Genetic identification and detection of human pathogenic Rhizopus species, a major mucormycosis agent, by multiplex PCR based on internal transcribed spacer region of rRNA gene

BACKGROUND: Mucormycosis is an invasive opportunistic infection caused by fungi belonging to the order Mucorales. Due to the lack of laboratory tests, the diagnosis of mucormycosis is notoriously difficult. Added with its rapid progression as well as the debilitated state of the patients who contract the disease, mortality is extremely high. OBJECTIVE: The goal of this study was to genetically identify human pathogenic Rhizopus species, a major mucormycosis agent, by the internal transcribed spacer (ITS) region of rRNA gene. METHODS: Primers were designed to identify five Rhizopus species known to cause human disease by multiplex PCR. PCR was done not only with test strains and clinical isolates, but also with clinical samples from cutaneous mucormycosis patients. Sporangiospore morphology was observed by scanning electron microscopy to confirm the correlation of phenotypic and genotypic features. RESULTS: Multiplex PCR identified five Rhizopus species including Rhizopus oryzae, where R. azygosporus could only be distinguished from R. microsporus by certain polymorphisms that were present in its sequence. When this multiplex PCR was applied to clinical samples from three mucormycosis patients (paraffin sections from all and sera from one patient), Rhizopus DNA corresponding to the isolated pathogens were specifically detected. CONCLUSION: While fungal DNA detection from clinical samples is a rigorously studied area, this is the first report to genetically identify and detect Rhizopus species from human mucormycosis specimens. This may expand the possibility of this multiplex PCR system not only to identify isolated fungi, but also as a screening method for visceral mucormycosis.     

Three patients with middle-age-onset hemochromatosis caused by novel mutations in the hemojuvelin gene

Hemochromatosis is a genetically heterogeneous condition. Mutations in the recently described hemojuvelin gene were found in patients with juvenile hemochromatosis, who usually manifest clinical signs of iron overload, including cardiomyopathy and hypogonadism, in their teens and early 20s. In this report, we describe three Japanese patients who showed typical clinical and hepatic histological damage compatible with hemochromatosis at around 50 years of age. Genetic analyses showed that all three patients carried mutations in the hemojuvelin gene. The first patient was homozygous for a novel mutation (745G > C [D249H]), and the second and third patients from the same family were homozygous for another novel mutation (934C > T [Q312X]). No mutations in their HFE, hepcidin, transferrin receptor 2, or ferroportin genes were found. One patient had chronic infection with Helicobacter pylori. The age at initial presentation of hemojuvelin-hemochromatosis occurs over a wider range than previously described.     

Changes in the characteristics and survival rate of hepatocellular carcinoma from 1976 to 2000: analysis of 1365 patients in a single institution in Japan

BACKGROUND: The authors analyzed changes in the characteristics and survival rate of patients with hepatocellular carcinoma (HCC) in the past 25 years. METHODS: Trends in clinical characteristics and survival rate of patients with HCC were evaluated retrospectively based on data from 1365 patients who were diagnosed, treated, and followed between 1976 and 2000. RESULTS: Between 1976-1995, the number of patients with smaller tumors, a less advanced tumor stage, and with a lower Child-Pugh class increased markedly. No differences were observed in the distributions of these three factors between the periods 1991-1995 and 1996-2000. The year of HCC diagnosis, tumor size, tumor stage, Child-Pugh class, and the kind of initial treatment received correlated significantly with patient survival rates by multivariate analysis. The year of HCC diagnosis was found to contribute independently to the improvement in patient survival rates. Using the Kaplan-Meier comparison, the time periods during which the highest patient survival rates occurred were found to be 1991-1995/1996-2000, 1986-1990, and 1976-1985, in that order. The authors did not observe a difference with regard to survival rates between patients in the 1991-1995 and 1996-2000 groups. CONCLUSIONS: The characteristics of patients with HCC changed dramatically from 1976 to 1995 (but not in the past 10 years) toward the earlier detection of HCC. This contributed to the improvement noted in patient survival rates during this period. The year of HCC diagnosis was found to be an independent factor for the improved survival rates by multivariate analysis. This indicated that the progress of treatment and care for patients with HCC contributed to the annual improvement in patient survival rates.     

Incidence and clinical features of Giardia lamblia

To determine the incidence and clinical features of Giardia lamblia infection, we studied 1790 patients at Kochi Medical School Hospital from April 1998 to July 2001. Fecal samples were examined microscopically by the direct smear method, direct immunofluorescent assay and by Kohn's one-step staining for G. lamblia cysts. Cysts of G. lamblia were found in 17 of 1,790(0.95%) stool samples, indicating that G. lamblia infection is not rare in Kochi. The most characteristic feature was that G. lamblia-positive cases were more frequent in the advanced age group(41-79 years old) and most of the subjects (except 2 cases) with G. lamblia had no history of traveling overseas. Four subjects had symptoms related to G. lamblia infection. Thus, more attention should be given to parasitic infections in laboratory stool examinations in order to detect cyst carriers as potential sources of infection.     

Adrenomedullin concentrations in early 2nd-trimester amniotic fluid: relation to preterm delivery and fetal growth at birth

OBJECTIVE: The purpose of this study was to evaluate whether adrenomedullin concentrations in the early 2nd-trimester amniotic fluid predict preterm delivery or fetal growth at birth. METHODS: The adrenomedullin concentrations in early 2nd-trimester amniotic fluid were measured in 70 pregnancies with term delivery and in 3 pregnancies with preterm delivery. Total and free adrenomedullin concentrations were measured from early 2nd-trimester amniotic fluid samples using an immunoradiometric assay. RESULTS: The amniotic fluid total adrenomedullin concentrations in women with preterm delivery were significantly higher (129.7 +/- 19.6 fmol/ml) than those in women with term delivery (92.5 +/- 28.2 fmol/ml; p < 0.05). There were no significant differences for amniotic fluid free adrenomedullin concentrations and free/total adrenomedullin ratios between the two groups. Total or free adrenomedullin concentrations in the early 2nd-trimester amniotic fluid showed an inverse correlation both with birth weight (r = 0.27, p < 0.05, and r = 0.21, p < 0.05) and height (r = 0.30, p < 0.05, and r = 0.28, p < 0.05). There were no correlations between placental weight and total or free adrenomedullin concentrations in the early 2nd-trimester amniotic fluid. CONCLUSIONS: These results suggest that adrenomedullin concentrations in the early 2nd-trimester amniotic fluid might be related to further in utero fetal growth and that high levels of adrenomedullin in the early 2nd-trimester amniotic fluid may be involved in the occurrence of preterm delivery.     

Persistent müllerian duct Syndrome in brothers

Persistent Müllerian duct syndrome is associated with cryptorchidism and transverse testicular ectopia. Such gonads are at an increased risk of malignant transformation. Furthermore, most patients have azoospermia. Here in we report about two brothers with persistent Müllerian duct syndrome. The diagnosis was made during surgical operation for testicular cancer in younger brother. In the other one, persistent Müllerian duct syndrome was diagnosed during examination for infertility.