ABSTRACT
BACKGROUND: Epidemics of aseptic meningitis due to enteroviruses has occurred annually in the late spring and early summer season. The early detection of such epidemics is important for the prevention of further infection. Enteroviruses consist of 67 serotypes but one or two serotypes may be the causative agents of epidemics in the year and several different serotypes involve sporadic cases. In order to discriminate epidemics from sporadic infection, the serotype should be determined. We evaluated the significance of the single-strand conformational polymorphism (SSCP) of reverse transcription-polymerase chain reaction (RT-PCR) products of 5'-untranslated region (UTR) for the determination of serotypes. METHODS: The specimens of patients were cultured with RD cell and the RT-PCR was performed in case of the positive cytopathic effect. For the amplification of 153-bp of 5'-UTR, primers (EN1: 5'-CTC CGG CCC CTG AAT GCG GCT AAT-3'; EN2: 5'-ATT GTC ACC ATA AGC AGC CA-3') were used. The RT-PCR products were denatured with 95% formamide at 95degrees C for 5 min and SSCP was performed. 12.5% polyacrylamide gel (49/1 acrylamide/bis) was made by using Mighty SmallTM SE245 Dual Gel Caster (Hoefer Scientific Instruments Inc., USA). Electrophoresis was done at 10 mA for 1.5 h, and silver nitrate stain was performed. The SSCP patterns were compared with serotypes determined by sequence analysis of VP1 region. RESULTS: Coxsackievirus A9 (two strains), coxsackievirus A16 (10), coxsackievirus B2 (two), coxsackievirus B3 (two), echovirus 3 (two), echovirus 11 (two), echovirus 16 (seven), echovirus 19 (two), echovirus 30 (three), and enterovirus 71 (six) showed the different SSCP patterns according to their serotypes. The same pattern was observed in the same serotype, except echovirus 30 showing two different patterns. CONCLUSIONS: The SSCP of RT-PCR products of 5'-UTR may be helpful to determine the serotype and discriminate epidemics from sporadic cases. This has the advantage to be able to test the specimens directly without the viral culture. But the serotype should be determined by other method such as neutralization or sequence analysis in case of the first isolate in the epidemic season and the stains showing the new SSCP patterns.
Subject(s)
Humans , Coloring Agents , Electrophoresis , Enterovirus B, Human , Enterovirus , Genotype , Meningitis, Aseptic , Polymorphism, Single-Stranded Conformational , Seasons , Sequence Analysis , Silver Nitrate , Natural ResourcesABSTRACT
PURPOSE: Leptin is a product of ob gene in adipose tissue. This is a important factor for regulation of body weight, especially body fat mass. So it is interested in correlation of leptin with obesity or various diseases related with obesity, particularly diabetes mellitus. It was demonstrated that leptin level was increased in type 2 diabetes and insulin was associated with increased leptin level. This study is to measure leptin levels in type 1 diabetes without insulin resistance, to validate whether insulin increases leptin level and to investigate correlation of leptin with degree of diabetic control and pubertal progression. METHODS: This study included 114 children(boys; 57, girls; 57) with type 1 diabetes, whose blood samples were taken to measure leptin level using RIA(radioimmunoassay). We observed the correlation of leptin with body mass index, HbA1c, insulin dose, duration of disease, and pubertal degree. RESULTS: Mean values of age, BMI, insulin dose, and HbA1c at the measurement of leptin were 13.0+/-5.04 years-old, 18.9+/-3.5kg/m2, 36U/day, and 8.9+/-2.1%, respectively. Thirty seven children were prepubertal and fifty seven children had pubertal characteristics and the rests were not confirmed. With increment of BMI, serum leptin levels were significantly increased in the all subjects, with the same correlation in pubertal female group, but not in pubertal male. There was a relatively weak correlation(r=0.29) of leptin levels with insulin doses. In prepubertal male, there was a significant positive correlation(r=0.55) of leptin with insulin dose, whereas in pubertal male there was shown negative correlation(r=-0.41). Female patients had a significantly higher leptin levels than male, irrespective of pubertal changes or BMI. There was not a significant correlation of leptin levels with duration of diabetes and HbA1c level. CONCLUSION: It is suggested that blood levels of insulin and sex hormones play an important roles in determining the levels of leptin.