Phenotypic traits, virulence-associated gene profile and genetic relatedness of Edwardsiella tarda isolates from Japanese eel Anguilla japonica in Korea.

UNLABELLED: Edwardsiella tarda is the predominant bacterium in farm-cultured eel in Korea. Here, we evaluated the heterogeneity of 37 E. tarda isolates derived from Japanese eel with various origins (olive flounder, common carp and ornamental fish) between 2003 and 2010. Regardless of origins, the biochemical characteristics of E. tarda isolates were homogenous except hydrogen sulfide production, citrate utilization and mannitol fermentation. Based on the phylogenetic analysis of 16S rRNA, E. tarda isolates could be classified into two subgroups and displayed a close relation with Edwardsiella ictaluri and Edwardsiella hosinae lineages, suggesting that the subgroup I has been a predominant type in the Jeonnam and Jeonbuk provinces. I-CeuI-based pulsed-field gel electrophoresis (PFGE) typing showed that the isolates from Japanese eels belonged to 11 pulsotypes, indicating that the presence of highly genomic diversity. Additionally, two isolates, ET-060 and ET-191, showed a high frequency of virulence genes (100%) and caused 90% and 60% mortality in Japanese eel, respectively. This finding suggests a substantial congruence of virulence gene profiles and pathogenicity. Our results demonstrate that the intraspecific diversity within E. tarda strains from Japanese eel has been in prior existence. SIGNIFICANCE AND IMPACT OF THE STUDY: Based on the biochemical characteristics, the phylogenetic property of the 16S rRNA gene and PFGE types of Edwardsiella tarda, we could identify the intraspecific diversity of isolates from Japanese eel, Anguilla japonica in Korea. In addition, this study describes the strong congruence of virulence-related genes and pathogenicity, suggesting that the virulence profile may be useful tool for prediction of pathogenicity.

Development of a stabilizer for lyophilization of an attenuated duck viral hepatitis vaccine.

The live attenuated vaccine against duck viral hepatitis currently available in Korea requires special freezers for storage and transportation with extra costs involved. The development of a lyophilization stabilizer for live attenuated duck viral hepatitis virus (DHV) vaccines, therefore, has been highly recommended for the wider application of the vaccines. Four conventional vaccine stabilizer formulations containing a disaccharide, such as lactose, trehalose, or sucrose, and new formulations containing sorbitol were tested for their efficacy in stabilizing a new attenuated DHV type 3 vaccine candidate under different storage temperatures, 4 and 37 degrees C. The vaccine virus and each stabilizer formulation were combined and submitted to lyophilization and the viability of the virus was measured in 7-d-old specific-pathogen-free chicken embryos by determining the 50% egg lethal dose. Stabilizer formulations containing 2, 4, or 8% sorbitol preserved the viability of the vaccine virus much better than the other stabilizer formulations and 2% sorbitol was the optimal concentration in a standard stabilizing buffer, phosphate glutamate gelatin (0.0038 M KH2PO4, 0.0071 M K2HPO4, 0.0049 M monosodium L-glutamate, and 0.5% gelatin). The results demonstrate that the stabilizer formulation containing 2% sorbitol and 0.5% gelatin can be used for convenient storage and transportation of live DHV vaccines.

The susceptibility of magpies to a highly pathogenic avian influenza virus subtype H5N1.

Korean wild magpies (Pica pica sericea) were intranasally inoculated with highly pathogenic avian influenza (A/Chicken/Korea/ES/03 virus) (H5N1), which was classified as clade 2.5. We estimated viral replication, death after infection, and histology and immunohistochemistry. This species was highly susceptible to severe infection; 100% of birds died within 5 to 8 d. The virus was detected from oropharyngeal (1 to 5 d postinfection) and cloacal (3 to 5 d postinfection) swabs from infected magpies. At necropsy, the prominent lesions were coalescing necrosis of the pancreas with enlargement of livers and spleens. Microscopically, pancreas, brain, heart, adrenal gland, and kidney were most consistently affected with necrotic and inflammatory changes, and viral antigen was frequently demonstrated in the parenchyma of these organs. As a result, Korean wild magpies were very susceptible to avian influenza (H5N1) virus.

The hydrodynamic filter separator for removal of urban storm runoff.

The majority of storm runoff pollution is trapped in particles smaller than 100 microm in diameter. Solid particles smaller than 100 microm in diameter are not easily separated by conventional types of hydrodynamic separator, and remain in suspension in overflow. To overcome this problem, a HDFS (hydrodynamic filter separator) has been developed for treatment of the microparticles in urban storm runoff. We conducted a laboratory scale study on treatable potential of microparticles using HDFS that combined HDS with perlite filter. To determine the efficiency for various operation conditions, a series of experiments was performed with different solids concentrations and surface loading rate. The operation ranges of surface loading rates were 100 to 2,800 m3/m2/day, and influent solids concentrations were varied from 800 to 1,900 mg/L. Also, the particle size distribution was monitored to investigate the effects of surface loading rates on the particle size. Results indicated that the HDFS-inside type showed greater efficiency at solids separation than the other type.

Highly pathogenic avian influenza in magpies (Pica pica sericea) in South Korea.

Highly pathogenic avian influenza (HPAI) is an extremely infectious, systemic viral disease of birds that produces high mortality and morbidity. HPAI was diagnosed in the three dead magpies (Pica pica sericea) submitted to the National Veterinary Research and Quarantine Service. At necropsy, the prominent lesions were multifocal or coalescing necrosis of the pancreas with enlargement of the livers and spleens. Microscopically, there were severely necrotizing pancreatitis and lymphocytic meningoencephalitis. Influenza viral antigen was also detected in areas closely associated with histologic lesions. Avian influenza virus was isolated from cecal tonsils and feces of the magpies. The isolated virus was identified as a highly pathogenic H5N1, with hemagglutinin proteolytic cleavage site deduced amino acid sequence of QREKRKKR/GLFGAIAG. To determine the pathogenicity of the isolate, eight 6-wk-old specific-pathogen-free chickens were inoculated intravenously with the virus, and all birds died within 24 hr after inoculation. This is the first report of HPAI in magpies.

Recent Korean isolates of duck hepatitis virus reveal the presence of a new geno- and serotype when compared to duck hepatitis virus type 1 type strains.

Duck hepatitis was first reported in 1985 in Korea. The complete nucleotide sequence of two past Korean isolates, DHV-HS and DHV-HSS, isolated in 1994 and 1995, and four recent Korean isolates, AP-03337, AP-04009, AP-04114 and AP-04203 isolated in 2003 and 2004, were determined. Phylogenetic analysis using the 3D protein sequence confirmed that the previously characterized duck hepatitis virus type 1 strains and the six Korean isolates described here constitute a monophyletic group and form two clades/genotypes in which all except the four recent Korean isolates form one group (A) and the recent Korean isolates of 2003 and 2004 constitute a second group (B). Phylogenetic analysis of the VP1 protein supported the division into two different groups. Antisera raised against viruses of group A showed significant neutralizing cross-reaction against a member of the same genotype but not to a strain of group B and vice versa. These results demonstrated that the two genotypes also could be regarded as two different serotypes.