Scuticociliatia is one of the most species-rich subclasses in the phylum Ciliophora. The evolutionary relationships among Scuticociliatia groups have long been very unclear due to the homogeneity of morphology and insufficiency of molecular data. With morphological and multi-gene-based molecular data presented here, the evolutionary phylogeny of several Scuticociliatia taxa that were hitherto especially poorly defined is analyzed and discussed. The results indicate: (1) all scuticociliates cluster into two well supported and one poorly supported group, representing three order-level taxa; (2) with the support of both morphological and molecular data, a new family Homalogastridae fam. nov. is proposed in the order Philasterida; (3) Parauronema is formally transferred to Uronematidae and Potomacus is treated as incertae sedis in the order Philasterida, therefore Parauronematidae is proposed to be a junior synonym of Uronematidae; (4) the genus Madsenia and the species Parauronema longum and Pseudocyclidium longum are treated as incertae sedis, while the genus Protophyra should be maintained in the family Ancistridae. In addition, the putative secondary structure of internal transcribed spacer 2 (ITS2) of representative taxa from the three orders of Scuticociliatia are analyzed, and consensus structures and nucleotide composition in each order are exhibited.

Psammophillic ciliates are an integral part of the foodweb despite being underrepresented in terms of molecular phylogeny and modern taxonomy. To investigate the karyorelictean group, sampling was conducted in interstitial marine habitats in China for ciliates living between the sand grains, resulting in an examination of the families Cryptopharyngidae Jankowski, 1980 and Kentrophoridae Jankowski, 1980. Three species, i.e., Cryptopharynx setigerus Kahl, 1928, Kentrophoros fasciolatus (Sauerbrey, 1928) Foissner, 1995 and K. fistulosus (Fauré-Fremiet, 1950) Foissner, 1995, are clearly recognized as being cosmopolitan, while other species await further recording. Phylogenetic analyses were carried out based on updated data. These revealed that the families Cryptopharyngidae and Kentrophoridae are closely related, and most genera studied are monophyletic, although Cryptopharynx qingdaoensis n. sp. is located within the Kentrophoridae branch. Brief revisions of two genera, namely Cryptopharynx Kahl, 1928 and Kentrophoros Sauerbrey, 1928, are provided including keys to the identification of nine species belonging to the former and 12 species belonging to the latter. One new genus, Parakentrophoros n. gen., and one new species, Cryptopharynx qingdaoensis n. sp., are described and a new combination, Parakentrophoros canalis (Wright, 1982) n. comb., is established. Finally, it appears that the subapical oral apparatus undergoes a gradual degeneration process from Cryptopharyngidae to Kentrophoridae.

Growth of the Pacific white shrimp Litopenaeus vannamei, the most important farmed crustacean, has consistently been a focal point for breeders. Over the past decades, some candidate genes for shrimp growth have been identified. However, further research is needed to elucidate the molecular regulatory mechanism of these genes. LvMmd2 was previously identified as a candidate gene that may inhibit the growth of L. vannamei. In this study, we analyzed the genotype and expression of the LvMmd2 gene in a breeding family and indicated its role as a growth-inhibiting gene. We found that LvMmd2 co-localized with its homolog LvPAQR3 at the Golgi apparatus. Using co-immunoprecipitation (Co-IP) and DUAL membrane system yeast two-hybrid (MbY2H), we indicated the interactions between LvMmd2 and LvPAQR3, LvPAQR3 and LvRaf1, as well as LvMmd2 and LvRho. These results suggest that LvMmd2 directly and indirectly regulates the Ras signaling pathway. Furthermore, we show that the LvMmd2 gene may indirectly affect the PI3K/AKT, insulin, and Hippo signaling pathways to regulate cell proliferation and differentiation via LvPAQR3 and LvRaf1. Through transcriptome and MbY2H analyses, we have also revealed the interaction between LvMmd2 and proteins involved in growth, immunity, protein transport, synthesis, and modification. These findings demonstrate the various molecular pathways through which LvMmd2 regulates L. vannamei growth. This study provides insights into the mechanism of shrimp growth regulated by Mmd2, enhances our understanding of LvMmd2 function, and highlights its potential application in shrimp breeding.

Declining populations and bottlenecks lead to the accumulation of deleterious mutations in fish populations. These processes also trigger genetic purging, which is a key genetic factor in reducing the deleterious burdens and increasing population viability. However, there is a lack of empirical evidence on the interaction between demographic history and the genome-wide pattern of deleterious variations. Here, we generated genome resequencing data of Eleutheronema rhadinum from China and Thailand, representing the major distribution of the species’ southern regions. E. rhadinum had exceptionally low genome-wide variability and experienced dramatic population expansions followed by continuous declines. The geographical divergence, which occurred~23,000 years ago, shaped different demographic trajectories and generated different regional patterns of deleterious mutations in China and Thailand populations. Several lines of evidence revealed that this geographical pattern of deleterious mutation was driven by the purging of highly deleterious mutations. We showed that purifying selection had inbreeding-associated fitness costs and was more efficient against missense mutations in the Thailand population, which had the lowest genetic burden of homozygous deleterious mutations. Multiple evolutionarily conserved protein domains were disrupted by the loss-of-function mutations, posing a high probability of gene functionality elimination. Moreover, thermal and salinity genes (Trpm3, Nek4, Gtf2f2, Cldn14) were identified in genomic divergence regions of E. rhadinum among China and Thailand populations. Our findings highlight the importance of demographic history factors shaping the geographical patterns of deleterious mutations. The results serve to deepen our understanding of the adaptive evolution and divergence of E. rhadinum with implications for other marine fish.

Determining 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.

Antarctic krill Euphausia superba, one of the most abundant species on the planet, is a keystone species of the Southern Ocean ecosystem. In the present study, we analyzed the RNA virome of Antarctic krill via metatranscription methods. The results showed that only 0.39% (49/12, 558) of the resultant unigenes could be assigned to known viral taxa, which were most similar to 17 known viruses, including nine invertebrate viruses, two vertebrate viruses, three protozoan viruses and three mycoviruses. However, most of the detected viruses possessed low amino acid similarity with counterparts in the viral databases. Penaeus vannamei picornavirus (PvPV; Family Picornaviridae) and covert mortality nodavirus (CMNV; Family Nodaviridae) were the two most abundant viruses in the Antarctic krill RNA virome. Notably, PvPV and CMNV are known pathogens to multiple aquatic animals according to epidemiological survey and exposure experiments, whereby PvPV positive krill caused clinical symptoms and histopathological lesions to P. vannamei and similarly, CMNV infection altered the swimming and feeding behavior of parent marine medaka Oryzias melastigma and caused tissue damage and even spinal curvature of the offspring. Results herein reveal, for the first time, the high abundance and taxonomic diversity of viruses in Antarctic krill while simultaneously highlighting the risk of an important virus reservoir to global aquaculture, and the potential impact on animals in the Antarctic ecosystem.

Nervous necrosis virus (NNV) is the etiological agent of viral encephalopathy and retinopathy in many fish species, including European sea bass (Dicentrachus labrax) and is of great economic losses to fish farmers. To solve this problem in fish production, antimicrobial peptides (AMPs) have been identified as potential candidates for NNV treatment in aquaculture. Hepcidin (Hamp) is one of the most promising AMPs. Thus, we aimed to evaluate the therapeutic application of Hamp synthetic peptide after NNV challenge. Hamp was able to significantly increase survival rates and ameliorate clinical signs of the disease, though the viral levels, determined by viral replication and immunolocalization, were not affected. Synthetic Hamp increased the immunoglobulin M (IgM) and AMP protein levels in serum and some tissues respect to the levels found in NNV-infected fish. However, Hamp peptide decreased the NNV-induced bactericidal activity. At the gene level, Hamp exerted anti-inflammatory properties, reducing the pro-inflammatory response orchestrated by NNV, probably preventing neuronal damage. Apart from this, Hamp up-regulated the expression of adhesion molecules that facilitated the recruitment of immune cells, namely T helper and B cells, probably to orchestrate the adaptive response. To conclude, Hamp immunomodulatory properties and therapeutic application against NNV are very promising for its use in aquaculture.

Obesity has become a worldwide health problem. Seeking natural products with anti-obesity activity from lots of fungi has drawn the attention of pharmacologists. In our study, dipenipenoids A and B (1 and 2), the first dimeric indole-diterpenoids with a rare C-20–C-22′ linkage, and their monomers (3 and 4), were isolated from a marine-derived Penicillium sp. CF-06 fungus from Suaeda salsa. The absolute configurations of 1–3 were assigned by the calculated TDDFT ECD method. The structure of 4 was verified by a single-crystal X-ray diffraction method for the first time. Interestingly, 1 and 2 displayed significant effects on the differentiation of 3T3-L1 adipocytes by down-regulating the expression of peroxisome proliferator-activated receptor gamma (PPARγ) and CCAAT/enhancer binding protein alpha (C/EBPα) proteins, while monomers 3 and 4 exhibited no activity. Molecular docking results explained the mechanism that the interaction between dimer 1 and PPARγ was stronger than that between monomer 3 and PPARγ. Our research could provide new insight for the discovery of anti-obesity drugs.

Zeaxanthin, an oxygenated carotenoid derivative with potent antioxidative properties, is produced by many organism taxa. Flavobacteriaceae are widely distributed in marine environments; however, the zeaxanthin biosynthesis property in this family remains incompletely explored. Here, we characterized zeaxanthin production by marine Flavobacteriaceae strains and elucidated underlying molecular mechanisms. Eight Flavobacteriaceae strains were isolated from the phycosphere of various dinoflagellates. Analyses of the zeaxanthin production in these strains revealed yields ranging from 5 to 3289 µg/g of dry cell weight. Genomic and molecular biology analyses revealed the biosynthesized zeaxanthin through the mevalonate (MVA) pathway diverging from the 2-C-methyl-d-erythritol-4-phosphate (MEP) pathway commonly observed in most Gram-negative bacteria. Furthermore, comprehensive genome analyses of 322 culturable marine Flavobacteriale strains indicated that the majority of Flavobacteriaceae members possess the potential to synthesize zeaxanthin using precursors derived from the MVA pathway. These data provide insight into the zeaxanthin biosynthesis property in marine Flavobacteriaceae strains, highlighting their ecological and biotechnological relevance.

Estuaries are usually characterized by strong spatial and temporal variability in water physicochemical conditions and are often largely affected by human activities. One important source of variability is caused by tides that can swiftly alter not only physicochemical conditions but also the abundance and composition of the biota. The effect of the diurnal tidal cycle on microbial community composition during different seasons remains uncertain, although this knowledge underlies having effective monitoring programs for water quality and potential identification of health risk conditions. In this study, we assessed the bacterioplankton community composition and diversity across four tidal water levels in a tropical estuary characterized by a mixed semidiurnal tide regime (i.e., two high and two low tides of varying amplitudes) during both dry and wet seasons. The bacterial community composition varied significantly among the four tidal levels, but only during the dry season, when the influence of the seawater intrusion was largest. Bacterial indicators’ taxa identified using the Indicator Value Index were found within Cyanobacteria, Actinobacteriota, Bacteroidota, and Proteobacteria. The indicator taxon Cyanobium sp. had a prominent presence across multiple tidal levels. The main predicted phenotypes of the bacterial communities were associated with potential pathogenicity, gram-negative, and biofilm formation traits. While there were no marked predicted phenotypic differences between seasons, pathogenic and gram-negative traits were more prevalent in the dry season, while biofilm formation traits dominated in the wet season. Overall, our findings underscore the intricate relationship between river hydrodynamics and bacterial composition variability and hint a significant human impact on the water quality of the Bangpakong River.

The interaction of riverine inputs and ocean current systems causes complex spatiotemporal variations in phytoplankton dynamics in marginal seas of the northwest Pacific Ocean, yet quantitative assessments of these variations and their causes remain limited. Here we evaluate phytoplankton biomass and community structure changes using lipid biomarkers, accompanying ocean circulation and nutrient variations in surface waters collected in spring and summer of 2017–2018 at 118 sites in the East China Sea off the Zhejiang coast. High biomass of diatoms, inferred from brassicasterol concentrations, shifted from the south in spring to the north in summer, while high dinoflagellate biomass, inferred from dinosterol concentrations, occurred mainly in the Changjiang (Yangtze) River plume and adjacent areas in both seasons. Seasonal variation in phytoplankton distribution was linked to the spatial extents of water masses such as the Changjiang Diluted Water (CDW) and the intrusion of the Kuroshio Subsurface Water (KSSW). A three end-member mixing model was applied to quantify water mass contributions. The results showed that an increase in the KSSW (from 0 to 40%) and a decrease in the CDW (from 100 to 20%) resulted in a significant (20%) increase in diatom proportions and a significant (20%) decrease in dinoflagellate proportions. Dinoflagellate proportions were highest in the CDW-dominated region, while diatoms and total phytoplankton biomass were higher in the CDW–KSSW mixing region and the KSSW-dominated region. This study highlights the dynamic response of the phytoplankton community to water mass changes in marginal seas that can aid coastal ecosystem management.

Bacterial community play an essential role in regulating water quality and the global biogeochemical cycle in aquatic ecosystems. However, how trophic interactions (i.e., biotic factors) regulate the diversity and composition of bacterial community in lake ecosystems remains unknown. Here, we employed DNA meta-barcoding of water samples to explore the impact of bacterivorous protozoans on the bacterial community. The results showed significant seasonal variations in the diversity and composition of both bacterial and protist communities. The composition of bacterivorous protozoans was identified as the primary predictor for the bacterial community alpha diversity in spring and summer, and for beta diversity in spring and autumn, indicating that biotic interactions play a greater role in driving the diversity of bacterial community across different seasons. Biological factors were more important than environmental factors for explaining the variations in the relative abundance of several bacterial genera (i.e., Pseudoxanthomonas, hgcI_clade, and Pseudorhodobacter). Network analyses showed that bacterial networks differed among seasons, and the autumn network exhibited the highest stability. Our findings indicated that the bacterial community stability was significantly affected by environmental factors, specifically SO₄^2–and PO₄^3–, rather than bacterivorous protozoans. Overall, our findings provide new perspectives on the role of trophic interactions in maintaining the structure of bacterial community in different seasons, and enhance our understanding of the bacterial community assembly in lake ecosystems.

Free trans-4-hydroxy-L-proline (T4LHyp), a non-proteinogenic amino acid, is mainly released from the degradation of collagen, hydroxyproline-rich glycoproteins, and some peptide antibiotics in nature. Although it has been known that some terrestrial bacteria utilize T4LHyp as carbon and nitrogen source via a T4LHyp gene cluster, which and how marine microorganisms catabolize T4LHyp still remains unclear. Here, five T4LHyp-utilizing marine bacterial strains, Halomonas sp. 5021, Salinicola sp. 4072, Alteromonas sp. 6022, Alteromonas sp. 5112 and Alteromonas sp. 30521, were isolated from hydrothermal vent sediment samples collected from the southwest Indian Ocean. While Halomonas sp. 5021 can utilize T4LHyp as both a nitrogen and carbon source, the other four strains can utilize T4LHyp as only a nitrogen source. Then, the T4LHyp catabolic mechanisms of Halomonas sp. 5021 and Salinicola sp. 4072, as a representative of the four strains, were further investigated by genomic, transcriptional, and biochemical analyses. Halomonas sp. 5021 adopts an intact T4LHyp gene cluster containing four enzymes to catabolize T4LHyp into NH₃ and α-ketoglutarate to provide nitrogen and carbon sources for its growth. Compared to Halomonas sp. 5021, Salinicola sp. 4072 lacks an α-KGSA dehydrogenase gene in the T4LHyp gene cluster and can only catabolize T4LHyp into NH₃ and α-ketoglutarate semialdehyde to provide a nitrogen source for its growth. Bioinformatic investigation showed that the 5021-like and 4072-like T4LHyp gene clusters are predominantly found in bacteria from Pseudomonadota, which are widely distributed in multiple marine habitats. Thus, Pseudomonadota bacteria are likely the dominant group to drive the recycling and mineralization of T4LHyp in the ocean.