It is commonly believed in the Western World that the more severe forms of gastroduodenal diseases like peptic ulcer are associated with infection by specific Helicobacter pylori strains classified as type I being considered to be more virulent than type II strains. However, in Korea, most of H. pylori isolates belong to type 1 strains regardless of virulence. Type I H. pylori strains differ from type II strains by the presence of the cag pathogenicity island (cag PAI) composed of a block of genes. In this study, the nucleotide sequence of cag PAI of the H. pylori Korean strain 51 was determined and compared with those of strains 26695 and J99 to assess the structural variation in the region and to evaluate its implication in the virulence of the H. pylori. The cag PAI of H. pylori strain 51 was smaller in size and in the number of constituting ORFs in comparison with 26695 and J99 strains. Although many cag orthologues were nearly identical one another with the similarity of 90% or more at the nucleotide and amino acid levels, there were some remarkable and significant differences in several cag genes among the three cag PAIs. Surprisingly, the percent similarities at amino acid level were lower than those at nucleotide level in one third of the ORFs. The two genes (cag7 and cagA) of strain 51 differed in sizes and deduced amino acid sequences from the corresponding genes of the other two strains. When comparing cagA ORF of H. pylori strain 51 with that of 8 non-Korean strains, phylogenetic tree revealed that the strain 51 formed a separate branch with the most far distances from the other strains except for a Japanese strain. The Cag7 protein of, strain 51 had a deletion in the repeat region II, suggesting a major change in the conformation and function of the protein.
Amino Acid Sequence ; Animals ; Asian Continental Ancestry Group ; Base Sequence ; Ecthyma, Contagious ; Genetic Variation ; Genomic Islands* ; Helicobacter pylori* ; Helicobacter* ; Humans ; Korea ; Open Reading Frames ; Peptic Ulcer ; Virulence* ; Western World

To investigate the seroprevalence of the Orientia tsutsugamushi infection of Apodemus peninsulae and genomic variations in O. tsutsugamushi isolates, 246 A. peninsulae were trapped in 14 mountainous areas approximately 500 meter above sea level in Korea during the period of 1997 and 2000. Seropositive rate of O. tsutsugamushi among A. peninsulae was 31.8% in Kyunggi, 8.2% in Chunbuk and 7.1% in Kangwon provinces by microimmunofluorescent test. The 56 kDa protein gene was amplified by PCR in the spleens of seropositive A. peninsulae. Two amplicons from seropositive A. peninsulae were sequenced and their phylogeny was analysed on the basis of sequence homology. The 56 kDa genes of A. peninsulae 98-12 strain and A. peninsulae 98-16 strain showed 98.7% nucleotide homology and 96.6% amino acid similarity. A. peninsulae 98-12 and A. peninsulae 98-16 strain were related to Kuroki, Boryong and Karp strains showing 93.3~92.2%, and 87.1~84.6% homologies in nucleotide and amino acids levels, respectively. In the phylogenetic analysis, A. peninsulae 98-12 and A. peninsulae 98-16 strain formed a distinct group with Boryong, Kuroki and Nishino strains and were clearly distinguished from other genetic groups. The results suggest that A. peninsulae might be an important reservoir of O. tsutsugamushi in Korea.
Amino Acids ; Animals ; Chungcheongnam-do ; Gangwon-do ; Gyeonggi-do ; Korea* ; Murinae* ; Orientia tsutsugamushi* ; Phylogeny ; Polymerase Chain Reaction ; Sequence Homology ; Seroepidemiologic Studies* ; Spleen

Coxiella burnetii is the etiological agent of Q fever, that may occur either acutely or the chronically. To understand the seroepidemiological patterns of C. burnetii infection in Korea, we examined a total of 3,178 sera from patients with acute febrile episodes by using indirect immunofluorescence assay (IFA) for detectable antibodies to C. burnetii and other eight rickettsial antigens. The IFA seropositivity>or=1:20 for C. burnetii phase II was 11.5% (368 out of 3,178 sera). The co-existence of antibodies to other rickettsial antigens was found in 216 out of the 368 positive sera. Thirty-seven point five percent (n=138) had antibodies to R. tsutsugamushi (cutoff>or=1:20), 16% (n=59) to Ehrlichia sennetsu, 14.9% (n=55) to Rickettsia typhi, 13.5% (n=50) to R. akari, 11.4% (n=42) to R. japonica, 8.9% (n=33) to R. prowazekii, 7.6% (n=28) to R. sibirica, and 6.7% (n=25) to R. conorii by IFA, respectively. These results are consistent with previous reports documenting diverse serum cross-reactivity in chronic Q fever. Therefore we excluded the samples that reacted to other rickettsial antigens at same or higher titers than to C. burnetii, resulting in the seropositive rate of 4.1%. The serological prevalence was 2% (n=64) when the conventional cut-off titer of 1:80 was used. Our results suggest that infections with C. burnetii are more prevalent than expected previously and should be differentially diagnosised for febrile illness occurring after exposure to ticks or other vectors.
Antibodies ; Coxiella burnetii* ; Coxiella* ; Diagnosis ; Fluorescent Antibody Technique, Indirect ; Humans ; Korea ; Neorickettsia sennetsu ; Prevalence ; Q Fever ; Rickettsia ; Rickettsia typhi ; Seroepidemiologic Studies* ; Ticks

To study the correlation of human papillomavirus (HPV) infection with clinical stage in cervical abnormalities, 17 cases of normal cervical tissue and 69 cases of abnormal cervical tissue (cervical intraepithelial neoplasia and invasive cervical cancer) was examined by PCR with HPV-specific consensus primers. One case (5.9%) of normal cervical tissue and 42 cases (60.9%) of abnormal cervical tissues harbored HPV. To investigate the integration of HPV genome in 24 cases of HPV 16 positive cervical cancer, E2 gene of HPV 16 was amplified. Integration of HPV 16 was found in 7 cases (29.2%) with E2 disruption. All samples with E2 disruption were from invasive cervical cancer. A multiplex PCR for the mapping of integrated HPV 16 genome with an anchor primer and indicator primers showed that 11 cases (45.8%) were disrupted somewhere in HPV genome but E6, E7, and LCR regions were conserved in all cases. Seven types of integrated HPV genome from long- (7,062 bp) to short-conserved type (3,204 bp) with various deletions were detected by the multiplex PCR. These results show that integration can be detected more accurately by multiplex PCR than by E2 PCR, and E2 disruption is not a critical event of integration
Consensus ; Genome ; Human papillomavirus 16 ; Humans* ; Multiplex Polymerase Chain Reaction ; Polymerase Chain Reaction ; Uterine Cervical Neoplasms*

Considerable effort has been directed at understanding the structure and function of HIV-1 envelope glycoproteins. It has been difficult to characterize HIV-1 envelope glycoproteins due to the limited availability of these proteins from virus particles or infected cells. To facilitate the structural and functional analysis of HIV-1 envelope glycoproteins, recombinant baculoviruses were generated to express wild type or mutant HIV-1 envelope glycoproteins. The gp160 precursor protein as well as the gp120 glycoprotein were detected in the cell infected with recombinant BacENVw.t containing wild type HIV-1 envelope gene. In the insect cells infected with recombinant BacENVc with mutations at the cleavage site of gp160, a precursor form of envelope glycoprotein was produced, but not secreted into the culture medium. However, the insect cells infected with recombinant BacENVc/t containing both mutations at the cleavage site and membrane spanning region produced mutant envelope glycoproteins that were efficiently secreted into the culture medium in the form of precursor. Therefore the recombinant HIV-1 envelope glycoproteins produced in this system would be useful as immunogens in the development of a vaccine against AIDS.
Baculoviridae* ; Glycoproteins* ; HIV* ; HIV-1* ; Humans* ; Insects ; Membranes ; Staphylococcal Protein A ; Virion

Envelope glycoprotein 1 (G1) and glycoprotein 2 (G2) of Hantaan (HTN) virus are believed to be major viral antigens that can induce neutralizing immunity against HTN virus infection. The purpose of this study is to clone and express G1 gene in an E. coli expression system. The truncated G1 gene (amino acid residues 35 to 123) of the HTN virus strain 76-118 was amplified by polymerase chain reaction (PCR). The 0.28 kb PCR product was cloned into pCR2.1 vector and named as pCGS1. The truncated G1 gene was excised from the pCGS1 and subcloned into the BamHI and SalI sites of pGEX-4T-2 and named pGGS1. The nucleotide sequence of the 0.28 kb truncated G1 gene was determined. It is revealed four non-silent nucleotide substitutions between the published sequence of strain HTN virus strain 76-118 and our stock of HTN virus strain 76-118 (passaged several times in our laboratory). The first G1 mutation was found to constitute an A to G nucleotide substitution, giving raise to an asparagine to serine mutation at residue 64. The second G1 mutation was found to constitute an A to C nucleotide substitution, giving raise to an lysine to threonine mutation at residue 112. The third G1 mutation was found to constitute an A to C nucleotide substitution, giving raise to an lysine to threonine mutation at residue 112. The fourth G1 mutation was found to constitute an G to A nucleotide substitution, giving raise to an glutamic acid to lysine mutation at residue 117. The truncated G1 gene was expressed as a 37 kDa protein fused to glutathione-S-transferase (GST). The GST fusion protein was purified by Glutathione Sepharose 4B affinity chromatography and reacted with the sera from patients of hemorrhage fever with renal syndrome (HFRS). One of 12 serum samples from HFRS patients was reactive with the 37 kDa fusion protein strongly. Three sera reacted moderately with the fusion protein. Six sera reacted only weakly with the protein, while remaing two were non-reactive. Control sera from patients with scrub typhus leptospirosis, or negative HFRS did not react with the recombinant fusion protein.
Antigens, Viral ; Asparagine ; Base Sequence ; Blotting, Western ; Chromatography, Affinity ; Clone Cells ; Fever ; Glutamic Acid ; Glutathione ; Glycoproteins* ; Hantaan virus* ; Hemorrhage ; Hemorrhagic Fever with Renal Syndrome ; Humans ; Leptospirosis ; Lysine ; Polymerase Chain Reaction ; Scrub Typhus ; Sepharose ; Serine ; Threonine

Large numbers of reports have shown that thermal injury (TI) causes a wide spectrum of defects in immune response that lead to a high susceptibility to various opportunistic infections. However, it is still a matter of debate whether TI induces Th2 polarization or global impairment in Th1/Th2 response. In this study, TI in a mouse model was induced by exposing shaved dorsal skin to boiling water and cytokine production was analyzed. At day 2 of injury, whole spleen cells and T cells were collected and then stimulated with an anti-CD3 antibody. The levels of cytokine secretion were determined by cytokine ELISA. Production of IFNgamma and IL 4 by whole spleen cells from injured mice were concurrently decreased when compared to those from sham-injured controls. Proportional changes in T, B, and T-subset cells were not accompanied. Using purified T cells devoid of accessory cells (AC), it was shown that those defects resulted primarily from lowered T cell potentials. By using mixed cultures of sham T and TI-AC and vice versa, it was revealed that AC also acted as inhibitor cells in IFNgamma and IL 4 production in less extent. Blockade of glucocorticoid signals rendered the T cells partially resistant to TI-induced inhibition in IFNgamma and but not IL 4 production. These results clearly demonstrate that TI induces overall suppression in Th1 and Th2 response through T cell dysfunction together with the inhibition of AC activity, and that reduction in only IFNgamma but not IL 4, production may be caused, in part, by corticosteroid hormone that is secreted prominently during trauma.
Animals ; Enzyme-Linked Immunosorbent Assay ; Mice ; Opportunistic Infections ; Skin ; Spleen ; T-Lymphocytes ; Water

Although Mycobacterium marinum is closely related to M. tuberculosis H37Rv (M. tbc) genomically, clinical outcome of human infection is quite different. The role of the host macrophage in determining differential pathologic responses was analyzed using an in vitro model of macrophage infection. By using subtractive hybridization, thirty-two differentially expressed genes were identified in the monocytic cell line U937 infected with M. tbc or M. marinum. Among them, IL-8 mRNA expression was more prominent in the M. tbc-infected U937 cells by Northern hybridization than in those infected with M. marinum. The IL-8 production was significantly lower in M. marinum-infected U937 or monocytes when compared with those infected by other strains of mycobacteria, such as M. tuberculosis H37Ra, M. bovis BCG or M. smegmatis. To identify possible mechanisms underlying these differences, changes in the expression of molecules such as nuclear factor-kappaB (NF-kappaB) involved in the signaling pathway activated by mycobacteria were assessed. U937 cells infected with M. tbc showed a significant degradation of IkBa proteins compared with M. marinum-infected U937 cells. Collectively, these results implicate distinct differences in IL-8 production in human macrophages infected with M. tbc or M. marinum, and suggest important role of IL-8 in the immunopathogenesis of tuberculosis.
Cell Line* ; Humans ; Interleukin-8* ; Macrophages ; Monocytes ; Mycobacterium bovis ; Mycobacterium marinum* ; Mycobacterium tuberculosis* ; Mycobacterium* ; RNA, Messenger ; Tuberculosis ; U937 Cells

Understanding human immune responses in chronic refractory tuberculosis (CRTB) is important for developing immunotherapy against the disease. The aim of this study was to examine cytokine responses [interferon (IFN)-gamma, tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, and IL-10] by peripheral blood mononuclear cells (PBMCs) in CRTB patients after in vitro stimulation with the 30-kDa or purified protein derivative (PPD) antigen (Ag). Most of the CRTB cases were multidrug-resistant (MDR) TB. The results were compared with those from early TB (E-TB) patients and healthy tuberculin reactors (HTR). IFN-gamma production was significantly depressed in both CRTB and E-TB groups compared with HTR. In response to the 30-kDa Ag, TNF-alpha levels were significantly depressed only in CRTB patients, while greatly increased in E-TB patients. In addition, IL-10 production was significantly increased in E-TB patients, and PBMC from both E-TB and CRTB patients secreted more IL-6 than HTR. IL-10 neutralization significantly increased TNF-alpha levels, whereas anti-TNF-alpha did not alter IL-10 induction significantly in PBMC from HTR and CRTB patients. Our findings suggest that CRTB patients have depression in both IFN-gamma and TNF-alpha reponses, which might play important roles during chronic M. tuberculosis infection.
Depression* ; Humans ; Immunotherapy ; Interferon-gamma* ; Interleukin-10 ; Interleukin-12 ; Interleukin-6 ; Interleukins ; Mycobacterium tuberculosis ; Tuberculin ; Tuberculosis* ; Tuberculosis, Multidrug-Resistant ; Tumor Necrosis Factor-alpha*

Candida albicans is a commensal yeast normally present in small numbers as normal oral flora. In a certain condition, however, the yeast may proliferate and/or become invasive resulting in oral candidiasis such as denture stomatitis, and may even cause life-threatening systemic candidiasis. The present study was undertaken to test whether polyphosphate (polyP), which has been shown to be a strong antibacterial agent against a variety of oral pathogens, has antifungal effect on C. albicans. C. albicans ATCC 90027 was grown in Sabouraud-Dextrose broth with or without polyP. Anti-C. albicans activity of polyPs with various chain lengths was determined by measuring the growth of candidal cells at 540 nm. polyPs with chain length of 3 (polyP3) or higher effectively inhibited the candidal growth when added at the very beginning of the culture, whereas orthophosphate and pyrophosphate failed to do so. At the concentration of 0.05 percent, all the polyPs tested inhibited candidal growth. The effect of polyP65 that showed stronger anti-candidal effect than others at the concentrations tested and of Calgon (hexametapolyphosphate, practical grade) was further examined. The concentration of 0.03 percent was enough for polyP65 and Calgon to suppress candidal growth throughout the 48-h incubation. PolyP65 added to the growing C. albicans at its exponential phase was as much effective in inhibiting the candidal growth as added at the very beginning of the culture. It was found that 93.8 and 96.9 percent of the yeast cells lost their viability when polyP65 was added to growing C. albicans at the concentrations of 0.03 and 0.05 percent, respectively. Intracellular nucleotide release from the candidal cells incubated with polyP65 was only slightly increased and the nucleotide release was not reversed by the addition of divalent metal ions like Mg++ and Ca++. Under the transmission electron microscope, although the majority of growing C. albicans cells appeared to be atypical in their shape in the presence of polyP65, only a small number of the cells were observed to be lysed. The overall results suggest that polyP has a strong fungicidal activity against C. albicans, in which chelation-mediated cell lysis may not play the major role, but other novel mechanisms that possibly affect the viability of the yeast may be involved. Since polyP also has a strong antibacterial effect on oral pathogens, it may well be used for the prevention and treatment of a variety of oral diseases caused by a wide spectrum of microorganisms including C. albicans.
Candida albicans ; Candidiasis ; Candidiasis, Oral ; Ions ; Phosphates ; Polyps ; Stomatitis, Denture ; Yeasts