RESUMEN

Prohibitins (PHBs) are ubiquitously expressed conserved proteins in eukaryotes that are associated with apoptosis, cancer formation, aging, stress responses and cell proliferation. However, the function of the PHBs in immune regulation has largely not been determined. In the present study, we identified PHB2 in the red swamp crayfish Procambarus clarkii. PHB2 was found to be widely distributed in several tissues, and its expression was significantly upregulated by white spot syndrome virus (WSSV) challenge. PHB2 significantly reduced the amount of WSSV in crayfish and the mortality of WSSV-infected crayfish. Here, we observed that PHB2 promotes the nuclear translocation of STAT by binding to STAT. After blocking PHB2 or STAT with antibodies or interfering with PHB2 or STAT, the expression levels of the antiviral genes ß-thymosin (PcThy-4) and crustin2 (Cru2) decreased. The gene sequence of PHB2 was analyzed and found to contain a nuclear introgression sequence (NIS). After in vivo injection of PHB2 with deletion of NIS (rΔNIS-PHB2), the nuclear translocation of STAT did not change significantly compared to that in the control group. These results suggest that PHB2 promoted the nuclear translocation of STAT through NIS and mediated the expression of antiviral proteins to inhibit WSSV infection.

Asunto(s)

Timosina , Virus del Síndrome de la Mancha Blanca 1 , Animales , Virus del Síndrome de la Mancha Blanca 1/fisiología , Astacoidea , Alimentos Marinos , Antivirales

RESUMEN

Hexavalent chromium (Cr(VI)) is a cytotoxic heavy metal pollutant that adversely affects all life forms. Interestingly, the crustacean Procambarus clarkii exhibits a relatively high tolerance to heavy metals. The underlying mechanisms remain unclear. In this study, we investigated the role of symbiotic bacteria in P. clarkii in alleviating Cr(VI)-induced damage and explored their potential mechanisms of action. Through transcriptomic analysis, we observed that Cr(VI) activated P. clarkii's antimicrobial immune responses and altered the bacterial composition in the hemolymph. After antibiotic treatment to reduce bacterial populations, Cr(VI)-induced intestinal and liver damage worsened, and crayfish exhibited lower levels of GSH/CAT/SOD activity. The Exiguobacterium, the symbiotic bacteria in the hemolymph of P. clarkii, were proved to be primary contributor to Cr(VI) tolerance. Further investigation suggested that it resists Cr(VI) through the activation of the ABC transporter system and the reduction of Cr(VI) via the reductase gene nfsA. To validate the role of Exiguobacterium in Cr(VI) tolerance, crayfish treated with antibiotics then supplemented with Exiguobacterium H6 and recombinant E. coli (with the nfsA gene), reduced Cr(VI)-induced ovarian damage. Overall, this study revealed that the symbiotic bacteria Exiguobacterium can absorb and reduce hexavalent chromium, mitigating Cr(VI)-induced damage in P. clarkii. These findings provide new insights into hexavalent chromium tolerance mechanisms in crustaceans.

Asunto(s)

Astacoidea , Metales Pesados , Animales , Escherichia coli , Hemolinfa , Cromo/toxicidad , Bacterias

RESUMEN

Viruses have evolved various strategies to achieve early infection by initiating transcription of their own early genes via host transcription factors, such as NF-κb, STAT, and AP1. How the host copes with this immune escape has been a topic of interest. Tripartite motif (TRIM) family proteins with RING-type domains have E3 ubiquitin ligase activity and are known as host restriction factors. Trim has been reported to be associated with phagocytosis and is also believed to be involved in the activation of autophagy. Preventing the virus from entering the host cell may be the most economical way for the host to resist virus infection. The role of TRIM in the early stage of virus infection in host cells remains to be further interpreted. In the current study, a crayfish TRIM with a RING-type domain, designated as PcTrim, was significantly upregulated under white spot syndrome virus (WSSV) infection in the red swamp crayfish (Procambarus clarkii). Recombinant PcTrim significantly inhibited WSSV replication in crayfish. RNAi targeting PcTrim or blocking PcTrim with an antibody promoted WSSV replication in crayfish. Pulldown and co-IP assays showed that PcTrim can interact with the virus protein VP26. PcTrim restricts the expression level of dynamin, which is involved in the regulation of phagocytosis, by inhibiting AP1 entry into the nucleus. AP1-RNAi effectively reduced the expression levels of dynamin and inhibited host cell endocytosis of WSSV in vivo. Our study demonstrated that PcTrim might reduce early WSSV infection by binding to VP26 and then inhibiting AP1 activation, resulting in reduced endocytosis of WSSV in crayfish hemocytes. Video Abstract.

Asunto(s)

Astacoidea , Virus del Síndrome de la Mancha Blanca 1 , Anticuerpos , Autofagia , Endocitosis , Fagocitosis , Proteínas de Motivos Tripartitos , Astacoidea/virología , Animales

RESUMEN

The hepatopancreas is an important digestive and immune organ in crustacean. There were low but stable numbers of microbes living in the hemolymph of crustacean, whereas the organs (including hepatopancreas) of crustacean were immersed in the hemolymph. It is very important to study the immune mechanism of the hepatopancreas against bacteria. In this study, a novel CTL (HepCL) with two CRDs, which was mainly expressed in the hepatopancreas, was identified in red swamp crayfish (Procambarus clarkii). HepCL binds to bacteria in vitro and could enhance bacterial clearance in vivo. Compared with the C-terminal CRD of HepCL (HepCL-C), the N-terminal CRD (HepCL-N) showed weaker bacterial binding ability in vitro and stronger bacterial clearance activity in vivo. The expression of some antimicrobial proteins, such as FLP, ALF1 and ALF5, was downregulated under knockdown of HepCL or blocked with Anti-HepCL after challenge with Vibrio in crayfish. These results demonstrated that HepCL might be involved in the antibacterial immune response by regulating the expression of antimicrobial proteins.

Asunto(s)

Crustáceos/inmunología , Crustáceos/metabolismo , Resistencia a la Enfermedad/inmunología , Hepatopáncreas/metabolismo , Interacciones Huésped-Patógeno/inmunología , Inmunidad Innata , Lectinas/metabolismo , Animales , Infecciones Bacterianas/veterinaria , Crustáceos/genética , Crustáceos/microbiología , Resistencia a la Enfermedad/genética , Expresión Génica , Interacciones Huésped-Patógeno/genética , Especificidad de Órganos/genética , Especificidad de Órganos/inmunología , Vibrio/inmunología

RESUMEN

Antimicrobial peptides (AMPs), most of which are small proteins, are necessary for innate immunity against pathogens. Anti-lipopolysaccharide factor (ALF) with a conserved lipopolysaccharide binding domain (LBD) can bind to lipopolysaccharide (LPS) and neutralize LPS activity. The antibacterial mechanism of ALF, especially its role in bacteria, needs to be further investigated. In this study, the antibacterial role of an anti-lipopolysaccharide factor (PcALF5) derived from Procambarus clarkii was analyzed. PcALF5 could inhibit the replication of the microbiota in vitro and enhance the bacterial clearance ability in crayfish in vivo. Far-western blot assay results indicated that PcALF5 bound to two proteins of E. coli (approximately 25 kDa and 15 kDa). Mass spectrometry (MS), far-western blot assay, and pull-down results showed that 30S ribosomal protein S4 (RPS4, 25 kD) interacted with PcALF5. Further studies revealed that another E. coli protein binding to PcALF5 could be the large mechanosensitive channel (MscL), which is reported to participate in the transport of peptides and antibiotics. Additional assays showed that PcALF5 inhibited protein synthesis and promoted the transcription of ribosomal component genes in E. coli. Overall, these results indicate that PcALF5 could transfer into E. coli by binding to MscL and inhibit protein synthesis by interacting with RPS4. This study reveals the mechanism underlying ALF involvement in the antibacterial immune response and provides a new reference for the research on antibacterial drugs.

Asunto(s)

Péptidos Catiónicos Antimicrobianos , Proteínas de Artrópodos , Astacoidea , Proteínas de Escherichia coli , Canales Iónicos , Proteínas Ribosómicas , Animales , Péptidos Catiónicos Antimicrobianos/metabolismo , Proteínas de Artrópodos/metabolismo , Astacoidea/inmunología , Astacoidea/microbiología , Escherichia coli/genética , Escherichia coli/inmunología , Escherichia coli/metabolismo , Proteínas de Escherichia coli/metabolismo , Interacciones Huésped-Patógeno/genética , Interacciones Huésped-Patógeno/inmunología , Canales Iónicos/metabolismo , Lipopolisacáridos/metabolismo , Biosíntesis de Proteínas/inmunología , Proteínas Ribosómicas/metabolismo

RESUMEN

Plasmodium infection in Anopheles is influenced by mosquito-derived factors. We previously showed that a protein in saliva from infected Anopheles, mosquito gamma-interferon-inducible lysosomal thiol reductase (mosGILT), inhibits the ability of sporozoites to traverse cells and readily establish infection of the vertebrate host. To determine whether mosGILT influences Plasmodium within the mosquito, we generated Anopheles gambiae mosquitoes carrying mosaic mutations in the mosGILT gene using CRISPR/CRISPR associated protein 9 (Cas9). Here, we show that female mosaic mosGILT mutant mosquitoes display defects in ovarian development and refractoriness to Plasmodium. Following infection by either Plasmodium berghei or Plasmodium falciparum, mutant mosquitoes have significantly reduced oocyst numbers as a result of increased thioester-containing protein 1 (TEP1)-dependent parasite killing. Expression of vitellogenin (Vg), the major yolk protein that can reduce the parasite-killing efficiency of TEP1, is severely impaired in mutant mosquitoes. MosGILT is a mosquito factor that is essential for ovarian development and indirectly protects both human and rodent Plasmodium species from mosquito immunity.

Asunto(s)

Anopheles/genética , Anopheles/parasitología , Malaria/parasitología , Oxidorreductasas actuantes sobre Donantes de Grupos Sulfuro/genética , Animales , Animales Modificados Genéticamente , Proteína 9 Asociada a CRISPR/genética , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas/genética , Culicidae/genética , Femenino , Masculino , Ratones , Mutación/genética , Plasmodium berghei/patogenicidad , Plasmodium falciparum/patogenicidad , Proteínas de Unión al ARN/genética , Vitelogeninas/genética

RESUMEN

Streptococcus agalactiae (S. agalactiae, GBS) infection has caused significant economic loss in the tilapia aquaculture, which is one of the most important commercial fish worldwide. Among the 10 serotypes of GBS, serotypes Ia, Ib, II and III were epidemic in tilapia while serotype IX has never been found in tilapia before. In this study, 80 strains isolated from moribund tilapia in China were identified as GBS. All the isolates have been classified as serotype III or serotype IX of GBS. Unexpectedly, the serotype IX has never been reported in fish, but it was epidemic in mammals. Antimicrobial resistance results showed that serotype IX but not III was resistant to streptomycin and erythromycin. Artificial infection results showed that both serotypes could cause serious pathological injuries in the infected tissues of tilapia. Furthermore, serotype IX instead of serotype III, mainly infected the brain of tilapia. The results will shed a new light on the epidemic and pathogenicity of GBS, and will pave a new way for the prevention of Streptococcosis in tilapia.

Asunto(s)

Antibacterianos/farmacología , Farmacorresistencia Bacteriana , Enfermedades de los Peces/microbiología , Enfermedades de los Peces/patología , Infecciones Estreptocócicas/veterinaria , Streptococcus agalactiae/efectos de los fármacos , Tilapia/microbiología , Animales , Encéfalo/microbiología , Encéfalo/patología , China , Eritromicina/farmacología , Serogrupo , Infecciones Estreptocócicas/microbiología , Infecciones Estreptocócicas/patología , Streptococcus agalactiae/clasificación , Streptococcus agalactiae/aislamiento & purificación , Streptococcus agalactiae/patogenicidad , Estreptomicina/farmacología

RESUMEN

Antimicrobial peptides (AMPs) play important roles in innate immunity against pathogens and lysozymes are a particularly type of AMP. Lysozymes are hydrolytic enzymes that are characterized by their ability to cleave the beta-(1,4)-glycosidic bond between N-acetylmuramic acid and N-acetylglucosamine in peptidoglycan, which is the major bacterial cell wall polymer. In this work, a lysozyme was identified from Procambarus clarkii and designated PcLys-i3. Quantitative RT-PCR was used to analyze the tissue distribution and expression profiles of PcLys-i3. PcLys-i3 was present in all tested tissues and had high expression levels in gills, stomach and intestine. The expression levels of PcLys-i3 were up-regulated in gills and intestine after challenge with Vibrio parahaemolyticus, Staphylococcus aureus and Aeromonas hydrophila. PcLys-i3 and PcFer proteins can enhance the bacterial elimination in crayfish, whereas the bacterial elimination was weakened when the expression level of PcLys-i3 or PcFer RNAs was suppressed by RNAi. Recombinant PcLys-i3 and PcFer significantly reduced the mortality of crayfish with bacterial infections. Further study found that PcLys-i3 could interact with PcFer in vitro. Finally, the PcLys-i3 and PcFer proteins could bind to bacteria and inhibit bacterial replication. These results suggest that both PcLys-i3 and PcFer play important roles in the antibacterial immunity of red swamp crayfish.

Asunto(s)

Antibacterianos/inmunología , Proteínas de Artrópodos/inmunología , Astacoidea/inmunología , Muramidasa/inmunología , Aeromonas hydrophila/inmunología , Aeromonas hydrophila/fisiología , Animales , Antibacterianos/metabolismo , Proteínas de Artrópodos/genética , Proteínas de Artrópodos/metabolismo , Astacoidea/genética , Astacoidea/microbiología , Perfilación de la Expresión Génica/métodos , Branquias/inmunología , Branquias/metabolismo , Branquias/microbiología , Interacciones Huésped-Patógeno , Mucosa Intestinal/metabolismo , Intestinos/inmunología , Intestinos/microbiología , Muramidasa/genética , Muramidasa/metabolismo , Interferencia de ARN , Staphylococcus aureus/inmunología , Staphylococcus aureus/fisiología , Regulación hacia Arriba , Vibrio/inmunología , Vibrio/fisiología

RESUMEN

Pacific white shrimp (Litopenaeus vannamei) is an important cultural species worldwide. However, Vibrio spp. infections have caused a great economic loss in Pacific white shrimp culture industry. The immune responses of Pacific white shrimp to the Vibrio spp. is not fully characterized. In this study, the transcriptomic profiles of L. vannamei hemocytes were explored by injecting with or without Vibrio parahaemolyticus. Totally, 42,632 high-quality unigenes were obtained from RNAseq data. Comparative genome analysis showed 2258 differentially expressed genes (DEGs) following the Vibrio challenge, including 1017 up-regulated and 1241 down-regulated genes. Eight DEGs were randomly selected for further validation by quantitative real-time RT-PCR (qRT-PCR) and the results showed that are consistent with the RNA-seq data. Due to the lack of predictable adaptive immunity, shrimps rely on an innate immune system to defend themselves against invading microbes by recognizing and clearing them through humoral and cellular immune responses. Here we focused our studies on the humoral immunity, five genes (SR, MNK, CTL3, GILT, and ALFP) were selected from the transcriptomic data, which were significantly up-regulated by V. parahaemolyticus infection. These genes were widely expressed in six different tissues and were up-regulated by both Gram negative bacteria (V. parahaemolyticus) and Gram positive bacteria (Staphylococcus aureus). To further extend our studies, we knock-down those five genes by dsRNA in L. vannamei and analyzed the functions of specific genes against V. parahaemolyticus and S. aureus by bacterial clearance analysis. We found that the ability of L. vannamei was significantly reduced in bacterial clearance when treated with those specific dsRNA. These results indicate that those five genes play essential roles in antibacterial immunity and have its specific functions against different types of pathogens. The obtained data will shed a new light on the immunity of L. vannamei and pave a new way for fighting against the bacterial infection in Pacific white shrimp.

Asunto(s)

Proteínas de Artrópodos/genética , Proteínas de Artrópodos/inmunología , Inmunidad Innata/genética , Penaeidae/genética , Penaeidae/inmunología , Vibrio parahaemolyticus/fisiología , Animales , Femenino , Perfilación de la Expresión Génica , Interacciones Huésped-Patógeno , Masculino , Distribución Aleatoria

RESUMEN

Hemocyanins (HMC): the copper-containing respiratory proteins present in invertebrate hemolymph, which plays many essential roles in the immune system. Currently, little is known about the HMC domains of Procambarus clarkii (P. clarkii) and their function in antimicrobial immune response. In this present study, we comparatively studied the expression pattern of native PcHMC with the three recombinant proteins of variable domains of crayfish hemocyanin (PcHMC-N, N-terminal domain of hemocyanin; PcHMC-T, tyrosinase domain of hemocyanin; PcHMC-C, C-terminal domain of hemocyanin). The results showed that three purified recombinant proteins had a strong binding to various bacteria and lipopolysaccharides that further highly agglutinated. The HMCs recombinant proteins showed strong antibacterial activity against V. parahaemolyticus and S. aureus by bacterial growth inhibition, phenoloxidase (PO) and phagocytosis assays. Specifically, rPcHMC1-T and rPcHMC1-C inhibited both the bacteria efficiently, rPcHMC1-T was highly upregulated the PO activity than the other recombinant proteins. Whereas, recombinant proteins pretreated crayfish hemocytes participated in phagocytosis activity, rPcHMC1-N and rPcHMC1-C proteins had a profound effect than the rPcHMC1-T on S. aureus and V. parahaemolyticus phagocytosis. The crayfish hemocyanin domains clearly exhibited antibacterial and phagocytic activities against both the bacteria, suggesting that its variable domains of hemocyanin have the different function on specific pathogen during the assault of pathogens.

Asunto(s)

Antibacterianos/farmacología , Proteínas de Artrópodos/farmacología , Astacoidea/inmunología , Astacoidea/microbiología , Fenómenos Fisiológicos Bacterianos , Hemocianinas/farmacología , Animales , Antibacterianos/química , Proteínas de Artrópodos/química , Hemocianinas/química , Lipopolisacáridos/farmacología , Peptidoglicano/farmacología , Dominios Proteicos , Ácidos Teicoicos/farmacología

RESUMEN

C-type lectins (CTLs) are characterized by the presence of a C-type carbohydrate recognition domain (CTLD) that by recognizing microbial glycans, is responsible for their roles as pattern recognition receptors in the immune response to bacterial infection. In addition to the CTLD, however, some CTLs display additional domains that can carry out effector functions, such as the collagenous domain of the mannose-binding lectin. While in vertebrates, the mechanisms involved in these effector functions have been characterized in considerable detail, in invertebrates they remain poorly understood. In this study, we identified in the kuruma shrimp (Marsupenaeus japonicus) a structurally novel CTL (MjCC-CL) that in addition to the canonical CTLD, contains a coiled-coil domain (CCD) responsible for the effector functions that are key to the shrimp's antibacterial response mediated by antimicrobial peptides (AMPs). By the use of in vitro and in vivo experimental approaches we elucidated the mechanism by which the recognition of bacterial glycans by the CTLD of MjCC-CL leads to activation of the JAK/STAT pathway via interaction of the CCD with the surface receptor Domeless, and upregulation of AMP expression. Thus, our study of the shrimp MjCC-CL revealed a striking functional difference with vertebrates, in which the JAK/STAT pathway is indirectly activated by cell death and stress signals through cytokines or growth factors. Instead, by cross-linking microbial pathogens with the cell surface receptor Domeless, a lectin directly activates the JAK/STAT pathway, which plays a central role in the shrimp antibacterial immune responses by upregulating expression of selected AMPs.

Asunto(s)

Proteínas de Artrópodos/metabolismo , Crustáceos/metabolismo , Lectinas Tipo C/metabolismo , Transducción de Señal , Animales , Proteínas de Artrópodos/inmunología , Crustáceos/microbiología , ADN Complementario/genética , Quinasas Janus/metabolismo , Lectinas Tipo C/química , Receptores de Reconocimiento de Patrones/metabolismo , Factores de Transcripción STAT/metabolismo , Regulación hacia Arriba

RESUMEN

Lysozymes are widely distributed immune effectors exerting muramidase activity against the peptidoglycan of the bacterial cell wall to trigger cell lysis. However, some invertebrate-type (i-type) lysozymes deficient of muramidase activity still exhibit antimicrobial activity. To date, the mechanism underlying the antimicrobial effect of muramidase-deficient i-type lysozymes remains unclear. Accordingly, this study characterized a novel i-type lysozyme, Splys-i, in the mud crab Scylla paramamosain. Splys-i shared the highest identity with the Litopenaeus vannamei i-type lysozyme (Lvlys-i2, 54% identity) at the amino acid level. Alignment analysis and 3D structure comparison show that Splys-i may be a muramidase-deficient i-type lysozyme because it lacks the two conserved catalytic residues (Glu and Asp) that are necessary for muramidase activity. Splys-i is mainly distributed in the intestine, stomach, gills, hepatopancreas, and hemocytes, and it is upregulated by Vibrio harveyi or Staphylococcus aureus challenge. Recombinant Splys-i protein (rSplys-i) can inhibit the growth of Gram-negative bacteria (V. harveyi, Vibrio alginolyticus, Vibrio parahemolyticus, and Escherichia coli), Gram-positive bacteria (S. aureus, Bacillus subtilis, and Bacillus megaterium), and the fungus Candida albicans to varying degrees. In this study, two binding assays and a bacterial agglutination assay were conducted to elucidate the potential antimicrobial mechanisms of Splys-i. Results demonstrated that rSplys-i could bind to all nine aforementioned microorganisms. It also exhibited a strong binding activity to lipopolysaccharide from E. coli and lipoteichoic acid and peptidoglycan (PGN) from S. aureus but a weak binding activity to PGN from B. subtilis and ß-glucan from fungi. Moreover, rSplys-i could agglutinate these nine types of microorganisms in the presence of Ca2+ at different protein concentrations. These results suggest that the binding activity and its triggered agglutinating activity might be two major mechanisms of action to realize the muramidase-deficient antibacterial activity. In addition, rSplys-i can hydrolyze the peptidoglycan of some Gram-positive bacteria because it exhibits weak isopeptidase activities in salt and protein concentration-dependent manner. This result indicates that such an isopeptidase activity may contribute to the muramidase-deficient antimicrobial activity to a certain degree. In conclusion, Splys-i is upregulated by pathogenic bacteria, and it inhibits bacterial growth by binding and agglutination activities as well as isopeptidase activity, suggesting that Splys-i is involved in immune defense against bacteria through several different mechanisms of action.

Asunto(s)

Antiinfecciosos/metabolismo , Proteínas de Artrópodos/genética , Braquiuros/inmunología , Candidiasis/inmunología , Mucosa Intestinal/metabolismo , Muramidasa/genética , Infecciones Estafilocócicas/inmunología , Staphylococcus aureus/inmunología , Vibriosis/inmunología , Vibrio/inmunología , Aglutinación , Animales , Proteínas de Artrópodos/metabolismo , Liasas de Carbono-Nitrógeno/metabolismo , Procesos de Crecimiento Celular , Clonación Molecular , Inmunidad Innata , Lipopolisacáridos/metabolismo , Muramidasa/metabolismo , Unión Proteica , Proteoglicanos/metabolismo , Alineación de Secuencia

RESUMEN

Streptococcosis due to the bacterium Streptococcus agalactiae (S. agalactiae) has resulted in enormous economic losses in aquaculture worldwide, especially in the tilapia culture industry. Previously, there were limited vaccines that could be employed against streptococcosis in tilapia. This study aimed to develop a vaccine candidate using the glyceraldehyde-phosphate dehydrogenase protein (GapA) of S. agalactiae encoded by the gapA gene. Tilapia were intraperitoneally injected with PBS, PBS + Freund's adjuvant, PBS + Montanide's adjuvant, GapA + Freund's adjuvant, GapA + Montanide's adjuvant, killed S. agalactiae whole cells (WC)+Freund's adjuvant, or killed S. agalactiae whole cells (WC)+ Montanide's adjuvant. They were then challenged with S. agalactiae, and the relative percentage survival (RPS) was monitored 14 days after the challenge. The highest RPSs were observed in the WC groups, with 76.7% in WC + Freund's adjuvant and 74.4% in WC + Montanide's adjuvant groups; these were followed by the GapA groups, with 63.3% in GapA + Freund's adjuvant and 45.6% in GapA + Montanide's adjuvant groups. The RPS of the PBS group was 0%, and those of PBS + Freund's adjuvant and PBS + Montanide's adjuvant groups were 6.7% and 3.3%, respectively. Additionally, the IgM antibody responses elicited in GapA groups and WC groups were significantly higher than those in PBS groups. Furthermore, the expressions of cytokine (IL-1ß and TNF-α) mRNAs in the GapA groups and WC groups were significantly higher than those in the PBS groups. Taken together, these results reveal that the GapA protein is a promising vaccine candidate that could be used to prevent streptococcosis in tilapia.

Asunto(s)

Adhesinas Bacterianas/inmunología , Cíclidos , Enfermedades de los Peces/prevención & control , Infecciones Estreptocócicas/veterinaria , Vacunas Estreptocócicas/inmunología , Streptococcus agalactiae/inmunología , Animales , Clonación Molecular , Escherichia coli/genética , Enfermedades de los Peces/inmunología , Enfermedades de los Peces/microbiología , Inyecciones Intraperitoneales/veterinaria , Organismos Modificados Genéticamente/genética , Distribución Aleatoria , Infecciones Estreptocócicas/inmunología , Infecciones Estreptocócicas/microbiología , Infecciones Estreptocócicas/prevención & control , Vacunas Estreptocócicas/genética , Vacunas de Subunidad/genética , Vacunas de Subunidad/inmunología , Vacunas Sintéticas/genética , Vacunas Sintéticas/inmunología

RESUMEN

Streptococcus agalactiae is a pathogen that causes severe anthropozoonosis within a broad range of hosts from aquatic animals to mammals, including human beings. Here, we describe the draft genome of S. agalactiae HZAUSC001, a low-virulent strain isolated from the gut of a moribund tilapia (Oreochromis niloticus) in China.

RESUMEN

Janus kinase (Jak) and signal transducers and activators of transcription (STAT) signaling pathway is associated in antiviral and antibacterial immune response. Previous studies primarily investigated the function of STATs in mammals. For most invertebrates, only one STAT was found in each species, such as STAT92E was found in Drosophila melanogaster. The studies, which focus on the functional difference between various STATs in the same species of invertebrate, are limited. In the present study, three STATs (HcSTAT1, HcSTAT2 and HcSTAT3) were identified in triangle shell pearl mussel, Hyriopsis cumingii. Phylogenetic analysis showed that HcSTAT1 and HcSTAT3 were clustered with Homo sapiens STAT5, and HcSTAT2 was clustered with Pinctada fucata STAT and Crassostea gigas STAT6. All three STATs could be detected in all tested tissues (hemocytes, hepatopancreas, gill, mantle and foot), and were induced expression when challenged with Staphylococcus aureus or Aeromonas hydrophilia in hemocytes and hepatopancreas. HcSTAT1 regulated the expression of HcDef, HcWAP, HcThe and HcTNF. The expression of HcWAP and HcTNF was down-regulated in HcSTAT2-RNAi mussel. And HcSTAT3 affected the expression of HcTNF. The study is the first report of different functions in antibacterial immune responses between STATs in mollusks.

Asunto(s)

Aeromonas hydrophila/fisiología , Regulación de la Expresión Génica/inmunología , Inmunidad Innata/genética , Factores de Transcripción STAT/metabolismo , Staphylococcus aureus/fisiología , Unionidae/genética , Unionidae/inmunología , Animales , Especificidad de Órganos , Filogenia , Factores de Transcripción STAT/genética , Análisis de Secuencia de ADN , Unionidae/microbiología

RESUMEN

High concentration of ammonia in aquatic system leads to detrimental effects on the health of aquatic animals. However, the mechanism underlying ammonia-induced toxicity is still not clear. To better understand the mechanism of ammonia toxicity effects on fish, juvenile grass carp was employed in the present study. RNA high-throughput sequencing technique was applied to analyze the total RNAs extracted from the liver of fish after 8 h post exposure to the water containing 2 mM NH4HCO3 which experimentally mimicked the high environmental ammonia (HEA). A total of 49,971,114 and 53,826,986 clean reads were obtained in control and 2 mM HEA group, respectively, in which there were 911 differently expressed genes (DEGs) including 563 up-regulated and 348 down-regulated genes. In addition, 10 DEGs were validated by quantitative PCR. These DEGs were involved in several pathways related with oxidative stress or apoptosis. Further analysis on oxidative stress, histopathology and cellular apoptosis in grass carp liver after HEA exposure revealed interesting findings. Increased reactive oxygen species (ROS) content and superoxide dismutase (SOD) activity together with the decreased catalase (CAT) activity were detected, which may be effected by DEGs and related pathways such as FOXO signaling pathway. The histopathology and TUNEL assays results confirmed that apoptosis was induced in liver when fish had suffered HEA. Combined with the results of transcriptomic experiments, c-Myc-Bax-Caspase9 apoptosis pathway could be involved in grass carp liver apoptosis induced by ammonia stress.

Asunto(s)

Amoníaco/toxicidad , Antioxidantes/metabolismo , Apoptosis/efectos de los fármacos , Carpas/metabolismo , Regulación de la Expresión Génica/efectos de los fármacos , Transcriptoma , Contaminantes Químicos del Agua/toxicidad , Amoníaco/metabolismo , Animales , Perfilación de la Expresión Génica/veterinaria , Inactivación Metabólica , Hígado/efectos de los fármacos , Hígado/metabolismo , Transducción de Señal/efectos de los fármacos , Contaminantes Químicos del Agua/metabolismo

RESUMEN

Eukaryotes have occasionally acquired genetic material through horizontal gene transfer (HGT). However, little is known about the evolutionary and functional significance of such acquisitions. Lysozymes are ubiquitous enzymes that degrade bacterial cell walls. Here, we provide evidence that two subclasses of bivalves (Heterodonta and Palaeoheterodonta) acquired a lysozyme gene via HGT, building on earlier findings. Phylogenetic analyses place the bivalve lysozyme genes within the clade of bacteriophage lysozyme genes, indicating that the bivalves acquired the phage-type lysozyme genes from bacteriophages, either directly or through intermediate hosts. These bivalve lysozyme genes underwent dramatic structural changes after their co-option, including intron gain and fusion with other genes. Moreover, evidence suggests that recurrent gene duplication occurred in the bivalve lysozyme genes. Finally, we show the co-opted lysozymes exhibit a capacity for antibacterial action, potentially augmenting the immune function of related bivalves. This represents an intriguing evolutionary strategy in the eukaryote-microbe arms race, in which the genetic materials of bacteriophages are co-opted by eukaryotes, and then used by eukaryotes to combat bacteria, using a shared weapon against a common enemy.

Asunto(s)

Bacteriófagos/enzimología , Bivalvos/genética , Muramidasa/genética , Animales , Antibacterianos/química , Antibacterianos/farmacología , Bacteriófagos/genética , Bivalvos/microbiología , Evolución Molecular , Duplicación de Gen , Transferencia de Gen Horizontal , Muramidasa/química , Muramidasa/farmacología , Filogenia , Conformación Proteica , Proteínas Virales/química , Proteínas Virales/genética , Proteínas Virales/farmacología

RESUMEN

Nervous necrosis virus (NNV), the causative agent of viral nervous necrosis (VNN) disease, has caused mass mortality of cultured marine and freshwater fish worldwide, resulting in enormous economic losses in the aquaculture industry. However, the molecular mechanisms underlying the pathogenicity of NNV are still poorly understood. In this study, the transcriptomic profiles of striped snakehead fish (Channa striatus) cells (SSN-1) infected with red-spotted grouper NNV (RGNNV) were investigated using deep RNA sequencing technique. From 254,955,234 raw reads, a total of 253,338,544 clean reads were obtained and they were assembled into 93,372 unigenes. Differentially expressed genes (DEGs) were identified from RGNNV-infected or mock-infected SSN-1 cells, including 1184 up-regulated and 1456 down-regulated genes at 3 h (h) post of infection (poi), and 1138 up-regulated and 2073 down-regulated genes at 24 h poi, respectively. These DEGs were involved in many pathways related to viral pathogenesis, including retinoic acid-inducible gene I (RIG-I) like receptors pathway, apoptosis pathway, oxidative phosphorylation, PI3K-Akt signaling pathway, and MAPK signaling pathway. Subsequent analysis focusing on the apoptosis pathway showed that the expression of Endonuclease G (EndoG) was up-regulated upon RGNNV infection at both 3 and 24 h poi. Therefore, EndoG gene was cloned and its function was further characterized. The results showed that over-expression of EndoG could also induce cellular apoptosis in SSN-1 cells, indicating that RGNNV infection might induce apoptosis of SSN-1 cells via EndoG-associated mitochondrial pathway. These results will shed a new light on the pathogenesis of NNV.

Asunto(s)

Apoptosis/genética , Enfermedades de los Peces/genética , Perciformes , Infecciones por Virus ARN/veterinaria , Transcriptoma/genética , Animales , Línea Celular , Endodesoxirribonucleasas/genética , Endodesoxirribonucleasas/metabolismo , Enfermedades de los Peces/inmunología , Proteínas de Peces/genética , Proteínas de Peces/metabolismo , Nodaviridae/fisiología , Infecciones por Virus ARN/genética , Infecciones por Virus ARN/inmunología

RESUMEN

In invertebrates, ficolin-like proteins (FLPs) play important roles in innate immunity against pathogens. Previous studies primarily investigated the functions of FLPs in immune recognition, activation, and regulation. However, limited research has examined the functions of FLPs as immune effectors. In this work, a ficolin-like protein was identified in red swam crayfish (Procambarus clarkii) and designated as PcFLP1. Quantitative RT-PCR and western blot were employed to analyze the distribution and expression profiles of PcFLP1 in the tissues of the crayfish. The results indicated that PcFLP1 was present in all tested tissues, including hemocytes, heart, hepatopancreas, gill, stomach, and mid-intestine. The expression level of PcFLP1 was up-regulated in hemocytes, hepatopancreas and mid-intestines of the crayfish challenged with Vibrio parahaemolyticus. Further study demonstrated that PcFLP1 could protect the hepatopancreatic cells of crayfish from V. parahaemolyticus infection. The recombinant PcFLP1 enhanced bacterial elimination in crayfish, whereas the antibacterial action was inhibited after PcFLP1 was knocked down. Furthermore, PcFLP1 could bound to bacteria and inhibited bacterial replication. These results demonstrated that PcFLP1 plays an important role in the anti-Vibrio immunity of red swamp crayfish.

Asunto(s)

Proteínas de Artrópodos/inmunología , Astacoidea/inmunología , Inmunidad Innata/inmunología , Lectinas/inmunología , Secuencia de Aminoácidos , Animales , Proteínas de Artrópodos/genética , Astacoidea/genética , Western Blotting , Técnica del Anticuerpo Fluorescente , Inmunidad Innata/genética , Lectinas/genética , Filogenia , Reacción en Cadena de la Polimerasa , Reacción en Cadena en Tiempo Real de la Polimerasa , Ficolinas

RESUMEN

Impaired phosphatase activity leads to the persistent activation of signal transducers and activators of transcription (Stat). In mammals, Stat family members are often phosphorylated or dephosphorylated by the same enzymes. To date, only one Stat similar to mammalian Stat5a/b has been found in crustaceans and there have been few studies in Stat signal regulation in crustaceans. Here, we report that ß-arrestin1 interacts with TC45 (45-kDa form of T cell protein tyrosine phosphatase) in the nucleus to attenuate Stat signaling by promoting dephosphorylation of Stat. Initially, we showed that Stat translocates into the nucleus to induce antimicrobial peptide (AMP) expression after bacterial infection. ßArr1 enters the nucleus of hemocytes and recruits TC45 to form the ßarr1-TC45-Stat complex, which dephosphorylates Stat efficiently. The interaction of TC45 with Stat decreased and Stat phosphorylation increased in ßarr1-silenced shrimp (Marsupenaeus japonicus) after challenge with Vibrio anguillarum. ßArr1 directly interacts with Stat in nucleus and accelerates Stat dephosphorylation by recruiting TC45 after V. anguillarum challenge. Further study showed that ßarr1 and TC45 also affect AMP expression, which is regulated by Stat. Therefore, ßarr1 and TC45 are involved in the anti-V. anguillarum immune response by regulating Stat activity negatively to decrease AMP expression in shrimp.

Asunto(s)

Péptidos Catiónicos Antimicrobianos/metabolismo , Proteína Tirosina Fosfatasa no Receptora Tipo 2/metabolismo , Factores de Transcripción STAT/metabolismo , beta-Arrestina 1/metabolismo , Animales , Chlorocebus aethiops , Decápodos/metabolismo , Mucosa Intestinal/metabolismo , Fosforilación , Unión Proteica , Dominios Proteicos , Proteína Tirosina Fosfatasa no Receptora Tipo 2/antagonistas & inhibidores , Proteína Tirosina Fosfatasa no Receptora Tipo 2/genética , Interferencia de ARN , ARN Mensajero/metabolismo , ARN Interferente Pequeño/metabolismo , Factores de Transcripción STAT/antagonistas & inhibidores , Factores de Transcripción STAT/genética , Transducción de Señal , Vibrio/fisiología , beta-Arrestina 1/antagonistas & inhibidores , beta-Arrestina 1/genética