Preimplantation genetic diagnosis for single gene disorders: experience with five single gene disorders

We report our experience of 14 preimplantation genetic diagnosis (PGD) cycles in eight couples carrying five different single gene disorders, during the last 18 months. Diagnoses were performed for myotonic dystrophy (DM), cystic fibrosis (CF) [ΔF508 and exon 4 (621+1 G>T)], fragile X and CF simultaneously, and two disorders for which PGD had not been previously attempted, namely neurofibromatosis type 2 (NF2) and Crouzon syndrome. Diagnoses for single gene disorders were carried out on ideally two blastomeres biopsied from Day 3 embryos. A highly polymorphic marker was included in each diagnosis to control against contamination. For the dominant disorders, where possible, linked polymorphisms provided an additional means of determining the genotype of the embryo hence reducing the risk of misdiagnosis due to allele dropout (ADO). Multiplex fluorescent polymerase chain reaction (F-PCR) was used in all cases, followed by fragment analysis and/or single-stranded conformation polymorphism (SSCP) for genotyping. Embryo transfer was performed in 13 cycles resulting in one biochemical pregnancy for CF, three normal deliveries (a twin and a singleton) and one early miscarriage for DM and a singleton for Crouzon syndrome. In each case the untransferred embryos were used to confirm the diagnoses performed on the biopsied cells. The results were concordant in all cases. The inclusion of a polymorphic marker allowed the detection of extraneous DNA contamination in two cells from one case. Knowing the genotype of the contaminating DNA allowed its origin to be traced. All five pregnancies were obtained from embryos in which two blastomeres were biopsied for the diagnosis. Our data demonstrate the successful strategy of using multiplex PCR to simultaneously amplify the mutation site and a polymorphic locus, fluorescent PCR technology to achieve greater sensitivity, and two-cell biopsy to increase the efficiency and success of diagnoses. Copyright © 2002 John Wiley & Sons, Ltd.     

One-year survey of opportunistic premise plumbing pathogens and free-living amoebae in the tap-water of one northern city of China

In this study, qPCR was used to quantify opportunistic premise plumbing pathogens (OPPPs) and free-living amoebae in 11 tap water samples collected over four seasons from a city in northern China. Results demonstrated that the average numbers of gene copies of Legionella spp. and Mycobacterium spp. were significantly higher than those of Aeromonas spp. (p?<?0.05). Legionella spp. and Mycobacterium spp. were 100% (44/44) positively detected while P. aeruginosa and Aeromonas spp. were 79.54% (35/44) and 77.27% (34/44) positively detected. Legionella pneumophila was only detected in 4 samples (4/44), demonstrating its occasional occurrence. No Mycobacterium avium or Naegleria fowleri was detected in any of the samples. The average gene copy numbers of target OPPPs were the highest in summer, suggesting seasonal prevalence of OPPPs. Average gene copy numbers of OPPPs in the taps of low-use-frequency were higher than in taps of high-use-frequency, but the difference was not significant for some OPPPs (p?>?0.05). Moderate negative correlations between the chlorine concentration and the gene copy numbers of OPPPs were observed by Spearman analysis (r_s ranged from ? 0.311 to ? 0.710, p?<?0.05). However, no significant correlations existed between OPPPs and AOC, BDOC, or turbidity. Moderate positive correlations were observed between the target microorganisms, especially for Acanthamoeba spp., through Spearman analysis (p?<?0.05). Based on our studies, it is proposed that disinfectant concentration, season, taps with different-use frequency, OPPP species, and potential microbial correlations should be considered for control of OPPPs in tap water.     

Investigation of powered 2-wheeler accident involvement in urban arterials by considering real-time traffic and weather data

Objective: Understanding the various factors that affect accident risk is of particular concern to decision makers and researchers. The incorporation of real-time traffic and weather data constitutes a fruitful approach when analyzing accident risk. However, the vast majority of relevant research has no specific focus on vulnerable road users such as powered 2-wheelers (PTWs). Moreover, studies using data from urban roads and arterials are scarce. This study aims to add to the current knowledge by considering real-time traffic and weather data from 2 major urban arterials in the city of Athens, Greece, in order to estimate the effect of traffic, weather, and other characteristics on PTW accident involvement.

Methods: Because of the high number of candidate variables, a random forest model was applied to reveal the most important variables. Then, the potentially significant variables were used as input to a Bayesian logistic regression model in order to reveal the magnitude of their effect on PTW accident involvement.

Results: The results of the analysis suggest that PTWs are more likely to be involved in multivehicle accidents than in single-vehicle accidents. It was also indicated that increased traffic flow and variations in speed have a significant influence on PTW accident involvement. On the other hand, weather characteristics were found to have no effect.

Conclusions: The findings of this study can contribute to the understanding of accident mechanisms of PTWs and reduce PTW accident risk in urban arterials.     

Applying real-time control to enhance the performance of nitrogen removal in CAST system

A bench-scale reactor（72 L） red with domestic sewage, was operated more than 3 months with three operation modes： traditional mode, modified mode and real-time control mode, so as to evaluate effects of the operation mode on the system performance and to develop a feasible control strategy. Results obtained from fixed-time control study indicate that the variations of the pH and oxidation-reduction potential（ORP） profiles can represent dynamic characteristics of system and the cycle sequences can be controlled and optimized by the control points on the pH and ORP profiles. A control strategy was, therefore, developed and applied to real-time control mode. Compared with traditional mode, the total nitrogen（TN） removal can be increased by approximately 16% in modified mode and a mean TN removal of 92% was achieved in real-time control mode. Moreover, approximately 12.5% aeration energy was saved in real- time control mode. The result of this study shows that the performance of nitrogen removal was enhanced in modified operation mode. Moreover, the real-time control made it possible to optimize process operation and save aeration energy.     

Prenatal diagnosis of autosomal dominant polycystic kidney disease using flanking DNA markers and the polymerase chain reaction

A prenatal diagnosis was carried out on a 9-week-old fetus at risk for autosomal dominant polycystic kidney disease (ADPKD). Ten members of the family were previously typed using five DNA markers linked to the PKD1 locus on chromosome 16, and one marker linked to the putative PKD2 locus on chromosome 2. The polymerase chain reaction (PCR) was used to amplify the D16S125 locus. Pairwise and multipoint lod scores indicated that the family was most likely segregating a PKD1 mutation. The fetus inherited the disease haplotype from the affected parent. Diagnostic accuracy was greater than 99 per cent, taking into account the possibility of genetic heterogeneity.     

SRDAAR-QNPP:a computer code system for the real-timedose assessment of an accident releasefor Qinshan Nuclear Power Plant

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Prenatal diagnosis of fragile X syndrome: Management of the male fetus with a premutation

Direct detection of the fragile X mutation by DNA analysis has greatly simplified prenatal diagnosis of this disease. However, women carrying a fragile X premutation may pass their expanded trinucleotide repeat to sons without expansion to a full mutation. Such sons are predicted to be intellectually normal. In this situation, the accuracy with which the fetal status can be inferred from analysis of chorionic villus sample (CVS) DNA is unclear. We describe such a case, in which it was felt necessary to proceed to fetal blood sampling despite technically unambiguous DNA results from the CVS. The lack of prospective data means that this dilemma may be expected to recur over the next few years when performing prenatal diagnosis on fragile X premutation carriers.     

Applicability of DNA isolated from syncytiotrophoblast vesicles to gene amplification and molecular analysis

Maternal contamination of fetal DNA represents a major problem when highly sensitive molecular techniques are used in the prenatal diagnosis of genetic diseases. For this reason, we have studied the possibility of using DNA isolated from syncytiotrophoblast vesicles as a target of gene amplification (PCR). Three PCR systems were selected which included a repetitive 149 bp fragment of the Y chromosome, the VNTR locus D1S80, and a portion of the β-globin gene. The results of these experiments indicate that DNA isolated from syncytiotrophoblast vesicles is free of maternal contamination and is suitable for gene amplification and DNA analysis.     

广州城市人工水系统中军团菌的发生和分布

综合运用平板分离法、半巢式PCR和EnviroAmp PCR反向杂交检测试剂盒，对广州市部分人工水系统中条件致病菌军团菌属细菌进行分离和检测．结果表明，虽然阳性率随着采样时间和地点的不同而有所差异，但总体来说，广州市人工水系统中存在较高程度的军团菌的污染，中央空调冷却塔水中军团菌阳性率平均为84％，个别路段甚至达到了100%，自来水池中则达到了35％，，但在后者的水样中没有检测到最常见的致病菌嗜肺军团菌．对各种培养方法的优劣及军团菌的季节性发生和分布进行了初步讨论．图2表2参16