Typing of Verotoxins by DNA Colony Hybridization with Poly- and Oligonucleotide Probes,a Bead-Enzyme-Linked Immunosorbent Assay,and Polymerase Chain Reaction

To identify the type of Verotoxins (VT) produced by Verocytotoxin-producing Escherichia coli (VTEC), a sensitive bead-enzyme-linked immunosorbent assay and polymerase chain reaction with common and specific primers to various VTs (VT1, VT2, VT2vha, VT2vhb, and VT2vp1) were developed. Together with colony hybridization tests with oligo- and polynucleotide probes, these methods were applied to VTEC isolates to type the VT produced. The toxin types of 26 of 37 strains were identified, but the reaction profiles in assays of the remaining 11 strains suggested the existence of new VT2 variants. The application of these identification procedures may be useful as a tool for clinical and epidemiological studies of VTEC infection.     

抗原捕获聚合酶链式反应（AC－PCR）直接分型检测单纯疱疹病毒

用抗单纯疱疹病毒（ＨＳＶ）型共同性ｇＣ和ｇＤ羊克隆抗体（ＭｃＡｂ）,包被即Ｅｐｐｅｎｄｏｒｆ管,捕捉ＨＳＶ,同时加入３个引物：一个是ＨＳＶ─１／ＨＳＶ─２型共同性上游引物,另两个分别是ＨＳＶ─１和ＨＳＶ─２型特异性下游引物。借此建立了能直接分型检测ＨＳＶ的抗原捕获聚合酶链式反应（ＡＣ─ＰＣＲ）。ＨＳＶ─１的扩增产物为４７７ｂｐ,ＨＳＶ─２的为３９９ｂｐ两型病毒经ＡＣ─ＰＣＲ扩增后产生分子量不同的ＤＮＡ片段,致使ＡＣ─ＰＣＲ能直接分型检测ＨＳＶ。ＨＳＶ─１和ＨＳＶ─２扩增产物的克隆和序列分析表明,本方法特异性好。用本法检测Ｂａｌｂ／ｃ幼鼠中枢神经系统ＨＳＶ感染的脑标本,进一步证实本方法不仅敏感、特异,而且分型准确。     

Microbes in food processing technology

Abstract: There is an increasing understanding that the microbial quality of a certain food is the result of a chain of events. It is clear that the microbial safety of food can only be guaranteed when the overall processing, including the production of raw materials, distribution and handling by the consumer are taken into consideration. Therefore, the microbiological quality assurance of foods is not only a matter of control, but also of a careful design of the total process chain. Food industry has now generally adapted quality assurance systems and is implementing the Hazard Analysis Critical Control Point (HACCP) concept. Rapid microbiological monitoring systems should be used in these cases. There is a need for rapid and simple microbiological tests which can be adapted to the technology and logistics of specific production processes. Traditional microbiological methods generally do not meet these high requirements. This paper discusses the tests, based on molecular biological principles, to detect and identify microbes in food-processing chains. Tests based on DNA technology are discussed, including in vitro DNA amplification like the polymerase chain reaction (PCR) method and identifications based on RFLP, RAPD and DNA fingerprinting analysis. PCR-haled methodology can be used for the rapid detection of microbes in food manufacturing environments. In addition, DNA fingerprinting methods are suitable for investigating sources and routes of microbial contamination in the food cycle.     

Insertion sequence (IS) hybridization supports classification of Rhizobium meliloti by phage typing

Sixty-one isolates of Rhizobium meliloti from two field sites which had been previously classified into 15 phage types on the basis of sensitivity to 16 typing phages, were subjected to insertion sequence (IS) hybridization using DNA probes for ISR m 3 and ISR m 5. Isolates from all but one phage type contained ISR m 3 (apparent copy no. 1–11) and all isolates contained ISR m 5 (apparent copy no. 3–11). The isolates were placed into 24 IS classes based on differences in their respective ISR m 3 and ISR m 5 hybridization profiles. At either field site, isolates representing different phage types possessed IS hybridization profiles that differed from each other, while those comprising a specific type had identical or closely related profiles. Isolates from one phage type were unusual since they did not react with any of the typing phages and were shown by IS hybridization to constitute a heterogeneous group. Evidence for spatial effects were provided by isolates from two of six types present at both sites which fell into separate IS classes on the basis of their site of origin. These data have ecological implications and suggest that for a particular site, phage typing may be employed for the rapid assessment of the genetic diversity among field isolates.     

Restriction Fragment Length Polymorphism (RFLP) of Genomic DNA of Moraxella (Branhamella) catarrhalis Isolates in a Hospital

Epidemiological typing, based on restriction fragment length polymorphism (RFLP) by pulsed-field gel electrophoresis (PFGE), was attempted for the 38 clinical isolates of Moraxella catarrhalis obtained at Shinshu University Hospital during the years 1987 and 1993. Digestion with SmaI or NotI generated well separable, 12 to 5 genomic DNA fragments ranging from 1,000 kb to 30 kb and the strains could be classified into 14 or 13 types, respectively. The electrophoretic profile differed with the strain in most of them and was hence useful to distinguish the each strain. Investigation for their RFLP have, however, suggested that majority of them, including the type strain ATCC25238, may have derived from a common ancestor.     

A hypervariable middle repetitive DNA sequence from citrus

The use of hypervariable sequences for DNA typing of plants is focussed on microsatellites and on amplification of regions defined by random (RAPD) or defined (AFLP) primers for PCR analysis of genomes. A hypervariable length of middle repetitive DNA has been isolated from citrus that contains no obvious hypervariable structures. The fingerprinting probe was shown to have an important commercial application in the separation of zygotic from nucellar progeny. A somatic variant of the sequence within one orange tree suggests that somatic variation in hypervariable markers may be a common event.     

Genome analysis of Pseudomonas aeruginosa by field inversion gel electrophoresis

Abstract The genome of Pseudomonas aeruginosa was analysed by digestion with rare-cutting restriction endonucleases and subsequent field inversion gel electrophoresis (FIGE). P. aeruginosa strain PAO and the 17 IATS strains were investigated. Each strain displayed a unique pattern of restriction fragments. Digestion with Dra I and Ssp I yielded, respectively 7–11 and 2–5 fragments of more than 130 kb in size, indicating the non-random occurrence of AT-rich sequences in the P. aeruginosa genome. The genome size of P. aeruginosa PAO was estimated to be (2.2 ± 0.3) × 10⁶ bp. The applications of DNA fingerprinting for gene cloning, construction of a physical chromosome map, and epidemiological studies, are discussed.     

乳腺癌分子分型的研究进展

乳腺癌是目前世界上女性最为常见的恶性肿瘤之一。随着个体化的乳腺癌的化疗、内分泌治疗和靶向治疗的广泛使用,乳腺癌的分子分型检测也因此得到高度重视和广泛使用。2013年St.Gallen国际乳腺癌会议上重新定义了乳腺癌的分子分型标准,使Luminal A型和Luminal B型乳腺癌的比例有所改变,导致原本分类为Luminal A型的部分患者重新分类为Luminal B型而使Luminal B型患者数量有所增加。人上皮生长因子受体-2(human epidermalgrowth factor receptor-2,HER2)型乳腺癌的靶向药物曲妥珠单抗与其他化疗药物的联合应用及不同时间应用的治疗效果有了进一步研究。雄激素受体(Androgen Receptor,AR)认为可用于常规评估三阴性乳腺癌,除此之外,三阴性乳腺癌的小亚型的分型也为三阴性乳腺癌的个体化治疗提供了新的思路。尽管乳腺癌分子分型方面已取得一定进展,并获得国际同行间的认可,给乳腺癌患者的治疗和预后复发预测提供了重要依据,但目前所有的分子标记物仍不能够满足治疗需求使其临床实用性仍旧具有局限性,因此还期待更多更深入的研究。     

我国若干地区蜡样芽孢杆菌723株噬菌体分型的初步报告

四年来,作者由水中获取蜡样芽孢杆菌噬菌体13株,旨在试作噬菌体分型。待检菌株共723株,由国内16个地区提供(其中食物中毒株121株;食品株602株)。分型结果显示:C29、C30、C24、C27、C19和A_4等型别占优势。中毒株