IS6110-限制性片段长度多态性DNA分型在结核分子流行病学研究中的应用

目的 建立宁夏、北京和上海等地结核分支杆菌发离株IS6110－RFLP DNA指纹图谱，观察其流行病学特征。方法 提取结核分支杆菌基因组DNA，经限制性内切酶PvuⅡ切割、琼脂糖凝胶电泳和Southern转印后，用荧光标记的IS6110DNA序列中245bp探针杂交，以核酸化学发光试剂盒探测荧光信号，比较各菌株指 IS6110拷贝数和带型，分析不同地理区域流行菌株的特点。结果 103例结核患的临床分离株，经245bp探针杂交后的指纹图谱显示大部分菌株含8－21个IS6110拷贝，在宁夏和北京地区流行的结核分支杆菌带型具有共同特点，并有成簇分布的现象，在上海分离株中发现1株零拷贝株和1株单拷贝株。结论 IS6110－RFLP DNA分型方法可用于我国流行的结核分支杆菌分子流行病学研究；宁夏分离株与北京分离株在基因上亲缘关系较相近。     

Molecular Typing in Public Health Laboratories: From an Academic Indulgence to an Infection Control Imperative

Using three Austrian case studies, the variegated applications of molecular typing in today''s public health laboratories are discussed to help illustrate preventive management strategies relying on DNA subtyping. DNA macrorestriction analysis by pulsed field gel electrophoresis has become the gold standard for subtyping of food borne pathogens like listeria, salmonella, campylobacter and Bacillus cereus. Using a Salmonella Mbandaka outbreak from the year 2010 as example, it is shown how the comparison of patterns from human isolates, food isolates, animal isolates and feed isolates can allow to identify and confirm a source of disease. An epidemiological connection between the simultaneous occurrence of tuberculosis in cattle and deer with cases of human tuberculosis due to Mycobacterium caprae in 2010 was excluded using mycobacterial interspersed repetitive units variable-number tandem repeats subtyping. Also in 2010, multilocus sequence typing with nonselective housekeeping genes, the so-called sequence based typing protocol, was used to elucidate connections between an environmental source (a hospital drinking water system) and a case of legionellosis. During the last decades, molecular typing has evolved to become a routine tool in the daily work of public health laboratories. The challenge is now no longer to simply type microorganisms, but to type them in a way that allows for data exchange between public health laboratories all over the world.     

Cloning of a novel specific SCAR marker for species identification in Lactobacillus pentosus

Identifying Lactobacillus species using only phenotypic and genotypic (16S rDNA sequence analysis) techniques yields inaccurate results. The objective of this study was to develop species-specific primers based on randomly amplified polymorphic DNA (RAPD) fingerprinting to distinguish species within the closely related Lactobacillus plantarum group. One of these primers, OPD-3, produced a species-specific band that was found only in the tested Lactobacillus pentosus. This specific fragment was isolated from agarose gel and ligated into a vector for DNA sequencing. A pair of primers, SpOPD3Lpen-F1/R1, that were highly specific sequence-characterized-amplified-regions (SCARs) were designed according to the nucleotide sequences of the specific RAPD marker. These primers were used for PCR analysis of the template DNA of the Lactobacillus strains, and a single 542 bp species-specific band was found only in L. pentosus. Using PCR, a novel species-specific primer pair is shown to rapidly, accurately and effectively distinguish L. pentosus from other species in the L. plantarum group of probiotic bacteria.     

光疗前后新生儿的DNA指纹分析

目的：了解光疗对新生儿的ＤＮＡ是否有损伤。方法：采用ＬＥ１１．８小卫星探针，经Ｓｏｕｔｈｅｒｎ印迹法检测接受光疗前后新生儿外周血的ＤＮＡ指纹谱带。结果：２２例接受光疗前后的新生儿ＤＮＡ指纹相比，均未见到任何谱带的改变。结论：光疗未引起新生儿的ＤＮＡ损伤，应用国产蓝光治疗器对高胆红素血症新生儿在４８ｈ内治疗是安全的。     

应用M13引物PCR指纹法分析假丝酵母菌氟康唑耐药性

目的探讨M13微卫星引物PCR指纹法用于假丝酵母菌氟康唑耐药性基因分型的可行性。方法采用纸片扩散法分析假丝酵母菌氟康唑耐药性,并进一步对41株假丝酵母菌进行M13引物PCR指纹分析,采用RAPD200软件邻接法(neighbour joining,NJ)聚类分析电泳带型模式。结果41例主要从痰液和阴道分泌物中分离的假丝酵母菌标本中中,氟康唑药物敏感11株(26.8%),依赖8例(19.5%),耐药22例(53.7%),PCR指纹分析至少可检测到12种不同的条带,条带数目从2条到12条不等,电泳带型模式与感染部位和氟康唑药物反应性有关。结论假丝酵母菌氟康唑耐药性菌珠所占比例较大,M13微卫星引物PCR指纹法可用于假丝酵母菌的分子流行病学调查和氟康唑耐药性基因分型。     

致病性小肠结肠炎耶氏菌DNA多态性

探讨致病性小肠结肠炎耶氏菌（简称：耶氏菌）DNA分子遗传特征。用聚合酶链反应（PCR）、DNA序列分析、随机扩增多态性DNA（RAPD）检测致病及非致病性耶氏菌10株。耶氏菌0：3、0：9、0：5，27血清型均含有致病基因粘附侵袭位点（ail),DNA指纹图谱也极其相似，与非致病型耶氏菌完全不同。然而，非致病性耶氏菌0：22血清型也含有ail基因，DNA指纹图谱在分子构像上部分与致病血清型相似。致病性耶氏菌具有相似的分子遗传特征，并表现为多态性。O：22血清型与致病性耶氏菌存在较近缘的种系进化关系。可能存在潜在致病性。     

An investigation into the use of restriction endonuclease analysis for the study of transmission of Actinomyces

Abstract DNA fingerprints of 28 reference strains of Actinomyces, comprising representatives of different species and serotypes, and 19 isolates recovered from 16 periodontal patients was performed. The aim was to determine the potential of the method for detecting strain differences in terms of discriminatory power and to evaluate its usefulness in the typing of Actinomyces strains for eco-epidemiological studies. Among the 17 restriction endonucleases tested, Bst EII, Pvu II and Sma I proved to be the most suitable for the genus Actinomyces restriction digest analysis. Visual comparisons of Bst EII, Pvu II and Sma I digest patterns of chromosomal DNA revealed clear differences within species but also within serotypes of Actinomyces that are otherwise identical. The method offers the qualities for use as an epidemiological tool for identifying sources and tracing routes of transmission of Actinomyces: stability, reproducibility, ease of preparation and interpretation and enough sensitivity for detection of differences between morphologically and serologically similar strains of Actinomyces.     

Down syndrome and male fertility: PCR-derived fingerprinting, serological and andrological investigations

Down syndrome, the most common birth defect causing mental retardation, is characterized by a specific phenotype including subfertility or sterility and hypogonadism in males. In contrast, several females with Down syndrome have borne offspring. Here, a male with trisomy 21 fathering an infant is described. This observation is verified by serological markers, DNA fingerprinting using different DNA micro-or minisatellites and andrological investigations.