Determination of vitamin B12 using the enzyme glycerol dehydrase.

Glycerol dehydrase is an enzyme that catalyzes dehydration of glycerol into beta-propionaldehyde. It requires 5'-deoxyadenosylcobalamin, one of the forms of vitamin B12, as a coenzyme. The enzyme is inactivated in vitro by all forms of vitamin B12 stoichiometrically. The objective of this study was to determine vitamin B12 content by utilizing the inactivation of the enzyme by vitamin B12. After various examinations, an excellent standard curve was obtained up to 1 pmol vitamin B12 using 14 mU of the enzyme per tube. Glycerol dehydrase does not respond to vitamin B12 if it is bound to haptocorrin, a vitamin B12-binding protein. This necessitates a procedure for extraction of vitamin B12 from samples before assay. The enzyme was less inactivated by 5'-deoxyadenosylcobalamin than any other form of vitamin B12. However, this did not matter because all forms of vitamin B12 were converted into cyanocobalamin during the extraction procedure cited above, which was performed in a buffer containing potassium cyanide.

[Clinical evaluation of an immunochromatography test for rapid diagnosis of influenza].

We evaluated a rapid diagnostic kit that detects influenza type A and B viral antigens by immunochromatography, Quick Vue Influenza Test (Quidel Corp., San Diego, CA, USA), with 425 specimens collected from patients with influenza-like symptoms at three hospitals between January and March 2001. The specimens included 184 nasal aspirates, 140 nasal swabs, and 101 throat swabs. The test correctly identified 179 of the 204 culture positive specimens and 203 of the 221 culture negative specimens; the sensitivity and specificity compared with the culture were 87.7% and 91.9%, respectively. The sensitivity of the test was 92.6% (112/121) for nasal aspirates, 83.7% (41/49) for nasal swabs, and 76.5% (26/34) for throat swabs, which is similar to the results for conventional rapid enzyme immunoassay kits for influenza virus infection. The sensitivity and specificity of the QuickVue Influenza Test were equivalent to those of Flu OIA (BioStar, Inc., Boulder, CO, USA), with the agreement of 84.2%. Although the QuickVue Influenza Test does not differentiate between influenza A and B viruses, the easy-to-use kit detects both types in the physician's office, allowing physicians to make a decision on prescription of neuraminidase inhibitor therapy during the initial visit.

Transferrin is not involved in the entry of 67Ga into hepatocytes from regenerating liver of partially hepatectomized rats.

Ever since the first observation of 67Ga accumulation in tumors and inflammatory lesions, 67Ga has been used to detect various tumors and inflammations. The aims of this study were to clarify whether or not transferrin is involved in the uptake of 67Ga by the regenerating liver after partial hepatectomy. The uptake of 67Ga by the liver of rats reached a maximum 2 d after partial hepatectomy. In order to inhibit the binding of 67Ga to transferrin in the blood, FeCl3 was administered 5 min before the injection of 67Ga. The administration of FeCl3 decreased the uptake of 67Ga by the liver of the partially hepatectomized rats, suggesting that transferrin is involved in the uptake by the liver. However, 67Ga was taken up only slightly by hepatocytes obtained from the liver of these rats. We conclude that transferrin is involved in the uptake of 67Ga by the liver tissue of partially hepatectomized rats but is not involved in its entry into the hepatocytes. Only a slight amount of gallium-67 enters the hepatocytes, and may accumulate primarily in the extracellular matrix of the liver tissue of these rats.

[Evaluation of the rapid detection test for influenza A and B viruses using neuraminidase activity].

The ZstatFlu test (ZymeTx, USA) is a rapid detection kit for influenza A and B viruses. This test is based upon the reaction between viral neuraminidase from influenza viruses and a chromogenic substrate. The clinical performance of the ZstatFlu test was determined by comparison with viral isolation in cell culture. A total of 176 respiratory specimens from 172 pediatric patients with influenza like illnesses during the 1998/99 season were tested. Influenza viruses were recovered from 97 specimens (type A: 6, type B: 91) in cell culture. ZstatFlu demonstrated 67.4% sensitivity (29/43) and 62.7% specificity (37/59) for throat swabs. Of the 22 ZstatFlu-positive, culture-negative throat swabs tested by RT-PCR, 18 were positive by RT-PCR. ZstatFlu showed 48.1% sensitivity (26/54) and 90.0% specificity (18/20) for nasopharyngeal aspirates. Of the two ZstatFlu-positive, culture-negative nasopharyngeal aspirates tested by HI titer of paired sera, one showed a 4-fold increase of HI titer. Nasopharyngeal aspirates therefore showed lower sensitivity than throat swabs at this test, different from EIA test kits such as Directigen FluA or FLU OIA. Overall, only 5 specimens were false positive by the ZstatFlu test. Therefore, this test demonstrated high specificity and positive predictive value. In conclusion, the ZstatFlu test is useful for the rapid detection of influenza A and B viruses to identify patients who need antiviral treatment.

The impact of winter epidemics of influenza and respiratory syncytial virus on paediatric admissions to an urban general hospital.

To demonstrate the impact of influenza epidemics on pediatric hospital admissions, admissions that were attributable to influenza and respiratory syncytial virus (RSV) infection to the pediatric ward of an urban general hospital in Japan were followed-up during a 4-month period from December to March 1991 through 1998. During the 1997-1998 influenza type A (H3N2) epidemic, a diagnosis of influenza type A (H3N2) was made in 26.3% of all patients admitted aged 15 years or lower. During the peak of the epidemic, as many as 50-70% of the admissions were attributable to influenza type A (H3N2). In the seven winters from 1991 to 1988, 14.0% of all admissions were associated with infection with influenza virus (mean age 4.4 years), and 17.5% were due to RSV. More patients were admitted to hospital for influenza than RSV infection in three of the seven seasons. Among the patients with influenza, 74.5% of the cases were previously healthy children. Influenza and RSV infection are leading causes of pediatric hospital admissions during the winter. Effective methods of prophylaxis are needed not only for high-risk patients, but for healthy young children.

[Optical immunoassay test for rapid detection of influenza A and B viruses: an evaluation].

The optical immunoassay test (FLU OIA, BioStar, USA) for rapid detection of influenza A and B viral antigens was compared with viral isolation in cell culture. A total of 103 respiratory specimens were tested on 75 pediatric patients with acute respiratory illnesses. Influenza viruses were recovered in 40 specimens (type A: 5, Type B: 35). FLU OIA demonstrated 80.0% sensitivity and 68.8% specificity for nasopharyngeal aspirates and 36.7% sensitivity and 83.9% specificity for throat swabs. We also tested FLU OIA, retrospectively, using 78 supernatant samples from pediatric patients with influenza A virus infection frozen after cell culture. FLU OIA demonstrated 91.4% sensitivity and 92.3% specificity for nasopharyngeal aspirates and 50.0% sensitivity and 91.7% specificity for throat swabs diluted in viral transport media. Nasopharyngeal aspirates showed higher sensitivity than throat swabs for detection of influenza virus by FLU OIA. We believe this rapid test kit is useful for the detection of influenza A and B viruses.

Congenital absence of the portal vein in a boy.

Congenital absence of the portal vein (CAPV) is a malformation that is generally thought to be limited to females. We encountered an 11-year-old boy with this malformation. In 17 previously reported cases of CAPV, 2 were male. Three male patients, including our case, were Abernethy type Ib malformation. They had no associated liver tumour and two had no additional congenital abnormality. Conversely, 13 of the 15 female patients had congenital abnormalities and 6 had liver mass lesions.

[Impact of influenza A virus infection as a cause of pediatric hospitalization and use of rapid antigen test of influenza A virus].

From the winter of 1997 to the spring of 1988, a severe epidemic caused by influenza A (H3N2) developed in Japan. During the epidemic (December 1997 to February 1998), 239 children were admitted to the pediatric ward of Nippon Kokan Hospital: as many as 76 (31.8%) were hospitalized with influenza A (H3N2) infection. Most were young healthy children (mean age, 3.7 years), suggesting the need of influenza immunization for this age group. 110 specimens of hospitalized children were examined by rapid antigen test of influenza A virus (Directigen Flu A, Becton Dickinson, USA). The rapid test showed about 90% sensitivity and specificity, as compared with isolation and serum hemagglutination inhibition test. Directigen Flu A proved to be a reliable, rapid screen for influenza A from symptomatic patients.

[Immunization with one dose of inactivated influenza vaccine].

In the winter of 1996, a total of 71 working adults from 20 to 60 (m = 37.3) years of age were immunized with one or two doses of inactivated influenza vaccine formulated in Japan. The reciprocal geometric mean hemagglutination inhibition antibody titers [mean HI] and the percentage with a hemagglutination inhibition titer of > or = x 128 [percent > or = x 128], against A/ Yamagata/32/89 (H1N1), A/Uhan/359/95 (H3N2), and B/Mie/1/93 were significantly elevated by either doses. The pre-/post-vaccination mean HI of H1N1 were 61.5/668.4 with one dose, and 85.7/637.3 with two doses. The percent > x 128 were 36.5/96.2 with one dose, and 47.4/94.7 with two doses. No significant differences were apparent between one and two doses. Similarly, nonsignificant differences were observed for the other two antigens contained in the vaccine. We observed the sufficient immune response to only one dose of inactivated influenza vaccine, which emphasizes the merit of one dose method of influenza immunization.

Efficacy of inactivated vaccine in preventing antigenically drifted influenza type A and well-matched type B.

OBJECTIVE: To evaluate the efficacy of currently used inactivated influenza vaccine during a severe epidemic caused by antigenically drifted influenza type A(H3N2) and well-matched type B viruses during the 1992-1993 season. DESIGN: Prospective nonrandomized controlled trial. SETTING: An urban general hospital pediatric asthma clinic in Japan. PARTICIPANTS: A total of 137 children with moderate to severe asthma (mean age, 7.0 years; range, 2 to 14 years). INTERVENTION: Eighty-five children received trivalent split-antigen vaccine containing A/Beijing/352/89 (H3N2) and B/Bangkok/163/90 (B/Panama/45/90-like strain). Fifty-two were unvaccinated. MAIN OUTCOME MEASURES: Protection against infection was determined using hemagglutination inhibition test and virus isolation. Clinical efficacy was estimated based on febrile episodes with antibody rise or virus isolation. RESULTS: Although marked antigenic drift in hemagglutinin was demonstrated in the epidemic virus (A/Beijing/32/92-like strain), the protection against influenza type A(H3N2) infection was 67.5% (P < .01). The protection against influenza type B infection was 43.7% (P < .01), although the epidemic influenza type B viruses were antigenically almost identical to the vaccine strain. Inactivated vaccine was not effective for protection against influenza type B infection in children younger than 7 years. High clinical effectiveness was demonstrated in children at least 7 years of age during the epidemic. CONCLUSIONS: Our data suggest that current inactivated vaccine is highly effective for protection against influenza type A(H3N2) virus infection regardless of antigenic drift. In contrast, the protective efficacy obtained by vaccination may not be sufficient against influenza type B virus infection, and especially in young children, it does not offer protection.

[Prediction of bone marrow function by the appearance rate of RNA-rich reticulocytes during the chemotherapy for hematologic malignancy].

Percentage of RNA-rich (High fluorescence Ratio, HFR), RNA-middle (Middle Fluorescence Ratio, MFR) and RNA-low (Low Fluorescence Ratio, LFR) reticulocytes was estimated by Sysmex R-1000 reticulocyte counter during the chemotherapy of 27 cases with acute leukemia and malignant lymphoma (36 courses). In the beginning of marrow suppression, HFR and/or MFR were decreased faster than LFR, neutrophils or platelets in 14 courses (38.9%). Simultaneous decrease of HFR, MFR and other blood cells were observed in 57.2%. In the recovery phase of bone marrow, faster increase of HFR and/or MFR was noted in 52.8% and simultaneous increase of these reticulocytes with other blood cells was observed in 9 courses (25%). Appearance rate of RNA-rich younger reticulocytes is a sensitive parameter for the prediction of bone marrow suppression and recovery during the chemotherapy for hematologic malignancy.

Increased activities of cytosol aminopeptidase and lactate dehydrogenase in serum originate from lymphocytes in necrotizing lymphadenitis.

In three pediatric patients with necrotizing lymphadenitis, cytosol aminopeptidase activity (c-AP; EC 3.4.11.1) in serum was markedly increased to 509, 417, and 191 U/L, respectively (normal range 25-60 U/L). Lactate dehydrogenase (LD; EC 1.1.1.27) was also increased, with LD-3 predominating. The increased concentrations of c-AP and LD presumably originated from the destruction of infected, activated lymphocytes, especially T lymphocytes. Necrotizing lymphadenitis is probably caused by a lymphocytotropic virus.

Serum amphotericin B concentration in a very premature infant with disseminated candidiasis.

A 1,040-g premature baby was diagnosed to have disseminated candidiasis and treated with amphotericin B (AMB) and 5-fluorocytosine. During the treatment, an unexpectedly large dose of AMB was infused unintentionally. AMB level was as high as 1.73 micrograms/mL soon after 5 mg/kg infusion instead of 0.5 mg/kg. However, it dropped rapidly to 0.83 micrograms/mL after 24 hours. AMB was detected in patient's serum at a higher level than minimal inhibitory concentration as long as one month after treatment was stopped. The patient showed liver dysfunction but no nephrotoxicity. The further studies are needed to establish safe and effective treatment regimen in premature infants with disseminated candidiasis.

Increased cytosol aminopeptidase and lactate dehydrogenase in serum originating from lymphocytes in measles and rubella infection.

We determined cytosol aminopeptidase (c-AP; EC 3.4.11.1) and lactate dehydrogenase (LDH) levels in serum; these enzymes are known to originate from lymphocytes in patients with measles and rubella. In patients with measles (n = 19), both enzyme levels increased markedly with the onset of rash: mean (+/- SD) c-AP was 269.7 +/- 103.5 U/L and LDH was 1149.5 +/- 255.2 U/L. In patients with rubella, activities of both enzymes increased mildly: c-AP (n = 18) was 81.6 +/- 24.4 U/L and LDH (n = 13) was 674.0 +/- 168.8 U/L. Increased c-AP and LDH levels in patients with measles and rubella presumably originate from the destruction of infected, activated lymphocytes, especially T lymphocytes.