RESUMEN
Edwardsiella anguillarum, a highly virulent species within the Edwardsiella genus, causes significant mortality in milkfish farms in Taiwan. This study aimed to investigate the comparison of milkfish susceptibility, a newly identified host species in Taiwanese aquaculture, with other species Nile tilapia (Oreochromis niloticus) and Asian seabass (Lates calcarifer), to E. anguillarum, elucidating its pathogenicity across both seawater and freshwater aquaculture environments. The results showed milkfish exhibited the highest mortality rate of 85% within 48 h of infection, whereas Nile tilapia exhibited a mortality rate of 70% between the second- and tenth-day post challenge, and seabass exhibited a mortality rate of 25% between the second- and sixth-day post challenge. Gross lesions observed in milkfish included splenomegaly and haemorrhage, whereas Nile tilapia exhibited signs of ascites, exophthalmia and brain haemorrhage. Seabass displayed spleen granulomas and haemorrhage at the injection site. Histopathological analysis revealed common features across all three species, including multifocal necrosis, bacterial presence in the necrotic areas, serositis and oedema. Asian seabass also exhibited chronic lesions in the form of splenic granulomas. This study highlights the high susceptibility of milkfish and Nile tilapia to E. anguillarum, emphasizing the urgent need for further investigation into targeted vaccine development for these fish species. These results not only deepen our understanding of the differing levels of pathogenicity among the three species but also offer valuable insights for improving disease prevention and management strategies in aquaculture, including those applied within polyculture systems and for the maintenance of aquaculture water environments.
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Cíclidos , Edwardsiella , Infecciones por Enterobacteriaceae , Enfermedades de los Peces , Animales , Enfermedades de los Peces/microbiología , Enfermedades de los Peces/patología , Enfermedades de los Peces/prevención & control , Infecciones por Enterobacteriaceae/veterinaria , Infecciones por Enterobacteriaceae/patología , Infecciones por Enterobacteriaceae/microbiología , Infecciones por Enterobacteriaceae/prevención & control , Edwardsiella/patogenicidad , Virulencia , Acuicultura , Taiwán , PecesRESUMEN
Persistent nocardiosis has prompted exploration of the effectiveness of heterologous approaches to prevent severe infections. We have previously reported the efficacy of a nucleic acid vaccine in protecting groupers from highly virulent Nocardia seriolae infections. Ongoing research has involved the supplementation of recombinant cholesterol oxidase (rCho) proteins through immunization with a DNA vaccine to enhance the protective capacity of orange-spotted groupers. Recombinant rCho protein exhibited a maturity and biological structure comparable to that expressed in N. seriolae, as confirmed by Western blot immunodetection assays. The immune responses observed in vaccinated groupers were significantly higher than those observed in single-type homologous vaccinations, DNA or recombinant proteins alone (pcD:Cho and rCho/rCho), especially cell-mediated immune and mucosal immune responses. Moreover, the reduction in N. seriolae occurrence in internal organs, such as the head, kidney, and spleen, was consistent with the vaccine's efficacy, which increased from approximately 71.4 % to an undetermined higher percentage through heterologous vaccination strategies of 85.7 %. This study underscores the potential of Cho as a novel vaccine candidate and a heterologous approach for combating chronic infections such as nocardiosis.
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Vacunas Bacterianas , Enfermedades de los Peces , Nocardiosis , Nocardia , Animales , Nocardiosis/veterinaria , Nocardiosis/prevención & control , Nocardiosis/inmunología , Nocardia/inmunología , Enfermedades de los Peces/prevención & control , Enfermedades de los Peces/inmunología , Vacunas Bacterianas/inmunología , Vacunas Bacterianas/administración & dosificación , Vacunas de ADN/inmunología , Vacunas de ADN/administración & dosificación , Lubina/inmunología , Colesterol Oxidasa/inmunología , Colesterol Oxidasa/genética , Proteínas Recombinantes/inmunología , Proteínas Recombinantes/administración & dosificaciónRESUMEN
Global ornamental fish transportation ranging from hours to days can produce multiple stress factors impact fish health and cause mortality. Clownfish, particularly Amphiprion ocellaris, are among the most traded saltwater ornamental fish. Vibrio includes several pathogenic strains that affect aquatic animals. Consequently, prophylactic treatment of the water or fish is recommended. In this study, six Vibrio strains including V. alginolyticus, V. parahaemolyticus and V. harveyi isolated from sick A. ocellaris and one V. harveyi strain from a sick East Asian fourfinger threadfin (Eleutheronema rhadinum) were tested for their sensitivity to a popular disinfectant, chlorine dioxide (ClO2). The results showed that 0.25 ppm ClO2 effectively suppressed five of the seven tested Vibrio strains for 24 h; however, 0.1 ppm ClO2 is safer for A. ocellaris. Meanwhile, ClO2 2.5 ppm reduced the bacterial counts to below 3.3 × 105 CFU/mL for 24 hours. The LC50 of ClO2 for A. ocellaris was 0.87 ppm at 10 min and 0.72 ppm at 24 h post treatment. Mild changes in water quality, including dissolved oxygen (DO), temperature and pH, were recorded during the trial. More research is necessary to understand the sensitivity of various aquatic animal pathogens to ClO2 and its toxicity to different aquatic animals.
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Compuestos de Cloro , Desinfectantes , Enfermedades de los Peces , Óxidos , Vibriosis , Vibrio , Compuestos de Cloro/farmacología , Animales , Óxidos/farmacología , Enfermedades de los Peces/microbiología , Enfermedades de los Peces/prevención & control , Vibrio/efectos de los fármacos , Desinfectantes/farmacología , Vibriosis/veterinaria , Vibriosis/prevención & control , Perciformes , Antibacterianos/farmacologíaRESUMEN
Nocardiosis, caused by Nocardia seriolae, has been a prominent disease in Southeast Asian aquaculture in the last three decades. This granulomatous disease reported in various fish species is responsible for significant economic losses. This study investigated the pathogenicity of N. seriolae in three cultured species in Taiwan: Nile tilapia (omnivore), milkfish (herbivore) and Asian seabass (carnivore). Administration of an infective dose of 1 × 106 CFU/ fish in tilapia, seabass and milkfish demonstrated mortalities of 100%, 90% and 75%, respectively. Additionally, clinical signs namely, granuloma and lesions displayed varying intensities between the groups and pathological scores. Polymerase chain reaction (PCR) amplification specific for N. seriolae was confirmed to be positive (432 bp) using NS1/NG1 primers. Post-mortem lesions revealed the absence of granulomas in tilapia and milkfish and their presence in the seabass. Interestingly, the gut in tilapia showed an influx of eosinophils suggesting its role during the acute stages of infection. However, post-challenge, surviving milkfish exhibited granulomatous formations, while surviving seabass progressed toward healing and tissue repair within sampled tissues. Overall, in conclusion, these results demonstrate the versatility in the immunological ability of individual Perciformes to contain this pathogen as a crucial factor that influences its degree of susceptibility.
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Cíclidos , Enfermedades de los Peces , Nocardiosis , Nocardia , Animales , Enfermedades de los Peces/microbiología , Enfermedades de los Peces/patología , Nocardia/patogenicidad , Nocardia/genética , Nocardia/aislamiento & purificación , Nocardiosis/veterinaria , Nocardiosis/microbiología , Taiwán , Acuicultura , Granuloma/veterinaria , Granuloma/microbiología , Granuloma/patologíaRESUMEN
Golden pompano (Trachinotus blochii) is a carnivorous teleost cultured in the Asia-Pacific region. Fish culture in high densities and numbers results in disease outbreaks, causing huge economic losses. Here, we collected cultured golden pompanos from 2021 to 2022 and identified the pathogens isolated from the diseased fish. Out of a total of 64 clinical cases observed in both sea cages and fish ponds, it was found that Nocardia seriolae was the predominant pathogen (26%), followed by Lactococcus garvieae (13%). Trichodina spp. was the most prevalent parasite in sea cages and earthen ponds (21%), while Neobenedenia spp. was the primary parasitic pathogen (16%) in sea cages. Given these findings, further investigations were conducted, including antibiotic susceptibility and pathogenicity tests specific to N. seriolae in golden pompanos. Antibiotic susceptibility tests of N. seriolae revealed that all strains were susceptible to doxycycline, oxytetracycline, florfenicol and erythromycin but resistant to amoxicillin and ampicillin. Additionally, a pathogenicity assessment was carried out by administering an intraperitoneal injection of 0.1 mL containing 107 CFU of N. seriolae per fish. The mortality rates observed varied between 40% and 90%, with the P2 strain exhibiting the highest level of virulence, resulting in a cumulative mortality of 90%. Therefore, disease outbreaks in fish can be minimized by developing effective treatments and prevention methods.
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Enfermedades de los Peces , Nocardiosis , Animales , Taiwán/epidemiología , Enfermedades de los Peces/epidemiología , Enfermedades de los Peces/prevención & control , Nocardiosis/epidemiología , Nocardiosis/veterinaria , Peces , Antibacterianos/farmacologíaRESUMEN
OBJECTIVE: As part of the National Disease Surveillance Program for Taiwanese Aquaculture, we investigated the causative agent of disease outbreaks in farmed Chicken Grunts Parapristipoma trilineatum. METHODS: In this study, outbreak cases on two separate farms were noticed in coastal Pingtung County, Taiwan. In total, 50 juvenile fish showing clinical signs (such as emaciation and erratic swimming behavior) and broodstock (two females and two males) from both farms were collected to perform gross lesion assessment, histopathological examination, and molecular identification of the pathogen. RESULT: Clinical symptoms were infected fish exhibited erratic swimming behavior, such as whirling and floating on the surface of the water. In the following months, cumulative mortality had reached 19% and 24%, respectively. The gross lesions in the infected fish included white oval cysts in the muscle, serosa of the internal organs, sclera of the eyes, and cerebral meninges. After conducting a wet mount examination of cysts using a light microscope, we observed a significant quantity of spores with morphological characteristics, suggesting their affiliation with the Myxosporea group. The spores were semiquadrate, with four tiny suture notches at the periphery; the mean spore length was 7.3 µm (SD = 0.5), and the mean spore width was 8.2 µm (SD = 0.6). The mean length and width of the pyriform polar capsules (nematocysts) were 3.6 µm (SD = 0.5) and 2.2 µm (SD = 0.5), respectively. The 18S and 28S ribosomal RNA sequences of these specimens were identical to those of Kudoa lutjanus. CONCLUSION: As this was the first time an outbreak of K. lutjanus in Chicken Grunts was confirmed, its reappearance with substantial mortality should serve as a warning to the aquaculture industry.
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Quistes , Enfermedades de los Peces , Myxozoa , Enfermedades Parasitarias en Animales , Femenino , Masculino , Animales , Pollos/genética , ADN Ribosómico/genética , Secuencia de Bases , ARN Ribosómico 18S/genética , Peces/genética , Myxozoa/genética , Brotes de Enfermedades/veterinaria , Quistes/epidemiología , Quistes/genética , Quistes/veterinaria , Enfermedades de los Peces/epidemiología , Filogenia , Enfermedades Parasitarias en Animales/epidemiologíaRESUMEN
Nocardiosis in aquatic animals caused by Nocardia seriolae is a frequently occurring serious infection that has recently spread to many countries. In this study, DNA vaccines containing potential bacterial antigens predicted using the reverse vaccinology approach were developed and evaluated in orange-spotted groupers. In silico analysis indicated that proteins including cholesterol oxidase, ld-transpeptidase, and glycosyl hydroxylase have high immunogenicity and are potential vaccine candidates. In vitro assays revealed the mature and biological configurations of these proteins. Importantly, when compared to a control PBS injection, N. seriolae DNA-based vaccines showed significantly higher expression of IL1ß, IL17, and IFNγ at 1 or 2 days, in line with higher serum antibody production and expression of other cellular immune-related genes, such as MHCI, CD4, and CD8, at 7 days post-immunization. Remarkably, enhanced immune responses and strong protective efficacy against a highly virulent strain of N. seriolae were recorded in DNA vaccine-cholesterol oxidase (pcD::Cho) injected fish, with a relative survival rate of 73.3%. Our results demonstrate that the reverse vaccinology approach is a valid strategy for screening vaccine candidates and pcD::Cho is a promising candidate that can boost both innate and adaptive immune responses and confer considerable protection against N. seriolae infection.
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Lubina , Enfermedades de los Peces , Nocardiosis , Vacunas de ADN , Animales , Vacunación Basada en Ácidos Nucleicos , Colesterol Oxidasa , Nocardiosis/prevención & control , Nocardiosis/veterinariaRESUMEN
Lactococcosis, caused by Lactococcus garvieae, is an acute hemorrhagic septicemia in fish recorded in marine and freshwater aquaculture during the summer months. In 2020-2021, several sea cage Pompano farms recorded sudden fish mortality events. Based on the results of phenotypic and biochemical tests, L. garvieae was predicted to be the cause. PCR with L. garvieae specific primers (pLG1 and pLG2) targeting the 16S rRNA region further confirmed the etiological agent as L. garvieae after amplifying an 1100 base pairs (bp) product. Furthermore, the 16S rRNA sequences of the two representative strains (AOD109-196-2B and AOD110-215-2B) shared 99.81% identity with L. garvieae (GenBank accession number: MT597707.1). The genetic profiles of the strains were classified using pulsed-field gel electrophoresis after digestion with SmaI and ApaI, which clustered our strains under the same pulsotype. Multiplex PCR targeting the capsule gene cluster and serotype-specific PCR collectively showed that the strains were non-capsulated; thus, they belonged to serotype I. An experimental infection was designed to fulfil Koch's postulates by infecting healthy Pompano with case-driven L. garvieae strains (AOD109-196-2B and AOD110-215-2B) with a cumulative mortality of 70%. Overall, L. garvieae infection in Pompano emphasizes the need for better monitoring and control procedures in aquaculture settings.
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Nocardia seriolae causes chronic nocardiosis in various marine and freshwater aquatic animals; however, grouper species have rarely been investigated. This study evaluated the pathogenicity of nocardiosis following N. seriolae infection in the orange-spotted grouper Epinephelus coioides. Nine identified genetic isolates of N. seriolae were tested in vivo using the intraperitoneal method and observed daily for 35 days. The most virulent isolate was then used to evaluate transmission through different routes (intraperitoneal IP, intramuscular IM, oral OR, and immersion IS) in the same fish model and was observed daily for 42 days. The results showed mild variation in virulence among N. seriolae isolates. AOD107132-2 K and OT103003-N11 strains displayed the highest and lowest risk virulence, respectively, based on the accumulation and kinetics of mortality. IM and IP administrations showed an early phase response with early mortality by 5 dpc (30%-100%), while slower kinetics of nocardiosis occurred in the OR and IS routes with slow mortality at 35 dpc (4%-8%). Histopathology revealed typical granulomas, confirming the progression of nocardiosis in the diseased fish. These results provide the basis for further studies on the virulence profile of N. seriolae in Taiwan and a well-suited route of administration in orange-spotted groupers for further prevention development.
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Elizabethkingia meningoseptica is a hazardous bacterium for agriculture production and human health. The present study identified E. meningoseptica from the bullfrog, human and reference strain BCRC 10677 by API 20NE, 50S ribosome protein L27 sequencing and pulse field gel electrophoresis to differentiate isolates of E. meningoseptica from aquatic animals and humans. All isolates from bullfrogs and humans were identified as E. meningoseptica by DNA sequencing with 98.8%-100% sequence identity. E. meningoseptica displayed significant genetic diversity when analysed using pulsed-field gel electrophoresis (PFGE). There were six distinct pulsotypes, including one pulsotype found in bullfrog isolates and five pulsotypes found in human isolates. However, E. meningoseptica from bullfrog exhibited one genotype only by PFGE. Overall, molecular epidemiological analysis of PFGE results indicated that the frog E. meningoseptica outbreaks in Taiwan were produced by genetically identical clones. The bullfrog isolates were not genetically related to other E. meningoseptica from human and reference isolates. This research provided the first comparisons of biochemical characteristics and genetic differences of E. meningoseptica from human and bullfrog isolates.
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Chryseobacterium , Enfermedades de los Peces , Infecciones por Flavobacteriaceae , Humanos , Animales , Rana catesbeiana , Taiwán/epidemiología , Infecciones por Flavobacteriaceae/epidemiología , Enfermedades de los Peces/epidemiología , Enfermedades de los Peces/tratamiento farmacológico , Chryseobacterium/genética , Genotipo , Electroforesis en Gel de Campo Pulsado/veterinaria , Antibacterianos/uso terapéuticoRESUMEN
The Chinese softshell turtle (CST; Pelodiscus sinensis) is a freshwater aquaculture species of substantial economic importance that is commercially farmed across Asia, particularly in Taiwan. Although diseases caused by the Bacillus cereus group (Bcg) pose a major threat to commercial CST farming systems, information regarding its pathogenicity and genome remains limited. Here, we investigated the pathogenicity of Bcg strains isolated in a previous study and performed whole-genome sequencing. Pathogenicity analysis indicated that QF108-045 isolated from CSTs caused the highest mortality rate, and whole-genome sequencing revealed that it was an independent group distinct from other known Bcg genospecies. The average nucleotide identity compared to other known Bcg genospecies was below 95%, suggesting that QF108-045 belongs to a new genospecies, which we named Bacillus shihchuchen. Furthermore, genes annotation revealed the presence of anthrax toxins, such as edema factor and protective antigen, in QF108-045. Therefore, the biovar anthracis was assigned, and the full name of QF108-045 was Bacillus shihchuchen biovar anthracis. In addition to possessing multiple drug-resistant genes, QF108-045 demonstrated resistance to various types of antibiotics, including penicillins (amoxicillin and ampicillin), cephalosporins (ceftifour, cephalexin, and cephazolin), and polypeptides, such as vancomycin.
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Bacillus anthracis , Bacillus , Tortugas , Animales , Bacillus/genética , Bacillus anthracis/genética , Bacillus cereus/genética , Genómica , Tortugas/genética , Tortugas/microbiología , Virulencia/genéticaRESUMEN
Streptococcus iniae is a Gram-positive bacterium and is considered a harmful aquaculture pathogen worldwide. In this study, S. iniae strains were isolated from East Asian fourfinger threadfin fish (Eleutheronema tetradactylum) reared on a farm in Taiwan. A transcriptome analysis of the head kidney and spleen was performed in the fourfinger threadfin fish 1 day after infection using the Illumina HiSeq™ 4000 platform for RNA-seq to demonstrate the host immune mechanism against S. iniae. A total of 7333 genes based on the KEGG database were obtained after the de novo assembly of transcripts and functional annotations. Differentially expressed genes (DEGs) (2-fold difference) were calculated by comparing the S. iniae infection and phosphate-buffered saline control group gene expression levels in each tissue sample. We identified 1584 and 1981 differentially expressed genes in the head kidney and spleen, respectively. Based on Venn diagrams, 769 DEGs were commonly identified in both the head kidney and spleen, and 815 and 1212 DEGs were specific to the head kidney and spleen, respectively. The head-kidney-specific DEGs were enriched in ribosome biogenesis. The spleen-specific and common DEGs were found to be significantly enriched in immune-related pathways such as phagosome, Th1, and Th2 cell differentiation; complement and coagulation cascades; hematopoietic cell lineage; antigen processing and presentation; and cytokine-cytokine receptor interactions, based on the KEGG database. These pathways contribute to immune responses against S. iniae infection. Inflammatory cytokines (IL-1ß, IL-6, IL-11, IL-12, IL-35, and TNF) and chemokines (CXCL8 and CXCL13) were upregulated in the head kidney and spleen. Neutrophil-related genes, including phagosomes, were upregulated post-infection in the spleen. Our results could offer a strategy for the treatment and prevention of S. iniae infection in fourfinger threadfin fish.
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Enfermedades de los Peces , Infecciones Estreptocócicas , Animales , Peces , Riñón Cefálico , Bazo , Streptococcus iniaeRESUMEN
Chronic disease following Nocardia seriolae infection in a wide range of aquatic animals has been reported in many Asian countries and recently in America and Mexico. This study aimed to investigate the epidemiological relationship among N. seriolae isolates in Taiwan by investigating their genotype and enzymatic activities. A total of 66 strains isolated from 14 known and four unknown host fish from five sites in Taiwan were characterized using five combined methods. High genotypic diversity was recognized among the isolates with 10 pulsotypes being identified from the pulsed-field gel electrophoresis method and 21 reptypes from the repetitive extragenic palindromic amplification method; however, no natural plasmids were detected in this bacterial population. Pulsotypes A8 and RI analysed by PFGE and repPCR, respectively, were found to be predominant within five sites in Taiwan over 17 years of isolation. Enzymatically, the majority of isolates displayed high leucine arylamidase, ß-glucosidase and α-glucosidase activities but were negative for lipase, α-galactosidase, ß-glucuronidase, N-acetyl-glucosaminidase, α-mannosidase and α-fucosidase activities. We identified a strong association between genotype and enzymatic activity since the majority of pulsotypes displayed the same type of enzymatic profile. This study provides comprehensive and potential epidemiological data, which will aid the fish farming activities and prevention method development.
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Enfermedades de los Peces , Nocardiosis , Nocardia , Animales , Taiwán/epidemiología , Enfermedades de los Peces/epidemiología , Enfermedades de los Peces/microbiología , Nocardia/genética , Nocardiosis/epidemiología , Nocardiosis/veterinaria , Nocardiosis/microbiología , Genotipo , Peces/microbiologíaRESUMEN
Piscine nocardiosis, caused by Nocardia seriolae, is a refractory granulomatous disease in South-East Asian aquaculture. This study investigates the virulence of nocardial lipids essential for pathogenesis among Actinomycetes. Petroleum ether (PE) was used to selectively delipidate two groups of N. seriolae, namely, live cell (LC) and killed cell (KC); resulting in delipidated live cell (DLC) and delipidated killed cell (DKC), respectively. Changes post-delipidation on genus characteristics, such as loss in acid-fast nature and resistance to lysozyme were observed. Transmission electron microscopy revealed notable changes in the lipid layer. Additionally, Lates calcarifer, Asian seabass intraperitoneally injected with LC and DLC had mortality rates of 90% and 50%, respectively, with the latter exhibiting a delay in mortality. Reverse-transcription quantitative PCR (RT-qPCR) analysis of host cytokines from the spleen and head kidney showed delipidation contributed to the induction of an immune response with increased transcriptional levels of interferon-γ (ifn-γ). Histopathological samples collected on day 7 post-inoculation displayed a varied granulomatous response between the treatment groups and scored for pathological changes. These findings affirm that the virulence of the lipids remains independent of the living state of the cell, significantly altering the immune and granulomatous responses in L. calcarifer to N. seriolae.
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Enfermedades de los Peces , Nocardiosis , Nocardia , Animales , Virulencia , Nocardiosis/veterinaria , Pared Celular , LípidosRESUMEN
Edwardsiella tarda (ET) and Edwardsiella anguillarum (EA) are the most harmful bacterial fish pathogens in Taiwan. However, there is confusion regarding the genotypic identification of E. tarda and E. piscicida (EP). Therefore, we used a novel Nanopore MinION MK1C platform to sequence and compare the complete genomes of E. piscicida and E. anguillarum. The number of coding genes, rRNA, and tRNA recorded for E. anguillarum and E. piscicida were 8322, 25, and 98, and 5458, 25, and 98, respectively. Ribosomal multilocus sequence typing (rMLST) for E. piscicida indicated 35 rps. The shared clusters between E. anguillarum and E. piscicida indicated several unique clusters for the individual genomes. The phylogenetic tree analysis for all complete genomes indicated that E. anguillarum and E. piscicida were placed into two species-specific genotypes. Distribution of subsystems for annotated genomes found that genes related to virulence, defence, and disease for E. anguillarum were 103 and those for E. piscicida were 60 and pathogenic islands (PI) were 498 and 225, respectively. Vaccine candidates were identified in silico from the core genes using high antigenic, solubility, and secretion probabilities. Altogether, the genome data revealed distinctive features between E. anguillarum and E. piscicida, which suggest different pathogenicity and thus the need for separate preventive strategies.
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Edwardsiella , Infecciones por Enterobacteriaceae , Enfermedades de los Peces , Nanoporos , Animales , Filogenia , Factores de Virulencia/genética , Taiwán , Enfermedades de los Peces/microbiología , Genómica , Infecciones por Enterobacteriaceae/microbiologíaRESUMEN
Vibrio harveyi is a Gram-negative bacterium that causes vibriosis in various aquaculture species, including the orange-spotted grouper (Epinephelus coioides). Bacterial flagellin is a potent pathogen-associated molecule that stimulates the innate and adaptive immune systems through toll-like receptor 5 (TLR5) signaling. In this study, we isolated V. harveyi flagellin A (VhFliA) gene from V. harveyi (originated from orange-spotted grouper) and investigated the in vivo activities of recombinant VhFliA protein. Multiple sequence alignment showed that the amino acid sequence of VhFliA has conserved domains of N- and C-terminals (D0 and D1) and a middle variable (MV) region. We produced the VhFliA recombinant protein (wild type (WT)-VhFliA) by Escherichia coli and investigated its in vivo biological activity. Additionally, we prepared the VhFliA recombinant proteins with deletion of domains (ΔMV-VhFliA and ΔD0MV-VhFliA) to identify the domain for biological activity in the orange-spotted grouper. WT and ΔMV-VhFliA induced the expression of inflammatory cytokines (IFNγ, IL-1ß, and IL-8) in groupers. However, ΔD0MV-VhFliA did not induce the expression of inflammatory cytokines. Additionally, to demonstrate the applicability of recombinant VhFliA to teleost species, we performed an in vivo assay of the recombinant proteins in koi carp (Cyprinus carpio). WT-VhFliA stimulates the expression of inflammatory cytokines (IL-1ß, IL-6, and IL-8) in carp. ΔMV-VhFliA did not upregulate IL-1ß and IL-6, whereas ΔD0MV-VhFliA induced expression in carp. These findings showed the potential of VhFliA as an effective immune stimulant adjuvant and comparative studies of flagellin - TLR5 signaling in teleosts.
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Carpas , Flagelina , Animales , Carpas/genética , Interleucina-6 , Interleucina-8 , Proteínas Recombinantes/genéticaRESUMEN
Edwardsiella spp. is a gram-negative, facultatively anaerobic, intracellular bacteria threatening the aquaculture industry worldwide. Noticeably, E. tarda is now genotypically classified into three distinct groups (E. tarda, E. piscicida and E. anguillarum), but morphologically, it is unclear due to varying degrees of virulence in different fish hosts. Hence, to reclassify E. tarda, we investigated differences in genotypes, phenotypes and pathogenicity. We collected Edwardsiella isolates from five different counties of Taiwan between 2017 and 2021. At first, gyrB gene was amplified for a phylogenetic tree from 40 isolates from different fish and one reference isolate, BCRC10670, from the human. Thirty-nine strains clustered into E. anguillarum, 1 strain into E. piscicida and 1 strain into E. tarda from human strain. Second, all isolates were characterized using various phenotypic (API 20E biochemical profiles) and genotypic (pulsed-field gel electrophoresis [PFGE], and virulence-related gene detection). SpeI digestion revealed 10 pulsotypes and I-CeuI into 7 pulsotypes. Virulent genes (citC, gadB, katB, mukF and fimA) confirmed in 35, 31, 28, 37 and 38 isolates, respectively. Finally, in vivo challenge test in milkfish (Chanos chanos) indicated the highest mortality from E. anguillarum. Overall, results revealed unique features with Edwardsiella spp. genotypes and pathogenicity, which are relevant to the host and provide useful insights for future vaccine development.
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Edwardsiella , Infecciones por Enterobacteriaceae , Enfermedades de los Peces , Animales , Edwardsiella/genética , Edwardsiella tarda/genética , Infecciones por Enterobacteriaceae/microbiología , Infecciones por Enterobacteriaceae/veterinaria , Enfermedades de los Peces/microbiología , Peces/microbiología , Humanos , Fenotipo , Filogenia , TaiwánRESUMEN
Between 2010 and 2015, nocardiosis outbreaks caused by Nocardia seriolae affected many permit farms throughout Vietnam, causing mass fish mortalities. To understand the biology, origin and epidemiology of these outbreaks, 20 N. seriolae strains collected from farms in four provinces in the South Central Coast region of Vietnam, along with two Taiwanese strains, were analysed using genetics and genomics. PFGE identified a single cluster amongst all Vietnamese strains that was distinct from the Taiwanese strains. Like the PFGE findings, phylogenomic and SNP genotyping analyses revealed that all Vietnamese N. seriolae strains belonged to a single, unique clade. Strains fell into two subclades that differed by 103 SNPs, with almost no diversity within clades (0-5 SNPs). There was no association between geographical origin and subclade placement, suggesting frequent N. seriolae transmission between Vietnamese mariculture facilities during the outbreaks. The Vietnamese strains shared a common ancestor with strains from Japan and China, with the closest strain, UTF1 from Japan, differing by just 220 SNPs from the Vietnamese ancestral node. Draft Vietnamese genomes range from 7.55 to 7.96 Mbp in size, have an average G+C content of 68.2â% and encode 7â602-7958 predicted genes. Several putative virulence factors were identified, including genes associated with host cell adhesion, invasion, intracellular survival, antibiotic and toxic compound resistance, and haemolysin biosynthesis. Our findings provide important new insights into the epidemiology and pathogenicity of N. seriolae and will aid future vaccine development and disease management strategies, with the ultimate goal of nocardiosis-free aquaculture.
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Enfermedades de los Peces , Nocardiosis , Animales , Acuicultura , Enfermedades de los Peces/epidemiología , Enfermedades de los Peces/genética , Genómica , Nocardia , Nocardiosis/epidemiología , Nocardiosis/veterinaria , Vietnam/epidemiologíaRESUMEN
Lactococcus garvieae is the etiological agent of Lactococcosis, an evolving disease affecting many fish species and causing significant economic losses worldwide. Assessing pathogen relatedness and bacterial population structure is critical for determining the epidemiology of L. garvieae infections and in establishing effective pathogen management methods. The previously published morphological and genetic studies point to a clonal population structure, as seen in other fish bacteria. In the present study, the pulsed-field gel electrophoresis (PFGE) method was utilized to define a population of 41 Taiwanese isolates from outbreaks with comparisons to four well-characterized non-Taiwanese isolates previously published. Two restriction enzymes (ApaI and SmaI) were utilized individually for PFGE analysis (cut-off value = 90.0%), revealing genetic heterogeneity across L. garvieae isolates, with ApaI and SmaI yielding 12 and seven distinct PFGE band patterns, respectively. The phylogenic analysis using internal transcribed spacer region clustered all L. garvieae isolates in the same clad. Furthermore, the electron microscopic results confirmed the absence of capsular gene cluster (CGC) in previously characterized Taiwanese vaccine strain (S3) from grey mullet. Overall, our findings emphasize the importance of analysing the morphological and genetic diversity in L. garvieae being correlated for proper taxonomic classification in vaccine strain selection and epidemiological studies.
Asunto(s)
Enfermedades de los Peces , Animales , Electroforesis en Gel de Campo Pulsado/veterinaria , Enfermedades de los Peces/microbiología , Peces , Genotipo , Lactococcus/genética , Microscopía ElectrónicaRESUMEN
For the first time, Longicollum pagrosomi Yamaguti, 1935, an acanthocephalan parasite, has been characterized with morphological, histopathological and molecular detail in farmed red snapper (Lutjanus erythropterus) in Taiwan. The diseased fish showed clinical signs of anorexia, emaciation and were gasping for air at the water's surface. Gross examination revealed extensive necrosis in the intestine and even penetration at the site of parasitic attachment, resulting in a large number of parasites being lodged in the peritoneal cavity, surrounded by black hyperplastic connective tissue. The parasites collected from the intestine were studied using optical microscopy, histopathology, scanning electron microscopy (SEM) and molecular phylogenetic analysis. They were creamy white in colour, and were separated into their proboscis, neck and metasoma (trunk) under optical microscopy. Histopathological examination revealed ovarian balls (floating ovaries) inside the ligament sac and eggs within the egg sorting apparatus. The SEM images revealed that the proboscis had 11-14 longitudinal rows with 9-12 recurrent, backward-facing, organized hooks. In the 18S and 28S phylogenetic tree, sequences of the specimens were identical to those of other Longicollum pagrosomi, in addition to the morphological features. Thus, we confirmed that the parasites belonged to Longicollum pagrosomi in this study.