Reproductive toxicity of hexabromocyclododecane in rotifer Brachionus plicatilis: Involvement of reactive oxygen species and calcium signaling pathways.

To assess the toxicity of Hexabromocyclododecane (HBCD), the population, individual, and cellular biochemical parameters of the rotifer Brachionus plicatilis exposed to different concentrations of HBCD were investigated. The results showed that the population growth rate, reproductive period, and offspring number in B. plicatilis significantly decreased under 324 µg/L and 648 µg/L HBCD. Antioxidant enzyme activity and mRNA expression of CAT and Mn-SOD were promoted at low concentrations (32 µg/L and 64 µg/L) and inhibited at high concentrations (324 µg/L and 648 µg/L), while MDA content accumulated continuously with increasing HBCD concentrations, indicating that HBCD induced oxidation imbalance in rotifers. Further evidence was provided by the correlation between DNA fragmentation and physiological changes. The increased intercellular concentration of Ca2+ and the expression of CaM mRNA suggested that HBCD activated pathways related to calcium signaling. In summary, the excessive production of ROS induced by HBCD was considered to be the main cause of reproductive toxicity.

Bdelloid rotifers deploy horizontally acquired biosynthetic genes against a fungal pathogen.

Coevolutionary antagonism generates relentless selection that can favour genetic exchange, including transfer of antibiotic synthesis and resistance genes among bacteria, and sexual recombination of disease resistance alleles in eukaryotes. We report an unusual link between biological conflict and DNA transfer in bdelloid rotifers, microscopic animals whose genomes show elevated levels of horizontal gene transfer from non-metazoan taxa. When rotifers were challenged with a fungal pathogen, horizontally acquired genes were over twice as likely to be upregulated as other genes - a stronger enrichment than observed for abiotic stressors. Among hundreds of upregulated genes, the most markedly overrepresented were clusters resembling bacterial polyketide and nonribosomal peptide synthetases that produce antibiotics. Upregulation of these clusters in a pathogen-resistant rotifer species was nearly ten times stronger than in a susceptible species. By acquiring, domesticating, and expressing non-metazoan biosynthetic pathways, bdelloids may have evolved to resist natural enemies using antimicrobial mechanisms absent from other animals.

Ice-inhabiting species of Bdelloidea Rotifera reveal a pre-Quaternary ancestry in the Arctic cryosphere.

Historical climate data indicate that the Earth has passed through multiple geological periods with much warmer-than-present climates, including epochs of the Miocene (23-5.3 mya BP) with temperatures 3-4°C above present, and more recent interglacial stages of the Quaternary, for example, Marine Isotope Stage 11c (approx. 425-395 ka BP) and Middle Holocene thermal maximum (7.5-4.2 ka BP), during which continental glaciers may have melted entirely. Such warm periods would have severe consequences for ice-obligate fauna in terms of their distribution, biodiversity and population structure. To determine the impacts of these climatic events in the Nordic cryosphere, we surveyed ice habitats throughout mainland Norway and Svalbard ranging from maritime glaciers to continental ice patches (i.e. non-flowing, inland ice subjected to deep freezing overwinter), finding particularly widespread populations of ice-inhabiting bdelloid rotifers. Combined mitochondrial and nuclear DNA sequencing identified approx. 16 undescribed, species-level rotifer lineages that revealed an ancestry predating the Quaternary (> 2.58 mya). These rotifers also displayed robust freeze/thaw tolerance in laboratory experiments. Collectively, these data suggest that extensive ice refugia, comparable with stable ice patches across the contemporary Norwegian landscape, persisted in the cryosphere over geological time, and may have facilitated the long-term survival of ice-obligate Metazoa before and throughout the Quaternary.

Range of trait variation in prey determines evolutionary contributions to predator growth rates.

Evolutionary and ecological dynamics can occur on similar timescales and thus influence each other. While it has been shown that the relative contribution of ecological and evolutionary change to population dynamics can vary, it still remains unknown what influences these differences. Here, we test whether prey populations with increased variation in their defence and competitiveness traits will have a stronger impact on evolution for predator growth rates. We controlled trait variation by pairing distinct clonal lineages of the green alga Chlamydomonas reinhardtii with known traits as prey with the rotifer Brachionus calyciforus as predator and compared those results with a mechanistic model matching the empirical system. We measured the impact of evolution (shift in prey clonal frequency) and ecology (shift in prey population density) for predator growth rate and its dependency on trait variation using an approach based on a 2-way ANOVA. Our experimental results indicated that higher trait variation, i.e., a greater distance in trait space, increased the relative contribution of prey evolution to predator growth rate over 3-4 predator generations, which was also observed in model simulations spanning longer time periods. In our model, we also observed clone-specific results, where a more competitive undefended prey resulted in a higher evolutionary contribution, independent of the trait distance. Our results suggest that trait combinations and total prey trait variation combine to influence the contribution of evolution to predator population dynamics, and that trait variation can be used to identify and better predict the role of eco-evolutionary dynamics in predator-prey systems.

Changes in life history parameters and expression of key genes of Brachionus plicatilis exposed to a combination of organic and inorganic ultraviolet filters.

The increasing use of ultraviolet filters has become an emerging contaminant on the coast, posing potential ecological risks. Rotifers are essential components of marine ecosystems, serving as an association between primary producers and higher-level consumers. These organisms frequently encounter ultraviolet filters in coastal waters. This study aimed to assess the comprehensive effects of organic ultraviolet filters, specifically 2-ethylhexyl-4-methoxycinnamate (EHMC), and inorganic ultraviolet filters, namely, titanium dioxide nanoparticles (TiO2 NPs), on the rotifer Brachionus plicatilis. We exposed B. plicatilis to multiple combinations of different concentrations of EHMC and TiO2 NPs to observe changes in life history parameters and the expression of genes related to reproduction and antioxidant responses. Our findings indicated that increased EHMC concentrations significantly delayed the age at first reproduction, reduced the total offspring, and led to considerable alterations in the expression of genes associated with reproduction and stress. Exposure to TiO2 NPs resulted in earlier reproduction and decreased total offspring, although these changes were not synchronised in gene expression. The two ultraviolet filters had a significant interaction on the age at first reproduction and the total offspring of rotifer, with these interactions extending to the first generation. This research offers new insights into the comprehensive effects of different types of ultraviolet filters on rotifers by examining life history parameters and gene expression related to reproduction and stress, highlighting the importance of understanding the impacts of sunscreen products on zooplankton health.

Is there potential for estradiol receptor signaling in lophotrochozoans?

Estrogen receptors (ERs) are thought to be the ancestor of all steroid receptors and are present in most lophotrochozoans studied to date, including molluscs, annelids, and rotifers. A number of studies have investigated the functional role of estrogen receptors in invertebrate species, although most are in molluscs, where the receptor is constitutively active. In vitro experiments provided evidence for ligand-activated estrogen receptors in annelids, raising important questions about the role of estrogen signalling in lophotrochozoan lineages. Here, we review the concordant and discordant evidence of estradiol receptor signalling in lophotrochozoans, with a focus on annelids and rotifers. We explore the de novo synthesis of estrogens, the evolution and expression of estrogen receptors, and physiological responses to activation of estrogen receptors in the lophotrochozoan phyla Annelida and Rotifera. Key data are missing to determine if de novo biosynthesis of estradiol in non-molluscan lophotrochozoans is likely. For example, an ortholog for the CYP11 gene is present, but confirmation of substrate conversion and measured tissue products is lacking. Orthologs CYP17 and CYP19 are lacking, yet intermediates or products (e.g. estradiol) in tissues have been measured. Estrogen receptors are present in multiple species, and for a limited number, in vitro data show agonist binding of estradiol and/or transcriptional activation. The expression patterns of the lophotrochozoan ERs suggest developmental, reproductive, and digestive roles but are highly species dependent. E2 exposures suggest that lophotrochozoan ERs may play a role in reproduction, but no strong dose-response relationship has been established. Therefore, we expect most lophotrochozoan species, outside of perhaps platyhelminths, to have an ER but their physiological role remains elusive. Mining genomes for orthologs gene families responsible for steroidogenesis, coupled with in vitro and in vivo studies of the steroid pathway are needed to better assess whether lophotrochozoans are capable of estradiol biosynthesis. One major challenge is that much of the data are divided across a diversity of species. We propose that the polychaetes Capitella teleta or Platyneris dumerilii, and rotifer Brachionus manjavacas may be strong species choices for studies of estrogen receptor signalling, because of available genomic data, established laboratory culture techniques, and gene knockout potential.

Recombination in bdelloid rotifer genomes: asexuality, transfer and stress.

Bdelloid rotifers constitute a class of microscopic animals living in freshwater habitats worldwide. Several strange features of bdelloids have drawn attention: their ability to tolerate desiccation and other stresses, a lack of reported males across the clade despite centuries of study, and unusually high numbers of horizontally acquired, non-metazoan genes. Genome sequencing is transforming our understanding of their lifestyle and its consequences, while in turn providing wider insights about recombination and genome organisation in animals. Many questions remain, not least how to reconcile apparent genomic signatures of sex with the continued absence of reported males, why bdelloids have so many horizontally acquired genes, and how their remarkable ability to survive stress interacts with recombination and other genomic processes.

Ionizing radiation responses appear incidental to desiccation responses in the bdelloid rotifer Adineta vaga.

BACKGROUND: The remarkable resistance to ionizing radiation found in anhydrobiotic organisms, such as some bacteria, tardigrades, and bdelloid rotifers has been hypothesized to be incidental to their desiccation resistance. Both stresses produce reactive oxygen species and cause damage to DNA and other macromolecules. However, this hypothesis has only been investigated in a few species. RESULTS: In this study, we analyzed the transcriptomic response of the bdelloid rotifer Adineta vaga to desiccation and to low- (X-rays) and high- (Fe) LET radiation to highlight the molecular and genetic mechanisms triggered by both stresses. We identified numerous genes encoding antioxidants, but also chaperones, that are constitutively highly expressed, which may contribute to the protection of proteins against oxidative stress during desiccation and ionizing radiation. We also detected a transcriptomic response common to desiccation and ionizing radiation with the over-expression of genes mainly involved in DNA repair and protein modifications but also genes with unknown functions that were bdelloid-specific. A distinct transcriptomic response specific to rehydration was also found, with the over-expression of genes mainly encoding Late Embryogenesis Abundant proteins, specific heat shock proteins, and glucose repressive proteins. CONCLUSIONS: These results suggest that the extreme resistance of bdelloid rotifers to radiation might indeed be a consequence of their capacity to resist complete desiccation. This study paves the way to functional genetic experiments on A. vaga targeting promising candidate proteins playing central roles in radiation and desiccation resistance.

A transcriptomic examination of encased rotifer embryos reveals the developmental trajectory leading to long-term dormancy; are they "animal seeds"?

BACKGROUND: Organisms from many distinct evolutionary lineages acquired the capacity to enter a dormant state in response to environmental conditions incompatible with maintaining normal life activities. Most studied organisms exhibit seasonal or annual episodes of dormancy, but numerous less studied organisms enter long-term dormancy, lasting decades or even centuries. Intriguingly, many planktonic animals produce encased embryos known as resting eggs or cysts that, like plant seeds, may remain dormant for decades. Herein, we studied a rotifer Brachionus plicatilis as a model planktonic species that forms encased dormant embryos via sexual reproduction and non-dormant embryos via asexual reproduction and raised the following questions: Which genes are expressed at which time points during embryogenesis? How do temporal transcript abundance profiles differ between the two types of embryos? When does the cell cycle arrest? How do dormant embryos manage energy? RESULTS: As the molecular developmental kinetics of encased embryos remain unknown, we employed single embryo RNA sequencing (CEL-seq) of samples collected during dormant and non-dormant embryogenesis. We identified comprehensive and temporal transcript abundance patterns of genes and their associated enriched functional pathways. Striking differences were uncovered between dormant and non-dormant embryos. In early development, the cell cycle-associated pathways were enriched in both embryo types but terminated with fewer nuclei in dormant embryos. As development progressed, the gene transcript abundance profiles became increasingly divergent between dormant and non-dormant embryos. Organogenesis was suspended in dormant embryos, concomitant with low transcript abundance of homeobox genes, and was replaced with an ATP-poor preparatory phase characterized by very high transcript abundance of genes encoding for hallmark dormancy proteins (e.g., LEA proteins, sHSP, and anti-ROS proteins, also found in plant seeds) and proteins involved in dormancy exit. Surprisingly, this period appeared analogous to the late maturation phase of plant seeds. CONCLUSIONS: The study highlights novel divergent temporal transcript abundance patterns between dormant and non-dormant embryos. Remarkably, several convergent functional solutions appear during the development of resting eggs and plant seeds, suggesting a similar preparatory phase for long-term dormancy. This study accentuated the broad novel molecular features of long-term dormancy in encased animal embryos that behave like "animal seeds".

Genomic and Functional Evaluation of Two Lacticaseibacillus paracasei and Two Lactiplantibacillus plantarum Strains, Isolated from a Rearing Tank of Rotifers (Brachionus plicatilis), as Probiotics for Aquaculture.

Aquaculture plays a crucial role in meeting the increasing global demand for food and protein sources. However, its expansion is followed by increasing challenges, such as infectious disease outbreaks and antibiotic misuse. The present study focuses on the genetic and functional analyses of two Lacticaseibacillus paracasei (BF3 and RT4) and two Lactiplantibacillus plantarum (BF12 and WT12) strains isolated from a rotifer cultivation tank used for turbot larviculture. Whole-genome sequencing (WGS) and bioinformatics analyses confirmed their probiotic potential, the absence of transferable antibiotic resistance genes, and the absence of virulence and pathogenicity factors. Bacteriocin mining identified a gene cluster encoding six plantaricins, suggesting their role in the antimicrobial activity exerted by these strains. In vitro cell-free protein synthesis (IV-CFPS) analyses was used to evaluate the expression of the plantaricin genes. The in vitro-synthesized class IIb (two-peptide bacteriocins) plantaricin E/F (PlnE/F) exerted antimicrobial activity against three indicator microorganisms, including the well-known ichthyopathogen Lactococcus garvieae. Furthermore, MALDI-TOF MS on colonies detected the presence of a major peptide that matches the dimeric form of plantaricins E (PlnE) and F (PlnF). This study emphasizes the importance of genome sequencing and bioinformatic analysis for evaluating aquaculture probiotic candidates. Moreover, it provides valuable insights into their genetic features and antimicrobial mechanisms, paving the way for their application as probiotics in larviculture, which is a major bottleneck in aquaculture.

Horizontal acquisition of a DNA ligase improves DNA damage tolerance in eukaryotes.

Bdelloid rotifers are part of the restricted circle of multicellular animals that can withstand a wide range of genotoxic stresses at any stage of their life cycle. In this study, bdelloid rotifer Adineta vaga is used as a model to decipher the molecular basis of their extreme tolerance. Proteomic analysis shows that a specific DNA ligase, different from those usually involved in DNA repair in eukaryotes, is strongly over-represented upon ionizing radiation. A phylogenetic analysis reveals its orthology to prokaryotic DNA ligase E, and its horizontal acquisition by bdelloid rotifers and plausibly other eukaryotes. The fungus Mortierella verticillata, having a single copy of this DNA Ligase E homolog, also exhibits an increased radiation tolerance with an over-expression of this DNA ligase E following X-ray exposure. We also provide evidence that A. vaga ligase E is a major contributor of DNA breaks ligation activity, which is a common step of all important DNA repair pathways. Consistently, its heterologous expression in human cell lines significantly improves their radio-tolerance. Overall, this study highlights the potential of horizontal gene transfers in eukaryotes, and their contribution to the adaptation to extreme conditions.

Highly efficient CRISPR-mediated gene editing in a rotifer.

Rotifers have been studied in the laboratory and field for over 100 years in investigations of microevolution, ecological dynamics, and ecotoxicology. In recent years, rotifers have emerged as a model system for modern studies of the molecular mechanisms of genome evolution, development, DNA repair, aging, life history strategy, and desiccation tolerance. However, a lack of gene editing tools and transgenic strains has limited the ability to link genotype to phenotype and dissect molecular mechanisms. To facilitate genetic manipulation and the creation of reporter lines in rotifers, we developed a protocol for highly efficient, transgenerational, CRISPR-mediated gene editing in the monogonont rotifer Brachionus manjavacas by microinjection of Cas9 protein and synthetic single-guide RNA into the vitellaria of young amictic (asexual) females. To demonstrate the efficacy of the method, we created knockout mutants of the developmental gene vasa and the DNA mismatch repair gene mlh3. More than half of mothers survived injection and produced offspring. Genotyping these offspring and successive generations revealed that most carried at least 1 CRISPR-induced mutation, with many apparently mutated at both alleles. In addition, we achieved precise CRISPR-mediated knock-in of a stop codon cassette in the mlh3 locus, with half of injected mothers producing F2 offspring with an insertion of the cassette. Thus, this protocol produces knockout and knock-in CRISPR/Cas9 editing with high efficiency, to further advance rotifers as a model system for biological discovery.

Core microbiome profiles and their modification by environmental, biological, and rearing factors in aquaculture hatcheries.

16S rRNA gene sequencing and bacteria- and genus-specific quantitative PCR was used to profile microbial communities and their associated functions in water, live feed (microalgae, Artemia, and rotifer), and European sea bass and gilthead sea bream larvae from hatcheries in Greece and Italy. The transfer to larvae of genus containing potential pathogens of fish was more likely with Artemia and rotifer than with microalgae or water, irrespective of geographic location. The presence of potentially pathogenic bacteria (Vibrio and Pseudoalteromonas) in the core microbiota of water, live feed, and fish larvae, the enrichment of different bacterial resistance pathways and biofilm formation, and the overall low beneficial bacteria load during larval ontogeny emphasizes the risk for disease outbreaks. The present data characterizing microbiota in commercial aquaculture hatcheries provides a baseline for the design of strategies to manage disease and to model or remediate potential adverse environmental impacts.

Back to the roots, desiccation and radiation resistances are ancestral characters in bdelloid rotifers.

BACKGROUND: Bdelloid rotifers are micro-invertebrates distributed worldwide, from temperate latitudes to the most extreme areas of the planet like Antarctica or the Atacama Desert. They have colonized any habitat where liquid water is temporarily available, including terrestrial environments such as soils, mosses, and lichens, tolerating desiccation and other types of stress such as high doses of ionizing radiation (IR). It was hypothesized that bdelloid desiccation and radiation resistance may be attributed to their potential ability to repair DNA double-strand breaks (DSBs). Here, these properties are investigated and compared among nine bdelloid species collected from both mild and harsh habitats, addressing the correlation between the ability of bdelloid rotifers to survive desiccation and their capacity to repair massive DNA breakage in a phylogenetically explicit context. Our research includes both specimens isolated from habitats that experience frequent desiccation (at least 1 time per generation), and individuals sampled from habitats that rarely or never experienced desiccation. RESULTS: Our analysis reveals that DNA repair prevails in somatic cells of both desiccation-tolerant and desiccation-sensitive bdelloid species after exposure to X-ray radiation. Species belonging to both categories are able to withstand high doses of ionizing radiation, up to 1000 Gy, without experiencing any negative effects on their survival. However, the fertility of two desiccation-sensitive species, Rotaria macrura and Rotaria rotatoria, was more severely impacted by low doses of radiation than that of desiccation-resistant species. Surprisingly, the radioresistance of desiccation-resistant species is not related to features of their original habitat. Indeed, bdelloids isolated from Atacama Desert or Antarctica were not characterized by a higher radioresistance than species found in more temperate environments. CONCLUSIONS: Tolerance to desiccation and radiation are supported as ancestral features of bdelloid rotifers, with a group of species of the genus Rotaria having lost this trait after colonizing permanent water habitats. Together, our results provide a comprehensive overview of the evolution of desiccation and radiation resistance among bdelloid rotifers.

m6A-mediated nonhomologous end joining (NHEJ) pathway regulates senescence in Brachionus plicatilis (Rotifera).

Epigenetic modifications play an important role in the regulation of senescence. N6-methyladenosine (m6A) is the most abundant modification of mRNA. However, the impact of m6A on senescence remains largely unknown at the animal individual level. Standard model organisms Caenorhabditis elegans and Drosophila melanogaster lack many gene homologs of vertebrate m6A system that are present in other invertebrates. In this study, we employed a small aquatic invertebrate Brachionus plicatilis which has been used in aging studies for nearly 100 years to study how m6A affects aging. Phylogenetic analysis confirmed that rotifers' m6A pathway has a conserved methyltransferase complex but no demethylases and the m6A reading system was more akin to that of vertebrates than that of D. melanogaster. m6A methyltransferases are highly expressed during development but reduces dramatically during aging. Knockdown of METTL3 results in decreased fecundity and premature senescence of rotifers. Furthermore, RT-qPCR analysis indicates a role for m6A in the nonhomologous end joining (NHEJ) pathway of DNA double-strand breaks (DSBs) repair. Altogether, our work reveals a senescence regulatory model for the rotifer METTL3-m6A-NHEJ pathway.

An experimental test of the growth rate hypothesis as a predictive framework for microevolutionary adaptation.

The growth rate hypothesis (GRH) posits that the relative body phosphorus content of an organism is positively related to somatic growth rate, as protein synthesis, which is necessary for growth, requires P-rich rRNA. This hypothesis has strong support at the interspecific level. Here, we explore the use of the GRH to predict microevolutionary responses in consumer body stoichiometry. For this, we subjected populations of the rotifer Brachionus calyciflorus to selection for fast population growth rate (PGR) in P-rich (HPF) and P-poor (LPF) food environments. With common garden transplant experiments, we demonstrate that in HP populations evolution toward increased PGR was concomitant with an increase in relative phosphorus content. In contrast, LP populations evolved higher PGR without an increase in relative phosphorus content. We conclude that the GRH has the potential to predict microevolutionary change, but that its application is contingent on the environmental context. Our results highlight the potential of cryptic evolution in determining the performance response of populations to elemental limitation of their food resources.

Changes in key life-history traits and transcriptome regulations of marine rotifer Brachionus plicatilis in eliminating harmful algae Phaeocystis.

Rotifers have great potential in controlling the harmful algae Phaeocystis blooms that frequently occur in coastal waters. To evaluate the effects of harmful algae on the key life-history traits of rotifer in eliminating Phaeocystis and reveal the underlying mechanism of these effects, we fed Brachionus plicatilis with Chlorella vulgaris and Phaeocystis globosa respectively, recorded the key life-history traits, and conducted transcriptomic analysis. Results showed that the rotifers feeding on P. globosa significantly decreased total offspring but obviously prolonged lifespan compared to those feeding on C. vulgaris, indicating that there was a trade-off between the reproduction and lifespan of rotifers feeding on algae with different nutrient contents. Nevertheless, rotifers can completely eliminate the population of P. globosa. The changes in the reproduction and lifespan of rotifers are highly correlated with algal key nutrition and the expression of some related genes. Transcriptomic analysis showed that the changes in the key life history traits of rotifers feeding on harmful algae are determined by regulating the expression of some key genes involved in the pathways of carbohydrate digestion and absorption, glycolysis, gluconeogenesis, unsaturated fatty acid biosynthesis, and environmental stress. Understanding the trade-off of the key life history traits of zooplankton in eliminating harmful algae from the underlying mechanism helps improve their application for controlling harmful algae.

Variation in heat shock protein 40 kDa relates to divergence in thermotolerance among cryptic rotifer species.

Genetic divergence and the frequency of hybridization are central for defining species delimitations, especially among cryptic species where morphological differences are merely absent. Rotifers are known for their high cryptic diversity and therefore are ideal model organisms to investigate such patterns. Here, we used the recently resolved Brachionus calyciflorus species complex to investigate whether previously observed between species differences in thermotolerance and gene expression are also reflected in their genomic footprint. We identified a Heat Shock Protein gene (HSP 40 kDa) which exhibits cross species pronounced sequence variation. This gene exhibits species-specific fixed sites, alleles, and sites putatively under positive selection. These sites are located in protein binding regions involved in chaperoning and may therefore reflect adaptive diversification. By comparing three genetic markers (ITS, COI, HSP 40 kDa), we revealed hybridization events between the cryptic species. The low frequency of introgressive haplotypes/alleles suggest a tight, but not fully impermeable boundary between the cryptic species.

Meiotic transmission patterns of additional genomic elements in Brachionus asplanchnoidis, a rotifer with intraspecific genome size variation.

Intraspecific genome size (GS) variation in Eukaryotes is often mediated by additional, nonessential genomic elements. Physically, such additional elements may be represented by supernumerary (B-)chromosomes or by large heterozygous insertions into the regular chromosome set. Here we analyze meiotic transmission patterns of Megabase-sized, independently segregating genomic elements (ISEs) in Brachionus asplanchnoidis, a planktonic rotifer that displays an up to two-fold intraspecific GS variation due to variation in size and number of these elements. To gain insights into the meiotic transmission patterns of ISEs, we measured GS distributions of haploid males produced by individual mother clones using flow cytometry and compared these distributions to theoretical distributions expected under a range of scenarios. These scenarios considered transmission biases resembling (meiotic) drive, or cosegregation biases, e.g., if pairs of ISEs preferentially migrated towards the same pole during meiosis. We found that the inferred transmission patterns were diverse and ranged from positive biases (suggesting drive) to negative biases (suggesting drag), depending on rotifer clone and its ISE composition. Additionally, we obtained evidence for a negative cosegregation bias in some of the rotifer clones, i.e., pairs of ISEs exhibited an increased probability of migrating towards opposite poles during meiosis. Strikingly, these transmission and segregation patterns were more similar among members of a genetically homogeneous inbred line than among outbred members of the population. Comparisons between early and late stages of haploid male embryonic development (e.g., young synchronized male eggs vs. hatched males) showed very similar GS distributions, suggesting that transmission biases occur very early in male development, or even during meiosis. Very large genome size was associated with reduced male embryonic survival, suggesting that excessive amounts of ISEs might be detrimental to male fitness. Altogether, our results indicate considerable functional diversity of ISEs in B. asplanchnoidis, with consequences on meiotic transmission and embryonic survival.

Phenotypic toxicity, oxidative response, and transcriptomic deregulation of the rotifer Brachionus plicatilis exposed to a toxic cocktail of tire-wear particle leachate.

Tire-wear particles (TWPs) are potential source of microplastic (MP) pollution in marine environments. Although the hazardous effects of MPs on marine biota have received considerable attention, the toxicity of TWPs and associated leachates remain poorly understood. Here, to assess the toxicity of TWP leachate and the underlying mechanisms of toxicity, the phenotypic and transcriptomic responses of the rotifer Brachionus plicatilis were assessed with chemistry analysis of a TWP leachate. Although acute toxicity was induced, and a variety of metals and polyaromatic hydrocarbons were detected in the leachate, levels were below the threshold for acute toxicity. The results of particle analysis suggest that the acute toxicity observed in our study is the result of a toxic cocktail of micro- and/or nano-sized TWPs and other additives in TWP leachate. The adverse effects of TWP leachate were associated with differential expression of genes related to cellular processes, stress response, and impaired metabolism, with further oxidative stress responses. Our results imply that TWPs pose a greater threat to marine biota than other plastic particles as they constitute a major source of nano- and microplastics that have synergistic effects with the additives contained in TWP leachate.