Comparative study for Isolation of Yersinia pseudotuberculosis from Water / 대한임상미생물학회지

BACKGROUND: Yersinia pseudotuberculosis is recognized throughout the world as a cause of water-or food born infections in human and animals. Although many attempts have been made to define optimal conditions for the isolation of the organism from water, their isolation yields remain low; therefore, we tried to find an effective method for the recovery of Y. pseudotuberculosis from water. METHODS: Water samples were deliberately contaminated with Y. pseudotuberculosis at various levels and then processed by the following three isolation METHODS: centrifugation, direct filtration, and intracellular culture. For the centrifugation method, the water samples were centrifuged at 5000 rpm for 1 hr and the final precipitates were inoculated in cefsulodin-irgasan-novobiocin(CIN) media. For the filtration method, the water samples were filtered by negative pressure and the filter papers were put directly on CIN media. For the intracellular culture method, the organisms were extracted from the HeLa cells that had been infected with Y. pseudotuberculosis and inoculated on CIN media. We also examined the efficacy of the filtration method after cold enrichment with a mixture of Y. pseudotuberculosis, Escherichia coli, and Citrobacter freundii. RESULTS: With the concentration of 3x10(2)/100 mL, Y. pseudotuberculosis was isolated only by the filtration method; however, none of the culture methods were good enough to recover the organism from the water sample when the concentration was 3x10/100 mL. With cold enrichment, however, the recovery was much more efficient; the organism grew after direct inoculation or after filter inoculation when the starting concentrations were 3x10(2)/100 mL or 3x10/100 mL, respectively. CONCLUSION: A combined use of direct filtration and filter inoculation after cold enrichment is the most effective method to yield Y. pseudotuberculosis isolation. The introduction of effective methods for the isolation of Y. pseudotuberculosis from untreated drinking water would increase the awareness by the public of the health hazard of spring water.

The Cryopreservation and Thawing of Red Blood Cells Using 25% Hydroxyethyl / 대한진단검사의학회지

BACKGROUND: The method using hydroxyethyl starch (HES) as a cryoprotectant is a simple, quick, and inexpensive way to make frozen red blood cells (RBCs) for an extended period. It needs neither sophisticated equipment nor skilled labor. As HES is a plasma expander, it does not have to be washed out before transfusion. But, it is not yet introduced in Korea to make frozen RBCs with HES. The aim of our study was to establish a method to cryopreserve RBCs using HES in Korea. METHODS: We rejuvenated RBCs (20 units) on the first day after expiration. They were washed three times with saline adenine glucose mannitol phosphate buffer solution (SAGMP), then their hemat-ocrits were adjusted up to 80%. The same weight of 25% HES was added to each RBC, to make a final mixture of 12.5% (wt/wt) HES-40% RBC. The blood bags were placed in a thin metal frame and frozen in liquid nitrogen. After 3 months, the blood bags were thawed in a 37 degrees Cwater bath. To evaluate the qualities of frozen-thawed RBCs, samples of HES-RBC mixtures before freezing and after thawing were tested for ATP, 2, 3-DPG, potassium, plasma hemoglobin, hemolysis %, and 30-minute saline stability (30-min SS). RESULTS: After thawing, the percentage (mean+/-SD) of RBC hemolysis was 1.6+/-0.4% (range, 1.1-2.6) and three units were over 2%; 30-min SS was 92.0+/-0.16% (88.4-93.8). Supernatant potassium and free hemoglobin were 15.6+/-3.5 mmol/L (10.9-23.0) and 215+/-62 mg/dL (139-318), respectively. The ATP and 2, 3-DPG of RBCs were 4.23+/-0.81 (3.31-6.51) micromol/g Hb and 8.46+/-1.62 (5.21-11.00) micromol/g Hb, respectively. CONCLUSIONS: Of the 20 units of RBCs tested, 17 units (85%) were within the criteria for transfusion (hemolysis88%, potassium<75 mmol/L) after thawing. We could successfully freeze and thaw RBCs utilizing 25% HES as a cryoprotectant.

Transfusion-transmitted Virus and TTV-like Mini Virus Infection in Blood Products / 대한진단검사의학회지

BACKGROUND: Transfusion-transmitted virus (TTV) and TTV-like mini virus (TLMV) are small DNA virus with single-stranded, closed circular, antisense genome infecting man. TTV and TLMV are trans-missible by transfusion. However there had been a few study about TTV prevalence and no study about prevalence in blood donors in Korea. There has been no study about the TTV and TLMV infection in blood products in Korea. The aim of this study was to gain the prevalence of two viruses in blood products. METHODS: A total of 150 plasma samples from blood products (each 50 units of Red blood cell, whole blood, and platelet concentrate) were tested. The samples are obtained from the segments of the blood products. TTV DNA was detected using polymerase chain reaction (PCR) with two sets of primers (A set and B set) and TLMV DNA was detected using nested PCR with primer set C. RESULTS: TTV DNA was detected in 85.3% (128/150) of blood products. TLMV DNA was detected in 41.3% (62/150) of blood products. Either TTV or TLMV was detected in a total of 140 blood products (92.3%) and both TTV and TLMV were detected in 50 products (33.3%). CONCLUSIONS: The blood products are frequently infected with TTV and (or) TLMV in Korea and they can be transmissible by blood products with high probability.

The Detection of Plasmodium vivax DNA, and Antibody using Filter Paper Blood / 대한진단검사의학회지

BACKGROUND: Filter paper blood samples are convenient for sampling, storage and transport, and widely used for diagnostic and epidemiological studies of Plasmodium species. Our study was aimed to establish antibody and DNA tests using filter paper and to evaluate the usefulness of filterpaper blood sample and storage duration at room temperature. METHODS: 50 L whole blood of 45 Plamsmodium vivax infected patients were spotted on filter paper, dried, and kept in a drawer at room temperature. The paired, whole blood samples were stored at -70degrees C freezer temperature. The filter paper samples were used for PCR at 12 and 24 months. The standard filter paper samples (5, 000, 500, 100, 50, 5, 1 parasite/microL) were made from 3 patients and used for polymerase chain reaction every 3 months until 2 years, antibody test filter paper samples were used every month for 8 months. As standard, -70 degrees C whole blood samples were used. RESULTS: P. vivax DNA was detected in all (45/45) of the filter paper samples by PCR during 24months of storage. Detection limit of PCR was 1 P. vivax/microL using standard filter paper samples for12 months of storage, and 5 P. vivax/microL samples after that. The P. vivax antibody was detected perfectly(43/43) from filter paper samples during 5 months of storage. After that, the antibody detectionrates were, respectively, 81.4% (35/43), 60.5% (26/43), 51.2% (22/43) at six, seven and eight months. CONCLUSIONS: Filter paper blood samples stored at room temperature remained stable for PCR and antibody studies for 1 year and 5 months, respectively.

The Comparsion of Polymerase Chain Reaction and Culture for Detection of Helicobacter pylori in Saliva / 대한진단검사의학회지

BACKGROUND: The presence of Helicobacter pylori DNA from saliva has been demonstrated by polymerase chain reaction; however, the culture of H. pylori is quite difficult. Because isolation of H. pylori strains from saliva plays an important role in epidemiological and pathogenic studies, our study was aimed to establish the H. pylori culture method from saliva and the compared detection limit between cultures after HCl-urea treatment and PCR. METHODS: A strain of H. pylori was cultured, diluted and mixed with saliva of an H. pylori-negative volunteer to make a final concentration of a 10(1)-10(5) colony forming unit (CFU)/mL saliva. Each concentration of samples was treated with various concentrations of HCl-urea (pH 1.2) to suppress other bacteria. Paired samples (HCl-urea treated and untreated) were streaked on horse blood agar and cultured for 3-10 days at 37 degrees C in microaerophilic condition. The DNA templates were extracted from the same concentrations of HCl-urea treated bacterial suspensions in saliva and PCR was performed. RESULTS: The detection limit of the culture after HCl-urea treatment was 10(3) CFU/mL of saliva and that of the PCR after HCl-urea treatment was 10(2) CFU/mL of saliva. CONCLUSIONS: A culture after HCl-urea treatment for H. pylori would be an effective method for isolation of H. pylori from saliva; however, this method is less sensitive than PCR as far as the detection limit is concerned. A combination of both methods would be an effective method for detecting H. pylori from saliva.

Molecular Relatedness between Isolates Yersinia pseudotuberculosis from a Patient and an Isolate from Mountain Spring Water

A 40-yr-old buddhist monk was admitted to the hospital with abdominal pain, fever, and confusion. He had a history of drinking untreated mountain spring water in his temple, and experienced the above symptoms for several days before admission. In past medical history, he had suffered from hepatic cirrhosis. Yersinia pseudotuberculosis was isolated from his blood and ascitic fluid. The mountain spring water that he had ingested was cultivated and Y. pseudotuberculosis was also isolated. For identification of pathogenic Y. pseudotuberculosis, each isolate from the three sources (blood, ascitic fluid, and drinking water) was also analysed for the inv gene for Y. pseudotuberculosis and the virF gene for virulent plasmid by PCR. All strains were positive for both the virF and the inv genes and also positive for autoagglutination test. For relationship study, each isolate from the three sources was also analysed with serotyping and restriction endonuclease analysis of virulence plasmid DNA (REAP) using BamHI. All belonged to the serotype 4b and REAP pattern D. Thus, all these findings supported that the mountain spring water was the source of the Y. pseudotuberculosis infection in this case.

The Preparation of Frozen Red Blood Cells and Deglycerolization of Frozen RBCs with Haemonetics V50plus / 대한수혈학회지

BACKGROUND: The method utilizing high concentration glycerol is a common way to make frozen red bool cells (RBCs) for extended period, but it requires special deglycerolization equipment for washing after thawing. For economics reason, we could not have a cell washer for wahing the frozen RBCs, so attempted to use Haemonetics V50plus that we have had. METHODS: Twelve fresh packed RBCs were cryopreserved with 40% glycerol method. After 3 months, the RBCs were thawed and washed with Haemonetics V50plus. For the evaluation of the procedure, RBC reovery rate, osmolarity, 30 min saline stability (SS), % hemolysis, ATP, 2,3-DPG, LDH, potassium, and plasma hemoglobin were tested at 24hrs after washing. RESULTS: The RBC recovery rate was 82.1 +/- 4.5% (75.8-89.2) and two units of Frozen RBCs were under 80%. The Hb ATP and Hb 2,3-DPG of RBCs were 5.2 +/- 0.6 micro mol/g Hb (3.9-6.0) and 13.0 +/- 2.1 micro mol/g Hb (8.8-15.1). The supernatant osmalrity, potassium, plasma Hb and LDH were 352 +/- 7mosmol/kg H2O (342~367), 0.8 +/- 0.2 mmol/L (0.5~1.2), 0.8 +/- 0.3 mg/dL (0.4~1.1), 352 +/- 7 U/L (342~367). The 30min SS was 98.9 +/- 0.8. CONCLUSION: We could successfully freeze, thaw, and wash the frozen RBCs with Haemonetics V50 plus.

Comparison of Detection of Escherichia coli O157 Between Culture After Acid Treatment and Polymerase Chain Reaction After Enrichment / 대한진단검사의학회지

BACKGROUND: The Polymerase Chain Reaction (PCR) for the Shiga toxin has been widely used for diagnosis of Shiga toxin-producing Escherichia coli (STEC) infection including Escherichia coli O157 (O157) instead of using a culture. However, bacteriological isolation must be followed for final diagnosis. Our study was aimed to compare the detection limit between the culture after the acid treatment and the PCR after enrichment. METHODS: The standard strain of O157 was cultured, diluted and mixed with the stool of normal adult in order to make a final concentration of the 10(1)-10(5) colony forming unit (CFU)/g of stool. Each concentration of samples was enriched in a trypticase soy broth for 6 hours at 42degrees C and treated with acid to suppress normal flora. Then it was streaked on cefixime-tellurite-sorbitol MacConkey (CT-SMAC) agar evenly and cultured for 18 hours at 37degrees C. The same concentrations of bacterial suspension in the stool were enriched in a Luria-Bertani (LB) broth overnight at 37degrees C. The centrifuged pellets from 1 mL of each concentration of the samples were boiled and DNA was extracted using the resin method and PCR was performed to amplify stx2. RESULTS: The detection limit for the culture after acid treatment was 10(3) CFU/g of the stool and that for PCR after enrichment was 101 CFU/g of the stool. CONCLUSIONS: Culture after acid treatment for O157 would be an effective method for isolation of O157 from a patient's stool. However, this method is less sensitive than the PCR after enrichment as far as detection limit is concerned. A combination of both methods would be an effective method for detecting O157 from patient stools.

The Extended Preservation of Outdated Red Blood Cells with Rejuvesol and Additive Solutions / 대한수혈학회지

BACKGROUND: Longer RBC storage will benefit blood banker by reduced outdating and patients by receiving fewer nonviable cells and RBC breakdown products as well as by having the opportunity to participate in markedly improved autologous storage system. METHODS: We rejuvenated whole blood 10 units and RBC 15 units, on 1-3 days after expiration, then they were divided into four, washed, and suspended with four kinds of solutions containing saline adenine glucose, mannitol, and phosphate, in closed system. For the evaluation of RBCs, 30 min saline stability (SS), 2 h SS, % hemolysis, ATP, 2,3-DPG, P50, LDH, potassium, and plasma hemoglobin were tested at every week during 42 days. RESULTS: On 35th day, 30 min SS of RBCs was higher than 88% and % hemolysis of RBCs was lower than 1% in solutions containing phosphate. ATP of RBCs was the 50% of the reference value of healthy persons and 2,3-DPG of RBCs was relatively higher in solutions containing phosphate (SAGMP1, 2, 3) than in solution without phosphate (SAGM). CONCLUSION: We could successfully rejuvenate outdated RBCs and extend the expiration date to additive 35 days, but for practical use, post transfusion RBC survival and safety should be evaluated in the future.

Comparison of Selective Media for Culture of Helicobacter pylori / 대한임상미생물학회지

BACKGROUNDS: Culture of Helicobacter pylori (H. pylori) from gastric biopsy specimens is a standard method with high specificity among H. pylori diagnostic tools and is also essential for antibiotic susceptibility test. The authors compared 5 selective media for H. pylori culture and tested fresh human serum instead of fresh animal blood as a media composite. METHODS: Gastric biopsy specimens from endoscopic examination were obtained from 50 patients (gastric ulcer:33, duodenal ulcer:12, stomach cancer:5) and they were finely minced with a tissue grinder. Specimens were inoculated onto 5 media (1. Columbia agar with 5% sheep blood, 2. Columbia agar with 10% human serum, 3. Thayer-Martin agar with 5% sheep blood, 4. T-M agar with 10% human serum, 5. T-M agar with 10% hemoglobin) and cultured for 3~7 days under microaerophilic condition. Gram stain, oxidase, catalase, and urease tests, were undertaken on typical colonies for diagnosis of H. pylori. Contamination by other organisms, number and size of H. pylori colonies were compared for each media. RESULTS: Positive culture rate of H. pylori was not significantly different among 4 media except TM agar with 10% hemoglobin. However T-M agar with 10% fresh human serum was considered as the best composition for culture of H. pylori because it had the least contaminating organisms and produced the largest colony sizes. CONCLUSIONS: These findings suggest that T-M agar with 10% fresh human serum can replace columbia agar with 5% sheep blood which has been commonly used for culture of H. pylori from gastric biopsy specimens. Fresh human serum, which is easily obtained in the clinical laboratory, can replace animal bloods in making media for H. pylori.

Nested Polymerase Chain Reaction Assay of Helicobacter pylori in Saliva / 대한임상미생물학회지

BACKGROUND: The aim of this study was to investigate the prevalence of Helicobacter pylori in the saliva of infected patients and the relation between H. pylori in the saliva and the severity of gastric infection. METHODS: Active gastric infection was determined by the 13C-urea breath test. Bacteria in saliva were detected by the nested polymerase chain reaction, using primer sets EHC-U/EHC-L and ET-5U/ET-5L. RESULTS: The PCR assay was able to detect as few as 5 H. pylori /mL. A total of 82 (71.9%) out of 114 patients with gastroduodenal diseases were positive by 13C-urea breath tests. Among these 82 patients, 21 (25.6%) were PCR positive in their saliva. H. pylori was present in the saliva of patients with highly active gastric infections (> 50 delta perrmille). No H. pylori was detected in saliva of patients with no active gastric infections. CONCLUSIONS: In patients with highly active gastric infections, H. pylori may be transmitted via saliva. The PCR assay of H. pylori in saliva is not useful for detecting gastric infection but may be a useful tool for the screening of highly infectious patients.

Comparison of serotypes, restriction enzyme analysis of plasmid DNA pattern and PFGE(pulsed-field gel electrophoresis) patterns of Yersinia pseudotuberculosis isolates in Korea / 대한임상병리학회지

BACKGROUND: Yersinia pseudotuberculosis, a member of genus Enterobactericeae, is a main etiologic organism of diarrhea in childhood. Because a mouse and a unchlorinated spring water are main reservoirs of Y. pseudotuberculosis, the strains from a contaminated spring water and mouse could be involved in human epidemic. The purpose of this study was to investigate a clonality between the strains from patients and those from an unchlorinated spring water and a mouse by restriction enzyme analysis of plasmid DNA and pulsed-field gel electrophoresis (PFGE). METHOD: We isolated 15 Y. pseudotuberculosis strains including 8 isolates from patients (S1-S8), 6 isolates from mountain water (W1-W6), 1 isolate from a mouse (M1) in northeast area of Seoul. Plasmid and chromosomal DNA of all strains were analyzed by REAP with Bam H1 restriction and by PFGE with Xba I restriction , respectively. RESULTS: Restriction enzyme analysis of plasmid DNA was classified into type B and type D. All 7 strains of serotype 15 were classified as type B and 8 strains of serotype 4b were classified as type D. PFGE were classified into 6 different types. Among them, strains of PFGE type I, II, III, IV belong to Y. pseudotuberculosis serotype 15 and Y. pseudotuberculosis 4b strains were classified into PFGE type V, VI. S1 and W1 were classified into PFGE type I . S8, W6 and M1 were classfied into PFGE type VI. CONCLUSIONS: PFGE revealed clonality among strains from patients, a water and a mouse. PFGE was more discriminative than REAP to characterize the Y. pseudotuberculosis outbreaks in Korea.

Korean Nationwide Surveillance of Antimicrobial Resistance of bacteria in 1997

Antimicrobial-resistant bacteria are known to be prevalent in tertiary-care hospitals in Korea. Twenty hospitals participated to this surveillance to determine the nationwide prevalence of resistance bacteria in 1997. Seven per cent and 26% of Escherichia coli and Klebsiella pneumoniae were resistant to 3rd-generation cephalosporin. Increased resistance rates, 19% of Acinetobacter baumannii to ampicillin/sulbactam, and 17% of Pseudomonas aeruginoa to imipenem, were noted. The resistance rate to fluoroquinolone rose to 24% in E. coli, 56% in A. baumannii and 42% in P. aeruginosa. Mean resistance rates were similar in all hospital groups: about 17% of P. aeruginosa to imipenem, 50% of Haemophilus influenzae to ampicillin, 70% of Staphylococcus aureus to methicillin, and 70% of pneumococci to penicillin. In conclusion, nosocomial pathogens and problem resistant organisms are prevalent in smaller hospitals too, indicating nosocomial spread is a significant cause of the increasing prevalence of resistant bacteria in Korea.

Epidemiological Investigation of Yersinia pseudotuberculosis Infection in Korean / 대한임상병리학회지

BACKGROUND: The purpose of this study is to prove that human infection of Yersinia pseudotuberculosis might be occurred by drinking of mountain spring water contaminated with wild mice excreta through epidemiological tool. METHOD: Y. pseudotuberculosis strains which were isolated from patient stools, mountain spring water and mice excreta were analysed by serotyping of O antigen and plasmid DNA profile (Restriction Endonuclease Analysis of Plasmic DNA analysis REAP) assay Also reservoir rate of Y. pseudotuberculosis was calculated from wild mice which were captured throughout Korean mountains. RESULTS: Reservoir rate of Y. pseudotuberculosis from wild mice in Korea was 0.85% and was not higher than that in other country. The analysis of 66 strains of Y. pseudotuberculosis showed that 36 strains of serotype 15, REAP B type, 24 strains of serotype 4b, REAP D type, and 1 strain of serotype 4b, REAP new unclassifiable type, but 5 strains didn't have plasmic (serotype 15:3, 11 :2) .Especially same 4b, D type of Y. pseudotuberculosis was isolated from patient stools, mountain spring water and wild mouse (Apodemus agrarius) excretion and this fact was considered that Y. pseudotuberculosis strains from 3 groups were closely correlated epidemiologically. Also serotype 15, REAP B strains were isolated from patient stools and mountain spring water, but were not isolated from wild mice yet and 15, B type was isolated from Korea only and considered as native Korean strain which had not isolated in other countries yet. CONCLUSIONS: Human Y. pseudotuberculosis infection in Korea was occurred by drinking of contaminated mountain spring water and A. agrarius was one of main reservoir which contaminates mountain spring waters in Korea, Also above antigenic distribution of Y. pseudotuberculosis would be useful for development of ELISA kit of Korean type.

Epidemiological Studies of Yersinia pseudotuberculosis Infection by Plasmid DNA Profile Assay / 감염

BACKGROUND: In order to investigate transmission route of Yersinia pseudotuberculosis infection in Korea, we tried epidemiological study among human strains, mountain spring water strain and wild mouse strain which were isolated in north eastern area of Seoul on spring in 1996. METHODS: Plasmid profile (Restriction Endonuclease Analysis of Virulence Plasmid DNA analysis: REAP) assay in addition to serotyping were performed among human strains, mountain spring water strain and wild mouse strains. RESULTS: All isolates were the same O serotype of 4b and the same REAP pattern of type D. CONCLUSION: These results suggested that wild mice (especially Apodemus agrarius) were one of main reservoir of Y. pseudotuberculosis in Korea and their fecal material might contaminate mountain spring water. Most of human infections of Y. pseudotuberculosis were originated from drinking of contaminated mountain spring waters in Korea.

A Case of Pernicious Anemia Associated with Autoimmune Thyroid Disease / 대한혈액학회지

Pernicious anemia is understood as an autoimmune disease and associated with various other autoimmune diseases, such as Graves` disease, primary hypothyroidism, thyroiditis and vitiligo. We report a case of pernicious anemia associated with autoimmune thyroiditis. A 40-year-old man was admitted to the Sanggye Paik Hospital due to general weakness, dyspnea on exertion and sore throat for 2 months. Eight years before admission, he had been treated with hyperthyroidism in other hospital. Examination revealed anemic conjunctivae, exophthalmos and smooth and beefy tongue. Laboratory tests showed 6.2g/dL of hemoglobin, 16.7% of hematocrit, 7,970/microliter of WBC, 152,000/microliter of platelets and 116.3fL of MCV. Reticulocyte index was 0.3%. Peripheral blood smear showed macrocytic red blood cells and hypersegmented neutrophils. The level of vitamin B12 was 139.2pg/mL and folic acid was in normal range. The result of schilling test was abnormal. Anti-parietal cell antibody was positive. The results of thyroid function tests were compatible with hypothyroidism and anti-microsomal antibody was positive. TBII was 9.8U/L. Treatment with vitamin B12 and thyroid hormone was started. Three months after treatment, he has been completely free of symptoms. Now he has been treated with thyroxine 0.2mg per day and adenosylcobalamine 1,000microgram per 2 month.

Bone marrow pathology of culture proven typhoid fever

The authors analysed bone marrow findings of sixteen cases of culture proven typhoid fever to reveal the pathologic changes according to the disease stage. The most frequent finding was chronic granulomatous inflammation (eight cases). Infection (bacteria) associated hemophagocytic syndrome (four cases), reactive marrow (two cases), and non specific findings (two cases) were also encountered. Granulocytic hyperplasia with hemophagocytosis appeared at the early stage and was followed by infection (bacteria) associated hemophagocytosis and granuloma in proliferative stage. In lysis (late) stage, granulomatous inflammation was noted. However, resolution of granulomatous inflammation was not distinct. Some nuclear debris and phagocytosis were remarkable in well-formed granulomas. Thrombocytopenia was the most remarkable peripheral blood finding at the time of biopsy. Anemia, leukopenia, and pancytopenia were also observed in descending order.

CK-MB isoenzyme quantitation in patients with acute myocardial infarction by electrophoresis and dry immunochemical method / 대한임상병리학회지

No abstract available.

In vitro quantitative clonal assay of CFU-megakaryocyte in mouse bone marrow / 대한임상병리학회지

No abstract available.

A Case of Congenital Cellular Neurilemoma

No abstract available.