Rapid detection of the hepatitis B virus YMDD mutant using TaqMan-minor groove binder probes

BACKGROUND: TaqMan-minor groove binder (MGB) probes were used in a real-time PCR-based assay for the rapid and accurate detection of hepatitis B virus (HBV) YMDD mutants. METHODS: TaqMan-MGB probes were designed to distinguish between wild-type (YMDD) and mutant (YVDD and YIDD) strains of HBV. The detection limit and sensitivity of the assay were determined using a dilution series of a mixture of wild-type and mutant plasmids. Serum samples collected from four patients with chronic mutant HBV infections during lamivudine therapy were analyzed using this method. RESULTS: The detection limit for YVDD and YIDD was 10 and 50 copies, respectively, whereas the sensitivity was 10% within a mixed virus population. In the clinical samples, mutant strains of HBV could be detected at levels <2.6 log copies/ml of HBV DNA. While 15 of the 21 samples tested by this method were positive for the YMDD mutant, direct sequencing and a reverse hybridization line probe assay (INNO-LiPA HBV DR v2) detected the mutant strain in only 11 and 9 samples, respectively. Moreover, the data for 6 samples analyzed by TA cloning were fully consistent with our TaqMan PCR results. CONCLUSIONS: We successfully established a sensitive and accurate assay for the YMDD mutant of HBV. This method may be useful for monitoring patients treated with lamivudine.     

A longitudinal model for the Mayo Clinical Score and its sub-components in patients with ulcerative colitis

Journal of Pharmacokinetics and Pharmacodynamics - Clinical trials in patients with ulcerative colitis (UC) face the challenge of high and variable placebo response rates. The Mayo Clinical Score...     

Individual-, family- and community-level determinants of full vaccination coverage among children aged 12–23 months in western Kenya

To identify individual-, family-, and community-level determinants of full vaccination status at most challenging areas in Kenya, we conducted a cross-sectional study among children aged 12–23 months and their mothers. 1965 children were involved in this research and their mothers completed a questionnaire. Middle or high knowledge of vaccination schedule (Adjusted Odds Ratio (AOR) = 2.69, 95%CI: 2.01–3.60 or AOR = 8.12, 95%CI:5.50–11.97), medium/long birth interval or first birth (AOR = 2.46, 95%CI: 1.29–4.69 or AOR = 1.84, 95%CI:1.10–3.09 or AOR = 2.14, 95%CI: 1.20–3.84), less than 5 children under five years old (AOR = 1.39, 95%CI: 1.04–1.88) and highest community health worker's (CHWs) performance (AOR = 2.20, 95%CI: 1.39–3.47) were significantly associated with complete vaccination status in the final multiple regression model. In addition, a interaction between literacy and wealth was significantly related in full vaccination status (AOR = 1.38, 95%CI: 1.08–1.75). Increased frequency and quality of CHW visits could be effective intervention to enhance vaccination coverage. Future interventions focusing on vaccination coverage should be given more attention especially to high risk group identified in this study.     

Prenatal developmental toxicity studies of 1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane (p, p'-DDT) in rats and rabbits

ABSTRACT  The studies were conducted in rats and rabbits to elucidate the potential developmental toxicity of p, p '-DDT in general accordance with the improved Japanese MAFF guidelines (12-Nousan-No. 8147,2–1–18, 2000). p, p '-DDT suspended in 1% aqueous solution of CMC was administered orally to pregnant Jcl:SD rats on gestational days (GD) 6–19 at a dose of 0,5, 25, or 100 mg/kg/day and to pregnant KbI: JW rabbits on GD 6–27 at a dose of 0,5,20, or 80 mg/kg/day. Maternal animals were killed on the day after the last day of administration for morphological examination of their fetuses with special attention to the reproductive organs.
Adverse effects on maternal animals were found only at the highest dose in both species; i.e. , clonic convulsion (2/24 in rats, 5/22 in rabbits), mortality (1/24 in rats), abortion or premature delivery (4/22 in rabbits), and reduced body weight gains and food consumption. However, the control and treated groups showed comparable values for the numbers of corpora lutea and implants, percent preimplantation losses, number of live fetuses, percent resorptions and fetal deaths, sex ratio, fetal body weights, and placental weights in both species, and anogenital distance and testicular histology in rats. Although fetal examination revealed slightly increased incidence of 27 presacral vertebrae in the highest dose group in rats, there was no treatment-related increase in the incidence of malformations in any of the species.
Based on these results, it is concluded that p, p '-DDT causes no malformations, including male reproductive organ abnormalities, in either rats or rabbits, although it results in an increased incidence of skeletal variations in rats at a maternally toxic dose.     

Predictive performance of the Bayesian analysis: Effects of blood sampling time,population parameters,and pharmacostatistical model

The present paper reports theoretical equations for the predictive performance of the Bayesian forecasting method. The precision of parameter estimates and predicted concentrations for an individual was described by general equations with the aid of a variance-covariance matrix of parameter estimates that involved the Bayes theorem. The equations were applied to assess the predictive performance of the one-point Bayesian method in association with blood sampling time, the population parameters, and the pharmacostatistical model. The simulation study showed that the prediction error in parameter estimates essentially depended upon the sampling time but the magnitude of dependency was affected by the size of inter-and intraindividual variances. With a smaller value of interindividual variance, the dependency on sampling time was less apparent. Effects of sampling time were further examined using clinical data obtained from 20 patients taking theophylline, and the results were in good agreement with the theoretical consideration. The present general equations are useful to investigate the sampling strategy as well as structural and variance modeling on the predictive performance of the Bayesian method.     

Gastrointestinal stromal tumor arising from anorectum: correlation of imprint cytology and radiologic imaging

The diagnosis of gastrointestinal stromal tumor (GIST) is generally established on histopathologic examination of surgical specimens. Gastrointestinal stromal tumor comprises a heterogenous group of neoplasms of the gastrointestinal tract previously referred to as leiomyomas, leiomyosarcomas, or schwannomas. Gastrointestinal stromal tumor arising from anorectum is a rare instance. We report a case of GIST for the correlation of imaging and cytologic features with immunocytochemical staining. A computed tomography and magnetic resonance imaging confirmed a 2-cm tumor growing into the rectal lumen. The central portion of the tumor showed T1-weighted imaging of low signal and suspected central necrosis by the T2-weighted imaging of high signal. Imprint cytology from excised tumors showed isolated or loosely aggregated spindle cells with scanty and fibrillary cytoplasmic processes, nuclear pleomorphism, fine granular chromatin, and irregular nuclear margins. Epithelioid tumor cells showed grooves with abundant cytoplasm and several round nucleoli. Both c-kit and CD34 antigen were positive with strong and diffuse stainability in smears as well as paraffin sections by immunoperoxidase staining. We suggest that the combined use of imaging diagnosis and cytology with immunocytochemical staining are useful initial diagnosis of GIST.     

Radiologic features of papillary adenoma and papillomatosis of the biliary tract

Background: The purpose of this study was to describe the imaging findings of papillary adenoma of the bile ducts. Methods: Imaging modalities including sonography, computed tomography, cholangiography, and endoscopic sonography obtained in five patients with papillary adenoma of the bile duct were retrospectively reviewed and correlated with pathologic findings. Results: In four cases, imaging findings were a dilatation of the bile ducts due to a protruding mass within the lumen of the common bile duct. In one case, the biliary tract obstruction was not due to the mass but to mucus secretion that was detected at sonography, endoscopic sonography, and cholangiography. Multiple lesions were observed in two cases at pathology and not detected preoperatively. All the lesions contained foci of in situ carcinoma or mild dysplasia. Conclusion: Imaging is useful in detecting bile duct tumors. Hypersecretion of mucus is rare but highly characteristic of bile duct adenoma. Received: 6 July 1995/Accepted: 18 August 1995     

Angiographically demonstrated coronary collaterals predict residual viable myocardium in patients with chronic myocardial infarction: a regional metabolic study

Angiographical demonstration of coronary collateral circulation may suggest the presence of residual viable myocardium. The development of coronary collaterals was judged according to Rentrop's classification in 37 patients with old anteroseptal myocardial infarction and 13 control patients with chest pain syndrome. The subjects with myocardial infarction were divided into 2 groups: 17 patients with the main branch of the left coronary artery clearly identified by collateral blood flow from the contralateral coronary artery [Coll(+)group, male/female 10/7, mean age 56.6 years]and 20 patients with obscure coronary trunk [Coll(-)group, male/female 16/4, mean age 54.9 years]. Thallium-201 myocardial scintigraphy and examination of local myocardial metabolism were carried out by measuring the flux of lactic acid under dipyridamole infusion load. Coronary stenosis of 99% or total occlusion was found in only 5 of 20 patients (25%)in the Coll(-)group but in 16 of 17 patients(94%)in the Coll(+)group(p < 0.001). Redistribution of myocardial scintigraphy was found in 11 of 15 patients(73%)in the Coll(+)group, but only 3 of 18 patients (17%)in the Coll(-)group(p < 0.01). The myocardial lactic acid extraction rate was--13.2 +/- 17.0% in the Coll(+)group, but 9.1 +/- 13.2% in the Coll(-)group(p < 0.001). These results suggest that coronary collateral may contribute to minimizing the infarct area and to prediction of the presence of viable myocardium.     

Activation of mitogen-activated protein kinases and p90 ribosomal S6 kinase in failing human hearts with dilated cardiomyopathy.

OBJECTIVE: A new member of the MAP kinase family, big MAP kinase-1 (BMK1), has been recently identified to promote cell growth and attenuate apoptosis. P90 ribosomal S6 kinase (p90RSK), one of the potentially important substrates of extracellular signal regulated kinase (ERK), regulates gene expression in part via phosphorylation of CREB and the Na(+)/H(+) exchanger. Recently, we have demonstrated that the activity of BMK1, Src (the upstream regulator of BMK1) and p90RSK was increased in hypertrophied myocardium induced by pressure-overload in the guinea pig. However, the abundance and activity of these kinases in human hearts are unknown. METHODS: In addition to the three classical MAP kinases (ERK, p38 kinase, and c-Jun NH(2)-terminal kinase (JNK)), we examined the protein expression and activity of Src, BMK1, and p90RSK in explanted hearts from patients with dilated cardiomyopathy (n=9). Normal donor hearts, which were not suitable for transplant for technical reasons, were used as controls (n=5). RESULTS: There were no significant differences in the levels of protein expression of these kinases between normal and failing hearts. ERK1/2 and p90RSK were activated in heart failure compared to control (P<0.01 and P<0.03, respectively), while the activity of p38 kinase was decreased (P<0.05) and the activity of JNK was unchanged in heart failure. By contrast, the activities of Src and BMK1 were significantly reduced in end-stage heart failure compared to normal donor hearts (P<0.05). CONCLUSION: These data suggest that multiple MAP kinases, p90RSK, and Src are differentially regulated in human failing myocardium of patients with idiopathic dilated cardiomyopathy and may be involved in the pathogenesis of this complex disease.     

A study on inoculum density and reproducibility of drug susceptibility testing by BACTEC MGIT 960

OBJECTIVE: The BACTEC MGIT 960 drug susceptibility system (MGIT AST) has been recently introduced in Japan. The issue of discordant MGIT results compared with the conventionally used Ogawa method has been raised. It has been speculated that discordant results might be due to MGIT inoculum density since there is no standardization step other than dilution of growth for tubes beyond 2 days after MGIT turns out to be positive. In this study, we examined the reproducibility of the MGIT AST system. MATERIALS AND METHODS: Nineteen sputum specimens from drug-resistant and susceptible pulmonary tuberculosis patients were processed with CCE pretreatment reagent (Japan BCG), inoculated into 3 MGIT tubes, and loaded into the MGIT 960. Inocula for MGIT AST were prepared 1, 3, and 5 days after MGIT tubes became positive. Cultures on day 3 and 5 were diluted 1: 5 with saline. Ten-fold dilutions from each positive culture were plated on Middlebrook 7H11 agar plates for CFU determination. MGIT AST results were compared with those of the conventional proportion method on Ogawa egg and Vite-spectrum (Kyokuto), or Pyrazinamidase (Pzase) assay and Kyokuto PZA test. RESULTS AND CONCLUSION: A total of 15 specimens were culture positive in all 3 tubes. Four of 19 cases were removed from the analysis because of negative cultures in one or more tubes. Three of 4 culture negative cases were MDR-TB. Colony counting showed the mean CFU/ml of inocula prepared from tubes 1, 3, and 5 days after MGIT tube became positive were 3.6 x 10(6), 1.6 x 10(6), 3.1 x 10(6), respectively. There was no significant difference although the CFU range was wide (8 x 10(4)-2 x 10(7)). MGIT AST results were consistent among 3 inocula. Moreover, overall concordance rates between MGIT AST and the conventional methods were over 90% for 5 first-line antituberculosis drugs. These results indicate that the BACTEC MGIT 960 system is very useful for rapid diagnosis of drug resistant tuberculosis.