Wood-Ljungdahl pathway found in novel marine Korarchaeota groups illuminates their evolutionary history.

Korarchaeota, due to its rarity in common environments, is one of the archaeal phyla that has received the least attention from researchers. It was previously thought to consist solely of strict thermophiles. However, our study provides genetic evidence for the presence of korarchaeal members in temperate subsurface seawater. Furthermore, a systematic reclassification of the Korarchaeota based on 16S rRNA genes and genomes has revealed three novel marine groups (Kor-6 to Kor-8) at the root of the Korarchaeota branch. Kor-6 contains microbes that are present in moderate temperatures. All three novel marine phyla possess genes for the Wood-Ljungdahl pathway, and Kor-7 and Kor-8 possess fewer genes encoding oxygen resistance traits than other korarchaeal groups, suggesting a distinct lifestyle for these novel phyla. Our results, together with estimations of Korarchaeota divergence times, suggest that oxygen availability may be one of the important factors that have influenced the evolution of Korarchaeota. IMPORTANCE Korarchaeota were previously thought to inhabit exclusively high-temperature environments. However, our study provides genetic evidence for their unexpected presence in temperate marine waters. Through analysis of publicly available korarchaeal reference data, we have systematically reclassified Korarchaeota and identified the existence of three previously unknown marine groups (Kor-6, Kor-7, and Kor-8) at the root of the Korarchaeota branch. Comparative analysis of their gene content revealed that these novel groups exhibit a lifestyle distinct from other Korarchaeota. Specifically, they have the ability to fix carbon exclusively via the Wood-Ljungdahl (WL) pathway, and the genomes within Kor-7 and Kor-8 contain few genes encoding antioxidant enzymes, indicating their strictly anaerobic lifestyle. Further studies suggest that the genes related to methane metabolism and the WL pathway may have been inherited from a common ancestor of the Korarchaeota and that oxygen availability may be one of the important evolutionary factors that shaped the diversification of this archaeal phylum.

Non-negligible roles of archaea in coastal carbon biogeochemical cycling.

Coastal zones are among the world's most productive ecosystems. They store vast amounts of organic carbon, as 'blue carbon' reservoirs, and impact global climate change. Archaeal communities are integral components of coastal microbiomes but their ecological roles are often overlooked. However, archaeal diversity, metabolism, evolution, and interactions, revealed by recent studies using rapidly developing cutting-edge technologies, place archaea as important players in coastal carbon biogeochemical cycling. We here summarize the latest advances in the understanding of archaeal carbon cycling processes in coastal ecosystems, specifically, archaeal involvement in CO2 fixation, organic biopolymer transformation, and methane metabolism. We also showcase the potential to use of archaeal communities to increase carbon sequestration and reduce methane production, with implications for mitigating climate change.

An Ancient Respiratory System in the Widespread Sedimentary Archaea Thermoprofundales.

Thermoprofundales, formerly Marine Benthic Group D (MBG-D), is a ubiquitous archaeal lineage found in sedimentary environments worldwide. However, its taxonomic classification, metabolic pathways, and evolutionary history are largely unexplored because of its uncultivability and limited number of sequenced genomes. In this study, phylogenomic analysis and average amino acid identity values of a collection of 146 Thermoprofundales genomes revealed five Thermoprofundales subgroups (A-E) with distinct habitat preferences. Most of the microorganisms from Subgroups B and D were thermophiles inhabiting hydrothermal vents and hot spring sediments, whereas those from Subgroup E were adapted to surface environments where sunlight is available. H2 production may be featured in Thermoprofundales as evidenced by a gene cluster encoding the ancient membrane-bound hydrogenase (MBH) complex. Interestingly, a unique structure separating the MBH gene cluster into two modular units was observed exclusively in the genomes of Subgroup E, which included a peripheral arm encoding the [NiFe] hydrogenase domain and a membrane arm encoding the Na+/H+ antiporter domain. These two modular structures were confirmed to function independently by detecting the H2-evolving activity in vitro and salt tolerance to 0.2 M NaCl in vivo, respectively. The peripheral arm of Subgroup E resembles the proposed common ancestral respiratory complex of modern respiratory systems, which plays a key role in the early evolution of life. In addition, molecular dating analysis revealed that Thermoprofundales is an early emerging archaeal lineage among the extant MBH-containing microorganisms, indicating new insights into the evolution of this ubiquitous archaea lineage.

Naringenin attenuates inflammation, apoptosis, and ferroptosis in silver nanoparticle-induced lung injury through a mechanism associated with Nrf2/HO-1 axis: In vitro and in vivo studies.

With the wide application of silver nanoparticles (AgNPs), their potential damage to human health needs to be investigated. Lung is one of the main target organs after inhalation of AgNPs. Naringenin has been reported to have anti-inflammatory and anti-oxidative properties. This study aims to evaluate the protective effects of naringenin against AgNPs-induced lung injury and determine the underlying mechanism. In in vivo experiments, AgNPs were intratracheally instilled into ICR mice (l mg/kg) to establish a lung injury model. These mice were then treated with naringenin by oral gavage (25, 50, 100 mg/kg) for three days. Naringenin treatment decreased the levels of white blood cells, neutrophils, and lymphocytes in the blood, ameliorated lung injury, suppressed the release of pro-inflammatory cytokines, normalized ferroptotic markers and prevented oxidative stress with elevating Nrf2 and HO-1 protein expressions in lung. In in vitro experiments, BEAS-2B cells were firstly treated with AgNPs (320 µg/mL) and then naringenin (25, 50, and 100 µM), respectively. Naringenin attenuated AgNPs-induced oxidative stress and inflammatory response. Moreover, naringenin attenuated AgNPs-induced apoptosis with modulated low BAX, CytC, cleaved Caspase9, cleaved Caspase3 but high Bcl2. Furthermore, naringenin effectively decreased ferroptotic markers and increased the protein expressions of Nrf2 and HO-1, as well as increased the nuclear translocation of Nrf2. Importantly, the anti-apoptotic and anti-ferroptotic effects of naringenin in BEAS-2B cells were found to be at least partially Nrf2-dependent. These results indicated that naringenin exerted anti-inflammation, anti-apoptosis, and anti-ferroptosis effects and protected against AgNPs-induced lung injury at least partly via activating Nrf2/HO-1 signaling pathway.

Genomic insights into versatile lifestyle of three new bacterial candidate phyla.

Metagenomic explorations of the Earth's biosphere enable the discovery of previously unknown bacterial lineages of phylogenetic and ecological significance. Here, we retrieved 11 metagenomic-assembled genomes (MAGs) affiliated to three new monophyletic bacterial lineages from the seawater of the Yap Trench. Phylogenomic analysis revealed that each lineage is a new bacterial candidate phylum, subsequently named Candidatus Qinglongiota, Candidatus Heilongiota, and Candidatus Canglongiota. Metabolic reconstruction of genomes from the three phyla suggested that they adopt a versatile lifestyle, with the potential to utilize various types of sugars, proteins, and/or short-chain fatty acids through anaerobic pathways. This was further confirmed by a global distribution map of the three phyla, indicating a preference for oxygen-limited or particle-attached niches, such as anoxic sedimentary environments. Of note, Candidatus Canglongiota genomes harbor genes for the complete Wood- Ljungdahl pathway and sulfate reduction that are similar to those identified in some sulfate-reducing bacteria. Evolutionary analysis indicated that gene gain and loss events, and horizontal gene transfer (HGT) play important roles in shaping the genomic and metabolic features of the three new phyla. This study presents the genomic insight into the ecology, metabolism, and evolution of three new phyla, which broadens the phylum-level diversity within the domain Bacteria.

Methods and efficacy of social support interventions in preventing suicide: a systematic review and meta-analysis.

QUESTION: Suicide is a global public and mental health problem. The effectiveness of social support interventions has not been widely demonstrated in the prevention of suicide. We aimed to describe the methods of social support interventions in preventing suicide and examine the efficacy of them. STUDY SELECTION AND ANALYSIS: We searched literature databases and conducted clinical trials. The inclusion criteria for the summary of intervention methods were as follows: (1) studies aimed at preventing suicide through method(s) that directly provide social support; (2) use of one or more method(s) to directly provide social support. The additional inclusion criteria for meta-analysis on the efficacy of these interventions included: (1) suicide, suicide attempt or social support-related outcome was measured; (2) randomised controlled trial design and (3) using social support intervention as the main/only method. FINDINGS: In total, 22 656 records and 185 clinical trials were identified. We reviewed 77 studies in terms of intervention methods, settings, support providers and support recipients. There was a total of 18 799 person-years among the ten studies measuring suicide. The number of suicides was significantly reduced in the intervention group (risk ratio (RR)=0.48, 95% CI 0.27 to 0.85). In 14 studies with a total of 14 469 person-years, there was no significant reduction of suicide attempts in the overall pooled RR of 0.88 (95% CI 0.73 to 1.07). CONCLUSIONS: Social support interventions were recommended as a suicide prevention strategy for those with elevated suicide risk.

Genomic and transcriptomic dissection of Theionarchaea in marine ecosystem.

Theionarchaea is a recently described archaeal class within the Euryarchaeota. While it is widely distributed in sediment ecosystems, little is known about its metabolic potential and ecological features. Here, we used metagenomics and metatranscriptomics to characterize 12 theionarchaeal metagenome-assembled genomes, which were further divided into two subgroups, from coastal mangrove sediments of China and seawater columns of the Yap Trench. Genomic analysis revealed that apart from the canonical sulfhydrogenase, Theionarchaea harbor genes encoding heliorhodopsin, group 4 [NiFe]-hydrogenase, and flagellin, in which genes for heliorhodopsin and group 4 [NiFe]-hydrogenase were transcribed in mangrove sediment. Further, the theionarchaeal substrate spectrum may be broader than previously reported as revealed by metagenomics and metatranscriptomics, and the potential carbon substrates include detrital proteins, hemicellulose, ethanol, and CO2. The genes for organic substrate metabolism (mainly detrital protein and amino acid metabolism genes) have relatively higher transcripts in the top sediment layers in mangrove wetlands. In addition, co-occurrence analysis suggested that the degradation of these organic compounds by Theionarchaea might be processed in syntrophy with fermenters (e.g., Chloroflexi) and methanogens. Collectively, these observations expand the current knowledge of the metabolic potential of Theionarchaea, and shed light on the metabolic strategies and roles of these archaea in the marine ecosystems.

Molting Alters the Microbiome, Immune Response, and Digestive Enzyme Activity in Mud Crab (Scylla paramamosain).

Molting is a crucial lifelong process in the growth, development, and reproduction of crustaceans. In mud crab (Scylla paramamosain), new exoskeleton, gills, and appendages are formed after a molting, which contributes to a 40 to 90% increase in body weight. However, little is currently known about the associations between molting and the dynamic changes of microbiota and physiological characteristics in mud crabs. In this study, the effects of molting on changes of the microbiome, immune response, and digestive enzyme activities in mud crabs were investigated. The results showed dynamic changes in the abundances and community compositions of crab-associated microbiota harboring the gills, subcuticular epidermis, hepatopancreas, midgut, and hemolymph during molting. Renewed microbiota was observed in the gills and midgut of crabs at the postmolt stages, which seems to be related to the formation of a new exoskeleton after the molting. A significant positive correlation between the expression of two antimicrobial peptide (AMP) genes (SpALF5 and SpCrustin) and the relative abundance of two predominant microorganisms (Halomonas and Shewanella) in hemolymph was observed in the whole molt cycle, suggesting that AMPs play a role in modulating hemolymph microbiota. Furthermore, digestive enzymes might play a vital role in the changes of microbiota harboring the hepatopancreas and midgut, which provide suitable conditions for restoring and reconstructing host-microbiome homeostasis during molting. In conclusion, this study confirms that molting affects host-associated microbiota and further sheds light on the effects on the immune response and the digestive systems as well. IMPORTANCE Molting is crucial for crustaceans. In mud crab, its exoskeleton is renewed periodically during molting, and this process is an ideal model to study the effects of host development on its microbiota. Here, multiple approaches were used to investigate the changes in microbial taxa, immune response, and digestive enzyme activity with respect to molting in mud crab. The results found that a renewed microbiota was generated in the gills and midgut of crab after a molt. A significant positive correlation between changes in the relative abundances of microbes (such as Halomonas and Shewanella) and the expression of AMP genes (SpALF5 and SpCrustin) was observed in the hemolymph of crabs during the whole molt cycle, suggesting the modulation of hemolymph microbes by AMPs. Furthermore, the digestive enzymes were found to participate in the regulation of microbiota in hepatopancreas and midgut, consequently providing a suitable condition for the restoration and reconstruction of host-microbiome homeostasis during the molting. This study confirms that molting affects the microbial communities and concomitantly influences the immune and digestive systems in mud crabs. This is also the first time the homeostasis of the host and microbiome, and the associations between molting and physiological characteristics in crustaceans, have been revealed.

Comparative genomic analysis reveals metabolic flexibility of Woesearchaeota.

The archaeal phylum Woesearchaeota, within the DPANN superphylum, includes phylogenetically diverse microorganisms that inhabit various environments. Their biology is poorly understood due to the lack of cultured isolates. Here, we analyze datasets of Woesearchaeota 16S rRNA gene sequences and metagenome-assembled genomes to infer global distribution patterns, ecological preferences and metabolic capabilities. Phylogenomic analyses indicate that the phylum can be classified into ten subgroups, termed A-J. While a symbiotic lifestyle is predicted for most, some members of subgroup J might be host-independent. The genomes of several Woesearchaeota, including subgroup J, encode putative [FeFe] hydrogenases (known to be important for fermentation in other organisms), suggesting that these archaea might be anaerobic fermentative heterotrophs.

Responses of sediment resistome, virulence factors and potential pathogens to decades of antibiotics pollution in a shrimp aquafarm.

Aquaculture ecosystem has become a hotspot of antibiotics resistance genes (ARGs) dissemination, owing to the abuse of prophylactic antibiotics. However, it is still unclear how and to what extent ARGs respond to the increasing antibiotic pollution, a trend as expected and as has occurred. Herein, a significant sediment antibiotic pollution gradient was detected along a drainage ditch after decades of shrimp aquaculture. The increasing antibiotic pollution evidently promoted the diversities and tailored the community structures of ARGs, mobile genetic elements (MGEs), virulence factors and pathogens. The profiles of ARGs and MGEs were directly altered by the concentrations of terramycin and sulphadimidine. By contrast, virulence factors were primarily affected by nutrient variables in sediment. The pathogens potentially hosted diverse virulence factors and ARGs. More than half of the detected ARGs subtypes non-linearly responded to increasing antibiotic pollution, as supported by significant tipping points. However, we screened seven antibiotic concentration discriminatory ARGs that could serve as independent variable for quantitatively diagnosing total antibiotic concentration. Co-occurrence analysis depicted that notorious aquaculture pathogens of Vibrio harveyi and V. parahaemolyticus potentially hosted ARGs that confer resistance to multiple antibiotics, while priority pathogens for humankind, e.g., Helicobacter pylori and Staphylococcus aureus, could have harbored redundant virulence factors. Collectively, the significant tipping points and antibiotic concentration-discriminatory ARGs may translate into warning index and diagnostic approach for diagnosing antibiotic pollution. Our findings provided novel insights into the interplay among ARGs, MGEs, pathogens, virulence factors and geochemical variables under the scenario of increasing antibiotic pollution.

Expanded diversity of Asgard archaea and their relationships with eukaryotes.

Asgard is a recently discovered superphylum of archaea that appears to include the closest archaeal relatives of eukaryotes1-5. Debate continues as to whether the archaeal ancestor of eukaryotes belongs within the Asgard superphylum or whether this ancestor is a sister group to all other archaea (that is, a two-domain versus a three-domain tree of life)6-8. Here we present a comparative analysis of 162 complete or nearly complete genomes of Asgard archaea, including 75 metagenome-assembled genomes that-to our knowledge-have not previously been reported. Our results substantially expand the phylogenetic diversity of Asgard and lead us to propose six additional phyla that include a deep branch that we have provisionally named Wukongarchaeota. Our phylogenomic analysis does not resolve unequivocally the evolutionary relationship between eukaryotes and Asgard archaea, but instead-depending on the choice of species and conserved genes used to build the phylogeny-supports either the origin of eukaryotes from within Asgard (as a sister group to the expanded Heimdallarchaeota-Wukongarchaeota branch) or a deeper branch for the eukaryote ancestor within archaea. Our comprehensive protein domain analysis using the 162 Asgard genomes results in a major expansion of the set of eukaryotic signature proteins. The Asgard eukaryotic signature proteins show variable phyletic distributions and domain architectures, which is suggestive of dynamic evolution through horizontal gene transfer, gene loss, gene duplication and domain shuffling. The phylogenomics of the Asgard archaea points to the accumulation of the components of the mobile archaeal 'eukaryome' in the archaeal ancestor of eukaryotes (within or outside Asgard) through extensive horizontal gene transfer.

CD36 as a Molecular Target of Functional DNA Aptamer NAFLD01 Selected against NAFLD Cells.

The aim of this study was to identify the target of nonalcoholic fatty liver disease (NAFLD) cell-specific aptamer NAFLD01 and investigate its effect on lipid metabolism in vitro. A distinct membrane protein of NAFLD cells pulled down by NAFLD01 was analyzed by mass spectrometry to determine target candidates, and affinity of NAFLD01 to target-protein-silent NAFLD cells was detected to validate it. Knockdown of CD36 abolished the binding of NAFLD01, and its binding affinity was associated with membrane-bound CD36. NAFLD01 affinity for NAFLD cells was proportional to the CD36 expression level. Moreover, compared to random sequences, NAFLD01 showed better recognition for both mouse and human tissue sections of NAFLD. Importantly, NAFLD01 could ameliorate liver fat deposition through interaction with CD36 in vitro. Therefore, aptamer NAFLD01 could act as an effective and safe targeted drug for NAFLD. NAFLD01 is the first reported CD36-specific aptamer. This aptamer can improve hepatocyte steatosis via specifically binding to CD36. This study provides a molecular tool to investigate the mechanism of CD36 in NAFLD.

Galangin Improved Non-Alcoholic Fatty Liver Disease in Mice by Promoting Autophagy.

BACKGROUND: Previous studies have shown that curcumin derivatives can improve the fatty degeneration of liver tissue that occurs in nonalcoholic fatty liver disease (NAFLD). However, the specific mechanism for that improvement remains unclear. We examined whether the curcumin derivative galangin could reduce the fatty degeneration of liver tissue in mice with NAFLD by inducing autophagy, from the perspective of both prevention and treatment. METHODS: C57BL/6J mice were randomly assigned to a prevention group (given galangin and a HFD simultaneously) or a treatment group (given galangin after being fed an HFD). The prevention group was treated with galangin (100 mg/kg/d) or an equal volume of normal saline (NS) while being fed an HFD. Some mice were treated with an autophagy inhibitor (3-methyladenine, 3-MA; 30 mg/kg/biwk, i.p.) while being fed an HFD and galangin. HepG2 cells were cultured in DMEM medium containing both free fatty acids and galangin. RESULTS: Galangin was found to reduce the fatty degeneration of liver tissue induced by eating an HFD at both the prevention and treatment levels, and that effect might be related to an enhancement of hepatocyte autophagy. Inhibition of autophagy by 3-MA blocked the protective effect of galangin on hepatic steatosis. At the cellular level, galangin reduced lipid accumulation and enhanced the level of hepatocyte autophagy. CONCLUSION: In vitro and in vivo studies showed that galangin cannot only improve pre-existing hepatic steatosis but also prevent the development of stenosis by promoting hepatocyte autophagy.

Alternative strategies of nutrient acquisition and energy conservation map to the biogeography of marine ammonia-oxidizing archaea.

Ammonia-oxidizing archaea (AOA) are among the most abundant and ubiquitous microorganisms in the ocean, exerting primary control on nitrification and nitrogen oxides emission. Although united by a common physiology of chemoautotrophic growth on ammonia, a corresponding high genomic and habitat variability suggests tremendous adaptive capacity. Here, we compared 44 diverse AOA genomes, 37 from species cultivated from samples collected across diverse geographic locations and seven assembled from metagenomic sequences from the mesopelagic to hadopelagic zones of the deep ocean. Comparative analysis identified seven major marine AOA genotypic groups having gene content correlated with their distinctive biogeographies. Phosphorus and ammonia availabilities as well as hydrostatic pressure were identified as selective forces driving marine AOA genotypic and gene content variability in different oceanic regions. Notably, AOA methylphosphonate biosynthetic genes span diverse oceanic provinces, reinforcing their importance for methane production in the ocean. Together, our combined comparative physiological, genomic, and metagenomic analyses provide a comprehensive view of the biogeography of globally abundant AOA and their adaptive radiation into a vast range of marine and terrestrial habitats.

Identification and molecular characterization of peroxiredoxin 6 from noble scallop Chlamys nobilis revealing its potent immune response and antioxidant property.

The 1-cyseine peroxiredoxin (Prx6) is an importantly antioxidant enzyme that protects cells from oxidative damage caused by excessive production of reactive oxygen species (ROS). In this study, we described the molecular characteristics of the noble scallop Chlamys nobilis peroxiredoxin 6 (designed as CnPrx6), immune responses and DNA protection activity of the recombinant protein. The complete ORF (696 bp) of CnPrx6 encoded a polypeptide (25.5 kDa) of 231 amino acids, harboring a conserved peroxidase catalytic center (41PVCTTE46) and the catalytic triads putatively involved in peroxidase and phospholipase A2 activities. The deduced amino acid sequence of CnPrx6 shared a relatively high amino acid sequence similarity (more than 50%). The qRT-PCR revealed that the CnPrx6 mRNA was constitutively expressed in all examined tissues, with the highest expression observed in adductor. Upon immunological challenge with Vibrio parahaemolyticus, lipopolysaccharides (LPS) and polyinosinic-polycytidylic acid (Poly I:C), the expression level of CnPrx6 mRNA was significantly up-regulated (P < 0.05). Furthermore, there was a significant difference (P < 0.05) in the expression level of CnPrx6 between golden and brown scallops. The purified recombinant CnPrx6 protein protected the supercoiled plasmid DNA from metal-catalyzed ROS damage. Taken together, these results indicated that the CnPrx6 may play an important role in modulating immune responses and minimizing DNA damage in noble scallop Chlamys nobilis.

A novel carotenoids-producing marine bacterium from noble scallop Chlamys nobilis and antioxidant activities of its carotenoid compositions.

Marine bacteria produce many bioactive compounds, including carotenoids. However, the quality of bacterium carotenoids is relatively unknown. Therefore, in this study, a novel carotenoids-producing bacterium Brevundimonas scallop Zheng & Liu was isolated from Chlamys nobilis. The genome of the isolate was analyzed, carotenoid compounds were screened using HPLC-MS and the carotenoid production in B. scallop was monitored. The results revealed that the genome of B. scallop contained a carotenoid synthesis gene cluster, which involved in astaxanthin and hydroxy-astaxanthin biosynthesis. The 2,2'-dihydroxy-astaxanthin was the major carotenoid produced by B. scallop. The optimum culture condition for the highest carotenoids production (1303.62 ± 61.06 µg/g dry cells) for B. scallop was at temperature and salinity of 20 °C and 3% salt, respectively, in 10 g/L glucose as carbon source. The results showed the B. scallop is a new carotenoids resource in marine bivalve, which has an excellent antioxidative activity and potential industrial use.

Comparison of Gut Microbiota Between Golden and Brown Noble Scallop Chlamys nobilis and Its Association With Carotenoids.

Many marine bivalves are regarded as healthy foods due to their high carotenoid content. Only plants and microorganisms have natural carotenoids biosynthesis ability, hence, animals such as bivalves must obtain carotenoids from their diets. Due to the filter-feeding behavior of bivalves, they have high diversity of gut microbes. However, the relationship between gut microbes and carotenoids has not been explored in mollusks. In the present study, the interaction between gut microbes and carotenoids in two polymorphic noble scallop Chlamys nobilis, golden scallops (designated GG) and brown scallops (designated BW), were studied. The gut of GG and BW showed statistically different bacteria communities. Results from 16S rRNA gene sequencing and qPCR analysis revealed that the gut of GG had significantly higher relative abundance of carotenoids-producing bacteria Brevundimonas, compared with BW. Moreover, HPLC-MS analysis showed that isolate Brevundimonas could produce astaxanthin. The current findings are very useful as they could form the basis for future studies in determining the relationship between gut microbiota and carotenoids absorption in bivalves.

Source of hemolymph microbiota and their roles in the immune system of mud crab.

Special innate immune mechanisms against pathogens are developed in marine invertebrates such as mud crab, which is also an economically important aquaculture species in many coastal countries. Hemolymph is a critical site in host immune response, but its source of microorganisms is less known. In this study, we provided a detailed investigation of the microorganisms inhabiting various body sites of healthy mud crabs, including hemolymph, midgut, gill, subcuticular epidermis and hepatopancreas. By using fluorescence microscopy and high-throughput sequencing of the bacterial 16S rRNA genes, various abundances and kinds of microorganisms were observed in the healthy mud crabs, of which some are potential pathogens to mud crab and human. The SourceTracker analysis and oral injection experiment confirm the hypothesis that hemolymph microorganisms are derived from the digestive systems of invertebrates with open circulatory systems, indicating that these microorganisms play vital roles in crab immune response. Moreover, physiological differences (gut length), behavioral characteristics (foraging behavior), diet preferences (herbivory), and/or sex hormones (testosterone) possibly determine the unique features of the crab-associated microbiota for both sexes. These findings also contribute to the development of appropriate microbial immunoenhancers, which has potential applications for improving quality and yield during crab aquaculture.

Metagenomics Reveals Microbial Diversity and Metabolic Potentials of Seawater and Surface Sediment From a Hadal Biosphere at the Yap Trench.

Hadal biosphere represents the deepest part of the ocean with water depth >6,000 m. Accumulating evidence suggests the existence of unique microbial communities dominated by heterotrophic processes in this environment. However, investigations of the microbial diversity and their metabolic potentials are limited because of technical constraints for sample collection. Here, we provide a detailed metagenomic analysis of three seawater samples at water depths 5,000-6,000 m below sea level (mbsl) and three surface sediment samples at water depths 4,435-6,578 mbsl at the Yap Trench of the western Pacific. Distinct microbial community compositions were observed with the dominance of Gammaproteobacteria in seawater and Thaumarchaeota in surface sediment. Comparative analysis of the genes involved in carbon, nitrogen and sulfur metabolisms revealed that heterotrophic processes (i.e., degradation of carbohydrates, hydrocarbons, and aromatics) are the most common microbial metabolisms in the seawater, while chemolithoautotrophic metabolisms such as ammonia oxidation with the HP/HB cycle for CO2 fixation probably dominated the surface sediment communities of the Yap Trench. Furthermore, abundant genes involved in stress response and metal resistance were both detected in the seawater and sediments, thus the enrichment of metal resistance genes is further hypothesized to be characteristic of the hadal microbial communities. Overall, this study sheds light on the metabolic versatility of microorganisms in the Yap Trench, their roles in carbon, nitrogen, and sulfur biogeochemical cycles, and how they have adapted to this unique hadal environment.

C-type lectin B (SpCTL-B) regulates the expression of antimicrobial peptides and promotes phagocytosis in mud crab Scylla paramamosain.

As pattern recognition receptors, C-type lectins (CTLs) play important roles in immune system of crustaceans through identifying and binding to the conservative pathogen-associated molecular patterns (PAMPs) on pathogen surfaces. In this study, a new CTL, SpCTL-B, was identified from the hemocytes of mud crab Scylla paramamosain. The full-length of SpCTL-B cDNA was 1278 bp with an open reading frame (ORF) of 348 bp. The predicted SpCTL-B protein contains a single carbohydrate-recognition domain (CRD). SpCTL-B transcripts were distributed in all examined tissues with the highest levels in hepatopancreas. After challenged with Vibrio parahaemolyticus, LPS, polyI:C and white spot syndrome virus (WSSV), the mRNA levels of SpCTL-B in hemocytes and hepatopancreas were up-regulated. The recombinant SpCTL-B (rSpCTL-B) purified by Ni-affinity chromatography showed stronger binding activities with Staphylococcus aureus, ß-hemolytic Streptococcus, Escherichia coli, Aeromonas hydrophila, Vibrio alginolyticus than those with V. parahaemolyticus and Saccharomyces cerevisiae. rSpCTL-B exhibited a broad spectrum of microorganism-agglutination activities against Gram-positive bacteria (S. aureus, ß-hemolytic Streptococcus) and Gram-negative bacteria (E. coli, V. parahaemolyticus, A. hydrophila, V. alginolyticus) in a Ca2+-dependent manner. The agglutination activities of rSpCTL-B could be inhibited by D-mannose and LPS, but not by d-fructose and galactose. The antimicrobial assay showed that rSpCTL-B exhibited the growth inhibition against all examined gram-positive bacteria and gram-negative bacteria. When SpCTL-B was silenced by RNAi, the bacterial clearance ability in mud crab was decreased and the transcript levels of five antimicrobial peptides (AMPs) (SpCrustin, SpHistin, SpALF4 (anti-lipopolysaccharide factor), SpALF5 and SpALF6) were significantly decreased in hemocytes. In our study, knockdown of SpCTL-B could down-regulate the expression of SpSTAT at mRNA transcriptional level and protein translational level in mud crab. Meantime, the phagocytosis rate and the expression of three phagocytosis related genes were declined after RNAi of SpCTL-B in hemocytes in mud crab. Collectively, our results suggest that SpCTL-B might play its roles as a pattern recognition receptor (PRR) in immune response towards pathogens infection through influencing the expression of AMPs and the phagocytosis of hemocytes in mud crab S. paramamosain.