• Journal of Environmental Health Sciences
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• v.32 no.4 s.91
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• pp.336-341
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• 2006

The biological wastewater treatment system is known to have an important role in reducing the quantify of enteric virus in water environments. To clarify the roles of activated sludge microbes in decreasing the virus infectivity, the behavior of the virus in bacteria, protozoa, and metazoa was examined by pure or mixed culture system using poliovirus type 1(Lsc, 2ab strain). In the bacterial culture systems, the virus infectivity in the liquid phase decreased by a reversible adsorption of the virus to the bacteria or bacterial flocs. On the other hand, in the protozoa and the metazoa culture systems using T. pyriformis and P. erythrophthalma, respectively, with a variety of bacterial strains as prey, the main virus decrease mechanism of reversible adsorption in early stage was changed to irreversible predation, which was not eluted in this study. The virus decrease was more effective in the P. erythrophthalma culture system, which had high predation and floc forming abilities. However, in the mixed culture system of Z. ramigera and P. erythrophthalma, the more rapid reversible adsorption of virus to Z. ramigera flocs preceded the irreversible predation of P. erythrophthalma.

• Journal of Plant Biotechnology
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• v.30 no.2
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• pp.155-160
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• 2003

Grapevine leafroll-associated virus 3(GLRaV-3) is one of the most severe pathogens for viral diseases found in Korea. This study was conducted to establish the virus-free stock production system for the virus disease control. The effects of thermotherapy, merestem culture and chemotheratpy to eliminate the GLRaV-3 in gratevines were tested. Thermotherapy at 37$\pm$2$^{\circ}C$ for 6∼8 weeks combined with 0.5∼1.0mm size of meristem culture method was the most effective for virus elimination. Thermotherapy alone was not effective. In chemotheratpy, DHT and Amantadine (20, 40mg/L) treatment in medium was more effective than Ribavirin to eliminate the GLRaV-3 in grapevine. However, Ribavirin spraying to potted was not available for virus elimination. Therefore, virus-free stock production system using the thermotherapy combined with shoot apical meristem culture was the most effective in grapevine.

• Journal of Plant Biotechnology
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• v.37 no.4
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• pp.370-378
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• 2010

This review describes the stratagies of development of virus-resistant Alstroemeria plants using the genetic modification system. Despite of increasing of its importance in cut flower market, improvements of some horticultuirally important traits such as fragrance, long vase-life, virus resistance and tolerance against abiotic stresses are lack of the breeding program in Alstroemeria. Of these traits, virus-resistance is quite difficult to develop in Alstroemeria plants due to the limitations of genetic variation in the existed germplasm. To extend the genetic variation, plant biotechnological techniques such as genetic transformation and tissue culture should be combined to develop virus-resistant line in Alstroemeria. In this review, several strategies for the generation of virus-resistance by using natural resistance genes, pathogen-derived genes and other sources including pathogen-derived proteins, virus-specific antibodies and ribosome-inactivating proteins are presented. Also, brief histories of breeding, tissue culture, and transformation system in Alstroemeria plants are described to inderstand of the application of transgenic approach for the development of virus-resistance in Alstroemeria species.

• 한국균학회소식:학술대회논문집
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• 2015.11a
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• pp.37-37
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• 2015

dsRNA was found in malformed cultures of Lentinula edodes strain FMRI0339, one of the three most popular sawdust cultivated commercial strains of shiitake, and was also found in healthy-looking fruiting bodies and actively growing mycelia. Cloning of the partial genome of the dsRNA revealed the presence of the RdRp sequence of a novel L. edodes mycovirus (LeV), and sequence comparison of the cloned amplicon showed an identical sequence to known RdRp genes of LeV found in strain HKA. The meiotic stability of dsRNA was examined by measuring the ratio of the presence of dsRNA among sexual monokaryotic progeny. More than 40% of the monokaryotic progeny still contained the dsRNA, indicating the persistence of dsRNA during sexual reproduction. Comparing the mycelia growth of monokaryotic progeny suggested that, although variations in the growth rate existed among progeny and virus infection was observed in highly actively growing progeny, there appeared to be a tendency toward a lower frequency of virus incidence in actively growing progeny. This study attempted to cure the edible mushroom L. edodes strain FMRI0339 of the L. edodes mycovirus (LeV) in order to obtain an isogenic virus-free fungal strain as well as a virus-infected strain for comparison. Mycelial fragmentation, followed by being spread on a plate with serial dilutions resulted in a virus-free colony. Viral absence was confirmed with gel electrophoresis after dsRNA-specific virus purification, Northern blot analysis, and PCR using reverse transcriptase (RT-PCR). Once cured, all of fungal cultures remained virus-free over the next two years. Interestingly, the viral titer of LeV varied depending on the culture condition. The titer from the plate culture showed at least a 20-fold higher concentration than that grown in the liquid culture. However, the reduced virus titer in the liquid culture was recovered by transferring the mycelia to a plate containing the same medium. In addition, oxygen-depleted culture conditions resulted in a significant decrease of viral concentration, but not to the extent seen in the submerged liquid culture. Although no $discernable phenotypic changes in colony morphology were observed, virus-cured strains showed significantly higher growth rates and mycelial mass than virus-infected strains. We were also explored effects of LeV on fruiting body formation and mushroom yield. The fruiting body formation yield of virus-free L. edodes was larger than virus-infected L. edodes. These results indicate that LeV infection has a deleterious effect on mycelial growth and fruiting body formation. In addition, we have been investigated host-parasite interaction between L. edodes and its mycovirus interaction to study viral mechanism by establishment of proteomics.

• Journal of Microbiology and Biotechnology
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• v.21 no.1
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• pp.100-108
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• 2011

A method for the rapid detection and quantification of Newcastle disease virus (NDV) produced in an animal cell culture-based production system was developed to enhance the speed of the NDV vaccine manufacturing process. A SYBR Green I-based real-time RT-PCR was designed with a conventional, inexpensive RT-PCR kit targeting the F gene of the NDV LaSota strain. The method developed in this study was validated for specificity, accuracy, precision, linearity, limit of detection (LOD), limit of quantification (LOQ), and robustness. The validation results satisfied the predetermined acceptance criteria. The validated method was used to quantify virus samples produced in an animal cell culture-based production system. The method was able to quantify the NDV samples from mid- or late-production phases, but not effective on samples from the early-production phase. For comparison with other quantifiable methods, immunoblotting, plaque assay, and tissue culture infectious dose 50 ($TCID_{50}$) assay were also performed with the NDV samples. The results demonstrated that the real-time RT-PCR method is suitable for the rapid quantification of virus particles produced in an animal cell-culture-based production system irrespective of viral infectivity.

• Journal of Microbiology and Biotechnology
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• v.9 no.4
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• pp.386-392
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• 1999

A large-scale culture of hepatitis A virus in human diploid MRC-5 cells was conducted. In a roller bottle culture, the virus was grown to a maximum titer in 3 weeks after infection. Over 95％ of the cell-associated virus was excreted after culturing the infected cells in suspension media without fetal bovine serum for 3 days. The cultured virus was inactivated with formalin, concentrated by ultrafiltration, and partially purified by ultracentrifugation in a non-ionic gradient medium of Renocal. Two separate peak fractions showing high anti-HAY ELISA titer were pooled and about 40％ of HAV antigen was recovered by this purification procedure. Of the partially purified vaccine, the protein pattern in SDS-PAGE and immunogenicity in mice were compared with a commercial HAV vaccine. In SDS-PAGE, the purified vaccine in this study and the commercial vaccine showed almost the same protein pattern. The seroconversion rate of the purified vaccine in mice was not different from that of the commercial vaccine. Therefore, we could prepare a good grade of HAV vaccine by a simple purification procedure although the purification itself was not completed.

• The Journal of the Korean Society for Microbiology
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• v.16 no.1
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• pp.83-89
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• 1981

Japanese encephalitis has been prevalent for long time in the Far East and many patients have been reported in both South East and Mid-West Asia recently. Recently, vaccine was used in prevention of this viral disease of man which was derived from formalin inactivated virus inoculated into mouse brain, but live attenuated active vaccine for human is not developed yet. Author inoculated Japanese encephalitis virus into several cell culture strains for development of live attenuated encephalitis virus strain and the results were as follows: 1. Japanese encephalitis virus was inactivated rapidly in cell free medium at $36^{\circ}C$ and totally inactivated by 72 hours. 2. In growth curve of Japanese encephalitis virus in HeLa cell cultures, maximal multiplication of the virus was occured at 4th day and virus multiplication was continued for at least 12 days. 3. After succeeding passage of the virus in HeLa cell cultures and human esophagus epithelial cell cultures, infectivity of virus for mice was disappeared from 2nd passage in HeLa cell cultures and 3rd passage in esophagus epithelial cell cultures. 4. In inoculation to monkey kidney epithelial cells and chick embryo cell cultures, infectivity of the virus for mice was continued after 10th passages.

• KSBB Journal
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• v.13 no.3
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• pp.231-237
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• 1998

An attenuated Japanese encephalitis virus (JEV) clone SA-14-14-2(Vero) was obtained through successive adaptation of a primary cell adapted strain, SA-14-14-2(PDK) to Vero cell, a continuous line of monkey kidney cells. The virus titer reached above the 107 plaque forming unit (pfu) per mL of culture supernatant with 3 passages in Vero cells and was maintained close to this level in the further passages. The optimum temperature for the virus growth was $35^{\circ}C$. Growth pattern of the virus indicated that optimum time for the virus harvest is 4 days post infection and the virus accomplished rapid initial propagation even in medium containing no serum supplement. The roller bottle (RB) system and the spinner flask (SF) system using micro-carrier (Cytodex-1) for the JEV cultivation were explored. When RB, SF, and T-flask culture system were compared, there was no significant difference in virus yield. Furthermore, the results indicated that virus could be harvested multiple times from 3 days to 9 days post infection; neither severe cytopathic effect (CPE) in the infected cells nor the decrease in the titer was observed on duration of 9 days.

• Korean Journal of Plant Resources
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• v.23 no.3
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• pp.233-241
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• 2010

In vitro elimination of Sweet potato leaf curl virus (SPLCV) from infected sweet potato is difficult due to low number of virus-free plants obtained from meristem tip culture and long growth period required for the virus detection. In this study, efficient production of the SPLCV-free sweet potato by in vitro therapy coupled with a PCR assay for virus detection was investigated. Infected shoots cultured on Murashige and Skoog medium were treated at three different temperatures for 7 weeks followed by meristem tip culture on the medium with or without ribavirin at 50 mg/L. The regenerated plantlets were tested for virus infection by a PCR assay. The results showed that the both heat- and cold-treatments, and addition of the ribavirin did not have significant effect on efficiency of the virus elimination. The meristem size, however, greatly affected the survival rate. Meristems sized over 0.4 mm survived better than smaller ones (0.2-0.3 mm). The PCR assay was approved to be a rapid, sensitive and reliable for the SPLCV detection in regenerated plantlets. Therefore, combination of cultivating meristem tips sized 0.4-0.5 mm on the medium at $22^{\circ}C$ without ribavirin and detection of SPLCV in the regenerated plantlets by the PCR assay was an efficient system for the SPLCV elimination from infected sweet potato.

• Korean Journal of Microbiology
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• v.55 no.1
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• pp.25-32
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• 2019

Influenza A (H1N1) virus caused a worldwide pandemic in 2009-2010 and still remains in seasonal circulation. Continuous surveillance activities are encouraged in the post pandemic phase to watch over the trend of occurrence every year, this is better to be done by a rapid and sensitive method for its detection. This study was conducted to detect proportions of occurrence of influenza A virus (H1N1) in patients with influenza-like illness. Samples from 500 patients with influenza or influenza-like clinical presentation were tested by real-time reverse transcription polymerase chain reaction (RT-PCR) and virus tissue culture. Among the total 500 participants, 193 (38.6%) were females and 307 (61.4%) males. Seventy-one patients (14.2%) were positive for H1N1 virus infection with real-time RT-PCR while 52 (10.4%) were positive by tissue culture. Non-statistically significant relation was found between age and gender with the positivity of H1N1. Sensitivity and specificity of real-time RT-PCR was 98.08% and 95.54%, respectively, in comparison to virus isolation with accuracy 95.8%. This study showed that H1N1 virus was responsible for a good proportion of influenza during the post-pandemic period. Real-time RT-PCR provides rapidity and sensitivity for the detection of influenza A virus (H1N1) compared with virus isolation and thus it is recommended as a diagnostic tool.