Identification of Actinobacillus actinomycetemcomitans: polymerase chain reaction amplification of IktA–specific sequences

Actinobacillus actinomycetemcomitans has been strongly implicated in the etiology of localized juvenile periodontitis. Techniques used in the identification of this periodontal pathogen include cultural, biochemical, immunological and DNA hybridization analysis. In this study, we report the use of polymerase chain reaction (PCR) to amplify unique sequences of A. actinomycetemcomitans. Specific oligo-nucleotide primers LKT2 and LKT3 were designed to hybridize to the A. actinomycetemcomitans lktA gene, which encodes leukotoxin, a putative A. actinomycetemcomitans virulence factor. The LKT2 and LKT3 primers amplified lktA-specific sequences from all 12 A. actinomycetemcomitans strains tested. In another set of experiments, 13 other bacterial species, most of which are normal residents of the oral cavity, were tested with these primers. These PCR amplifications also contained 2 additional primers, RRN4 and RRN5, which served as positive controls; RRN4 and RRN5 were designed to amplify specific sequences of eubac-terial 16S ribosomal DNA (rDNA). PCR amplifications of all bacterial species tested, including A. actinomycetemcomitans , yielded 16S rDNA-specific DNA fragments. Furthermore, each bacterial species tested, with the exception of A. actinomycetemcomitans , failed to amplify lktA sequences. The LKT and RRN primers were used in further PCR experiments to detect A. actinomycetemcomitans directly from gingival fluid samples. The results clearly demonstrate the simplicity, rapidity, specificity and accuracy of the LKT primers in the identification of A. actinomycetemcomitans.     

广州市一起志贺氏菌痢暴发的分子流行病学分析

本文对61株福氏2a(Shigella flexneri 2a)分离株进行了耐药谱、质粒图谱、菌落原位杂交和Southern blot分析。结果显示:从广州1992年暴发的一起菌痢流行中分离的30株暴发株和当年在广州分离的28株散发株的耐药谱和质粒图谱相同,而且菌落原位杂交证明它们均有4.7kb的ipaBC基因,Southern blot进一步指出了上述分离株的ipa BC基因位于7.6kb,2.5kb和1.6kb的EcoR Ⅰ片段。从分子水平证明了1992年广州市一起志贺氏菌痢(幅氏2a)暴发为单一克隆株引起,该克隆为当地1992年福氏菌的优势克隆。而福州1991年分离的3株同型菌分属不同的克隆。     

黄花败酱皂甙的分离鉴定

从黄花败酱根茎中用柱层析方法分离出一单体化合物，经化学反应和光谱分析鉴定为齐墩果酸－3－β－0－α－L－鼠李吡喃糖基（1→2）－α－L－阿拉伯吡喃糖甙，即β-常春藤素（β-hederin）．     

抗氧化性样品的剖析

从一份抗氧化性样品中分离出三个组分（Ⅰ、Ⅱ和Ⅲ）。组分Ⅰ经元素分析和ＩＲ光谱鉴定证明是双苯胺棉酚，这个鉴定结果和先前的１ＨＮＭＲ鉴定结果是一致的。组分Ⅱ通过元素分析、质谱和ＩＲ光谱鉴定证明是三聚氰胺。组分Ⅲ，ＩＲ光谱鉴定证明是次磷酸二氢纳。分离过程的平均回收率为（９７．２±３．２）％．     

Simple and rapid identification of low level hepatitis B virus DNA by the nested polymerase chain reaction

A rapid and sensitive method has been developed to detect hepatitis B virus DNA (HBV) by nested polymerase chain reaction (PCR) technique with primers specific for the surface and core regions in capillary thermal cycler within 80 min. The lower limit for detection by present PCR method is 10⁻⁵ pg of recombinant HBV DNA which is equivalent to that determined by one round of PCR amplification and Southern blot hybridization analysis. When boiled HBV positive serum was serially diluted 10-fold, HBV DNA was successfully determined in 1 μl≈10⁻³ μl of serum. HBV DNA was detected by present method in 69 clinical samples including HBsAg positives and negatives by enzyme-linked immunosorbent assay (ELISA). When serum samples were amplified by nested PCR using surface and core region primers, HBV DNAs were detected in 37 of 69 samples (53.6%) and 18 of 69 samples (26.1%), respectively. These results can inform the infectious state of HBsAg positive pateints. A simple and rapid nested PCR protocol by using boiled serum as DNA template has been described for the clinical utility to determine HBV DNA in human serum.     

Typing of consecutive methicillin-resistant Staphylococcus aureus isolates from intensive care unit patients and staff with pulsed-field gel electrophoresis

Six consecutive methicillin-resistant Staphylococcus aureus (MRSA) isolates obtained in a 2-month period from tracheal aspirates of six intensive care unit (ICU) patients with nosocomial pneumonia and two MRSA isolates from nasal carriers among staff were typed to determine whether one or more strains were involved and whether nasal carriage was the source of the outbreak. Pulsed-field gel electrophoresis (PFGE) of chromosomal DNA was used to type MRSA isolates. The typing showed that the outbreak was caused by a single epidemic MRSA clone. The MRSA strain isolated from the staff was unrelated to the outbreak strain and was therefore not the source of the outbreak in this study. The source was apparently the index patient followed by transfer of MRSA to other patients on medical equipment or on the hands of staff who did not adhere strictly to infection control measures.     

A Bayesian approach to validating STR multiplex databases for use in forensic casework

A previous paper in this journal has described the conventional statistical analysis of three databases (Caucasian, Afro-Caribbean and Asians from the Indian subcontinent) where individuals are typed at six short tandem repeat (STR) loci. This paper presents a Bayesian analysis of the same data and the approach is centred on the concept of estimating coancestry coefficients from mixed databases. Posterior distributions for the three databases are presented and discussed and the consequences of implementing bootstrap estimation procedures are also shown. Received: 18 November 1996 / Received in revised form: 21 April 1997     

High pressure liquid chromatography and electrospray ionization mass spectrometry are advantageously integrated into a two‐levels approach to detection and identification of haemoglobin variants

Detecting and correctly identifying haemoglobin (Hb) variants is typically achieved by a two‐levels laboratory approach. We report our experience in dealing with 91 Hb variants, including a number of frequent and a few rare variants. Screening included akaline agarose gel electrophoresis (AGE), ion‐exchange automated high‐performance liquid chromatography (HPLC) and a test for deoxyhaemoglobin solubility. Identification was based on electrospray ionization–mass spectrometry (ESI–MS). Our results confirmed the advantages of HPLC over AGE for screening, because of the occurrence of some electrophoretically ‘silent’ variants. ESI–MS permitted the definitive identification of 90 of the 91 variants included in the study, in some cases (e.g. HbS) through the application of a simple protocol (direct injection of the sample), in other cases requiring the application of more demanding procedures (purification of the variant chain and peptide analysis after enzymatic or chemical cleavage). In an additional case (Hb J‐Oxford), ESI–MS assay did not lead to definitive identification, but gave indications for designing the appropriate primers to focus DNA sequence analysis on the specific region of the gene. Deoxyhaemoglobin solubility test was positive only in the presence of HbS. We conclude that HPLC and ESI–MS are advantageously integrated into a two‐level analytical system for the detection and confirmation of variant Hbs.     

中药石斛显微鉴定研究Ⅲ

本文报道了剑叶石斛Dendrobium acinaciforme Foxb等8种“石斛”的显微鉴定。主要比较其表皮和角质层,皮下层细胞,维管束、外侧纤维群,木质部导管,内侧纤维群等。并附有这8种石斛茎横切面图。