Use of the chemical cleavage of mismatch method for prenatal deficiency diagnosis of alpha-1-antitrypsin

The most common mutation in alpha-1-antitrypsin deficiency, conversion of a G to an A at base 9989 (PI-Z), was detected with the chemical cleavage of mismatch method, demonstrating the power of the method for prenatal diagnosis. Exon V of the gene was amplified using the polymerase chain reaction and heteroduplexes were formed to test for the presence of the mutation. The predicted C mismatch was readily detectable with hydroxylamine, and by making the probe from the chorionic villus sample it was possible to determine that the fetus was heterozygous, not homozygous, for the mutation.     

Prenatal diagnosis of medium-chain acyl-CoA dehydrogenase (MCAD) deficiency in a family with a previous fatal case of sudden unexpected death in childhood

Medium-chain acyl-CoA dehydrogenase (MCAD) deficiency is a potentially fatal inherited disease with a carrier frequency of approximately 1:100 in most Caucasian populations. The disease is implicated in sudden unexpected death in childhood. A prevalent disease-causing point mutation (A985G) in the MCAD gene has been characterized, thus rendering diagnosis easy in the majority of cases. Since the clinical spectrum of MCAD deficiency ranges from death in the first days of life to an asymptomatic life, there are probably other genetic factors—in addition to MCAD mutations—involved in the expression of the disease. Thus, families who have experienced the death of a child from MCAD deficiency might have an increased risk of a seriously affected subsequent child. In such a family we have therefore performed a prenatal diagnosis on a chorionic villus sample by a highly specific and sensitive polymerase chain reaction (PCR) assay for the G985 mutation. The analysis was positive and resulted in abortion. We verified the diagnosis by direct analysis on blood spots and other tissue material from the aborted fetus and from family members.     

Detection of Class I and II integrons for the assessment of antibiotic and multidrug resistance among Escherichia coli isolates from agricultural irrigation waters in Bulacan,Philippines

Contaminated irrigation water may greatly affect not only the quality of produce but also the people exposed to it. In this study, agricultural irrigation waters in Bulacan, Philippines were assessed and found to be contaminated with Escherichia coli (E. coli) ranging from 0.58 to 4.51 log₁₀ CFU/mL. A total of 79 isolates of E. coli were confirmed through polymerase chain reaction (PCR) amplifying the uidA gene and were tested for phenotypic resistance using 10 antimicrobials through the Kirby–Bauer disc diffusion method. Forty-six isolates (58.22%) were noted to be multidrug resistant (MDR) with high resistance rate to cephalothin, tetracycline, streptomycin, ampicillin, trimethoprim, nalidixic acid, and chloramphenicol. Moreover, this study also examined the prevalence of Class I and II integrons accounting to 67.39% and 17.39%, respectively, of the MDR E. coli strains using multiplex PCR. The results imply that the agricultural water used in Bulacan is contaminated with the fecal material of man or other animals present in the area, and the presence of MDR bacteria, which pose a potential threat to individuals in these areas, is alarming. In addition, detection of integrons could be a good marker for the identification of MDR isolates. Lastly, this study could develop strategies for the proper management of farming sites leading to the detection of food-borne pathogens and prevention of infectious diseases.     

天津沿岸海水中创伤弧菌的PCR检测和定量

采用普通PCR方法和SYBR Green实时定量PCR方法,根据创伤弧菌16S rDNA和23S rDNA基因间隔序列(internal transcribed spacer,ITS)设计引物,对渤海湾天津沿岸海水中的创伤弧菌进行了定性和定量检测.8个样品采集自天津市汉沽区沿岸海水,采集时间分别为2008年7月、10月和2009年3月、5月.PCR检测结果表明,这些样品都能扩增出276 bp的ITS序列,这些序列和GenBank上的同源序列(DQ462478)的相似性为96%～98%.用SYBR Green实时定量PCR方法测定了8个海水样品,样品中创伤弧菌的浓度为7.12×10~3～8.40×10~5 个/L,表明天津沿岸海水存在严重的创伤弧菌污染.     

Investigation into the effect of high concentrations of volatile fatty acids in anaerobic digestion on methanogenic communities

A study was conducted to determine whether differences in the levels of volatile fatty acids (VFAs) in anaerobic digester plants could result in variations in the indigenous methanogenic communities. Two digesters (one operated under mesophilic conditions, the other under thermophilic conditions) were monitored, and sampled at points where VFA levels were high, as well as when VFA levels were low. Physical and chemical parameters were measured, and the methanogenic diversity was screened using the phylogenetic microarray ANAEROCHIP. In addition, real-time PCR was used to quantify the presence of the different methanogenic genera in the sludge samples. Array results indicated that the archaeal communities in the different reactors were stable, and that changes in the VFA levels of the anaerobic digesters did not greatly alter the dominating methanogenic organisms. In contrast, the two digesters were found to harbour different dominating methanogenic communities, which appeared to remain stable over time. Real-time PCR results were inline with those of microarray analysis indicating only minimal changes in methanogen numbers during periods of high VFAs, however, revealed a greater diversity in methanogens than found with the array.     

北京地区菜田土壤抗生素抗性基因的分布特征

为研究北京地区菜田土壤抗生素抗性基因(antibiotic resistance genes,ARGs)的污染状况和分布特征,采集了11个长期施用粪肥蔬菜基地的温室土壤和大田土壤,进行了抗生素以及ARGs种类和丰度的检测.结果表明,菜田土壤中四环素类抗生素残留量最高,其次为磺胺类抗生素,温室土壤抗生素残留普遍高于大田土壤.温室和大田土壤磺胺类抗性基因sul1、sul2和四环素类抗性基因tet L的检出率均为100%.温室土壤的Ⅰ类整合子(int I1)检出率比大田土壤高1.5倍.定量PCR的检测结果表明,菜田土壤中sul2和tet L的相对丰度介于10-4~10-2之间,温室土壤sul2和tet L的相对丰度普遍高于大田土壤.sul2的相对丰度与磺胺二甲嘧啶和强力霉素的含量显著正相关(P0.05),tet L相对丰度与抗生素含量无明显相关性(P0.05).本研究结果对于掌握北京地区农田土壤ARGs的污染现状,以及从ARGs角度评估农艺措施具有重要的指导意义.     

基于实时控制技术的CANON工艺稳定性运行

为保证出水水质稳定并提高氨氮去除率,实现CANON工艺的优化,利用SBR反应器进行了基于实时控制技术的CANON工艺稳定性研究.试验过程中,温度控制在30℃±1℃,pH 7~8,通过反应过程中氨氮、硝态氮、亚硝态氮和pH、DO、ORP的变化规律,制定了可行的实时控制策略.结果表明,进水氨氮浓度为917~1 540 mg·L~(-1)时,以6 mg·L~(-1)的剩余氨氮浓度作为控制参数可以满足工艺稳定性的要求,但氨氮传感器存在费用昂贵和误差较大等问题.采用pH、DO和ORP曲线的平台和特征点作为自控参数,可以维持CANON工艺的长期稳定运行,保证氨氮去除率平均维持在99%以上且出水水质稳定.     

石油污染土壤生物修复过程中氮循环功能基因的动态检测

氮循环相关功能微生物在土壤发挥其生态功能中起着重要的作用.为定量分析其中的固氮细菌、反硝化细菌和硝化细菌在石油污染土壤生物修复过程中的演变情况,采用了实时定量PCR技术对其相关功能基因nifH、narG和amoA的拷贝数进行检测.结果表明,污染土壤中nifH、narG和amoA基因拷贝数及其占总16S rRNA基因拷贝数的比例远低于正常土壤,修复后土壤中的同类分析结果与正常土壤接近.表明石油污染物破坏了氮循环相关菌落结构,而生物修复则使其得以恢复.进一步分析了不同修复方式下氮循环功能基因的恢复情况以及土壤中石油烃降解率.也表明同时添加秸秆和菌剂具有最好的修复效果,处理40 d后其nifH、narG和amoA基因拷贝数（以干土计）分别恢复到2.68×106、1.71×106和8.54×104g-1,石油烃降解率达到48%.投加真菌-细菌复合菌剂的效果优于只投加细菌菌剂的效果.本研究结果表明氮循环功能基因的动态监测可以从基因和物种水平上反映土壤修复的效果,为土壤修复的监控和效果评估提供参考.     

短程深度脱氮中试工艺的低温启动和维持

试验采用有效体积为7.0 m3的SBR中试反应器处理生活污水,考察了低温条件下短程深度脱氮工艺的启动方法及稳定性.利用基于鼓风机频率(BF)和pH的实时控制策略对SBR系统进行实时控制.结果表明,SBR系统在低温条件(11～16℃)和50 d内快速启动短程深度脱氮工艺,亚硝酸盐积累率从19.8%上升到90%.在中低温条件(9～28℃)下短程深度脱氮性能维持长达550 d,亚硝酸盐积累率始终维持在95%左右,出水TN浓度维持在3 mg·L-1,TN去除率维持在95%以上.由此可见,低温下启动SBR短程深度脱氮工艺并长期维持完全可行.