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John Hwa Lee,Yeong Hwan Choi,Min Su Choi,Jin Hur 한국예방수의학회 2013 예방수의학회지 Vol.37 No.4
We previously developed a novel attenuated Salmonella Typhimurium (S.Typhimurium) △lon△cpxR vaccine. This study was conducted in order to examine whether this vaccine could effectively protect growing piglets against Salmonella infection. Pregnant sows in group A were primed and boosted with the vaccine, whereas pregnant sows in group B received sterile PBS-sucrose. After farrowing, newborn piglets in groups A and B were challenged with a wild type virulent S. Typhimurium at three weeks of age. During the study, serum IgG titers of piglets in group A were significantly higher than those of piglets in group B (P<0.001). In addition, clinical signs were observed in 5.9% of piglets in group A during the entire experimental period after the challenge, while diarrhea was observed in 81.6% of piglets in group B. These results indicate that vaccination of the pregnant sows resulted in effective protection in piglets against Salmonella infection.
Lee, John Hwa,Hur, Jin,Stein, Barry D. Elsevier 2008 The veterinary journal Vol.176 No.2
<P><B>Abstract</B></P><P>The aims of this study were: (1) to examine whether or not enterohemorrhagic <I>Escherichia coli</I> O26 and O111 (EHEC O26 and O111) are involved in neonatal calf diarrhea; (2) to determine the specific age periods at which the calves are vulnerable to these organisms, and (3) to reveal the biochemical, genetic and cytotoxic characteristics of the isolates. The study investigated the occurrence of EHEC O26 and O111 in calves associated with or without diarrhea. A total of 442 diarrheic and non-diarrheic young calves from 115 different farms were examined. Of the 257 calves with diarrhea, 37 (14.4%) and 32 (12.5%) tested positive for EHEC O26 and EHEC O111, respectively. Of the 185 non-diarrheic calves, 14 (7.6%) and 11 (5.9%) tested positive for EHEC O26 and EHEC O111, respectively. EHEC O26 and O111 were recovered from 14/69 (20%) and 11/69 (16%) diarrheic calves <2-weeks-old, respectively, and no EHEC O26 and O111 were detected in the non-diarrheic claves of this age group, suggesting that EHEC O26 and O111 are possible causes of the disease in infected neonatal calves. However, there were similar rates of occurrence in the diarrheic and non-diarrheic calves in the older animals (particularly, aged >10 weeks). PCR analysis showed that the isolates carried various virulence genes such as <I>Ehly</I>, <I>eae</I>, <I>stx1</I> and <I>stx2</I>, which highlight the potential importance of these attributes for the infection, colonization and the possible pathogenesis of calf diarrhea. Cytotoxicity analysis revealed that many of the EHEC isolates showed high cytotoxicity to Vero cells, re-emphasizing the potential for cattle being a direct source of EHEC infections in humans.</P>
Importance of Nucleotides Adjacent to the Core Region of Diphtheria tox Promoter/Operator
( John Hwa Lee ) 한국미생물 · 생명공학회 2002 Journal of microbiology and biotechnology Vol.12 No.4
Diphtheria toxin repressor (DtxR) binds to approximately 30 to 35-bp regions containing an interrupted 9-bp inverted repeat within a 19-bp core sequence. The core sequence is fairly conserved and critical for DtxR binding. The flanking regions that are consisted of 5 to 8 more of nucleotides from the core are also required for DtxR binding. The nucleotides in both flanking regions are A-T rich. To examine whether the A-T nucleotides in both flanking regions from the core have significant roles for DtxR binding, a DNA fragment was constructed based on the diphtheria tox promoter/operator, and DNA fragments with substitution of A and T nucleotides in the flanking regions to G and C were also constructed. To assess the effect of these substitutions on binding of DtxR and repressibility by DtxR, β-galactosidase activity from lacZ fused to the region was assessed. Gel mobility shift of the region by purified DtxR was also examined. The DNA fragments containing the mutations in the flanking regions still exhibited repression and mobility shift with DtxR. The core segment with the mutation is still, therefore, recognized by DtxR. Nonetheless, the results from the assays indicated that the substitution significantly decreased repression of the operator by DtxR in vivo under high-iron condition and decreased binding of DtxR to the operator. These results suggest that A and T nucleotides for both flanking regions are preferred for the binding of DtxR.
Short Communications : Efficacy of two virulence gene-deleted Salmonella
( John Hwa Lee ),( Yeong Hwan Choi ),( Min Su Choi ),( Jin Hur ) 한국예방수의학회 2013 예방수의학회지 Vol.37 No.4
We previously developed a novel attenuated Salmonella Typhimurium (S.Typhimurium) △lon△cpxR vaccine. This study was conducted in order to examine whether this vaccine could effectively protect growing piglets against Salmonella infection. Pregnant sows in group A were primed and boosted with the vaccine, whereas pregnant sows in group B received sterile PBS-sucrose. After farrowing, newborn piglets in groups A and B were challenged with a wild type virulent S. Typhimurium at three weeks of age. During the study, serum IgG titers of piglets in group A were significantly higher than those of piglets in group B (P<0.001). In addition, clinical signs were observed in 5.9% of piglets in group A during the entire experimental period after the challenge, while diarrhea was observed in 81.6% of piglets in group B. These results indicate that vaccination of the pregnant sows resulted in effective protection in piglets against Salmonella infection.
( John Hwa Lee ) 대한인수공통전염병학회 2018 창립총회 및 학술대회 초록집 Vol.2018 No.1
Human typhoid fever caused by Salmonella Typhi still poses a severe global disease burden in developing countries despite the availability of commercial vaccines. In this study, we constructed a non-living parenteral S. Typhi Ty21a vaccine candidate by employing a lambda (λ) phage-derived holin-endolysin system to efficiently construct bacterial ghosts. The lysis plasmid pJHL464, which contains an R lysis cassette that is stringently regulated by dual promoters, was introduced into an asd gene-deleted S. Typhi Ty21a strain. The in vitro expression of endolysin in the subsequent JOL1675 vaccine construct when grown under lysis inducible conditions was validated by immunoblotting. In scanning electron microscopy analysis, surface transmembrane tunnels and a collapsed body were visualized in the ghosts. Following 48 h of lysis, no viable JOL1675 cells were recovered, indicating that complete lysis of the cells was achieved. Subcutaneous immunization with the ghost significantly increased IgG and secretory IgA titers in mice during the observational period. Further, potent serum bactericidal activity (SBA) was exhibited by serum antibodies induced after JOL1675 immunization at 6 weeks post immunization in the presence of complement. The immunological data indicate that antigenic characteristics on the surface of resultant ghosts were minimally altered during the lysis process. Collectively, the incorporation of λ phage holin-endolysin-mediated lysis constructs into S. Typhi provides an efficient alternative strategy to develop a safe non-living typhoid vaccine candidate.