RESUMEN
Takifugu rubripes is a highly valued cultured fish in Asia, while pathogen infections can result in severe diseases and lead to substantial economic losses. Toll-like receptors (TLRs), as pattern recognition receptors, play a crucial role on recognition pathogens and initiation innate immune response. However, the immunological properties of teleost-specific TLR23 remain largely unknown. In this study, we investigated the biological functions of TLR23 (TrTLR23) from T. rubripes, found that TrTLR23 existed in various organs. Following bacterial pathogen challenge, the expression levels of TrTLR23 were significantly increased in immune related organs. TrTLR23 located on the cellular membrane and specifically recognized pathogenic microorganism. Co-immunoprecipitation and antibody blocking analysis revealed that TrTLR23 recruited myeloid differentiation primary response protein (MyD88), thereby mediating the activation of the ERK signaling pathway. Furthermore, in vivo showed that, when TrTLR23 is overexpressed in T. rubripes, bacterial replication in fish tissues is significantly inhibited. Consistently, when TrTLR23 expression in T. rubripes is knocked down, bacterial replication is significantly enhanced. In conclusion, these findings suggested that TrTLR23 played a critical role on mediation TLR23-MyD88-ERK axis against bacterial infection. This study revealed that TLR23 involved in the innate immune mechanism, and provided the foundation for development disease control strategies in teleost.
Asunto(s)
Enfermedades de los Peces , Proteínas de Peces , Inmunidad Innata , Factor 88 de Diferenciación Mieloide , Takifugu , Receptores Toll-Like , Animales , Proteínas de Peces/genética , Proteínas de Peces/inmunología , Takifugu/inmunología , Takifugu/genética , Enfermedades de los Peces/inmunología , Factor 88 de Diferenciación Mieloide/genética , Factor 88 de Diferenciación Mieloide/metabolismo , Factor 88 de Diferenciación Mieloide/inmunología , Inmunidad Innata/genética , Receptores Toll-Like/genética , Receptores Toll-Like/inmunología , Receptores Toll-Like/metabolismo , Sistema de Señalización de MAP Quinasas/inmunología , Regulación de la Expresión Génica/inmunología , Edwardsiella/fisiología , Edwardsiella/inmunología , Vibrio/fisiologíaRESUMEN
Video surveillance is widely used in monitoring environmental pollution, particularly harmful dust. Currently, manual video monitoring remains the predominant method for analyzing potential pollution, which is inefficient and prone to errors. In this paper, we introduce a new unsupervised method based on latent diffusion models. Specifically, we propose a spatio-temporal network structure, which better integrates the spatial and temporal features of videos. Our conditional guidance mechanism samples frames of input videos to guide high-quality generation and obtains frame-level anomaly scores, comparing generated videos with original ones. We also propose an efficient compression strategy to reduce computational costs, allowing the model to perform in a latent space. The superiority of our method was demonstrated by numerical experiments in three public benchmarks and practical application analysis in coal mining over previous SOTA methods with better AUC, of at most over 3%. Our method accurately detects abnormal patterns in multiple challenging environmental monitoring scenarios, illustrating the potential application possibilities in the environmental protection domain and beyond.
RESUMEN
Complement C1q domain containing protein (C1qDC) is a vital recognition molecule and has an important effect on immunity. The C1qDCs exhibit opsonic activity in fish, while the mechanisms of C1qDCs in activation complement still remain unclear. This study explored immunological characteristics of a C1qDC from Japanese flounder (Paralichthys olivaceus) (PoC1qDC). PoC1qDC consists of 296 amino acid residues, possessing a collagen domain and a C1q domain. According to our results, PoC1qDC was expressed in 9 diverse tissue samples and showed up-regulation after bacterial challenge. Recombinant PoC1qDC (rPoC1qDC) activated normal serum bactericidal and hemolytic activities by interaction with Japanese flounder IgM, but not enhanced the complement activity of C3-depeleted serum. rPoC1qDC was significantly bound to various bacterial species and agglutination activity against Edwardsiella piscicida and Streptococcus iniae. Furthermore, rPoC1qDC showed direct interaction with peripheral blood leucocytes while enhancing phagocytic and chemotactic activity. When PoC1qDC was overexpressed in Japanese flounder before E. piscicida infection, bacterial replication was significantly inhibited in fish tissues. Consistently, when PoC1qDC expression in Japanese flounder was knocked down, bacterial replication was significantly enhanced. The above findings first suggested the role of PoC1qDC in teleost in mediating complement activation by interaction with IgM, which can positively influence bacterial infection.
Asunto(s)
Infecciones por Enterobacteriaceae , Enfermedades de los Peces , Lenguado , Animales , Bacterias , Activación de Complemento , Colágeno , Inmunoglobulina M , Proteínas de Peces/química , Edwardsiella tarda/fisiologíaRESUMEN
Interleukin 8 (IL8) is a CXC chemokine that plays a crucial role on promoting inflammatory response and immune regulation. In teleost, IL8 can induce the migration and activation of immune cells. However, the biological functions of IL8 are still unknown in Takifugu rubripes. In this study, we examined the biological characteristics of TrIL8 in T. rubripes. TrIL8 is composed of 98 residues and contained a chemokine CXC domain. We found that the TrIL8 expression was detected in diverse organs and significantly increased by Vibrio harveyi or Edwardsiella tarda challenge. The recombinant TrIL8 (rTrIL8) exhibited significantly the binding capacities to 8 tested bacteria. In addition, rTrIL8 could bind to peripheral blood leukocytes (PBL), and increased the expression of immune gene, resistance to bacterial infection, respiratory burst, acid phosphatase activity, chemotactic activity, and phagocytic activity of PBL. In the presence of rTrIL8, T. rubripes was enhanced the resistance to V. harveyi infection. These results indicated that TrIL8 is a chemokine and involved in the activation of immune cells against bacterial infection in teleost.
Asunto(s)
Infecciones Bacterianas , Takifugu , Animales , Interleucina-8 , Secuencia de Aminoácidos , Proteínas de Peces/química , Leucocitos , Factores Inmunológicos/metabolismo , Quimiocinas/metabolismo , Antibacterianos/metabolismoRESUMEN
In mammals, complement factor I (CFI) is a serine protease in serum and plays a pivotal role in the regulation of complement activation. In the presence of cofactor, CFI cleaves C3b to iC3b, and further degrades iC3b to C3c and C3d. In teleost, the function of CFI is poorly understood. In this study, we examined the immunological property of CFI from Japanese flounder (Paralichthys olivaceus) (PoCFI), a teleost species with important economic value. PoCFI is composed of 597 amino acid residues and possesses a trypsin-like serine protease (Tryp) domain. We found that PoCFI expressions occurred in nine different tissues and were upregulated by bacterial challenge. Recombinant PoCFI-Tryp (rPoCFI-Tryp) inhibited complement activation and degraded C3b in serum. rPoCFI-Tryp exhibited apparent binding capacities to a board-spectrum of bacteria and inhibited bacterial growth. These results provide the first evidence to indicate that CFI in teleost negatively regulates complement activation via degradation C3b, and probably plays a role in host immune defense against bacterial infection.
Asunto(s)
Activación de Complemento , Factor I de Complemento/inmunología , Enfermedades de los Peces/inmunología , Lenguado/microbiología , Serina Endopeptidasas , Animales , Antibacterianos/inmunología , Bacterias , Factor I de Complemento/genética , Factor I de Complemento/metabolismo , Enfermedades de los Peces/microbiología , Lenguado/genética , Lenguado/inmunología , Regulación de la Expresión Génica , Unión ProteicaRESUMEN
In this study, we examined the function of a Japanese flounder (Paralichthys olivaceus) microRNA (miRNA), pol-miR-363-3p. We found that pol-miR-363-3p targets an ubiquitin-specific protease (USP), USP32. USP is a family of deubiquitinating enzymes essential to the functioning of the ubiquitin proteasome system. In mammals, USP32 is known to be associated with cancer and immunity. In fish, the function of USP32 is unknown. We found that flounder USP32 (PoUSP32) expression was detected in the major tissues of flounder, particularly intestine. In vitro and in vivo studies showed that pol-miR-363-3p directly regulated PoUSP32 in a negative manner by interaction with the 3'UTR of PoUSP32. Overexpression of pol-miR-363-3p or interference with PoUSP32 expression in flounder cells significantly blocked Streptococcus iniae infection. Consistently, in vivo knockdown of pol-miR-363-3p or overexpression of PoUSP32 enhanced dissemination of S. iniae in flounder tissues, whereas in vivo knockdown of PoUSP32 inhibited S. iniae dissemination. In addition, pol-miR-363-3p knockdown also significantly promoted the tissue dissemination of the viral pathogen megalocytivirus, which, as well as S. iniae, regulated pol-miR-363-3p expression. Together these results revealed an important role of pol-miR-363-3p in flounder immune defense against bacterial and viral infection.
Asunto(s)
Enfermedades de los Peces/inmunología , Peces Planos/inmunología , Inmunidad Innata/genética , MicroARNs/inmunología , Ubiquitina Tiolesterasa/genética , Animales , Infecciones por Virus ADN/inmunología , Infecciones por Virus ADN/veterinaria , Enfermedades de los Peces/genética , Proteínas de Peces/genética , Proteínas de Peces/inmunología , Peces Planos/genética , Iridoviridae/fisiología , MicroARNs/genética , Infecciones Estreptocócicas/inmunología , Infecciones Estreptocócicas/veterinaria , Streptococcus iniae/fisiología , Ubiquitina Tiolesterasa/inmunologíaRESUMEN
BACKGROUND: Suboptimal asthma control during pregnancy may impact perinatal outcomes. U.S. guidelines recommend questionnaires to assess asthma control including the Asthma Control Test (ACT). It is unknown in a research setting to what extent recall differs by the time between symptom occurrence and the administration of the questionnaire. METHODS: Between 2009-2014, 196 pregnant asthmatic women were recruited by the Organization of Teratology Information Specialists (OTIS) MotherToBaby Pregnancy Studies. Participants were administered the ACT at enrollment, gestational weeks 20 and 32, and shortly after delivery. The same women were also administered the ACT retrospectively at approximately 6 months postpartum. RESULTS: The Pearson correlation coefficients between the in-pregnancy and retrospective continuous ACT scores for the 1st, 2nd and 3rd trimesters were: 0.67 (95% CI: 0.58, 0.74), 0.61 (0.52, 0.70) and 0.65 (0.56, 0.72), respectively. When dichotomized into well-controlled asthma (ACT score ≥ 20) versus otherwise, the chi-square test for all three trimesters resulted in p values <0.0001. Cohen's Kappa statistics for the same dichotomized scores were 0.51, 0.45 and 0.40 for each trimester respectively. There was no evidence that adverse outcome of pregnancy (recall bias) influenced postpartum responses. CONCLUSIONS: The retrospectively recalled ACT score obtained postpartum was substantially different compared to in-pregnancy administration of the same questionnaire which could reflect test-retest variability as well as attenuation of recall. Documentation of the magnitude and direction of these differences could be useful in interpretation of the impact of asthma control when the ACT is used in retrospective case-control studies for pregnancy outcomes.
Asunto(s)
Asma/fisiopatología , Recolección de Datos/métodos , Recolección de Datos/normas , Complicaciones del Embarazo/fisiopatología , Teléfono , Adulto , Asma/terapia , Femenino , Humanos , Recuerdo Mental , Embarazo , Complicaciones del Embarazo/terapia , Estudios Retrospectivos , Factores SocioeconómicosRESUMEN
CD59 is a complement regulatory protein that inhibits the formation of membrane attack complex of complement. In this study, we examined the expression and activity of tongue sole (Cynoglossus semilaevis) CD59 (CsCD59). CsCD59 possesses the conserved structural features of CD59 and shares 33%-46% sequence identities with other fish CD59. Expression of CsCD59 was high in liver, spleen, and muscle, and was stimulated by infection of bacterial pathogens. Recombinant CsCD59 (rCsCD59) exhibited an apparent inhibition effect on the activation of tongue sole serum complement. ELISA and microscopy detected binding of rCsCD59 to a number of Gram-negative and Gram-positive bacteria. Interaction with rCsCD59 did not affect bacterial viability but significantly enhanced bacterial resistance against the killing effect of fish serum. Together these results indicate that fish CD59 may to some degrees facilitate a general escape of bacteria from complement-mediated immunity.
Asunto(s)
Antígenos CD59/genética , Proteínas Inactivadoras de Complemento/genética , Enfermedades de los Peces/genética , Proteínas de Peces/genética , Peces Planos , Infecciones por Bacterias Gramnegativas/veterinaria , Infecciones por Bacterias Grampositivas/veterinaria , Animales , Secuencia de Bases , Antígenos CD59/metabolismo , Proteínas Inactivadoras de Complemento/metabolismo , Enfermedades de los Peces/inmunología , Enfermedades de los Peces/microbiología , Proteínas de Peces/metabolismo , Regulación de la Expresión Génica , Bacterias Gramnegativas/fisiología , Infecciones por Bacterias Gramnegativas/genética , Infecciones por Bacterias Gramnegativas/inmunología , Infecciones por Bacterias Gramnegativas/microbiología , Bacterias Grampositivas/fisiología , Infecciones por Bacterias Grampositivas/genética , Infecciones por Bacterias Grampositivas/inmunología , Infecciones por Bacterias Grampositivas/microbiología , Alineación de Secuencia/veterinariaRESUMEN
Edwardsiella tarda, a bacterial pathogen to farmed fish as well as humans, possesses the genes of two lysozyme inhibitors, ivy and mliC (ivy(Et) and mliC(Et)). We recently studied IvyEt and found it to be implicated in E. tarda virulence. In the present study, we characterized MliC(Et) in comparison with Ivy(Et) in a turbot model. MliC(Et) contains the FWSKG motif and two cysteines (C33 and C98) that are highly conserved in subgroup 1 MliCs but are of unknown functional importance. To examine the essentialness of these conserved structural features, recombinant MliC(Et) (rMliC) and its mutants bearing C33S and W79A (of the FWSKG motif) substitutions were prepared. Subsequent analysis showed that rMliC (i) inhibited lysozyme-induced lysis of a Gram-positive bacterium, (ii) reduced serum-facilitated lysozyme killing of E. tarda, and (iii) when introduced into turbot, promoted bacterial dissemination in fish tissues. The C33S mutation had no influence on the activity of rMliC, while the W79A mutation slightly but significantly enhanced the activity of rMliC. Knockout strains of either mliC(Et) or ivy(Et) were severely attenuated for the ability of tissue invasion, host lethality, serum survival, and intracellular replication. The lost virulence of the mliC transformant (TXΔmliC) was restored by complementation with an introduced mliC(Et) gene. Compared to the Δivy(Et) or ΔmliC(Et) single-knockout strains, the ΔmliC(Et) Δivy(Et) double-knockout strain was significantly impaired in most of the virulence features. Together, these results provide the first evidence that the conserved cysteine is functionally dispensable to a subgroup 1 MliC and that as a virulence factor, MliC(Et) most likely works in a concerted and parallel manner with Ivy.
Asunto(s)
Proteínas Bacterianas/genética , Edwardsiella tarda/patogenicidad , Infecciones por Enterobacteriaceae/veterinaria , Peces Planos/microbiología , Monocitos/microbiología , Muramidasa/antagonistas & inhibidores , Animales , Secuencia de Bases , Infecciones por Enterobacteriaceae/inmunología , Infecciones por Enterobacteriaceae/patología , Enfermedades de los Peces/microbiología , Regulación Bacteriana de la Expresión Génica , Técnicas de Inactivación de Genes , Riñón/citología , Proteínas Recombinantes/genética , Proteínas Recombinantes/farmacología , Análisis de Secuencia de ADN , Factores de Virulencia/genéticaRESUMEN
Megalocytivirus, a DNA virus belonging to the Iridoviridae family, is a severe pathogen to a wide range of marine and freshwater fish. In this study, using turbot (Scophthalmus maximus) as a host model, we examined the immunoprotective property of one megalocytivirus gene, ORF75, in the form of DNA vaccine (named pORF75). Immunofluorescence microscopy and RT-PCR analysis showed that P444, the protein encoded by ORF75, was naturally produced in the tissues of turbot during megalocytivirus infection, and that the vaccine gene in pORF75 was expressed in fish cells transfected with pORF75 and in the tissues of turbot immunized with pORF75. Following vaccination of turbot with pORF75, a high level of survival (73%) was observed against a lethal megalocytivirus challenge. Consistently, viral replication in the vaccinated fish was significantly inhibited. Immune response analysis showed that pORF75-vaccinated fish (i) exhibited upregulated expression of the genes involved in innate and adaptive immunity, (ii) possessed specific memory immune cells that showed significant response to secondary antigen stimulation, and (iii) produced specific serum antibodies which, when co-introduced into turbot with megalocytivirus, blocked viral replication. Furthermore, whole-genome transcriptome analysis revealed that ORF75 knockdown altered the transcription of 43 viral genes. Taken together, these results indicate that ORF75 encoded a highly protective immunogen that is also a global transcription regulator of megalocytivirus.
Asunto(s)
Iridoviridae , Proteínas Virales/inmunología , Vacunas Virales , Animales , Anticuerpos Antivirales/sangre , Infecciones por Virus ADN/genética , Infecciones por Virus ADN/prevención & control , Enfermedades de los Peces/genética , Enfermedades de los Peces/prevención & control , Peces Planos/sangre , Peces Planos/genética , Peces Planos/virología , Expresión Génica , Bazo/metabolismo , Transcripción Genética , Proteínas Virales/genéticaRESUMEN
Chemokines are a large, diverse group of small cytokines that can be classified into several families, including the CC chemokine family, which plays a pivotal role in host defense by inducing leukocyte chemotaxis under physiological and inflammatory conditions. Here we studied 9 CC chemokines from half-smooth tongue sole (Cynoglossus semilaevis). Phylogenetic analysis divided these chemokines into four groups. The tissue specific expression patterns of the 9 chemokines under normal physiological conditions varied much, with most chemokines highly expressed in immune organs, while some other chemokines showing high expression levels in non-immune organs. In addition, the 9 chemokines exhibited similar or distinctly different expression profiles in response to the challenge of virus and intracellular and extracellular bacterial pathogens. These results indicate that in tongue sole, CC chemokines may be involved in different immune responses as homeostatic or inflammatory chemokines.
Asunto(s)
Quimiocinas CC/genética , Enfermedades de los Peces/genética , Enfermedades de los Peces/inmunología , Proteínas de Peces/genética , Peces Planos , Transcriptoma , Animales , Quimiocinas CC/metabolismo , Infecciones por Virus ADN/genética , Infecciones por Virus ADN/inmunología , Infecciones por Virus ADN/microbiología , Infecciones por Virus ADN/veterinaria , ADN Complementario/genética , ADN Complementario/metabolismo , Edwardsiella tarda/fisiología , Infecciones por Enterobacteriaceae/genética , Infecciones por Enterobacteriaceae/inmunología , Infecciones por Enterobacteriaceae/microbiología , Infecciones por Enterobacteriaceae/veterinaria , Enfermedades de los Peces/microbiología , Enfermedades de los Peces/virología , Proteínas de Peces/metabolismo , Iridoviridae/fisiología , Filogenia , ARN Mensajero/genética , ARN Mensajero/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa/veterinaria , Análisis de Secuencia de ADN/veterinaria , Vibrio/fisiología , Vibriosis/genética , Vibriosis/inmunología , Vibriosis/microbiología , Vibriosis/veterinariaRESUMEN
Edwardsiella tarda is a Gram-negative bacterium with a broad host range that includes a wide variety of farmed fish as well as humans. E. tarda has long been known to be able to survive in host serum, but the relevant mechanism is unclear. In this study, we investigated the fundamental question, i.e. whether E. tarda activated serum complement or not. We found that (i) when incubated with flounder serum, E. tarda exhibited a high survival rate (87.6%), which was slightly but significantly reduced in the presence of Mg(2+); (ii) E. tarda-incubated serum possessed strong hemolytic activity and bactericidal activity, (iii) compared to the serum incubated with a complement-sensitive laboratory Escherichia coli strain, E. tarda-incubated serum exhibited much less chemotactic activity, (iv) in contrast to the serum incubated with live E. tarda, the serum incubated with heat-inactivated E. tarda exhibited no apparent hemolytic capacity. Taken together, these results indicate for the first time that E. tarda circumvents serum attack by preventing, to a large extent, complement activation via the alternative pathway, and that heat-labile surface structures likely play an essential role in the complement evasion of E. tarda.
Asunto(s)
Activación de Complemento , Edwardsiella tarda/inmunología , Infecciones por Enterobacteriaceae/veterinaria , Enfermedades de los Peces/inmunología , Peces Planos , Animales , Ensayo de Actividad Hemolítica de Complemento/veterinaria , Infecciones por Enterobacteriaceae/inmunología , Infecciones por Enterobacteriaceae/microbiología , Enfermedades de los Peces/microbiologíaRESUMEN
Chemokines are a large family of chemotactic cytokines. Based on the arrangement of the first two cysteine residues, chemokines are divided into four groups, one of which is the CC chemokine group. In this study, we characterized a CC chemokine, CsCCL21, from half-smooth tongue sole (Cynoglossus semilaevis), and analyzed its activity. CsCCL21 contains two conserved N-terminal cysteine residues in a NCCL motif and is phylogenetically related to the CCL19/21/25 subgroup of CC chemokines. CsCCL21 was constitutively expressed in nine tissues and significantly upregulated by bacterial and viral infection. The recombinant CsCCL21 (rCsCCL21) induced migration of peripheral blood leukocytes. When the two conserved cysteine residues in the NCCL motif were mutated, the chemotactic activity of rCsCCL21 was abolished. rCsCCL21 enhanced the resistance of tongue sole against bacterial infection, but the mutant protein with NCCL mutation lacked this antibacterial effect. Taken together, these results suggest that CsCCL21 is a functional CC chemokine with the ability to recruit leukocytes and is involved in antibacterial immunity in a manner that requires the conserved NCCL motif.
Asunto(s)
Quimiocina CCL21/genética , Enfermedades de los Peces/genética , Enfermedades de los Peces/inmunología , Proteínas de Peces/genética , Peces Planos , Inmunidad Innata , Secuencia de Aminoácidos , Animales , Quimiocina CCL21/química , Quimiocina CCL21/metabolismo , Infecciones por Virus ADN/genética , Infecciones por Virus ADN/inmunología , Infecciones por Virus ADN/veterinaria , Edwardsiella tarda/fisiología , Infecciones por Enterobacteriaceae/genética , Infecciones por Enterobacteriaceae/inmunología , Infecciones por Enterobacteriaceae/veterinaria , Escherichia coli/fisiología , Proteínas de Peces/química , Proteínas de Peces/metabolismo , Iridoviridae/fisiología , Filogenia , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Alineación de Secuencia/veterinaria , Regulación hacia Arriba , Vibrio/fisiología , Vibriosis/genética , Vibriosis/inmunología , Vibriosis/veterinariaRESUMEN
Hepcidin is an antimicrobial peptide and a regulator of iron homeostasis. In turbot (Scophthalmus maximus), two types of hepcidins have been identified, which share approximately 50% sequence identity. In this study, we examined the antimicrobial potentials of the two hepcidins in the form of synthesized peptides, SmHep1P and SmHep2P. We found that SmHep1P and SmHep2P exhibited apparent bactericidal activities against both Gram-positive and Gram-negative bacteria in a dose-dependent manner. The bactericidal effect of SmHep1P was stronger against Gram-positive bacteria, while the bactericidal effect of SmHep2P was stronger against Gram-negative bacteria. Fluorescence and electron microscopy showed that both peptides were able to bind to the target bacterial cells and alter the surface structure of the cells. In vitro studies showed that SmHep1P and SmHep2P reduced bacterial invasion into cultured fish cells. In vivo studies showed that turbot administered with SmHep1P and SmHep2P exhibited significantly enhanced resistance against bacterial and viral infection. In both in vivo and in vitro studies, the antimicrobial activities of SmHep2P were in most cases significantly stronger than those of SmHep1P. Together these results indicate that the two hepcidins of turbot most likely possess antimicrobial properties and play a role in the innate immune defense against bacterial and viral pathogens.
Asunto(s)
Bacterias/efectos de los fármacos , Peces Planos/metabolismo , Regulación de la Expresión Génica/inmunología , Hepcidinas/metabolismo , Hepcidinas/farmacología , Animales , Células Cultivadas , Hepcidinas/genéticaRESUMEN
Pentraxins are a family of evolutionarily conserved proteins that play an important part in innate immunity. C-reactive protein (CRP) is a member of the pentraxin family and in humans is known to be the major acute phase protein. In this work, we report the identification and analysis of a CRP, CsCRP, from half-smooth tongue sole (Cynoglossus semilaevis). CsCRP is composed of 228 amino acid residues and possesses a Pentraxin/CRP domain. Expression of CsCRP occurred in a wide range of tissues and was upregulated by pathogen infection in kidney, spleen, blood, and, in particular, liver. Following bacterial infection, CsCRP level in blood rose rapidly within 12 h and was approximately 3.8 fold of that of the basal level. Purified recombinant CsCRP (rCsCRP) was able to interact with Gram-negative and Gram-positive bacteria including those of pathogenic nature in a dose-dependent manner. When peripheral blood leukocytes (PBL) were infected with bacterial pathogen in the presence of rCsCRP, the respiratory burst and phagocytic capacity of the cells were increased to significant extents. Taken together, these results indicate that CsCRP is an acute phase protein that plays a role in innate immune defense against bacterial infection.
Asunto(s)
Bacterias/inmunología , Proteína C-Reactiva/inmunología , Peces Planos/inmunología , Regulación de la Expresión Génica/inmunología , Inmunidad Innata/inmunología , Leucocitos/inmunología , Análisis de Varianza , Animales , Bacterias/metabolismo , Secuencia de Bases , Proteína C-Reactiva/metabolismo , Clonación Molecular , Peces Planos/metabolismo , Peces Planos/microbiología , Datos de Secuencia Molecular , Fagocitosis/inmunología , Plásmidos/genética , Estructura Terciaria de Proteína , Reacción en Cadena en Tiempo Real de la Polimerasa/veterinaria , Estallido Respiratorio/inmunología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/veterinaria , Análisis de Secuencia de ADN/veterinariaRESUMEN
Thioredoxin (Trx) is a redox protein that plays an important role in anti-oxidative stress and redox signaling. In a recent report, we observed that the Trx of tongue sole (Cynoglossus semilaevis), CsTrx1, is a reductase and an antioxidant with immunostimulatory property. In the current work, we studied the biological effect of a truncated CsTrx1, CsTrx80, which is composed of the N-terminal 80 residues of CsTrx1. We found that recombinant CsTrx80 (rCsTrx80) purified from Escherichia coli exhibited no insulin disulfide reductase activity; however, unlike rCsTrx1, which showed no apparent chemotactic activity, rCsTrx80 was able to bind to the head kidney monocytes of tongue sole and caused cellular migration in a dose-dependent manner. Mutation of the two cysteine residues in the highly conserved CXXC motif had no effect on the chemotactic activity of rCsTrx80. Like rCsTrx1, rCsTrx80 induced the expression of immune relevant genes in the target cells, but the induction patterns differed from that induced by rCsTrx1. In vivo study showed that when tongue sole were administered with rCsTrx80 before being infected with a bacterial pathogen, bacterial dissemination and colonization in fish tissues were significantly reduced compared to control fish. Taken together, these results indicate for the first time that a Trx80 derived from fish Trx possesses chemotactic and immunoregulatory properties which promote host resistance against bacterial infection.
Asunto(s)
Infecciones por Enterobacteriaceae/veterinaria , Enfermedades de los Peces/genética , Enfermedades de los Peces/inmunología , Proteínas de Peces/genética , Peces Planos , Tiorredoxinas/genética , Animales , Quimiocinas/genética , Quimiocinas/metabolismo , Quimiotaxis , Edwardsiella tarda/fisiología , Infecciones por Enterobacteriaceae/genética , Infecciones por Enterobacteriaceae/inmunología , Infecciones por Enterobacteriaceae/microbiología , Escherichia coli/genética , Enfermedades de los Peces/microbiología , Proteínas de Peces/metabolismo , Regulación de la Expresión Génica , Riñón Cefálico/inmunología , Monocitos/inmunología , Reacción en Cadena en Tiempo Real de la Polimerasa , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Tiorredoxinas/metabolismoRESUMEN
Peptidoglycan recognition proteins (PGRPs) are a family of innate immune molecules that recognize bacterial peptidoglycan. PGRPs are highly conserved in invertebrates and vertebrates including fish. However, the biological function of teleost PGRP remains largely uninvestigated. In this study, we identified a PGRP homologue, SoPGLYRP-2, from red drum (Sciaenops ocellatus) and analyzed its activity and potential function. The deduced amino acid sequence of SoPGLYRP-2 is composed of 482 residues and shares 46-94% overall identities with known fish PGRPs. SoPGLYRP-2 contains at the C-terminus a single zinc amidase domain with conserved residues that form the catalytic site. Quantitative RT-PCR analysis detected SoPGLYRP-2 expression in multiple tissues, with the highest expression occurring in liver and the lowest expression occurring in brain. Experimental bacterial infection upregulated SoPGLYRP-2 expression in kidney, spleen, and liver in time-dependent manners. To examine the biological activity of SoPGLYRP-2, purified recombinant proteins representing the intact SoPGLYRP-2 (rSoPGLYRP-2) and the amidase domain (rSoPGLYRP-AD) were prepared from Escherichia coli. Subsequent analysis showed that rSoPGLYRP-2 and rSoPGLYRP-AD (i) exhibited comparable Zn(2+)-dependent peptidoglycan-lytic activity and were able to recognize and bind to live bacterial cells, (ii) possessed bactericidal effect against Gram-positive bacteria and slight bacteriostatic effect against Gram-negative bacteria, (iii) were able to block bacterial infection into host cells. These results indicate that SoPGLYRP-2 is a zinc-dependent amidase and a bactericide that targets preferentially at Gram-positive bacteria, and that SoPGLYRP-2 is likely to play a role in host innate immune defense during bacterial infection.
Asunto(s)
Amidohidrolasas/genética , Amidohidrolasas/inmunología , Bacterias Grampositivas/metabolismo , Infecciones por Bacterias Grampositivas/enzimología , Infecciones por Bacterias Grampositivas/veterinaria , Perciformes/inmunología , Perciformes/microbiología , Amidohidrolasas/química , Secuencia de Aminoácidos , Animales , Péptidos Catiónicos Antimicrobianos/química , Péptidos Catiónicos Antimicrobianos/genética , Péptidos Catiónicos Antimicrobianos/inmunología , Secuencia de Bases , Proteínas Portadoras/genética , Proteínas Portadoras/inmunología , Perfilación de la Expresión Génica , Regulación Enzimológica de la Expresión Génica , Infecciones por Bacterias Grampositivas/inmunología , Datos de Secuencia Molecular , Proteínas Recombinantes/metabolismo , Alineación de Secuencia , ZincRESUMEN
Invasin is an outer membrane protein that is known to mediate entry of enteric bacteria into mammalian cells. In this study, we analyzed the function and immunoprotective potential of the invasin Inv1 from Edwardsiella tarda, a serious fish pathogen that can also infect humans. In silico analysis indicated that Inv1 possesses a conserved N-terminal DUF3442 domain and a C-terminal group 1 bacterial Ig-like domain. Subcellular localization analysis showed that Inv1 is exposed on cell surface and could be recognized by specific antibodies. Mutation of inv1 had no effect on bacterial growth but attenuates overall bacterial virulence and impaired the ability of E. tarda to attach and invade into host cells. Consistent with these observations, antibody blocking of Inv1 inhibited E. tarda infection of host cells. To examine the immunoprotective potential of Inv1, recombinant Inv1 (rInv1) corresponding to the DUF3442 domain was purified and used to vaccinate Japanese flounder (Paralichthys olivaceus). The results showed that rInv1 induced strong protection against lethal-dose challenge of E. tarda. ELISA analysis showed that rInv1-vaccinated fish produced specific serum antibodies that could enhance the serum bactericidal activity against E. tarda. Taken together, these results indicate that Inv1 is a surface-localized virulence factor that is involved in host infection and can induce effective immunoprotection when used as a subunit vaccine.
Asunto(s)
Adhesinas Bacterianas/inmunología , Adhesinas Bacterianas/metabolismo , Vacunas Bacterianas/inmunología , Edwardsiella tarda/metabolismo , Enfermedades de los Peces/prevención & control , Lenguado , Secuencia de Aminoácidos , Animales , Anticuerpos Antibacterianos/sangre , Adhesión Bacteriana/fisiología , Infecciones por Enterobacteriaceae/prevención & control , Infecciones por Enterobacteriaceae/veterinaria , Ensayo de Inmunoadsorción Enzimática , Enfermedades de los Peces/microbiología , Anotación de Secuencia Molecular , Mutación , Transporte de ProteínasRESUMEN
Complement plays an important role in the innate immune system, and it comprises about 35 individual proteins. In mammals, complement is activated via three different pathways, the classical pathway, the alternative pathway, and the lectin pathway. All three activation pathways produce C3-convertase in different forms. C3-convertase cleaves C3 to C3a and C3b and initiates a cascade of cleavage and activation, eventually resulting in the formation of the membrane attack complex. Complement activation results in the generation of activated fragments that are involved in microbial killing, phagocytosis, inflammatory reactions, immune complex clearance, and antibody production. Although the complement system has been studied extensively in mammals, complement is less well understood in teleosts. This review summarizes the current knowledge of the teleost complement components involved in phagocytosis, chemotaxis, and cell lysis. We report the characterized complement components in various teleost species. In addition, we provide a comprehensive compilation of complement regulators, and this information is used to analyze the role of complement regulators in pathogen infection. The influence of complement receptors on the immune responses of teleosts is reviewed. Finally, we propose directions for future study of the molecular evolution, structure, and function of complement components in teleosts. This review provides new insights into the complement system of recognition and defense, and such knowledge is essential for the development of new immune strategies in aquaculture.
Asunto(s)
Complemento C3 , Complejo de Ataque a Membrana del Sistema Complemento , Animales , Complejo Antígeno-Anticuerpo , Activación de Complemento , Complemento C3/metabolismo , Convertasas de Complemento C3-C5/metabolismo , Complemento C3b , Lectinas , Mamíferos , Receptores de Complemento/metabolismoRESUMEN
Edwardsiella tarda is a well-known bacterial pathogen with a broad range of host, including fish, amphibians, and mammals. One eminent virulence feature of E. tarda is its strong ability to resist the killing of host serum complement, but the involving mechanism is unclear. In this report, we identified E. tarda TraT as a key player in both complement resistance and cellular invasion. TraT, a surface-localized protein, bound and recruited complement factor H onto E. tarda, whereby inhibiting complement activation via the alternative pathway. TraT also interacted with host CD46 in a specific complement control protein domain-dependent manner, whereby facilitating the cellular infection and tissue dissemination of E. tarda. Thus, by acting as an anti-complement factor and a cellular infection promoter, TraT makes an important contribution to the complement evasion and systemic infection of E. tarda. These results add insights into the pathogen-host interaction mechanism during E. tarda infection.