes.

Filter no more: A modified plankton sampler for rapid in‐water eDNA capture

Abstract The combination of an efficient sampling method and high‐throughput analysis of environmental DNA (eDNA) can be a powerful approach for characterising biodiversity across aquatic ecosystems. Plankton net tows are one of the oldest, simplest, and least expensive methods for seston and eDNA collection, but require laborious filtration steps which often lead to clogging and/or the introduction of contaminants. In this study, we used a cruising speed net (CSN) device enabling the collection of seston‐derived eDNA at 5 knots speed combined with a novel modified cod‐end with 20 μm nylon mesh inserts enabling eDNA capture while towing. We compared the performance of the CSN sampling protocol with the original conventional filtration of water sample versus the modified cod‐end. Samples were collected in parallel horizontal tows along New Zealand's North‐Eastern coastline. Concentrated water was filtered on conventional 5 μm cellulose acetate membranes, while the 20 μm nylon mesh inserts were immediately isolated post‐towing. Metabarcoding of bacterial 16S rRNA, eukaryotic nuclear 18S rRNA and mitochondrial COI genes, revealed no significant difference in alpha diversity between filtration techniques. In terms of community composition, a clear and significant shift could be observed between sampling sites and environments. Significant differences could be detected between filtration methods for 16S and COI markers, likely driven by fine‐scale differences at more turbid sheltered sites. Nonetheless, each technique could detect shifts in communities between sites and environments with similar sensitivity. Our results demonstrate the promising potential of the modified cod‐end to enable practical and cost‐effective isolation of eDNA‐derived biodiversity data from any vessel types (at ≤5 knots) across a large range of aquatic ecosystems and biogeographic scales.

Ataluren improves myelopoiesis and neutrophil chemotaxis by restoring ribosome biogenesis and reducing p53 levels in Shwachman-Diamond syndrome cells.

Shwachman-Diamond syndrome (SDS) is characterized by neutropenia, exocrine pancreatic insufficiency and skeletal abnormalities. SDS bone marrow haematopoietic progenitors show increased apoptosis and impairment in granulocytic differentiation. Loss of Shwachman-Bodian-Diamond syndrome (SBDS) expression results in reduced eukaryotic 80S ribosome maturation. Biallelic mutations in the SBDS gene are found in ~90% of SDS patients, ~55% of whom carry the c.183-184TA>CT nonsense mutation. Several translational readthrough-inducing drugs aimed at suppressing nonsense mutations have been developed. One of these, ataluren, has received approval in Europe for the treatment of Duchenne muscular dystrophy. We previously showed that ataluren can restore full-length SBDS protein synthesis in SDS-derived bone marrow cells. Here, we extend our preclinical study to assess the functional restoration of SBDS capabilities in vitro and ex vivo. Ataluren improved 80S ribosome assembly and total protein synthesis in SDS-derived cells, restored myelopoiesis in myeloid progenitors, improved neutrophil chemotaxis in vitro and reduced neutrophil dysplastic markers ex vivo. Ataluren also restored full-length SBDS synthesis in primary osteoblasts, suggesting that its beneficial role may go beyond the myeloid compartment. Altogether, our results strengthened the rationale for a Phase I/II clinical trial of ataluren in SDS patients who harbour the nonsense mutation.

Precipitation seasonality and soil pH drive the large-scale distribution of soil invertebrate communities in agricultural ecosystems.

Soil invertebrates contribute significantly to vital ecosystem functions such as the breakdown of organic matter and cycling of essential nutrients, but our knowledge of their large-scale distribution in agricultural systems is limited, which hinders our ability to robustly predict how they will respond to future global change scenarios. Here, we employed metabarcoding analysis of eukaryotic 18S rRNA genes to examine the diversity and community composition of invertebrates in 528 sorghum rhizosphere and bulk soils, collected from 53 experimental field sites across China. Our results revealed that Nematoda, Arthropoda, and Annelida were the dominant soil invertebrate groups in agroecosystems. Among all climatic and soil parameters we examined, precipitation seasonality (i.e. the irregular distribution of precipitation during a normal year) had the strongest relationship with the richness of soil invertebrates, with an increase in soil invertebrate richness predicted with increasing precipitation seasonality. Mean annual precipitation and soil pH were the most important predictors of soil invertebrate community structure, with numerous invertebrate phylotypes showing either significantly positive or negative relationships with these two variables. Our findings suggest that shifts in precipitation patterns and soil pH, induced by future climate change and agricultural practices, will have important consequences for the distribution of soil invertebrate communities, with implications for agricultural ecosystem sustainability.

Promise and limitations of 18S genetic screening of extracted fecal DNA from wild capuchins

Genomic screening of fecal DNA provides insight into diet, parasite infection dynamics, and other aspects of the ecology and pathogens of wild populations. Here, we amplify and sequence the V4/V5 regions of the eukaryotic 18S ribosomal RNA gene from fecal DNA of wild capuchin monkeys (Cebus imitator). We collected 94 fecal samples from 26 individuals, each sampled 1-4 times across a 19-month period and examined the eukaryotic diversity in 63 of these samples which had sufficient numbers and quality of reads during downstream analyses. We found a total of 234 distinct amplicon sequence variants (ASVs) classified as Eukaryotes in our samples. Of these, 66 were assigned to the phylum Nematoda. 64 ASVs are from taxa that possibly parasitize monkeys or their food items: 33 were assigned to lungworms (Superfamily Metastrongyloidae; genus Angiostrongylus), two to the genus Strongyloides, and one to the genus Austrostrongylus. The remaining 28 ASVs were assigned to nematodes that likely parasitize plants and/or invertebrates that the monkeys consume. Taken together with past dietary and coprological study of the same primate population, our results suggest that invertebrate consumption and parasitic infection, especially by lungworms, is common and widespread among this population of wild monkeys. We also discuss limitations of our approach, including the amplification of off-target ASVs, and make suggestions for future research. Overall, 18S screening shows promise for identifying various components of the capuchin gastrointestinal eukaryotic ecosystem, including parasitic helminths, and its utility will increase with the improvement of genetic databases.

Structure of the actively translating plant 80S ribosome at 2.2 Å resolution

In plant cells, translation occurs in three compartments: the cytosol, the plastids and the mitochondria. While the structures of the (prokaryotic-type) ribosomes in plastids and mitochondria are well characterized, high-resolution structures of the eukaryotic 80S ribosomes in the cytosol have been lacking. Here the structure of translating tobacco (Nicotiana tabacum) 80S ribosomes was solved by cryo-electron microscopy with a global resolution of 2.2 Å. The ribosome structure includes two tRNAs, decoded mRNA and the nascent peptide chain, thus providing insights into the molecular underpinnings of the cytosolic translation process in plants. The map displays conserved and plant-specific rRNA modifications and the positions of numerous ionic cofactors, and it uncovers the role of monovalent ions in the decoding centre. The model of the plant 80S ribosome enables broad phylogenetic comparisons that reveal commonalities and differences in the ribosomes of plants and those of other eukaryotes, thus putting our knowledge about eukaryotic translation on a firmer footing.

Selecting 16S rRNA Primers for Microbiome Analysis in a Host-Microbe System: The Case of the Jellyfish Rhopilema nomadica.

Amplicon sequencing of the 16S rRNA gene is extensively used to characterize bacterial communities, including those living in association with eukaryotic hosts. Deciding which region of the 16S rRNA gene to analyze and selecting the appropriate PCR primers remains a major decision when initiating any new microbiome study. Based on a detailed literature survey of studies focusing on cnidarian microbiomes, we compared three commonly used primers targeting different hypervariable regions of the 16S rRNA gene, V1V2, V3V4, and V4V5, using the jellyfish Rhopilema nomadica as a model. Although all primers exhibit a similar pattern in bacterial community composition, the performance of the V3V4 primer set was superior to V1V2 and V4V5. The V1V2 primers misclassified bacteria from the Bacilli class and exhibited low classification resolution for Rickettsiales, which represent the second most abundant 16S rRNA gene sequence in all the primers. The V4V5 primer set detected almost the same community composition as the V3V4, but the ability of these primers to also amplify the eukaryotic 18S rRNA gene may hinder bacterial community observations. However, after overcoming the challenges possessed by each one of those primers, we found that all three of them show very similar bacterial community dynamics and compositions. Nevertheless, based on our results, we propose that the V3V4 primer set is potentially the most suitable for studying jellyfish-associated bacterial communities. Our results suggest that, at least for jellyfish samples, it may be feasible to directly compare microbial community estimates from different studies, each using different primers but otherwise similar experimental protocols. More generally, we recommend specifically testing different primers for each new organism or system as a prelude to large-scale 16S rRNA gene amplicon analyses, especially of previously unstudied host-microbe associations.

Assessing the Fate of Superparamagnetic Iron Oxide Nanoparticles Carrying Usnic Acid as Chemical Cargo on the Soil Microbial Community.

In the present study we evaluate the effect of superparamagnetic iron oxide nanoparticles (SPIONs) carrying usnic acid (UA) as chemical cargo on the soil microbial community in a dystrophic red latosol (oxysol). Herein, 500 ppm UA or SPIONs-framework carrying UA were diluted in sterile ultrapure deionized water and applied by hand sprayer on the top of the soil. The experiment was conducted in a growth chamber at 25 °C, with a relative humidity of 80% and a 16 h/8 h light-dark cycle (600 lx light intensity) for 30 days. Sterile ultrapure deionized water was used as the negative control; uncapped and oleic acid (OA) capped SPIONs were also tested to assess their potential effects. Magnetic nanostructures were synthesized by a coprecipitation method and characterized by scanning and transmission electron microscopy (SEM and TEM), X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), zeta potential, hydrodynamic diameter, magnetic measurements, and release kinetics of chemical cargo. Uncapped and OA-capped SPIONs did not significantly affect soil microbial community. Our results showed an impairment in the soil microbial community exposed to free UA, leading to a general decrease in negative effects on soil-based parameters when bioactive was loaded into the nanoscale magnetic carrier. Besides, compared to control, the free UA caused a significant decrease in microbial biomass C (39%), on the activity of acid protease (59%), and acid phosphatase (23%) enzymes, respectively. Free UA also reduced eukaryotic 18S rRNA gene abundance, suggesting a major impact on fungi. Our findings indicate that SPIONs as bioherbicide nanocarriers can reduce the negative impacts on soil. Therefore, nanoenabled biocides may improve agricultural productivity, which is important for food security due to the need of increasing food production.

Primer design for the amplification of the ammonium transporter genes from the uncultured haptophyte algal species symbiotic with the marine nitrogen-fixing cyanobacterium UCYN-A1.

The multiple symbiotic partnerships between closely related species of the haptophyte algae Braarudosphaera bigelowii and the nitrogen-fixing cyanobacteria Candidatus Atelocyanobacterium thalassa (UCYN-A) contribute importantly to the nitrogen and carbon cycles in vast areas of the ocean. The diversity of the eukaryotic 18S rDNA phylogenetic gene marker has helped to identify some of these symbiotic haptophyte species, yet we still lack a genetic marker to assess its diversity at a finer scale. One of such genes is the ammonium transporter (amt) gene, which encodes the protein that might be involved in the uptake of ammonium from UCYN-A in these symbiotic haptophytes. Here, we designed three specific PCR primer sets targeting the amt gene of the haptophyte species (A1-Host) symbiotic with the open ocean UCYN-A1 sublineage, and tested them in samples collected from open ocean and near-shore environments. Regardless of the primer pair used at Station ALOHA, which is where UCYN-A1 is the pre-dominant UCYN-A sublineage, the most abundant amt amplicon sequence variant (ASV) was taxonomically classified as A1-Host. In addition, two out of the three PCR primer sets revealed the existence of closely-related divergent haptophyte amt ASVs (>95% nucleotide identity). These divergent amt ASVs had higher relative abundances than the haptophyte typically associated with UCYN-A1 in the Bering Sea, or co-occurred with the previously identified A1-Host in the Coral Sea, suggesting the presence of new diversity of closely-related A1-Hosts in polar and temperate waters. Therefore, our study reveals an overlooked diversity of haptophytes species with distinct biogeographic distributions partnering with UCYN-A, and provides new primers that will help to gain new knowledge of the UCYN-A/haptophyte symbiosis.