Diseased Olive Flounder Research Articles

Draft genome sequence of multiple antibiotic-resistant Vibrio harveyi isolated from diseased olive flounder (Paralichthys olivaceus) farmed in South Korea.

Vibrio harveyi strain 22FBVib0145 was isolated from a diseased olive flounder farmed in Jeju, Korea. Here, we report the draft genome sequence of this strain. It is 6,238,277 bp in length with a G + C content of 44.8%.

Complete Genome Sequences of Streptococcus parauberis Serotype Ia, Ib/Ic, and II Isolates from Diseased Olive Flounder in South Korea.

We report the complete genome sequences of three isolates of Streptococcus parauberis, representing serotypes Ia, Ib/Ic, and II, which were isolated from diseased olive flounder (Paralichthys olivaceus) in South Korea.

Phenotypic differences between Edwardsiella piscicida and Edwardsiella anguillarum isolates in Japan.

Most studies refer to Edwardsiella tarda as the causative agent of edwardsiellosis in the cultured marine and freshwater fish species in Japan; however, our previous study was the first to genetically classify an E. tarda-like isolate from diseased olive flounder as E. piscicida and that from diseased red sea bream as E. anguillarum. This study aimed to understand the phenotypical differences between the two species. The API 50 CH test showed that L-arabinose and D-mannitol were fermented by 7 E. anguillarum isolates, but not by 14 E. piscicida isolates; the difference is likely to be useful for the recognition of E. piscicida, although it was not common among previously-reported E. anguillarum isolates. The use of the defined medium consisting of M9 minimal medium supplemented with 43 components revealed that the growth of E. piscicida and E. anguillarum was reduced in the defined medium without methionine and iron sulfate. Individually, the growth of E. piscicida was reduced in the defined medium without phenylalanine, tyrosine, alanine, or nicotinic acid, whereas the growth of E. anguillarum was reduced without serine, cysteine, leucine, threonine, or isoleucine. Tyrosine and alanine were present in higher concentrations in the olive flounder serum and threonine and isoleucine were present in higher concentrations in the red sea bream serum, compared with each other, suggesting that E. piscicida and E. anguillarum may have the advantage of growth opportunity in their hosts. Finally, the minimal defined medium for the two species was determined to enable safe vaccine development.

Draft Genome Sequences of Two Edwardsiella piscicida Strains, JF1307 and JF1411, Isolated from Diseased Olive Flounder (Paralichthys olivaceus) Cultured in Japan.

Edwardsiella piscicida strains JF1307 and JF1411 were isolated from cultured olive flounder that were diagnosed as being infected with edwardsiellosis. The draft genome sequences of the two isolates comprise 3,882,000 bp and 3,827,424 bp with G+C contents of 59.5% and 59.6%, respectively.

Complete Genome Sequence of Multiple-Antibiotic-Resistant Streptococcus parauberis Strain SPOF3K, Isolated from Diseased Olive Flounder (Paralichthys olivaceus).

ABSTRACTHere, we report the complete genome sequence of multiple-antibiotic-resistant Streptococcus parauberis strain SPOF3K, isolated from the kidney of a diseased olive flounder in South Korea in 2013. Sequencing using a PacBio platform yielded a circular chromosome of 2,128,740 bp and a plasmid of 23,538 bp, harboring 2,123 and 24 protein-coding genes, respectively.

Genotype and virulence of Streptococcus iniae isolated from diseased olive flounder Paralichthys olivaceus in Korea

In the study, we characterized 29 Streptococcus iniae isolates from diseased olive flounder Paralichthys olivaceus in Korea from 2000 to 2005. Biochemical characteristics of 29 isolates using API 20 strep were identical. Through analysis of repetitive sequence-based PCR (rep-PCR) using BoxA primer and random amplified polymorphic DNA using p14 primer, 29 isolates of S. iniae were divided into two genotypes. The isolates were divided into two clusters by comparison of genetic distance using a sequence of the capsular polysaccharide D gene that was consistent with genotyping by the rep-PCR. The isolates belonging to genotype 1 in rep-PCR analysis showed a high virulence in the flounder, while the isolates belonging to genotype 2 were relatively low in virulence. Therefore, a correlation between the genotype and the virulence of S. iniae isolates has been identified.

Genome Sequence of Streptococcus parauberis Strain KCTC11980, Isolated from Diseased Paralichthys olivaceus

Streptococcus parauberis is a coccoid, nonmotile, alpha-hemolytic, Gram-positive bacterium of the Streptococcaceae family. Streptococcus parauberis strain KCTC11980 was isolated from the kidney of a diseased olive flounder collected from an aquaculture farm on Jeju Island in 2010. The 2.12-Mb genome sequence consists of 44 large contigs in 16 scaffolds and contains 2,214 predicted protein-coding genes, with a G+C content of 35.4%.

Microscopic Image Recognition-based Fish Disease Diagnosis System

This paper presents a microscopic image recognition-based fish disease diagnosis system capable of faster treatment of diseased fish in order to prevent the spread of disease. The system is composed of three steps: 1) extracting the pathogen area from the microscopic images of diseased fish tissue by applying image processing techniques such as noise reduction, edge detection, and object detection, 2) confirming the pathogen and making the final diagnosis by feature extraction, and correlation matching compared to the registered pathogen database, and 3) proving the information of diagnosed disease, treatment and prevention methods by a connected web server through the Internet and SMS message by mobile communication. The designed fish disease diagnosis system was implemented to diagnose 3 parasitic diseases of olive flounder. The system can support fish farmers and veterinarians by providing easy and rapid diagnosis of diseased olive flounder, guidance in the use of appropriate drugs and a suitable treatment method for the diagnosed disease.

Determination of epidemiological tetracycline MIC cut-off value for Vibrio ichthyoenteri

Normalized resistance interpretation (NRI) analysis for tetracycline was applied to generate information on the epidemiological cut-off value for Vibrio ichthyoenteri isolated from diseased olive flounder (Paralichthys olivaceus) larvae. Thus, 42 strains of V. ichthyoenteri were used to determine minimum inhibitory concentration (MIC) values of tetracycline using Etest. Also, 11 tetracycline resistance related genes were investigated by PCR method. Most tetracycline-resistant strains harbored both tetB and tetM with a few exceptions. NRI-derived mean and 2 SD above the mean of theoretical normal distributions of susceptible isolates were 0.33 mg/L and 1.66 mg/L, respectively. The epidemiological cut-off value for V. ichthyoenteri from the calculations could be set to S <TEX>${\leq}$</TEX> 2 mg/L. Of the 42 strains, 15 were classified as non-wild type (NWT), and MIC values of the NWT strains vary regardless of tetB and tetM detection, suggesting that there may be other mechanisms involved in tetracycline resistance in this Vibrio species.

Small subunit ribosomal RNA and mitochondrial cytochrome c oxidase subunit 1 gene sequences of 21 strains of the parasitic scuticociliate Miamiensis avidus (Ciliophora, Scuticociliatia)

The scuticociliate Miamiensis avidus is a histophagous parasite that causes high mortality in cultured marine fishes. Small subunit ribosomal RNA (SSU rRNA) and mitochondrial cytochrome c oxidase subunit 1 (cox1) genes were analyzed for 21 strains of M. avidus isolated from diseased olive flounder (Paralichthys olivaceus), ridged-eye flounder (Pleuronichthys cornutus), and spotted knifejaw (Oplegnathus fasciatus) in Korea and Japan (collected in 2003-2007). Analysis of SSU rRNA gene sequences (1,759 bp) indicates they are very conserved with less than 0.17% (3 nucleotides) differences suggesting that SSU rRNA are useful to identify M. avidus; however, the cox1 gene (900 bp) has higher variations with intraspecific divergences up to 5.67% (51 nucleotides). A distance tree of cox1 gene sequences based on a neighbor-joining analysis can separate 21 strains into five cox1 types (two heterogeneous clusters and three individual branches). The cox1-type matches with serotype of strains but do not reflect geographical origins, host species, or pathogenicity.

Phenotypic characteristics ofStreptococcus iniaeandStreptococcus parauberisisolated from olive flounder (Paralichthys olivaceus)

The etiological agents of streptococcosis were isolated from diseased olive flounder collected on the Jeju island of Korea. A total of 151 bacterial isolates were collected between 2003 and 2006. The isolates were examined using various phenotypic and proteomic analyses, including sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), immunoblotting, and glycoprotein assays. In addition, isolates were grown on blood agar to assess hemolytic activity, and biochemical assays were performed using the API20 Strep kit. Our results revealed that all isolates were nonmotile, Gram-positive cocci that displayed negative catalase and oxidase activities. Multiplex PCR assays revealed that 43% and 57% of the isolates were Streptococcus iniae and Streptococcus parauberis, respectively. These results were consistent with those of the SDS-PAGE and immunoblot analyses using whole-cell lysates of bacterial isolates. Significant differences were observed with respect to the Voges-Proskauer, pyrrodonyl arylamidase, alkaline phosphatase, and hemolytic activities of the S. iniae and S. parauberis isolates. Isolates of S. iniae displayed uniform profiles in the immunoblot and glycoprotein assays; however, immunoblot assays of S. parauberis isolates (using a chicken IgY antibody raised against a homologous isolate) revealed three distinct antigenic profiles. Our findings suggest that S. parauberis and S. iniae are endemic pathogens responsible for the development of streptococcosis in olive flounder.

Pathogenicity of Streptococcus parauberis to Olive Flounder Paralichthys olivaceus

Streptococcus parauberis (stain SNUFPC-050803), isolated from diseased olive flounder Paralichthys olivaceus in Jeju Island, Korea, was evaluated for its pathogenicity to healthy juvenile flounder (29.3 g in average body weight). When challenged with the isolate by intraperitoneal injection with tenfold serial dilutions of 4.5 × 10-106 CFU/fish, the cumulative mortality ranged from 10% to 80% within 14 days except for 4.5 × 10 CFU/fish and control with no mortality. Disease signs were hemorrhage around the mouth, eyes and pectoral fins, pale and friable liver with hepatomegaly and ascitic fluid in the peritoneal cavity. These signs were similar to those of naturally affected fish. S. parauberis was reisolated and identified by PCR method, which confirmed the pathogenicity of the bacterium to olive flounder.

Isolation and characterization ofStreptococcussp. from diseased flounder (Paralichthys olivaceus) in Jeju Island

Streptococcus sp. is gram-positive coccus that causes streptococcal infections in fish due to intensification of aquaculture and caused significant economic losses in fish farm industry. A streptococcal infection occurred from cultured diseased olive flounder (Paralichthys olivaceus) in May, 2005 at a fish farm in Jeju Island, Korea. The diseased flounder exhibited bilateral exophthalmic eyes and rotten gills; water temperature was 16~18℃ when samples were collected. Of the 22 fish samples collected, 3 samples were identified as Lactococcus garvieae and 18 samples were identified as Streptococcus parauberis by culture-based, biochemical test. Serological methods such as slide agglutination, hemolysis and antimicrobial susceptibility test were also used as well as multiplex PCR-based method to simultaneously detect and confirm the pathogens involved in the infection. S. parauberis and L. garvieae have a target region of 700 and 1100 bp., respectively. One fish sample was not identified because of the difference in the different biochemical and serological tests and was negative in PCR assay. In the present study, it showed that S. parauberis was the dominant species that caused streptococcosis in the cultured diseased flounder.

넙치 (Paralichthys olivaceus) 병어에서 분리된 연쇄상구균의 다양성

To evaluate the biological diversity of fish pathogenic streptococci, 35 strains isolated from diseased olive flounder (Paralichtys olivaceus), were analyzed using a random amplified polymorphic DNA (RAPD) technique with the oligonucleotide commercial primer 6 (Amersham Biosciences). Api 20 Strep test, drug resistance and artificial infection were carried out for further characterization of the isolates. RAPD fingerprints showed similar pattern in 25 strains (about of 35 isolates) and these strains were designed as RA group 1. Similarities greater than were obtained when the Dice coefficient was applied among the isolates of RA 1. On the other hand, the reference Streptococcus iniae showed a similar RAPD profile to the isolates with similarity levels of Rh I was suggested to be the dominant group isolated from olive flounder suffering from streptococcosis. However, the isolates of Rh 1 group were not classified into the same species by the Api 20 Strep identification system. There was no peculiarity in drug resistance patterns of Rh I group isolates against 7 antibacterial agents. However, only 3 of 25 isolates showed oxytetracycline (OTC) resistance and OTC might be a useful chemotherapeutic agent in controlling the streptococcosis by strains of RA I group in olive flounder. Fish injected intraperitoneally with CFU of an isolate of Rh I and RA III group showed accumulative mortality for 20 days, respectively ( in control or Rh II). However luther comparative studies about differences in virulence between isolates are needed.