• Journal of fish pathology
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• v.18 no.2
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• pp.147-156
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• 2005

To obtain antimicrobial algae against fish pathogenic bacteria, we screened 80% methanolic extracts of 30 algae using fish pathogenic bacteria, Staphylococcus sp., Streptococcus sp., Edwardsiella tarda and Vibrio anguillarum. Among them, Corallina officinalis, Dumontia simplex, Gloipeltis furcata, Grateloupia lanceolata and Grateloupia turuturu were effective for growth inhibition of a Gram-positive bacterium, Staphylococcus sp.. Sargassum thunbergii and Polysiphonia morrowii exhibited significant inhibitory effects against the growth of Gram-negative bacteria, both E. tarda and V. anguillarum. Moreover, antimicrobial activity-guided fractionation for P. morrowii extract yielded significantly active 90% methanolic fraction. This fraction significantly inhibited the growth of E. tarda exhibiting a MIC of 1 mg/ml. In addition, its antimicrobial activity was stable under various pH conditions.

• Journal of fish pathology
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• v.2 no.2
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• pp.83-90
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• 1989

Edwardsiella tarda, the causative organism of Edwardsiellosis, was isolated from the water and the eel, Anguilla japonica of the recirculating eel culture ponds in 1988. E. tarda was always isolated from the water of eel culture ponds in the number of log 2.30~log 4.78 CFU/$m{\ell}$ without distinguishable seasonal characteristics. The counts of isolated E. tarda were changed with the health condition of the eel, i. e., a number of E. tarda were isolated from all checked organs, intestine contents, liver, spleen and kidney of the diseased eel, however, it was hardly isolated from spleen and kidney of the healthy eel. The isolates were classified according to o-agglutinating test into three serotypes (I, II, III). There were no distinct differences in the composition of the serotypes between intestine contents of eel and pond water samples(I, 52.4~53.3% ; II, 20.0~23.8% ; III, 4.8~6.7%). Serotype I was ascendant in the spleen(66.7%) and the kidney(73.7%) samples. By experimental challenge, most of serotype I proved highly virulent to eel in comparison with the other serotypes tested.

• Journal of fish pathology
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• v.12 no.1
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• pp.24-31
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• 1999

Optimization and standardization of solid phase enzyme immunoassay were done for the diagnosis of edwardsiellosis in fish. The analyzed degree of immobilized antibody on surface of solid phase with peroxidase saturation method showed the optimized result by using partially purified $50{\mu}g/ml$ of rabbit anti-E. tarda Edk-2 antibody in sodium bicarbonate buffer for overnight incubation to cover the surface of polystyrene beads. Optimized immunoreaction was observed in the treatment of $50{\mu}g/ml$ of biotin conjugated antibody followed extravidin-peroxidase diluted 1 : 2,000 in PBS. The detectable concentrations of the this method were $1{\times}10^5$ cells/ml and $1{\times}10^5$ cells/ml expressed as the source of antigen amount for EDTA extraction and heat extraction, respectively. High cross-reaction of solid phase ELISA with the prepared rabbit and-E. tarda Edk-2 was observed against E. tarda strains isolated from flounder suffering from edwardsiellosis in aquatic farms of Korea. It suggested that the potential of this solid phase of ELISA technique is very powerful for the application to different strains of E. tarda isolated in farms of many different areas.

• Journal of Microbiology and Biotechnology
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• v.20 no.12
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• pp.1735-1743
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• 2010

The full-length genes gyrB (2,415 bp), parC (2,277 bp), and parE (1,896 bp) in Edwardsiella tarda were cloned by PCR with degenerate primers based on the sequence of the respective quinolone resistance-determining region (QRDR), followed by elongation of 5' and 3' ends using cassette ligation-mediated PCR (CLMP). Analysis of the cloned genes revealed open reading frames (ORFs) encoding proteins of 804 (GyrB), 758 (ParC), and 631 (ParE) amino acids with conserved gyrase/topoisomerase features and motifs important for enzymatic function. The ORFs were preceded by putative promoters, ribosome binding sites, and inverted repeats with the potential to form cruciform structures for binding of DNA-binding proteins. When comparing the deduced amino acid sequences of E. tarda GyrB, ParC, and ParE with those of the corresponding proteins in other bacteria, they were found to be most closely related to Escherichia coli GyrB (87.6% identity), Klebsiella pneumoniae ParC (78.8% identity), and Salmonella Typhimurium ParE (89.5% identity), respectively. The two topoisomerase genes, parC and parE, were found to be contiguous on the E. tarda chromosome. All 18 quinolone-resistant isolates obtained from Korea thus far did not contain subunit alternations apart from a substitution in GyrA (Ser83$\rightarrow$Arg). However, an alteration in the QRDR of ParC (Ser84$\rightarrow$Ile) following an amino acid substitution in GyrA (Asp87$\rightarrow$Gly) was detected in E. tarda mutants selected in vitro at $8{\mu}g/ml$ ciprofloxacin (CIP). A mutant with a GyrB (Ser464$\rightarrow$Leu) and GyrA (Asp87$\rightarrow$Gly) substitution did not show a significant increase in the minimum inhibitory concentration (MIC) of CIP. None of the in vitro mutants exhibited mutations in parE. Thus, gyrA and parC should be considered to be the primary and secondary targets, respectively, of quinolones in E. tarda.

• Fisheries and Aquatic Sciences
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• v.25 no.4
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• pp.243-249
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• 2022

Bacillus SW1-1 is a probiotic isolated from shrimp intestines. We investigated the effects of Bacillus SW1-1 coated diets on the growth, feed utilization, innate immunity, hematological parameters and resistance to Edwardsiella tarda in olive flounder (Paralichthys olivaceus). A commercial diet was used as the control (AP0) and two other diets were prepared by coating 0.25% (AP25) or 0.50% (AP50) probiotic powder which contains 1.0 × 10⁷ CFU/g Bacillus SW1-1. Four replicate groups of olive flounder (153 ± 2 g) were fed one of the diets for 12 weeks. Growth performance and feed utilization of the fish were not significantly affected by the dietary Bacillus SW1-1. After the challenge with E. tarda, AP50 group showed significantly higher survival than AP0 and AP25 groups. Innate immunity and anti-oxidant capacity of the fish were not significantly affected after the feeding trial. However, after the E. tarda challenge, the innate immune parameters (immunoglobulin, lysozyme and anti-protease) were significantly improved in fish fed AP25 and AP50 diets compared to those in fish fed AP0 diet. After the challenge test, significantly lower glucose level was observed in AP50 group compared to AP0 group. These results indicate that dietary supplementation of Bacillus SW1-1 could increase the disease resistance of olive flounder against E. tarda infection. The optimum coating levels of Bacillus SW1-1 needs to be further elucidated.

• Journal of fish pathology
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• v.33 no.2
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• pp.111-118
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• 2020

The genus Edwardsiella belonging to the family Enterobacteriaceae is a member of Gram-negative rod-shaped bacteria that cause disease in diverse aquatic organisms such as fish, amphibians and reptiles as well as avians and mammals including human throughout the world. This genus had been composed of three species, E. hoshinae, E. ictaluri and E. tarda, but recent researches erected two novel species, E. anguillarum and E. piscicida that were conventionally identified as E. tarda. In this study, we isolated seven strains belonging to the genus Edwardsiella from freshwater fishes that had been reared at inland fish farms in South Korea and investigated their biochemical characteristics and molecular phylogenetic relationships. The seven strains showed typical characteristics of four Edwardsiella species, E. anguillarum, E. ictaluri, E. piscicida and E. tarda, by biochemical analyses of Gram staining, indole and hydrogen sulfide (H2S) production, and API (Analytic Profile Index) 20E test. Molecular phylogenetic analyses inferred from DNA sequence data of both 16S ribosomal RNA (rRNA) and DNA gyrase subunit B (gyrB) genes were congruent with the biochemical characteristics. As a result, both biochemical and molecular phylogenetic analyses identified four strains isolated from three Anguilla species as E. anguillarum, E. piscicida and E. tarda, two strains from Pelteobagrus fulvidraco and Silurus asotus as E. ictaluri, and one strain from Moroco oxycephalus as E. piscicida. In this study, we isolated and successfully identified recently newly erected species, E. anguillarum and E. piscicida in addition to historically notorious pathogenic species, E. ictaluri and E. tarda. In the future study, systematic and comprehensive monitoring of the four Edwardsiella species are required for studying differences in pathogenicity among freshwater fishes.

• Journal of fish pathology
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• v.24 no.3
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• pp.297-305
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• 2011

A trap identification system for isolating functional sequences to allow the constitutive expression of foreign protein from Edwardsiella tarda was developed. Using the green fluorescent protein (GFP) reporter-based trap system, various functional sequences to drive heterologous expression of the GFP were selectable in Escherichia coli host. However from the bioinformatic sequence analysis, all the segments predicted as regulatory regions were not native promoters actually existing upstream of endogenous E. tarda genes. Instead, a number of non-authentic sequences, possibly resulted from the random shuffling and/or intermolecular ligation were also proven to be able to display a potent GFP expression in the recombinant E. coli. Further analysis with selected clones showed that both authentic and non-authentic sequences could function in as a constitutive promoter, leading quite a consistent and stable GFP expression after repetitive subcultures. Microscopic examination also confirmed the uniform pattern of GFP expression in every host bacterium. Semi-quantitative assay of GFP showed that there was no clear relationship between expression levels and organizational features of the promoters trapped. Functional promoter-like elements achieved in the present study could be a good starting material for multivalent genetic engineering of E. tarda in order to produce recombinant vaccines in a cost-effective fashion.

• Journal of fish pathology
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• v.7 no.2
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• pp.119-126
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• 1994

To investigate effects of water temperature on immune response of flounder, Paralichthys olivaceus, against Edwardsiella tarda, fish were immunized with formalin killed E. tarda antigen, and humoral immune response of these fish were observed. At lower water temperature (12 and $15^{\circ}C$), the antibody appeared 2 to 3 weeks after injection of formalin killed E. tarda antigen and the maximum agglutination titer was 16 and 32, respectively. However at higher water temperature (20 and $23^{\circ}C$), the antibody appeared one week after injection and the maximum agglutination titer was about 2,048. Once produced agglutination titer was sensitively responsed to variation of water temperature and showed that this phenomenon had also a similar tendency under natural condition. And it showed that agglatination titer of flounder immunized with formalin killed E. tarda maintained above 19 months.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.41 no.3
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• pp.188-192
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• 2008

Methyl gallate isolated from bark of the tree Rhus verniciflua Stokes has significant antimicrobial activity against the fish pathogenic bacteria Edwardsiella tarda and Vibrio anguillarum. To evaluate the antimicrobial activity of gallate derivatives, eight alkyl gallates were tested. Ethyl gallate and propyl gallate had the highest activities, with MICs of $15.6-31.3{\mu}g/mL$ against E. tarda. For V. anguillarum, propyl gallate and butyl gallate were highly effective, with MICs of $7.81-31.3{\mu}g/mL$. When used in combination with antibiotics, methyl gallate exhibited synergistic effects with oxytetracycline against E. tarda and with norfloxacin against V. anguillarum. These results suggest that short-chain alkyl gallates can be used as alternatives to antibiotics against the fish pathogenic bacteria.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.47 no.1
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• pp.23-30
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• 2014

The present study investigated the effects of dietary Edwardsiella tarda (E. tarda) specific bacteriophage (phage) and Bacillus subtilis (B. subtilis) mixture on innate immune responses and antibacterial activity of Nile tilapia, Oreochromis niloticus. In a dietary experiment, tilapia were fed the control diet (C), a phage-only supplemented diet (P), a B. subtilis only supplemented diet (B), or a B. subtilis and phage mixed diet (B+P). A respiratory burst and significant increase in lysozyme activity (P<0.05) were noted in the B+P group, as compared to other groups after 4 days of feeding. The B group showed a significant (P<0.05) increase in respiratory burst and lysozyme activity versus the C and P groups, whereas no significant increases (P<0.05) were observed in the P and C groups. $ACH_{50}$ was significantly up-regulated in the B+P group versus other groups after 8 days of feeding (P<0.05). In vivo antibacterial activity was significantly enhanced in the B+P fed group, as compared to other groups (P<0.05) after 7 days of E. tarda challenge. A significant (P<0.05) increase in antibacterial activity was seen in the B group, as compared to C or P groups after 14 days of feeding. These results suggest that a B. subtilis and phage mixture could be utilized as an alternative to antibiotics in the control of fish diseases caused by E. tarda.