Spread of an Enterococcus faecalis sequence type 6 (CC2) clone in patients undergoing selective decontamination of the digestive tract

Enterococcus faecalis (E. faecalis) is a common cause of nosocomial infection in immunocompromised patients. The presence and dissemination of high‐risk clonal complexes, such as CC2, is an ongoing problem in hospitals. The aim of this work was to characterize 24 E. faecalis isolates from ICU patients undergoing selective decontamination of the digestive tract (SDD) by phenotypical (antimicrobial susceptibility) and genotypical (presence of virulence genes, RAPD‐PCR and MLST) methods. Our results showed high prevalence of the ST6 E. faecalis clone (91.6%), especially adapted to the hospital environment, with a multidrug resistance pattern and a multitude of putative virulence genes. In addition, ST179 (4.2%) and ST191 (4.2%) were detected. By RAPD–PCR analysis, the 22 isolates identified as ST6 showed six different DNA patterns, while the two remaining isolates, ST179 and ST191, showed two additional profiles. CC2 is a known clonal complex with high adaptability to hospital environment and worldwide distribution. The high prevalence of the ST6 clone in the studied population could be related to the presence of gentamicin in the SDD mixture since most strains were gentamicin resistant. Consequently, strict surveillance should be applied for rapid detection and control of this clone to prevent future spread outside the ICU.     

酸枣仁与其混淆品及伪品的RAPD分析

目的 对酸枣仁及其混淆品和伪品进行DNA指纹图谱的鉴别研究，并对分子生物学在中药材鉴别中存在的一些问题进行探讨。方法 应用随机扩增DNA多态性技术对酸枣仁及其易混淆品、伪品进行DNA指纹图谱多态性的比较。结果 DNA指纹图谱显示，各样品间具有不同的遗传关系，根据这些差别成功地进行了酸枣仁及其易混淆品、伪品的鉴别。结论 RAPD可以有效地鉴别酸枣仁与其混淆品及伪品。同时对RAPD中存在的问题进行了讨论。     

柴胡属5种植物RAPD分析与分类鉴定

目的 探讨正晶柴胡与同属近似种包括柴胡Bupleurum chinense、狭叶柴胡B.scozonerifolium、竹叶柴胡B.marginatum、小叶黑柴胡B.smithii var.parvifolium、大叶柴胡B.longiradiatum5种柴胡属植物鉴定的分子证据。方法 CTAB法提取柴胡属原植物总DNA，以随机引物进行非特异扩增。结果 用随机引物OPA－1（5‘－CAGGCCCTTC－3‘），OPD－8（5‘－GTGTGCCCCA－3‘）和OPD－11（5‘－AGCGCCATTG－3‘）能有效地鉴定这5种柴胡属植物。结论 RAPD法可准确鉴定正品柴胡及其近似种。     

山医群体中国地鼠近交E家系RAPD分析

目的　分析山医群体中国地鼠E家系的遗传纯度。方法　应用经过筛选的 3 1条随机引物对中国地鼠E家系 12只个体基因组DNA进行RAPD扩增 ,计算近交个体间的相似系数。结果　所有样品的相似系数0 943 1到 0 9978之间变动 ,平均相似系数为 0 9749,聚类分析得到了这些个体的同源树资料。结论　山医群体E家系有较高的遗传纯度     

食品中污染酵母菌的RAPD分类鉴定

目的探索采用RAPD技术对食品污染酵母进行分类鉴定的可行性。方法采用RAPD技术对从食品中分离到的22株污染酵母菌进行分子分型并以UPGMA法构建以上菌株的带型关系系统树状图。结果22株污染酵母菌总共扩增出11种不同的指纹图谱，经聚类分析可分为2个聚类群。结论RAPD技术可以作为一种有效、快速、简便的对污染酵母菌进行分类鉴定的手段。     

RAPD法对肺炎克雷白杆菌的基因分型

为了探讨肺炎克雷白杆菌的基因分型，我们用煮沸法制备细菌基因组ＤＮＡ，进行随机引物－聚合物酶链反应，所有１８个肺炎克雷白杆菌菌株均产生了特异性的，可重复的扩增条带，提示：ＲＡＰＤ方法可用于肺炎克雷白杆菌的分型鉴定和进行分子流行病学研究。     

Assignment of RFLP,RAPD and isoenzyme markers to Aspergillus nidulans chromosomes,using chromosome-substituted segregants of a hybrid of A. nidulans and A. quadrilineatus

Chromosome-substituted haploid segregants were selected from among the benomyl-induced progeny of an interspecific hybrid produced by polyethylene-glycol-induced fusion of protoplasts of an Aspergillus nidulans master strain and an A. quadrilineatus auxotrophic mutant. These segregants were examined by RFLP, RAPD, and isoenzyme analysis. The A. nidulans ribosomal repeat unit was assigned to chromosome V, while the benA and the pyrG genes were assigned to linkage groups VIII and I, respectively, of A. nidulans. None of the other cloned genes tested (gdhA, amdS and 25s rRNA) showed polymorphism between the two parents. The method was also used to assign RAPD markers and isoenzyme bands of -arylesterase, phosphatases, NAD-dependent malate dehydrogenase, and cellulase, to A. nidulans chromosomes and/or to their A. quadrilineatus equivalents. The isoenzyme and DNA sequences assigned to chromosomes could be used to saturate the genetic map of A. nidulans, or could serve as starting points for the construction of a genetic map of A. quadrilineatus. No method affording the same possibilities has been described so far in Aspergilli. This chromosome-assay method may be a useful alternative to pulsed-field-gel electrophoretic procedures for the assignment of molecular markers to chromosomes.     

Outcrossing in the homothallic oomycete,Pythium ultimum,detected with molecular markers

The oomycete Pythium ultimum is homothallic, thus a single isolate completes the sexual stage in pure culture. It has been generally assumed that homothallic oomycetes are predominantly inbreeding. In P. ultimum, antheridia occasionally develop from hyphae not directly connected to the oogonium and appear to participate in fertilization, suggesting a possible mechanism for outcrossing. We have used molecular markers to confirm that outcrossing can occur between isolates of P. ultimum. Genetic markers based on randomly amplified polymorphic DNA (RAPD) and restriction fragment length polymorphisms (RFLP) were used to distinguish isolates in a collection of P. ultimum. Two isolates displaying a high level of polymorphism were mixed and placed on media which allows the development of the sexual stage. RAPD markers were used to screen single oospore progeny to identify potential hybrids between the two parental isolates. Subsequent self-fertilization of one putative F₁ yielded a F₂ population which demonstrated segregation and independent assortment of RAPD and RFLP markers. A similar strategy was used to show that an isolate which is incapable of producing oospores in pure culture can outcross when mixed with a homothallic isolate. These results suggest that other homothallic oomycetes may be capable of outcrossing, and sexual reproduction may, therefore, play an important role in the generation of variation in homothallic oomycetes.     

Evidence of two genetic entities inBothriocephalus funiculus (Cestoda) detected by arbitrary-primer polymerase chain reaction random amplified polymorphic DNA fingerprinting

The genetic diversity of two samples of Cestoda (Bothriocephalus funiculus, Renaud and Gabrion, 1984) parasitizing two sympatric teleostean species was assessed using random amplified polymorphic DNA (RAPD). A total of 72Bothriocephalus were analyzed individually, and electrophoretic analysis of the amplification products of 65 primers among the 68 tested revealed monomorphic patterns, reflecting the close genetic relatedness within and between the parasites of the two samples. However, 3 primers showed polymorphic patterns at 6 RAPD sites. Analysis of the distribution of these genomic fragments, assuming random mating, showed strong linkage disequilibria (only 8 genetic combinations were observed among the 32 expected). Two genetic entities displaying a high degree of host specificity were evidence within our two samples ofB. funiculus. This powerful molecular technique can be used as a diagnostic tool in studies concerning the biodiversity of related genetic entities and could have broad applications in parasitology.     

Relationships among genotypes, virulence and clinical forms of Sporothrix schenckii infection

This study explored the relationships among genotypes, virulence and clinical forms of Sporothrix schenckii. Genomic DNA from isolates of S. schenckii, collected from different clinical forms of sporotrichosis, was amplified by randomly amplified polymorphic DNA (RAPD). Suspensions of different isolates of S. schenckii were inoculated into healthy BALB/c mice to compare their virulence, and the numbers and distribution of spores were determined by histological analysis. RAPD analysis indicated that the isolates from different clinical forms of sporotrichosis belonged to different genotypes. The mice inoculated with isolates from disseminated sporotrichosis showed an earlier onset of illness and more severe lesions than those inoculated with isolates from lymphocutaneous sporotrichosis, which, in turn, showed an earlier onset of illness and more severe lesions than those inoculated with isolates from fixed cutaneous sporotrichosis. Healthy BALB/c mice injected with isolates from disseminated sporotrichosis died within 10 days, whereas isolates from lymphocutaneous sporotrichosis and fixed cutaneous sporotrichosis failed to cause death. Histologically, mice inoculated with isolates from disseminated sporotrichosis had more spores than those inoculated with isolates from lymphocutaneous sporotrichosis and fixed cutaneous sporotrichosis. Thus, different genotypes may be associated closely with the virulence of different clinical forms of S. schenckii infection.