We have developed a simple, rapid, and reliable protocol for the small-scale purification of DNA and RNA from, e.g., human serum and urine. The method is based on the lysing and nuclease-inactivating properties of the chaotropic agent guanidinium thiocyanate together with the nucleic acid-binding properties of silica particles or diatoms in the presence of this agent. By using size-fractionated silica particles, nucleic acids (covalently closed circular, relaxed circular, and linear double-stranded DNA; single-stranded DNA; and rRNA) could be purified from 12 different specimens in less than 1 h and were recovered in the initial reaction vessel. Purified DNA (although significantly sheared) was a good substrate for restriction endonucleases and DNA ligase and was recovered with high yields (usually over 50%) from the picogram to the microgram level. Copurified rRNA was recovered almost undegraded. Substituting size-fractionated silica particles for diatoms (the fossilized cell walls of unicellular algae) allowed for the purification of microgram amounts of genomic DNA, plasmid DNA, and rRNA from cell-rich sources, as exemplified for pathogenic gram-negative bacteria. In this paper, we show representative experiments illustrating some characteristics of the procedure which may have wide application in clinical microbiology.

The significance of BKV infections relative to infections by generally tested respiratory agents was investigated in children with acute respiratory disease. Paired sera from 177 children admitted to a hospital for acute respiratory disease were tested for significant rises in antibodies. Sera from seven patients showed a seroconversion to BKV and clinical signs of acute upper respiratory tract infection were exhibited by each of these patients. BKV infections were present in 8% of the patients with upper respiratory tract disease while seroconversions to adenovirus (2%), influenza A virus (1%), parainfluenza virus (5%), RS virus (6%) and mycoplasma pneumoniae (1%) were observed in 15% of the patients with upper respiratory tract disease. BKV was isolated from the urine of one child with tonsillitis with a concomitant seroconversion to BKV. Tonsils from children with recurrent attacks of acute respiratory disease were tested for the presence of BKV DNA by hybridization with a cloned genomic 32P-labeled DNA of prototype BKV. Five of twelve tonsil DNAs showed hybridization with BKV DNA. Each tonsil showing hybridization with BKV DNA contained multiple nonintegrated copies of the BKV genome per diploid amount of host cell DNA. Attempts to recover infective BKV by transfection of primary human embryonic cells with tonsil DNAs or by co-cultivation of tonsillar cells with primary human embryonic cells were unsuccessful.

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Sequential serum samples from 13 homosexual men who seroconverted for antibodies to human immunodeficiency virus (HIV) were tested for HIV antigen. In one of these men, who developed the acquired immune deficiency syndrome (AIDS), HIV antigenaemia preceded the onset of AIDS by more than a year and persisted throughout the course of the disease. This antigenaemia was accompanied by the disappearance of IgG antibody reactivity to the major HIV core protein p24. In none of the 12 others, who all remained without serious disease, were serum concentrations of HIV antigen detected, except on one occasion in one man. All their serum samples showed strong IgG antibody reactivity to p24.Nine children who were infected with HIV in 1981 by plasma transfusion from a single donor were also followed up for HIV

Longitudinal IgG recognition patterns of viral proteins were studied in 15 men who had seroconverted for lymphadenopathy associated virus/human T lymphotropic virus (LAV/HTLV-II). Antibodies to the major viral core protein p24, which is a cleavage product ofthe gag gene encoded precursorprotein pr55, appeared first. These were soon followed by antibodies to pr65 and more gradually by antibodies to the other gag gene encoded cleavage product p18, the env gene encoded transmembrane glycoprotein gp4l, the env gene encoded glycoproteins gp65 and gpllO, and the putative pol gene product p33.In 13 subjects who remained healthy the reactivity to the differept proteins increased or stabilised with time, while in two men who developed acquired immune deficiency syndrome (AIIDS) the reactivity, most noticeably to gag encoded proteins, diminished before or at the-onset of symptoms.