Neuropeptides are essential neuronal signaling molecules that orchestrate animal behavior and physiology via actions within the nervous system and on peripheral tissues. Due to the small size of biologically active mature peptides, their identification on a proteome-wide scale poses a significant challenge using existing bioinformatics tools like BLAST. To address this, we have developed NeuroPeptide-HMMer (NP-HMMer), a hidden Markov model (HMM)-based tool to facilitate neuropeptide discovery, especially in underexplored invertebrates. NP-HMMer utilizes manually curated HMMs for 46 neuropeptide families, enabling rapid and accurate identification of neuropeptides. Validation of NP-HMMer on Drosophila melanogaster, Daphnia pulex, Tribolium castaneum and Tenebrio molitor demonstrated its effectiveness in identifying known neuropeptides across diverse arthropods. Additionally, we showcase the utility of NP-HMMer by discovering novel neuropeptides in Priapulida and Rotifera, identifying 22 and 19 new peptides, respectively. This tool represents a significant advancement in neuropeptide research, offering a robust method for annotating neuropeptides across diverse proteomes and providing insights into the evolutionary conservation of neuropeptide signaling pathways.

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.

Artificial light at night (ALAN) may pose threat to rotifer Brachionus plicatilis. Additionally, the food of rotifer, i.e. algal community composition, often fluctuates. Thus, we selected five wavelengths of ALAN (purple, blue, green, red, white) and a three-colored light flashing mode (3-Flash) to test their impacts on life history traits of B. plicatilis with different food experiences, including those feeding Chlorella vulgaris (RC) or Phaeocystis globosa (RP). Results indicated purple ALAN promoted RC development, white ALAN inhibited RC development, while 3-Flash and white ALAN promoted RP development. Under red and white ALAN, RP increased fecundity but decreased lifespan. High-quality food enhanced rotifer\'s resistance to the impact of ALAN on lifespan. ALAN and food experience interacted on B. plicatilis. The effect of blue ALAN has less negative effects on B. plicatilis, based on hierarchical cluster analysis. Such findings are helpful to evaluate the potential impact of ALAN on marine zooplankton.

Vibrio species are naturally found in estuarine and marine ecosystems, but are also recognized as significant human enteropathogens, often linked to seafood-related illnesses. In aquaculture settings, Vibrio poses a substantial risk of infectious diseases, resulting in considerable stock losses and prompting the use of antimicrobials. However, this practice contributes to the proliferation of antimicrobial-resistant (AMR) bacteria and resistance genes. Our investigation aimed to explore the potential of biological agents such as bacteriophage CH20 and endolysin LysVPp1 in reducing Vibrio bacterial loads in both rotifer and fish larvae. LysVPp1\'s lytic activity was assessed by measuring absorbance reduction against various pathogenic Vibrio strains. Phage CH20 exhibited a limited host range, affecting only Vibrio alginolyticus GV09, a highly pathogenic strain. Both CH20 and LysVPp1 were evaluated for their effectiveness in reducing Vibrio load in rotifers or fish larvae through short-setting bioassays. Our results demonstrated the significant lytic effect of endolysin LysVPp1 on strains of Vibrio alginolyticus, Vibrio parahaemolyticus, and Vibrio splendidus. Furthermore, we have showcased the feasibility of reducing the load of pathogenic Vibrio in live feed and fish larvae by using a non-antibiotic-based approach, such as lytic phage and endolysin LysVPp1, thus contributing to the progress of a sustainable aquaculture from a One Health perspective.

Microplastics (MPs) are a major concern in marine ecosystem because MPs are persistent and ubiquitous in oceans and are easily consumed by marine biota. Although many studies have reported the toxicity of MPs to marine biota, the toxicity of environmentally relevant types of MPs is little understood. We investigated the toxic effects of fragmented polyethylene terephthalate (PET) MP, one of the most abundant MPs in the ocean, on the marine rotifer Brachionus koreanus at the individual and molecular level. No significant rotifer mortality was observed after exposure to PET MPs for 24 and 48 h. The ingestion and egestion assays showed that rotifers readily ingested PET MPs in the absence of food but not when food was supplied; thus, there were also no chronic effects of PET MPs. In contrast, intracellular reactive oxygen species levels and glutathione S-transferase activity in rotifers were significantly increased by PET MPs. Transcriptomic and metabolomic analyses revealed that genes and metabolites related to energy metabolism and immune processes were significantly affected by PET MPs in a concentration-dependent manner. Although acute toxicity of PET MPs was not observed, PET MPs are potentially toxic to the antioxidant system, immune system, and energy metabolism in rotifers.

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.

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.

Terrestrial microinvertebrates provide important carbon and nutrient cycling roles in soil environments, particularly in Antarctica where larger macroinvertebrates are absent. The environmental preferences and ecology of rotifers and tardigrades in terrestrial environments, including in Antarctica, are not as well understood as their temperate aquatic counterparts. Developing laboratory cultures is critical to provide adequate numbers of individuals for controlled laboratory experimentation. In this study, we explore aspects of optimising laboratory culturing for two terrestrially sourced Antarctic microinvertebrates, a rotifer (Habrotrocha sp.) and a tardigrade (Acutuncus antarcticus). We tested a soil elutriate and a balanced salt solution (BSS) to determine their suitability as culturing media. Substantial population growth of rotifers and tardigrades was observed in both media, with mean rotifer population size increasing from 5 to 448 ± 95 (soil elutriate) and 274 ± 78 (BSS) individuals over 60 days and mean tardigrade population size increasing from 5 to 187 ± 65 (soil elutriate) and 138 ± 37 (BSS) over 160 days. We also tested for optimal dilution of soil elutriate in rotifer cultures, with 20-80% dilutions producing the largest population growth with the least variation in the 40% dilution after 36 days. Culturing methods developed in this study are recommended for use with Antarctica microinvertebrates and may be suitable for similar limno-terrestrial microinvertebrates from other regions.

The unfavorable phenomenon of activated sludge bulking that occurs in sewage treatment plants (WWTPs) is caused by the over-proliferation of filamentous bacteria that should be limited by the Lecane rotifers that feed on them; however, predatory, rotiferovorous fungi that often inhabit WWTPs pose a real threat to these organisms. To solve this problem, we investigated the interaction of the fungus Clonostachys rosea, which is a known Biological Control Agent (BCA) and the predacious Zoophagus sp. in simplified laboratory culture conditions. The presence of C. rosea in the cultures reduced the number of active traps, thus translating into a much smaller number of rotifers being caught. The mycelium of C. rosea was labeled with a red fluorescent protein (RFP). The life cycle of C. rosea that were attacking Zoophagus sp. (hunting for rotifers) is described. C. rosea spores germinate into single-celled forms and penetrate the interior of the Zoophagus mycelium where they feed on the cytoplasm. Then is the mycelium produced abundantly and forms conidiophores. This type of life strategy has not been known before. The obtained results demonstrated the potential of C. rosea as a BCA that can be used to protect rotifers in the event of an infection of activated sludge by the predatory fungi that threaten the rotifer population.

Characterizing swimming behavior can provide a holistic assessment of the health, physiology and ecology of microfaunal species when done in conjunction with measuring other biological parameters. However, tracking and quantifying microfauna swimming behavior using existing automated tools is often difficult due to the animals\' small size or transparency, or because of the high cost, expertise, or labor needed for the analysis. To address these issues, we created a cost-effective, user-friendly protocol for behavior analysis that employs the free software packages HitFilm and ToxTrac along with the R package \'trajr\' and used the method to quantify the behavior of rotifers. This protocol can be used for other microfaunal species for which investigators may face similar issues in obtaining measurements of swimming behavior.