Detection of DNA from infectious laryngotracheitis virus by colourimetric analyses of polymerase chain reactions

A combination of the polymerase chain reaction and a novel ELISA-type DNA colourimetric assay (developed from studies with a retrovirus from man) was used in a preliminary study to detect DNA from avian infectious laryngotracheitis virus. The method is sensitive, specific and easy to perform. Since it can be readily adapted for the detection of DNA from other sources it could be useful for the identification of a variety of pathogens from other species of veterinary importance.     

Hepatitis C viraemia in adults with type 2 autoimmune hepatitis.

Sera from 14 patients with type 2 autoimmune hepatitis (anti-LKM1 positive) were investigated for evidence of hepatitis C virus (HCV) infection. Antibodies to HCV were detected in 13 patients by both commercial and "in-house" ELISAs and also by a second generation recombinant immunoblot assay. Nine of the 13 (69%) anti-HCV positive patients were shown to be viraemic by a polymerase chain reaction-based assay for serum HCV RNA. Neither anti-HCV nor serum HCV RNA were detected in any of 6 controls with primary biliary cirrhosis or in 39 healthy blood donors. These findings strongly suggest a role for HCV in the pathogenesis of type 2 autoimmune hepatitis.     

Paraffin wax-embedded material as a source of DNA for the detection of somatic genetic changes

Loss of genetic material, corresponding to chromosomal deletions, has been detected in a wide range of tumours and may indicate the position of a tumour suppressor gene. In order to identify the position of such a gene more precisely, many tumour samples must be studied until a minimum consensus deletion is characterized. This process is particularly necessary for lung tumours in which the deletion in chromosome 3, seen with such high frequencies in all histological subtypes, is almost always large. We have recently described the use of the polymerase chain reaction (PCR) for restriction fragment length polymorphism (RFLP) analysis of DNA isolated from small bronchial biopsies of lung tumours. In this study we adapted this technique to allow genotyping of DNA isolated from paraffin wax-embedded material (PWEM) microdissected from glass slides. We have investigated 12 lung tumours at polymorphic loci on chromosome 3 and showed allelic loss in all samples. In adapting PCR–RFLP analysis for DNA isolated from PWEM, we have concentrated on those approaches which might be adaptable to routine clinical practice. Somatic genetic changes are now being identified in many tumour types, and this information is expected to be of diagnostic and prognostic significance.     

Genotypic analysis of sequential genital herpes simplex virus type 1 (HSV-1) isolates of patients with recurrent HSV-1 associated genital herpes

Clinical recurrences of Herpes simplex virus type 1 (HSV-1)-associated genital herpes are thought to be caused by reactivation of latent endogenous HSV-1. However, the possibility of reinfection with exogenous HSV-1 cannot be excluded. This study aimed to determine the incidence of genital HSV-1 superinfection in patients by investigating the genotype of sequential HSV-1 isolates obtained from the same anatomical site of patients with clinical recurrences of genital HSV-1 recurrent genital herpes. Sequential genital HSV-1 isolates were genotyped by PCR amplification of the hypervariable regions located within the HSV-1 genes US1 and US12. Whereas the sequential HSV-1 isolates in 11 of the 13 patients studied had the same genotypes, the sequential isolates of 2 patients showed a different genotype. The data suggest that HSV-1-induced recurrent genital herpes can be associated with genital reinfection with an exogenous HSV-1 strain.     

Rapid determination of zygosity and common aneuploidies from amniotic fluid cells using quantitative fluorescent polymerase chain reaction following genetic amniocentesis in multiple pregnancies

Following second-trimester twin amniocentesis, we used quantitative fluorescent polymerase chain reaction (QF-PCR) assays and polymorphic small tandem repeats (STR) for rapid determination of zygosity and common aneuploidies from amniotic fluid (AF) cells in four pregnancies with like-sex twins, fused placentae and inconclusive chorionicity. The first and the second cases were suspected to have inadvertent sampling of the same amniotic cavity twice. The first case showed a dizygotic (DZ) pattern and repeat amniocentesis was thus avoided. The second case was monozygotic (MZ) and was complicated by discordant fetal growth and twin-twin transfusion syndrome. The third case was associated with a co-twin malformation, occipital encephalocele. DNA studies revealed MZ twinning with a discordant structural defect. The fourth case was associated with co-twin abnormalities of cystic hygroma and hydrops fetalis. DNA studies showed DZ twinning with discordant structural and chromosomal defects. The QF-PCR assay with STR has the advantages of rapid determination of zygosity and common aneuploidies in AF cells. This simple test appears to be useful in the instances of possible inadvertent puncture of the same amniotic cavity twice during amniocentesis and of discordant fetal structural and/or chromosomal abnormalities following genetic amniocentesis in multiple pregnancies with uncertain chorionicity.     

Clonorchis sinensis: molecular cloning and localization of myosin regulatory light chain

One cDNA clone was purified from an adult Clonorchis sinensis cDNA library, and its deduced polypeptide sequence was found to be homologous with myosin regulatory light chain (MRLC) of invertebrates and vertebrates. Two amino-acid residues, Thr and Ser, were conserved at the phosphorylation sites that regulate the function of MRLCs. Recombinant C. sinensis MRLC (rCsMRLC) protein was produced and purified from Escherichia coli, and mouse anti-CsMRLC immune sera recognized a protein of molecular weight 24 kDa from a soluble protein preparation of C. sinensis. The CsMRLC protein was immunohistochemically localized to the muscle fibers of the subtegumental muscle layer and to the muscles of oral and ventral suckers. However, the rCsMRLC protein proved to be less useful antigen for the serodiagnosis of human clonorchiasis.The nucleotide sequence reported herein was submitted to GenBank and assigned accession number AY519356.     

脂质、血红蛋白、胆红素标本对乙肝病毒DNA荧光定量测定的影响

目的:探讨脂血、高胆红素和溶血标本对乙肝病毒DNA(HBVDNA)荧光定量测定结果的影响。方法:将乙肝大三阳高脂血和非脂血、溶血血清和未溶血血清同时作HBVDNA荧光定量检测;将HBVDNA阴性黄疸血清和HBVDNA阴性正常血清与来自同一份乙肝大三阳血清混合,在相同条件下进行HBVDNA荧光定量。结果:乙肝大三阳溶血与未溶血样本HBVDNA含量都在同一数量级。乙肝大三阳高脂血的HBVDNA含量明显低于对照标本。高黄疸血清、正常对照血清与相同的HBVDNA阳性模板组合后所测得的HBVDNA结果无差异。结论:脂血对HBVD-NA定量测定有严重干扰;溶血样本、高胆红素样本对HBVDNA测定结果无影响。     

应用聚合酶链反应技术检测肝组织中的HBV－DNA

采用聚合酶链反应（ＰＣＲ）技术，对４２例肝活切组织石蜡切片中乙型肝炎病毒（ＨＢＶ）ＤＮＡ进行检测，并与乙肝表面抗原（ＨＢｓＡｇ）的免疫组织化学及血清学检测进行比较，ＨＢＶ－ＰＣＲ阳性率为７３．８％，高于组织及血清ＨＢｓＡｇ阳性率（分别为５９．５％和５０．０％）。３例病理形态呈肝炎改变，而血清ＨＢｓＡｇ（─）的肝组织中有２例检出ＨＢＶ－ＤＮＡ，提示ＰＣＲ的高度敏感性和准确性。８３．３％的门脉性肝硬变和８７．５％的肝细胞癌组织中ＨＢＶ－ＰＣＲ呈阳性，进一步证实了上述两病与ＨＢＶ的关系密切。我们还发现肝细胞淤胆患者ＨＢＶ感染率较高，ＨＢＶ－ＤＮＡ及组织ＨＢｓＡｇ阳性比例各为６／９和４／８。     

Histopathology of gastric carcinoids: a survey of 42 cases

An unselected series of 42 gastric carcinoids has been reviewed. Clinically the tumours simulated common gastric lesions including ulcer, polyp and carcinoma. No endocrine symptoms were identified. The tumours were most frequent in the body of the stomach and in 25% in that site were multiple. Morphologically most tumours when classified according to Soga (1974) demonstrated a mixed growth pattern. Six tumours displayed an atypical morphology (type D): they were larger and metastasized more frequently than the rest of the tumours. Six tumours contained a few scattered argentaffinic cells but the others were negative indicating negligible serotonin secretion in only a few cases. The Grimelius argyrophilic reaction was positive in most cells in all tested tumours except in three, two of which showed atypical morphology (type D). It is suggested that gastric carcinoids with a type D morphology or a minority cell population of argyrophil cells are dedifferentiated carcinoids which are biologically nearer to gastric carcinomas. The most frequent clinicopathological correlation was achlorhydria linking pernicious anaemia and gastric carcinoids. This indicates pathogenetic similarities between gastric carcinoids and gastric carcinomas.     

Favourable associations between the myosin heavy-chain and light-chain isoforms in human skeletal muscle

Histochemical and biochemical analyses were performed in order to examine the relationship between myosin light-chain (LC) isoforms and fibre-type distributions in whole human skeletal muscle. Muscle biopsies were obtained from the vastus lateralis muscle in six healthy men, and analysed for the relative area occupied by each fibre type (percentage of fibre type area) and the molar ratio of each LC isoform. The percentage of type I fibre area was positively correlated with the molar ratio of slow LC (LC1s and LC2s) to total LC. The regression line was located below the line of unity. Also, the ratio of percentage of type II fibre area to that of type II fibre area was positively correlated with the molar ratio of the fast alkali LC LC1f to fast alkali LCs LC1f and LC3f. These results support previous study, having shown that in human skeletal muscle some type I fibres express various amounts of fast LC in addition to slow LC and suggest that fast myosin heavy-chain HCII a is favourably associated with LC1f, whereas HCIIb is favourably associated with LC3f.