• Journal of fish pathology
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• v.7 no.1
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• pp.37-46
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• 1994

A total 103 strains of Edwardsiella tarda was isolated from eel culture-ponds and examined for drug resistance, distribution and transferabilities of R plasmids. The drugs used were lincomycin(LM), penicillin(PM), sulfamethazine(SF), sulfadimethoxine(SD), cephalosprin(CP), chloramphenicol(CH), streptomycin(SM), oxytetracycline(OT), ampicillin(AP), oxolinic acid(OA), kanamycin(KM), amikacin(AK), gentamicin(GM) and enrofloxacin(EF). Two strains were resistant to the all drugs used, and all isolates were multiply resistant to drugs(at least 8 among 14 drugs), mostly restricted to LM(103 strains), PM(103 strains), SD(103 strains), SF(103 strains), CP(102 strains), CM(101 strains), SM(100 strains), OT(94 strains), AM(92 strains), OA(80 strains), KM(60 strains), AK(30 strains), GM(19 strains) and EF(14 strains), in combination at high degree showing 34 different drug resistant patterns. The most frequently encountered patterns were LM PM SD SF CP CH SM OT AP OA KM(22 strains, 22.4%) followed by LM PM SD SF CP CH SM OT AP OT(12 strains, 11.7%). LM PM SD SF CP CH SM OT AP(7 strains, 6.8%), LM PM SD SF CP CH SM OT AP OA KM AK GM(6 strains, 5.8%) and LM PM SD SF CP CH SM OT AM OX KM AK GM(6 strains, 5.8%). Transfer experiment of drug resistance showed that of 103 resistant strains, 100 strains(97%) transferred part or all resistance to all drugs, indicating that most isolates carried conjugally transferrable R plasmids determining multiple drugs. The most frequently observed transferarble R plasmids were LM PM SD SF CP CH SM OT AP OA(10 strains), LM PM SD SF CP CH SM OT AP OX KM(7 strains) and LM PM SD SF CP CH AP OA(7 strains). These results sugested that eel culture-ponds were highly contaminated with different strains of Edwardsiella tarda, and that contaminated bacteria might be highly multiple resistant strains to drugs, carring transferable R plasmids.

• Journal of fish pathology
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• v.22 no.3
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• pp.253-262
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• 2009

We conducted bacteriological and histopathological analysis for olive flounder Paralichthys olivaceus after experimental infection with Streptococcus parauberis (FP2284) isolated from diseased olive flounder under different stress conditions. Experimental challenge was performed in healthy flounder (40.4 g in average body weight) by intraperitoneal (i.p.) injection with $2{\times}10^{8}$ CFU/fish under normal (no stress) or netting (for 2 min, twice/day) stress condition. The cumulative mortalities of no-stress and netting stress group were 70% and 95%, respectively. The most prevalent changes observed in experimentally infected flounder were darkness of skin and inflammation of the heart. Severe pericarditis, myocarditis and fibrosis were observed in the heart of the affected flounder. The results of viable counts showed the number of bacteria of the heart tissue was maintained over the $10^{4}$ CFU $g^{-1}$ heart for 3 weeks after inoculation. Histological lesions of the heart was more extensive and gradual decrease in bacterial numbers of heart tissue was delayed under stress condition.

• Journal of fish pathology
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• v.25 no.3
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• pp.151-158
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• 2012

Diseased eel (Anguilla bicolor) displayed severe hemorrhages in the gills, and congestion and swelling in the liver. During the epizootic, the water temperature was $28^{\circ}C$ and the morality rates were about 5%. No parasites were found on the gills and skin. Bacteria were not cultured from any internal organs using TSA or SS agar at $28^{\circ}C$ for 48 hrs. Histopathologically, the gills showed epithelial hyperplasia in the base of secondary gill lamellae and hemorrhages in the capillaries. Some cells in the proliferated interlamellar epithelia exhibited marginal hyperchromatosis. And severe vacuolated changes in the parenchymal cells and congestion in the central veins were observed in the liver. The specific amplicon (396 bp) was detected from gills and opercula of affected eel PCR using Anguillid herpesvirus-1 (AngHV-1) -specific primer sets HVAPOLVPSD (5-'GTG TCG GGC TTT GTG GTG C-3') and HVAPOLOOSN (5'-CAT GCC GGG AGT CTT TTT GAT-3'). Sequencing analysis of the amplicon demonstrated that this gene was 99% homologous to the AngHV-1 sequence deposited in GenBank. This is the first report of AngHV-1 outbreak in the farmed shortfin eels (A. bicolor) in Korea. When diseased fish were maintained for 10 days at water temperatures of $32^{\circ}C$ and $35^{\circ}C$, the cumulative mortalities were 100% and 10%, respectively. Even though the AngHV-1 genome in the gills from the eel kept at $35^{\circ}C$ was detected using PCR, the structure of gill filaments was similar with that of normal fish. Increasing the water temperature to $35^{\circ}C$ was an effective way to diminish the mortality of AngHV-1 affected eel.

• Journal of fish pathology
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• v.32 no.1
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• pp.37-43
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• 2019

Due to the rapid global expansion of aquaculture industry during the past decades, production volume of aquatic organisms has accordingly grown. This has been accompanied by a rise in the incidence of diseases in aquatic organisms, leading to an increased use of aquatic medicines. While aquatic medicines are essential for the prevention and treatment of diseases of aquatic organisms, misuse can cause evolution of antibiotic-resistant bacteria and pose serious problems to the safety and hygiene of fisheries products. In this study, we surveyed and compared, by analyzing data compiled by all national control centers for aquatic animal diseases in South Korea, to estimate the amount of aquatic medicines sold in different regions between 2012 through 2015. Since we also examined the marketing routes of aquatic medicines, this basic data can be utilized for policy implementation to improve drug safety. In the future research, however, it will be clearly necessary to estimate directly the amounts of drugs actually applied to aquaculture organisms. The current sales figure data should be helpful for preparation of an effective system for aquatic drug use management.

• Journal of fish pathology
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• v.28 no.2
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• pp.63-69
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• 2015

The biological characteristics of Bacillus sp.CPB-St as a probiotic strain to control fish streptococcosis was determined. Based on 16S rRNA sequencing, Bacillus sp.CPB-St was identified as Bacillus pumilus and named B. pumilus CPB-St (Abbreviated as CPB-St). Growth inhibitory activity of CPB-St against Streptococcus spp. was examined at three different incubation temperatures ($20^{\circ}C$, $25^{\circ}C$, and $30^{\circ}C$) and three culture media (NA, TSA, and BHIA) based on the diameter of inhibition zone. Its activity (inhibition zone of 11~29 mm) at $20^{\circ}C$ was higher than that (12~21 mm) at $30^{\circ}C$. Its activity (29 mm) in NA media was the same as that (29 mm) in TSA media. However, it was higher than that (22 mm) in BHIA media. The inhibitory activity of CPB-St against Streptococcus spp. was high at pH7. However, its activity was the same at salinity of 0.5% to 3%. CPB-St showed maximum growth after incubation at $25^{\circ}C$ for 48 h. To use CPB-St as probiotics, settlement studies in fish intestine and its efficacy through feeding are needed. CPB-St was highly resistant to gastric juice at pH4 and flounder's bile salt as well as deoxycholic acid at $300{\mu}g/ml$. CPB-St showed optimal viability in 1% NaCl. It showed similar growth in 0% to 7% NaCl. CPB-St could tolerate $-20^{\circ}C$ and $-70^{\circ}C$ for 45 min. There was no difference in the growth of the strain between room temperature and $4^{\circ}C$. Fish diet supplemented with CPB-St could be stored at low temperature without cell loss. Therefore, CPB-St might be used as probiotics to control streptococcosis of fish.

• Annals of Clinical Microbiology
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• v.18 no.2
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• pp.37-43
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• 2015

Background: Tuberculosis is globally the most important cause of death from single pathogen. Rapid and accurate identification of mycobacteria is essential for the control of tuberculosis. We evaluated a fluorescence in situ hybridization (FISH) method using peptide nucleic acid (PNA) probes for the differentiation of Mycobacterium tuberculosis complex (MTB) and nontuberculous mycobacteria (NTM) in direct smears of sputum specimens. Methods: The cross-reactivity of MTB- and NTM-specific PNA probes was examined with reference strains of M. tuberculosis ATCC 13950, Mycobacterium kansasii ATCC 12479, Mycobacterium fortuitum ATCC 6841, several clinical isolates of mycobacteria (Mycobacterium abscessus, Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium gordonae and Mycobacterium chelonae), and 11 frequently isolated respiratory bacterial species other than mycobacteria. A series of 128 sputa (89 MTB culture positive, 29 NTM culture positive, and 10 under treatment culture negative) with grades of trace to 4+ were used to evaluate the performance of the method. Results: The MTB- and NTM-specific PNA probes showed specific reactions with the reference strains of MTB and M. kansasii and clinical isolates of mycobacteria except M. fortuitum ATCC 6841, and no cross-reactivity with other tested bacteria. The PNA probe-based FISH assay for detection of MTB had a sensitivity and specificity of 100%, respectively. The sensitivity and specificity of the NTM-specific PNA probe was 100%. The smear grades of the PNA FISH test were same as with those of the fluorescence AFB stain in 2+ or higher grade. Conclusion: Detection and differentiation based on PNA FISH is sensitive and accurate for detecting mycobacteria and for differentiating MTB from NTM in clinical sputum smears.

• Journal of Food Hygiene and Safety
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• v.21 no.3
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• pp.189-195
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• 2006

This research was designed to evaluate the nutritional an microbiol quality assessment of Chungmukimbab purchased from market in Tongyeoung area. Contents of calories, calcium, iron, thiamin and riboflavin in ordinary kimbab and Chungmukimbab were lower than the recommended levels of Korean adult men. So, We suggested that a fruit, beverage and ffod stuff were supplemented to maintain nutritional balance. Total aerobic bacteria and coliform group of just prepared ordinary kimbab and Chungmukimbab samples from market were not significantly different, showing approximately $5.50{\pm}0.38 log_{10} CFU/g,\;2.10{\pm}0.47log_{10}MPN/100g$ in ordinary kimbab, $5.61{\pm}0.42log_{10}CFU/g,\;1.75{\pm}0.34 log_{10} MPN/100g$ in Chungmukimbab, respectively. Total aerobic bacteria of law ingredients of chungmukimbab sample were 3 to $4 log_{10}CFU/g$ in kimbab, seasoning squid and radish roots kimchi, 4 to $5 log_{10}CFU/g$ in boiled fish paste. The coliform groups were 1 to $2 log_{10}$ MPN/100 g in kimbab, seasoning squid and radish roots kimchi, 2 to $3 log_{10}$ MPN/100g in boiled fish paste. Detection rate of E. coli and Staphylococcus aureus counts were 10.0, 12.5% in Chungmu-kimbab, 15.0, 10.0% in seasoning squid, 0, 10.0% in radish roots kimchi respectively, not detected in boiled fish paste samples. During storage at $15^{\circ}C$ for 24 hours, total aerobic counts and coliform groups in ordinary kimbab and Chungmukimbab were increased by the 1.94, $0.97log_{10}CFU/g$, 0.60, 0.72 log10 MPN/100g respectively. Total aerobic counts of Chungmukimbab ingredients increased $0.83{\sim}l.33 log_{10}CFU/g$ at different time

• Korean Journal of Fisheries and Aquatic Sciences
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• v.28 no.6
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• pp.812-813
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• 1995

Lactococcal cells are nutritionally fastidious and thus, generally cultured either in milk or M17 medium (Terzaghi and Sandine, 1975). In this study, Lactococcus cremoris wild-type (KH) and its less­proteolytic mutant (KHA1) cells were grown on the M17 medium or with modified M17 medium by replicated parallel experiments. The modified M17 medium had the same composition as M17 medium, except that lactose was replaced by glucose. Analyses of culture-broth samples, in which the M17 and the modified M17 media were used, were conducted by high-performance liquid chromatography (HPLC). But, working with these media created noisy problems in analyses of samples. Therefore, a new semi-synthetic medium was developed on the basis of nutritional requirements (Morishita et al., 1981). The composition of the semi-synthetic medium determined on the basis of the nutritional requirements and the composition of milk, is presented in Table 1. The composition of M17 medium is also presented and compared in the table. L. cremoris KH and KHA1 cells were grown again on the new synthetic medium containing glucose or lactose. The broth samples were then drawn and analyzed by HPLC. Clearer separations of fermented products were achieved from the new medium than those with the M17 and the modified M17 media. In comparison with the M17 or the modified M17 media, growth on the new medium was good (Kim et al, 1993). Additional fermentations were also carried out at a controlled pH of 7.0, where enhanced growth of lactococcal cells was obtained. In the fermentations, samples were also analyzed for the concentrations of sugar and lactic acid. The results showed that the new synthetic medium was as good as or better than the M 17 and the modified M 17 media. This is because casein hydrolysate in the synthetic medium provided a ready supply of amino acids and peptides for L. cremoris KH and KHA1 cells. Lactic acid bacteria (LAB) including Lactococcal cells have been known to be an effective means of preserving foods, at the same time as giving particular tastes in fields of dairy products. LAB also have always occupied an important place in the technology of sea products, and marine LAB have known to be present in traditional fermented products (Ohhira et al, 1988). To apply the new synthetic medium to marine LAB, two different LAB were isolated from pickled anchovy and pollacks caviar and were grown on the new media in which various concentrations of NaCl $(3, 5, 7 and 10\%)$ added. They were also grown on the medium solution in natural seawater $(35\%o\;salinity)$ and on the solution of natural seawater itself, too. As seen in Fig. 1, Marine LAB were grown best on the synthetic medium solution in natural seawater and the higher concentrations of NaCl were added to the medium, the longer lag-phase of growth profile appeared. Marine LAB in natural seawater were not grown well. From these results, the synthetic medium seems good to cultivate cells which are essential to get salted fish aged. In this study, it showed that the new synthetic medium provided adequate nutrition for L. cremoris KH and KHA1 cells, which have been used as cheese starters (Stadhouders et al, 1988). Using this new medium, the acid production capability of starter cultures could be also measured quantitatively. Thus, this new medium was inferior to the M17 or the modified M17 medium in culturing the cheese starters and in measuring fermentation characteristics of the starter cells. Moreover, this new medium found to be good for selected and well-identified marine LAB which are used in rapid fermentations of low-salted fish.

• Journal of fish pathology
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• v.29 no.1
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• pp.35-43
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• 2016

In comparison to the numbers of such studies of fish, few studies have been carried out on the immunity, physiology and ecology of abalone, while studies on abalone disease are also extremely rare. Moreover, mass mortality of cultured abalone due to pathogenic bacteria has not been reported in the southern coast of Korea. However, Vibrio-like bacteria have been isolated from dead abalone, which indicates that a review is required in order to determine the cause of abalone mortality. Use of an antimicrobial agent to minimize the damage caused by disease in abalone farms is common, but the therapeutic effects are insignificant. Demand for probiotics has increased, but research on the development of probiotics for use in abalone culture is very rare. Therefore, the present study isolated KC16-2 from fermented kimchi soup and investigated the characteristics of the isolate as a candidate probiotic bacterium in abalone. KC16-2 was identified as Bacillus amyloliquefaciens (B. amyloliquefaciens KC16-2) based on its biochemical properties and 16S rRNA gene sequence. B. amyloliquefaciens KC16-2 showed inhibitory effects against the growth of various vibrios in vitro, and kept the numbers constant until four days after inoculation in marine water at a temperature of $15{\sim}25^{\circ}C$, indicating the possible use of KC16-2 as a probiotic, except in the winter. The growth of KC16-2 was inhibited by bile salt, but the numbers increased over time suggesting the bacteria were still alive in the abalone's digestive tract. Abalone fed with a diet including KC16-2 for 12 weeks showed good growth, but showed no significant differences from the control group. However, the mortality of the abalone supplied the probiotic diet was reduced to half that of the control group in a challenge test with Vibrio tubiashii. Therefore, we suggest that B. amyloliquefaciens KC16-2 could be used as a probiotic bacterium for control of the mortality of abalone caused by opportunistic pathogenic vibrios.

• Journal of Life Science
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• v.25 no.5
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• pp.568-576
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• 2015

The marine microalga Pavlova viridis can grow fast and has the ability to accumulate essential nutrients for culturing marine animals, such as EPA and DHA, and it has been used as food for raring larval fish and prawn. The symbiotic relationship between the flagellate microalga Pavlova viridis and its associated bacteria was investigated. An axenic culture of P. viridis was obtained by repeated treatment of the microalga with an antibiotic cocktail. The axenic status was confirmed after sub-culturing three times in a sterile f/2 medium without an antibiotic. The axenic alga was then co-inoculated with five bacteria, arbitrarily designated as I1–I5, isolated from the alga to test the growth promotion of the algae. All bacterial strains promoted the growth of P. viridis, and bacterial isolate I3 was the most effective among the five bacteria tested. The cell number of P. viridis in the co-culture with I3 was significantly higher than that of the control culture. A sequence analysis of the 16S rRNA gene isolated from I3 revealed a 97% nucleotide sequence similarity to that of Citrobacter sp. The growth of strain I3 was also significantly enhanced by co-culturing with P. viridis, indicating a symbiotic relationship between the microalga and its associated bacterium. The association between the microalga and bacterium was confirmed by scanning electron microscopy.