Identification, Expression and Evolutional Analysis of Two cyp19-like Genes in Amphioxus.

The mechanism of sex determination and differentiation in animals remains a central focus of reproductive and developmental biology research, and the regulation of sex differentiation in amphioxus remains poorly understood. Cytochrome P450 Family 19 Subfamily A member 1 (CYP19A1) is a crucial sex differentiation gene that catalyzes the conversion of androgens into estrogens. In this study, we identified two aromatase-like genes in amphioxus: cyp19-like1 and cyp19-like2. The cyp19-like1 is more primitive and may represent the ancestral form of cyp19 in zebrafish and other vertebrates, while the cyp19-like2 is likely the result of gene duplication within amphioxus. To gain further insights into the expression level of these two aromatase-like, we examined their expression in different tissues and during different stages of gonad development. While the expression level of the two genes differs in tissues, both are highly expressed in the gonad primordium and are primarily localized to microsomal membrane systems. However, as development proceeds, their expression level decreases significantly. This study enhances our understanding of sex differentiation mechanisms in amphioxus and provides valuable insights into the formation and evolution of sex determination mechanisms in vertebrates.

Bmp8a deletion leads to obesity through regulation of lipid metabolism and adipocyte differentiation.

The role of bone morphogenetic proteins (BMPs) in regulating adipose has recently become a field of interest. However, the underlying mechanism of this effect has not been elucidated. Here we show that the anti-fat effect of Bmp8a is mediated by promoting fatty acid oxidation and inhibiting adipocyte differentiation. Knocking out the bmp8a gene in zebrafish results in weight gain, fatty liver, and increased fat production. The bmp8a-/- zebrafish exhibits decreased phosphorylation levels of AMPK and ACC in the liver and adipose tissues, indicating reduced fatty acid oxidation. Also, Bmp8a inhibits the differentiation of 3T3-L1 preadipocytes into mature adipocytes by activating the Smad2/3 signaling pathway, in which Smad2/3 binds to the central adipogenic factor PPARγ promoter to inhibit its transcription. In addition, lentivirus-mediated overexpression of Bmp8a in 3T3-L1 cells significantly increases NOD-like receptor, TNF, and NF-κB signaling pathways. Furthermore, NF-κB interacts with PPARγ, blocking PPARγ's activation of its target gene Fabp4, thereby inhibiting adipocyte differentiation. These data bring a signal bridge between immune regulation and adipocyte differentiation. Collectively, our findings indicate that Bmp8a plays a critical role in regulating lipid metabolism and adipogenesis, potentially providing a therapeutic approach for obesity and its comorbidities.

The evolution and immunomodulatory role of Zc3h12 proteins in zebrafish (Danio rerio).

Zc3h12 family is an important RNA-binding protein family regulating mRNA of inflammatory cytokines in mammals. However, there are few studies on their post-transcriptional level regulation of inflammatory cytokines in fish. Here, we investigated the evolution of zebrafish Zc3h12 family and explored their immunomodulatory role. Phylogenetic and syntenic analysis indicated the number of zc3h12 family members had increased ranging from a single member in invertebrates to a single copy of four members in mammals. As the most evolutionarily diverse group of vertebrates, the number of zc3h12 family members was more complex and diverse in the teleost, each member experienced different fates and followed different rules in multiple rounds of whole-genome duplication events. Thereinto, zebrafish contained three zc3h12 genes, among which zc3h12aa and zc3h12ab were duplicated from the same gene. Zebrafish Zc3h12 family could recognize the 3'-UTR regions of inflammatory cytokines through binding to the specific RNA secondary structure and negatively regulate their expression. Deletion of either Zc3h12 domains or mutation of the key amino acid in RNAase domain attenuated their modulatory effect, suggesting both domain and RNAase activity are important to the immunomodulatory role. These results elucidated the evolution of Zc3h12 family and uncovered Zc3h12-mediated post-transcriptional regulation of cytokines in zebrafish.

BMP8B Activates Both SMAD2/3 and NF-κB Signals to Inhibit the Differentiation of 3T3-L1 Preadipocytes into Mature Adipocytes.

Bone morphogenetic protein 8B (BMP8B) has been found to regulate the thermogenesis of brown adipose tissue (BAT) and the browning process of white adipose tissue (WAT). However, there is no available information regarding the role of BMP8B in the process of adipocyte differentiation. Here, we showed that BMP8B down-regulates transcriptional regulators PPARγ and C/EBPα, thereby impeding the differentiation of 3T3-L1 preadipocytes into fully mature adipocytes. BMP8B increased the phosphorylation levels of SMAD2/3, and TP0427736 HCl (SMAD2/3 inhibitor) significantly reduced the ability of BMP8B to inhibit adipocyte differentiation, suggesting that BMP8B repressed adipocyte differentiation through the SMAD2/3 pathway. Moreover, the knockdown of BMP I receptor ALK4 significantly reduced the inhibitory effect of BMP8B on adipogenesis, indicating that BMP8B triggers SMAD2/3 signaling to suppress adipogenesis via ALK4. In addition, BMP8B activated the NF-κB signal, which has been demonstrated to impede PPARγ expression. Collectively, our data demonstrated that BMP8B activates both SMAD2/3 and NF-κB signals to inhibit adipocyte differentiation. We provide previously unidentified insight into BMP8B-mediated adipogenesis.

Cd248a and Cd248b in zebrafish participate in innate immune responses.

CD248, also known as endosialin or tumor endothelial marker 1, is a type I single transmembrane glycoprotein. CD248 has been demonstrated to be upregulated in cancers, tumors and many fibrotic diseases in human and mice, such as liver damage, pulmonary fibrosis, renal fibrosis, arthritis and tumor neovascularization. However, no definite CD248 orthologs in fish have been documented so far. In this study, we report the identification of cd248a and cd248b in the zebrafish. Both the phylogenetic analysis and the conserved synteny strongly suggested that zebrafish cd248a and cd248b are orthologs of the human CD248. Both cd248a and cd248b exhibited similar and dynamic expression pattern in early development, both genes had weak maternal expression, the zygotic transcripts were first seen in anterior somites and head mesenchyme, then shifted to eyes and head mesenchyme, later expanded to branchial arches, and gradually declined with development. The expression profiles of cd248a and cd248b were upregulated upon LPS (Lipopolysaccharide) challenge. Both Cd248a protein and Cd248b protein were localized on the cell membrane and cytoplasm, and overexpression of cd248a and cd248b induced the expression of pro-inflammatory cytokines, in vitro and in vivo. Moreover, deficiency of cd248a or cd248b both downregulated the expression of pro-inflammatory cytokines and upregulated anti-inflammatory cytokine. Additionally, loss of cd248a or cd248b both downregulated the expression of pro-inflammatory cytokines after LPS treatment. Taken together, these results indicated that cd248a and cd248b in zebrafish were involved in immune response and would provide further information to understand functions of Cd248 protein in innate immunity of fish.

Bacterial-agglutinating and opsonic activities of RIPK1 in zebrafish.

Many studies have demonstrated that receptor interacting protein kinase-1 acts as a crucial mediator in the regulation of immune response, but evidence remains lacking for its direct interaction with bacteria. In this study, we found that challenge with lipopolysaccharide and lipoteichoic acid resulted in a significantly increased transcriptional expression of receptor interacting protein kinase-1 in zebrafish, suggesting the receptor interacting protein kinase-1 is implicated in anti-infectious responses. In accordance, we found that recombinant receptor interacting protein kinase-1 was not only able to bind to Gram-negative and -positive bacteria via interaction with lipopolysaccharide and lipoteichoic acid, but also agglutinate both Gram-negative and -positive bacteria in a Ca2+-dependent manner.

Evolutionary and Expression Analysis of MOV10 and MOV10L1 Reveals Their Origin, Duplication and Divergence.

MOV10 and MOV10L1 both encode ATP-dependent RNA helicases. In mammals, MOV10 and MOV10L1 participate in various kinds of biological contexts, such as defense of RNA virus invasion, neuron system, germ cell and early development. However, mov10 and mov10l1 in zebrafish are obscure and the evolutionary relationships of mov10 among different species remain unclear. In this study, we found MOV10 and MOV10L1 had some variations despite they possessed the conserved feature of RNA helicase, however, they may originate from a single ancestor although they shared limited homology. A single MOV10L1 gene existed among all species, while MOV10 gene experienced lineage-specific intra-chromosomal gene duplication in several species. Interestingly, the mov10 gene expanded to three in zebrafish, which originating from a duplication by whole genome specific duplication of teleost lineage followed by a specific intra-chromosome tandem duplication. The mov10 and mov10l1 showed distinct expression profiles in early stages, however, in adult zebrafish, three mov10 genes exhibited similar diverse expression patterns in almost all tissues. We also demonstrated mov10 genes were upregulated upon virus challenge, highlighting they had redundant conserved roles in virus infection. These results provide valuable data for the evolution of MOV10 and MOV10L1 and they are important to the further functional exploration.

Amphioxus ribosomal proteins RPS15, RPS18, RPS19 and RPS30-precursor act as immune effectors via killing or agglutinating bacteria.

Previous studies show that some ribosomal proteins perform immune effector functions via killing bacteria directly. However, it remains largely unknown about other effector functions of ribosomal proteins during a bacterial infection. In this study, we expressed and purified four ribosomal proteins of the amphioxus Branchiostoma japonicum, termed rBjRPS15, rBjRPS18, rBjRPS19 and rBjRPS30-precursor (rBjRPS30P). They all exhibited bactericidal activity against Gram-positive Staphylococcus aureus, and with the exception of rBjRPS19 and rBjRPS30P, were capable of killing Gram-negative Escherichia coli. Importantly, rBjRPS15, rBjRPS19 and rBjRPS30P were able to agglutinate S. aureus in the presence of Mg2+, but none of them could agglutinate E. coli even in the presence of Mg2+ or Ca2+. Moreover, the S. aureus agglutination was achieved by the binding of these three proteins to the peptidoglycan component of the bacterial cell wall. This is the first report showing that some ribosomal proteins possess bacterial agglutinating activity, and these data provide a new angle to the roles of ribosomal proteins in immune defense.

Zebrafish Ism1 is a novel antiviral factor that positively regulates antiviral immune responses.

Isthmin1 (Ism1), first identified as a secreted protein in Xenopus embryos in 2002, has been shown to perform multiple biological functions, but little is known currently regarding its role in immunity. Here we show that the expression of ism1 is inducible by challenge with Grass carp reovirus (GCRV) in zebrafish, suggesting involvement of Ism1 in antiviral response. We then demonstrate that recombinant Ism1 (rIsm1) reduces the cytopathic effect in the cells infected by GCRV, promotes the expression of type I IFN gene and IFN-inducible antiviral protein Mxa gene, and reduces the virus quantity in virus-infected cells and host. We also show that rIsm1 promotes the expression of tbk1, irf3 and irf7, suggesting it promotes the expression of type I IFN gene and Mxa gene via induction of Tbk1-Irf3-Ifn pathway. These data together indicate that Ism1 is a new immune-relevant factor functioning in antiviral immune response, and provides a target for controlling viral infection.

Bmp8a is an essential positive regulator of antiviral immunity in zebrafish.

Bone morphogenetic protein (BMP) is a kind of classical multi-functional growth factor that plays a vital role in the formation and maintenance of bone, cartilage, muscle, blood vessels, and the regulation of adipogenesis and thermogenesis. However, understanding of the role of BMPs in antiviral immunity is still limited. Here we demonstrate that Bmp8a is a newly-identified positive regulator for antiviral immune responses. The bmp8a-/- zebrafish, when infected with viruses, show reduced antiviral immunity and increased viral load and mortality. We also show for the first time that Bmp8a interacts with Alk6a, which promotes the phosphorylation of Tbk1 and Irf3 through p38 MAPK pathway, and induces the production of type I interferons (IFNs) in response to viral infection. Our study uncovers a previously unrecognized role of Bmp8a in regulation of antiviral immune responses and provides a target for controlling viral infection.

Identification of ribosomal protein L30 as an uncharacterized antimicrobial protein.

Several ribosomal proteins have been shown to adopt for an antimicrobial function as antimicrobial proteins (AMPs). However, information as such is rather limited and their mode of action remains ill-defined. Here we demonstrated that amphioxus RPL30, BjRPL30, was a previously uncharacterized AMP, which was not only capable of binding Gram-negative and Gram-positive bacteria via interaction with LPS, LTA and PGN but also capable of killing the bacteria. We also showed that the residues positioned at 2-46 formed the core region for the antimicrobial activity of BjRPL30. Notably, both the hydrophobic ratio and net charge as well as 3D structures of the residues corresponding to BjRPL302-27 and BjRPL3023-46 from both eukaryotic and prokaryotic RPL30 proteins were closely similar to those of BjRPL302-27 and BjRPL3023-46, suggesting the antibacterial activity of RPL30 was highly conserved. This was further corroborated by the fact that the synthesized counterparts human RPL5-30 and RPL26-49 also had antibacterial activity. We show that the recombinant protein BjRPL30 executes antimicrobial function in vitro by a kind of membranolytic action including interaction with bacterial membrane through LPS, LTA and PGN as well as induction of membrane depolarization. Finally, we found that neither BjRPL30 nor its truncated form BjRPL302-27 and BjRPL3023-46 had hemolytic activity towards human red blood cells, making them promising lead molecules for the design of novel AMPs against bacteria. Altogether, these indicated that RPL30 is a member of AMP which has ancient origin and is highly conserve throughout evolution.

A novel hepatic lectin of zebrafish Danio rerio is involved in innate immune defense.

ASGPR (asialoglycoprotein receptor, also known as hepatic lectin) was the first identified animal lectin, which participated in a variety of physiological processes. Yet its detailed immune functions are not well studied in lower vertebrates. After reporting a zebrafish hepatic lectin (Zhl), we identified a novel hepatic lectin (zebrafish hepatic lectin-like, Zhl-l) in zebrafish. The zhl-l was mainly expressed in liver in a tissue specific manner. And challenge with LPS/LTA induced a significant change of zhl-l expression. What's more, recombinant C-type lectin domain (rCTLD) of Zhl-l had the activity of agglutinating and binding to both Gram-negative and Gram-positive bacteria. It promoted the phagocytosis of bacteria by carp macrophages. Moreover, rCTLD could bind to insoluble lipopolysaccharide (LPS), lipoteichoic acid (LTA) and peptidoglycan (PGN) independent of Ca2+, which was inhibited by galactose. Interestingly, Zhl-l was located in the membrane, and its overexpression could upregulate the production of pre-inflammatory cytokines. Taken together, these results indicated that Zhl-l played a role in immune defense, and would provide further information to understand functions of C-type lectin family and the innate immunity in vertebrates.

Invasive pulmonary aspergillosis secondary to microwave ablation: a multicenter retrospective study.

PURPOSE: Invasive pulmonary aspergillosis (IPA) is a life-threatening complication of microwave ablation (MWA) during the treatment of primary or metastatic lung tumors. The purpose of this study was to investigate the clinical, radiological and demographic characteristics and treatment responses of patients with IPA after MWA. MATERIALS AND METHODS: From January 2011 to January 2016, all patients who were treated by MWA of their lung tumors from six health institutions were enrolled in this study. Patients with IPA secondary to MWA were identified and retrospectively evaluated for predisposing factors, clinical treatment, and outcome. RESULTS: The incidence of IPA secondary to lung MWA was 1.44% (23/1596). Of the 23 patients who developed IPA, six died as a consequence, resulting in a high mortality rate of 26.1%. Using computed tomography (CT), pulmonary cavitation was the most common finding and occurred in 87.0% (20/23) of the patients. Sudden massive hemoptysis was responsible for one-third of the deaths (2/6). Most patients (22/23) received voriconazole as an initial treatment, and six patients with huge cavities underwent intracavitary lavage. Finally, 17 patients (73.9%) achieved treatment success. CONCLUSIONS: Lung MWA may be an additional host risk factor for IPA, particularly in elderly patients with underlying diseases and in patients who have recently undergone chemotherapy. Early and accurate diagnosis of IPA after MWA is critical for patient prognosis. Voriconazole should be given as the first-line treatment as early as possible. Bronchial artery embolization or intracavitary lavage may be required in some patients.

Protective effect of hydrogen-rich water on liver function of colorectal cancer patients treated with mFOLFOX6 chemotherapy.

The present study was conducted to investigate the protective effect of hydrogen-rich water on the liver function of colorectal cancer (CRC) patients treated with mFOLFOX6 chemotherapy. A controlled, randomized, single-blind clinical trial was designed. A total of 152 patients with CRC were recruited by the Department of Oncology of Taishan Hospital (Taian, China) between June 2010 and February 2016, among whom 146 met the inclusion criteria. Subsequently, 144 patients were randomized into the treatment (n=80) and placebo (n=64) groups. At the end of the study, 76 patients in the hydrogen treatment group and 60 patients in the placebo group were included in the final analysis. The changes in liver function after the chemotherapy, such as altered levels of alanine aminotransferase (ALT), aspartate transaminase (AST), alkaline phosphatase, indirect bilirubin (IBIL) and direct bilirubin, were observed. The damaging effects of the mFOLFOX6 chemotherapy on liver function were mainly represented by increased ALT, AST and IBIL levels. The hydrogen-rich water group exhibited no significant differences in liver function before and after treatment, whereas the placebo group exhibited significantly elevated levels of ALT, AST and IBIL. Thus, hydrogen-rich water appeared to alleviate the mFOLFOX6-related liver injury.

Identification of neuroglobin as a novel player in anti-bacterial responses in amphioxus.

Theoretical considerations support various functions of neuroglobin (Ngb), but further studies are required for full characterization of these functions. In this study, we identified the presence of a single Ngb gene, BjNgb, in the amphioxus Branchiostoma japonicum. BjNgb was expressed in various tissues including the notochord, gonads (ovary and testis) and gill, and up-regulated significantly in response to the challenge with LPS and LTA, suggesting involvement in immune response of amphioxus against bacterial infection. In accord, we demonstrated for the first time that recombinant BjNgb (rBjNgb) not only interacted with the Gram-positive and negative bacteria as well as their conserved surface components LPS and LTA, but also enhanced the phagocytosis of bacteria by macrophages. Collectively, these data suggest that BjNgb is a novel player in amphioxus, via functioning as a pattern recognition molecule and an opsonin.

Identification and functional characterization of a novel member of low-density lipoprotein receptor-related protein (LRP)-like family in amphioxus.

Low-density lipoprotein receptor-related protein (LRP) is a group of important endocytic receptors contributing to binding ligands and maintaining internal environment. In this study, we identified a soluble LRP-like molecule in the amphioxus B. japonicum, BjLRP, with an uncharacterized domain structure combination of LY-EGF-CRD-EGF-CRD. It was mainly expressed in the gill, muscle, notochord and testis, and was significantly up-regulated following the challenge with bacteria. Recombinant BjLRP was capable of interacting with both Gram-negative and positive bacteria as well as PAMPs including lipopolysaccharide (LPS), lipoteichoic acid (LTA) and peptidoglycan (PGN). Interestingly, recombinant LY peptide was also able to bind to the Gram-negative and positive bacteria as well as the PAMPs LPS, LTA and PGN. By contrast, none of recombinant EGF1, EGF2, CRD1 and CRD2 had affinity to the bacteria and the PAMPs. In addition, BjLRPΔLY had no affinity to the PAMPs, although BjLRPΔLY showed slight affinity to the bacteria. These suggest that the interaction of BjLRP with the bacteria and PAMPs was primarily attributable to the LY domain. It is clear that BjLRP is a novel pattern recognition protein capable of identifying and interacting with invading bacteria in amphioxus.

Demonstration of a Functional Kisspeptin/Kisspeptin Receptor System in Amphioxus With Implications for Origin of Neuroendocrine Regulation.

Amphioxus belongs to the Cephalochordata, which is the most basal subphylum of the chordates. Despite many studies on the endocrine system of amphioxus, key information about its regulation remains ambiguous. Here we clearly demonstrate the presence of a functional kisspeptin/kisspeptin receptor (Kiss-Kissr) system, which is involved in the regulation of reproduction in amphioxus. Evolutionary analyses revealed large expansion of Kiss and Kissr (gpr54) genes in amphioxus, and they might represent the ancestral type of the Kiss/gpr54 genes in chordates. Amphioxus Kiss was obviously expression at the cerebral vesicle and the Hatschek pit, whereas amphioxus gpr54 messenger RNA (mRNA) was abundantly present in nerve cord, ovary, and testes. Amphioxus GPR54-Like1 (GPR54L-1) was shown to be located on the cell membrane. The synthetic amphioxus Kiss-like (KissL) peptides were capable of activating the amphioxus GPR54L-1 with different potencies, hinting the interaction between Kiss and GPR54. Moreover, the expression of amphioxus gpr54 mRNA was significantly decreased during low or high temperature extremes. Importantly, the injection of amphioxus KissL could cause an elevation of zebrafish blood luteinizing hormone level and induce the expression of amphioxus gpb5, a gene encoding the ancestral type of vertebrate pituitary glycoprotein hormones. Also, the expression levels of BjkissL-2 or Bjgpr54L-1 were downregulated after spermiation or spawning. Collectively, the amphioxus Kiss-Kissr system has a correlation with the regulation of reproduction. Our studies provide insights into the functional roles and evolutionary history of the Kiss-Kissr system, as well as the origin of the vertebrate neuroendocrine axis for controlling reproduction.

Functional characterization of avidins in amphioxus Branchiostoma japonicum: Evidence for a dual role in biotin-binding and immune response.

Avidin is well known for its high affinity to biotin and has been found in many egg-laying vertebrate species. However, little is known about avidin in invertebrate species to date. Here we clearly showed the presence of two avidin genes, Bjavidin1 and Bjavidin2, in the amphioxus Branchiostoma japonicum, the first ones in non-vertebrate animals. We also showed that the expression of both Bjavidin1 and Bjavidin2 were inducible by progesterone, LTA and LPS. Moreover, we demonstrated for the first time that in addition to biotin-binding, the recombinant proteins rBjAVIDIN1 and rBjAVIDIN2 were not only able to interact with Gram-positive and negative bacteria as well as their conserved surface components LTA and LPS but also to enhance phagocytosis of bacteria by macrophages, suggesting that BjAVIDIN1 and BjAVIDIN2 both function as pattern recognition receptors and opsonins. It is thus clear that avidin may play a dual role in biotin-binding and immune response.

Identification of Ly2 members as antimicrobial peptides from zebrafish Danio rerio.

The emergence of multidrug-resistant (MDR) microbes caused by overuse of antibiotics leads to urgent demands for novel antibiotics exploration. Our recent data showed that Ly2.1-3 (a novel lymphocyte antigen 6 (Ly6) gene cluster) were proteins with cationic nature and rich in cysteine content, that are characteristic of antimicrobial peptides (AMPs) and their expression were all significantly up-regulated after challenge with lipopolysaccharide (LPS). These strongly suggested that Ly2.1-3 are potential AMPs, but firm evidence are lacking. Here, we clearly showed that the recombinant proteins of Ly2.1-3 were capable of killing Gram-negative bacteria Aeromonas hydrophila and Escherichia coli, while they had little bactericidal activity against the Gram-positive bacteria Staphylococcus aureus and Bacillus subtilis We also showed that recombinant proteins Ly2.1-3 (rLy2.1-3) were able to bind to the Gram-negative bacteria A. hydrophila, E. coli and the microbial signature molecule LPS, but not to the Gram-positive bacteria S. aureus and B. subtilis as well as the microbial signature molecule LTA. Moreover, the Scatchard analysis revealed that rLy2.1-3 could specifically bind to LPS. Finally, we found that Ly2.1-3 were not cytotoxic to mammalian cells. All these together indicate that Ly2.1-3 can function as AMPs.

Molecular cloning, expression, purification and characterization of vitellogenin in scallop Patinopecten yessoensis with special emphasis on its antibacterial activity.

Vitellogenin (Vg), the major precursor of the egg-yolk proteins, has been found to play an immune role in fish and protochordate amphioxus, however, no study on the immune function of Vg in invertebrates has ever been studied before. In this study, the complete cDNA of Vg was identified from the scallop Patinopecten yessoensis (termed PyVg). The cDNA contained an open reading frame (ORF) of 6888 bp, encoding a polypeptide of 2295 amino acid protein, which had an N-terminal signal peptide followed by the mature Vg. The mature Vg had the domains Vitellogenin_N, domain of unknown function 1943 (DUF1943) and von Willebrand factor type D domain (VWD) as well as the consensus cleavage site (R-X-R/K-R) and conserved motif (KTIGNAG). Tissue distribution assay revealed that PyVg transcripts were predominantly present in the ovary and hepatopancreas, and its expression profile in ovary well reflected the annual cycle of vitellogenesis. Interestingly, bacterial challenge caused a significant change in PyVg expression, hinting an involvement of PyVg in the acute phase response in P. yessoensis. Consistently, recombinant DUF1943 and VWD domains both could interact with LTA and LPS on bacterial wall, and purified native PyVg displayed a broad-spectrum antibacterial activity against both Gram-negative (Escherichia coli and Vibrio anguillarum) and Gram-positive bacteria (Staphylococcus aureus). Overall, these data indicate that Vg is a pattern recognition molecule with bacterial growth-inhibiting activity in the scallop.