The ability to introduce recombinant DNA molecules back into mycobacteria would greatly increase the potential of molecular genetic approaches for the study of mycobacteria as well as for the use in clinical purposes. We have initiated the construction of vectors that facilitates the introduction of recombinant DNA into mycobacteria. The vector was designed to contain replicons for multiplication in mycobacteria and Escherichia coli, a promoter for gene expression, a drug resistant gene for selecting transformants, and a few restriction enzyme sites for convenient cloning. Constructed Mycobacterium-E. coli shuttle vector named p YMC (hsp60) was shown to transform M. smegmatis at high efficiency and maintain plasmid at stable level. The ability of the vector to express cloned foreign gene was also monitored by measuring the expressed level of luciferase gene which was used as a reporter. High level of luciferase activity in M. smegmatis with pYMC (hsp60:luc) was detected confirming successful construction of Mycobacterium-E. coli shuttle vector.
Clone Cells ; Cloning, Organism ; DNA, Recombinant ; Escherichia coli* ; Escherichia* ; Gene Expression ; Genetic Vectors* ; Luciferases ; Molecular Biology ; Mycobacterium* ; Plasmids ; Replicon

The 38 kDa protein of Mycobacterium tuberculosis, which was known previously as antigen 5, has been extensively used in the serodiagnosis of tuberculosis. In an attempt to develop and evaluate a serodiagnostic test using the antigen, we expressed the 38 kDa protein in BCG and its seroreactivity was compared to that expressed in Escherichia coli. The coding region of the 38 kDa protein was amplified by PCR, and the gene was cloned into a Mycobacterium-E. coli shuttle expression vector pYMC-his and pQE30 expression vector and expressed in BCG and E. coli, respectively. Both recombinant 38 kDa proteins showed strong seroreactivity against pooled serum from tuberculosis patients. There was no significant difference in seroreactivity between the two recombinant antigens in sera from the far advanced tuberculosis patients. However, of 25 tuberculosis patients graded as ""minimal"" by chest X-ray, 5 (20.0%) were seropositive by r38 kDa expressed in E. coli, while 8 (32.0%) by that expressed in BCG. Likewise, higher seroreactivity by r38 kDa expressed in BCG was found in sera from the moderately advanced tuberculosis. This study thus indicates that the recombinant 38 kDa expressed in BCG is more effective than that expressed in E. coli in detecting antibodies to the native 38 kDa protein of M. tuberculosis in sera from minimally affected tuberculosis patients.
Antibodies ; Clinical Coding ; Clone Cells ; Escherichia coli ; Humans ; Mycobacterium bovis* ; Mycobacterium tuberculosis* ; Mycobacterium* ; Polymerase Chain Reaction ; Serologic Tests* ; Thorax ; Tuberculosis*

No Abstract Available.
Brucella abortus* ; Brucella* ; Immunity, Humoral* ; Jeju-do* ; Recombinant Proteins*

Direct surgical repair of complex intracranial vascular lesions is difficult. Sometimes the neurosurgery is performed under circulatory arrest, profound hypothermia and barbiturates cerebral protection. Total ischemia is tolerated for 30~60 minutes because oxygen requirements of the brain decrease exponentially as body temperature is lowered. We experienced that this technique was successfully used for inoperable basilar artery aneurysm. We reviewed the surgical and anesthetic considerations of basilar artery aneurysm.
Aneurysm* ; Barbiturates ; Body Temperature ; Brain ; Hypothermia* ; Intracranial Aneurysm ; Ischemia ; Neurosurgery ; Oxygen

As an effort to develop a rapid and sensitive diagnostic test, we produced previously a monoclonal antibody (MAb) specific to the lipoarabinomannan (LAM) antigen and used in a sandwich ELISA for detection of mycobacterial antigens in sputum. In this study, we attempted to improve the antigen detection assay by combination of af5nity-purified antibodies against Mycobacterium tuberculosis soluble antigen and anti-LAM MAb. With the new assay, the LAM antigen was detectable as low as 2 ng/ml, and none of 10 gram-negative and gram-positive organisms gave significant absorbance, thus indicating the specific detection of mycobacterial antigens. Sputum samples from 62 patients who were suspected having tuberculosis and from 37 healthy controls were examined. The sensitivity of the antigen detection assay ranged from 0% in the 1+ culture group to 78.8% in the 3+ culture group. When the results were combined, 15 of 24 culture-positive samples were antigen-positive, thus giving an overall sensitivity of 62.5%. The overall specificity was 96.0%, when all the culture-negative samples were combined. The results thus demonstrate that the antigen detection assay can provide a rapid supplemental information for the diagnosis of pulmonary diagnosis.
Antibodies ; Diagnosis* ; Diagnostic Tests, Routine ; Enzyme-Linked Immunosorbent Assay ; Humans ; Mycobacterium tuberculosis* ; Mycobacterium* ; Sensitivity and Specificity ; Sputum* ; Tuberculosis ; Tuberculosis, Pulmonary*

PURPOSE: To identify MR predictors of bone cement leakage in patients receiving percutaneous vertebroplasty. MATERIALS AND METHODS: Percutaneous vertebroplasties were performed in 45 vertebras (T7; one, T8; two, T10; two, T11; two, T12; eight, L1; fifteen, L2; eight, L3; five, L4; two) in 35 patients (age 52-83). The procedure was performed using an 11 G Jamshidi needle, which was inserted into the target by the bipedicular approach. Kyphoplasty, unilateral pedicular approach and extrapedicular approach cases were excluded. Shortly after the procedure, all patients underwent a noncontrast CT covering the vertebroplasty sites. A retrospective study was performed to determine whether cement leakage is related to any of following MR findings: presence of cortical disruption of the vertebral body, severity of body compression (proportion of abnormal to normal vertebral body volumes), bone cement amount, bone cement amount/severity of body compression ratio, proportion of low-signal area in a vertebral body on T1 weighted image, presence of either vacuum or cystic portion below a linear dark signal in a fractured vertebra, and the location of dark signal intensity line in a vertebral body. Logistic discrimination model stepwise method was used in the statistical analysis. RESULTS: On post-vertebroplasty CT scan, bone cement leakage was detected in or around 29 vertebrae (64%), including 11 vertebrae (24%) where leakage was found in the epidural space or radial vein. No patients displayed any neurological symptoms or signs. The most frequent site of bone cement leakage was the anterior external vertebral venous plexus (49%). Endplate cortical bone disruption was related to an increased risk of intervertebral bone cement leakage (p<0.05). Bone cement leakage tended to occur less frequently when there is a vacuum or cystic change below the dark linear signal intensity in a fractured vertebra (p<0.05). No other MR findings showed a statistically significant correlation with bone cement leakage. CONCLUSION: On pre-vertebroplasty MR imaging, vertebral endplate cortical bone disruption and vacuum or cystic change below dark linear signal intensity in fractured vertebra showed a significant correlation with bone cement leakage.
Bone Cements ; Discrimination (Psychology) ; Epidural Space ; Humans ; Kyphoplasty ; Magnetic Resonance Imaging ; Needles ; Retrospective Studies ; Spine ; Tomography, X-Ray Computed ; Vacuum ; Veins ; Vertebroplasty*

PURPOSE: We assessed the efficacy of radical prostatectomy (RP) in prostate cancer patients with preoperative prostate-specific antigen (PSA) levels > or = 20 ng/ml and no distant metastases. MATERIALS AND METHODS: The records of 132 prostate cancer patients undergoing RP with preoperative PSA levels > or =20 ng/ml and no distant metastases were reviewed. Sixty-six patients received preoperative neoadjuvant hormonal therapy. Pathologic and clinical outcomes were compared between the groups with PSA of 20-40 ng/ml and > or =40 ng/ml. RESULTS: There were no statistical differences in age, prostate volume, or the frequency of neoadjuvant hormonal therapy between the two groups. The PSA > or =40 ng/ml group had a higher RP Gleason score, tumor stage, and extracapsular extension. After a mean follow-up of 47.0 months, 73 (55.3%) patients had PSA progression with a median time of 37.0 months. Fifty-six patients received adjuvant hormonal therapy, and 19 received salvage external beam radiation therapy. Clinical disease progression developed in 10 patients (7.6%). During follow-up, 8 patients died, 1 of prostate cancer and 7 of other causes. Preoperative PSA was a significant predictor of PSA progression and time to PSA progression after RP, whereas there were no differences in distant metastasis, local recurrence, hormone-refractory prostate cancer progression, and overall or prostate cancer-specific death between the 2 groups. CONCLUSIONS: On the basis of the favorable postoperative outcomes of RP in patients with preoperative PSA > or =20 ng/ml and no distant metastases, we suggest that RP has a role in treating these high-risk prostate cancer patients and that preoperative PSA is a significant predictor of postoperative PSA progression.
Disease Progression ; Follow-Up Studies ; Humans ; Neoplasm Grading ; Neoplasm Metastasis ; Prostate ; Prostate-Specific Antigen ; Prostatectomy ; Prostatic Neoplasms ; Recurrence ; Treatment Outcome

In the present study, we made an attempt to compare polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) with PCR-direct sequence analysis for their accuracy and sensitivity in detecting resistance to rifampicin (RMP). A total of 32 clinical isolates of Mycobacterium tuberculosis including 22 resistant and 10 sensitive isolates, whose drug susceptibility have been tested by conventional proportion method, were analyzed by using PCR-SSCP and PCR-sequence analysis. Among 22 RMP resistant isolates, 16 isolates showed SSCP profiles different from that of a RMP sensitive control strain, M. tuberculosis H37Rv indicating the possible existence of a sequence alteration in this region of the rpoB gene, while 6 resistant isolates displayed SSCP profiles indistinguishable from the sensitive control strain. On the other hand, all of 10 RMP sensitive isolates showed SSCP profiles similar to that of the sensitive control strain. Therefore, overall agreement rste between conventional proportion method and PCR-SSCP reached 81%. Subsequently, all of 32 clinical isolates were subjected to sequence analysis. The results from the sequence analysis revealed that all of 22 resistant isolates indeed contain mutations in the stretch of 81 bp region of rpoB gene, while none of 10 sensitive isolates contain any sequence alterations. Therefore, this study suggests that PCR-sequence analysis works more efficiently and accurately than PCR-SSCP analysis for rapid screening of RMP-resistant M. tuberculosis clinical isolates.
Hand ; Mass Screening ; Mycobacterium tuberculosis* ; Mycobacterium* ; Polymerase Chain Reaction* ; Polymorphism, Single-Stranded Conformational ; Rifampin* ; Sequence Analysis* ; Tuberculosis

Following the Korean War, presence of foreign troops in Korea, economic difficulty and changing moral Standards have brought about the development of concentrations of prostitutes around army camps known as "villes". The problems with venereal disease among the prostitutes living in the villes and consequently among soldiers has been a problem which has perplexed both American and Korean health authorities. Research into the problems and efforts to solve the V.D. problem have yielded only fragmentary results. Fortunately, with the cooperation of the U.S. Army Research and Development Group, an extensive research project has been undertaken to determine the root causes and best methods to prevent venereal disease in the villes around American military installations. The villes around large military installations were selected as the target of this research. In particular, the Pyeongtaek ville, the Gunsan "Silver Town" area and the "Texas Town" area of Busan were studied. The study took two years, from April, 1973 to March, 1975. Microbiological and serological studies were undertaken to determine the venereal disease infection rate and the efficacy of different antibiotics both in treatment and in sensitivity tests. In addition to the microbiological and serological studies other factors related to venereal diseases control such as social environment, health facilities, knowledge of venereal disease and its control were studied. The following results were obtained. 1. An overall gonorrheal morbidity rate of 14.1% was found among the 1,282 girls studied. This shows e rather dramatic rise compared to other studies. The majority of the increase is attributed to an actual increase of the V.D. rate, but more precise methods of isolation and culture using Thayer-Martin media also contributed to the increase recorded. 2. In Pyeongtaek, a gonorrhea morbidity rate of 5.39/0 (endocervical), the lowest in any of the villes was recorded. The Pyeongtaek police of enforced hospitalization until cure was effected tended to eliminate contact with infected girls and also make the girls more aware of and concerned about prevention of venereal disease. The Texas town prostitutes in Busan had a 15.5% (endocervical) morbidity rate, the highest of any of the areas. This might be attributed to sexual contact with many foreign sailors. 3. In culturing for gonorrhea, it was found that rectal as well as cervical cultures are necessary. Rectal cultures revealed 71 cases(39.2% to total positive cases) some of which were negative by endocervical culture, and endocervical cultures yielded 181 positives among the 1,282 cases surveyed. 4. A total of 988 serological tests for syphilis were carried out. By the V.D.R.L. method, 11.5% reactive cases were recorded, Kolmer C-F showed 5.9% reactive and RPCF test showed a 4.3% reactive rate. The RPR card test was used to retest 151 persons who were demonstrated to be V.D.R.L. (in prostitutes and general entertainers) reactive and confirmatory reactive results were shown in 74.0% of the cases. The FTA-ABS test was done on 57 RPCF tes/; reactive cases and showed confirmatory reactive results in 96.5% of the cases. It was concluded that the RPR card test, although it is less sensitive than the V.D.R.L test was both simple and effective for use in mass screening. However it is suggested that RPR card test rectors be reconfirmed using either the RPCF test or the FTA-ABS test. 5. Penicillin treatment failed to cure 40% of the gonorrhea patients in clinical tests and 16% of the strains used in the sensitivity tests showed resistance to penicillin. Therefore, penicillin was judged to be rather ineffective against gonorrhea. Ampicillin, however, is still quite effective, and spectinomycin was the most effective antibiotic against gonorrhea. 6. The socio-medical study of prostitutes revealed that their average age 25.5 years and the average number of years of education was 6. 8, implying the completion of primary school. Living conditions were generally poor, for example, in Pyeongtaek, 37% of the girls did not have running water and depended on well water. Data on the toilet facilities revealed that 78% of the prostitutes living in the control area in Wanweoldong, Busan were still using traditional Korean privy style toilets, and in Pyeongtaek 91% also were using privy toilets. Over two-thirds of the prostitutes only bathed once or twice a week. Therefore, improvement of living conditions was considered a critical problem. 7. Generally speaking, the girls had some know/edge of syphilis and gonorrhea and the measures necessary to prevent these diseases. However, over half of the girls abused antibiotics in venereal disease treatment and prevention attempts and only 65% of them required their partner to use a condom. Therefore, much improvement is needed in this area. 8. A survey of knowledge about contraceptive measures showed that 96% of the girls had good knowledge about oral contraceptives, 90 knew about condoms and 88% had knowledge of the douche method. In actual practice however, 95% of the girls used the douche method and 62% used condoms for contraception.
Ampicillin ; Anti-Bacterial Agents ; Baths ; Busan ; Condoms ; Contraception ; Contraceptives, Oral ; Education ; Female ; Fluorescent Treponemal Antibody-Absorption Test ; Gonorrhea ; Gyeonggi-do ; Health Facilities ; Hospitalization ; Humans ; Jeollabuk-do ; Korea* ; Korean War ; Mass Screening ; Military Personnel ; Penicillins ; Police ; Running ; Serologic Tests ; Sex Workers* ; Sexually Transmitted Diseases* ; Social Conditions ; Social Environment ; Spectinomycin ; Syphilis ; Texas ; Toilet Facilities

BCG has been one of the vehicles for multi-recombinant vaccine. However, low transformation efficiency of BCG with plasmid DNA hampered studies involving expression of foreign antigens in BCG. In an effort to determine the optimal conditions, this study was initiated to investigate factors involved in the transformation of BCG with a Mycobacterium-Escherichia coli shuttle vector, pYUB18, by electroporation. Mycobacterium bovis BCG (strain 1173P2) was grown in Middlebrook (M) 7H9 broth containing albumin-dextrose-catalase and 0.05% tween 80, and transformed BCG was grown in M7H10 agar containing kanamycin for counting viable cells. Pretreatment of BCG with 10 mM CaCl2 improved the transformation efficiency, but overnight incubation of BCG with 1% glycine did not. The transformation efficiency in BCG also varied depending on voltage, resistance, and DNA concentration. The maximum transformation efficiency was obtained when the infinity resistance, 12.5 Kv/cm, and 100 ng of DNA were used, and reached 1.4 x 10(5) CFU/microgram of plasmid DNA, which is about 3-100 times greater than those from previous reports. The transformation conditions described in this study, therefore, will give us a better position for employing BCG as a vehicle for developing multi-recombinant vaccines.
Calcium Chloride/pharmacology ; Comparative Study ; DNA/metabolism ; Electrophysiology ; Electroporation* ; Escherichia coli/genetics* ; Genetic Vectors* ; Glycine/pharmacology ; Mycobacterium/genetics* ; Mycobacterium bovis/genetics* ; Osmolar Concentration ; Transformation, Bacterial/physiology* ; Transformation, Bacterial/drug effects