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.

The objective of the present study was the development of a diagnostic reverse transcription (RT)-PCR for the specific detection of enterovirus (EV) RNA in clinical specimens controlled by an internal control (IC) RNA. The IC RNA contains the same primer binding sites as EV RNA but has a different probe region. The IC RNA was packaged into an MS2 phage core particle (armored) and was added to the clinical sample to allow monitoring of both extraction efficiency and RT-PCR efficiency. Serial dilutions of the IC RNA were made, and the detection limit of the RT-PCR was tested in a background of EV RNA-negative cerebrospinal fluid. The sensitivity and specificity of the RT-PCR assay were tested by using all 64 known EV serotypes, several non-EV serotypes, and two Quality Control for Molecular Diagnostics ( The human enteroviruses (EVs) are members of the family Picornaviridae, are ubiquitous, and are mainly enterically transmitted. EVs have traditionally been identified by serotype-specific antisera in a virus-neutralizing test, and 66 EV types are known to infect humans (19). The 66 EV serotypes were initially recognized and divided into five major groups: polioviruses (PV; types 1 to 3), coxsackieviruses A (CVAs; types 1 to 22 and 24), coxsackieviruses B (CVBs; types 1 to 6), echoviruses (types 1 to 7, 9, 11 to 27, and 29 to 33), and EV types 68 to 71 (17). Recent molecular analyses have proved that echovirus types 22 and 23 are genetically distinct from the members of the genus Enterovirus and have been reclassified in a separate genus, Parechovirus, in the family Picornaviridae (13,20,23).Infections with EVs cause a wide range of clinical outcomes, such as asymptomatic infections, aseptic meningitis (meningeal inflammation in the absence of a bacterial pathogen), encephalitis, paralytic poliomyelitis, and myocarditis. Although the majority of EV infections do not cause significant disease, infection can cause serious illness, especially in infants and immune-compromised patients. EV infections are the most common cause of aseptic meningitis and account for 80 to 90% of all cases of central nervous system infections for which a possible causative agent is identified (24). In the neonate, aseptic meningitis-induced complications and poor outcomes of EV infections generally occur within the first 2 days of life (1, 2). Aseptic meningitis in immune-competent adults is characterized by sudden onset of fever, but neurological abnormalities are rare, and both short-term and long-term outcomes are generally good. Encephalitis caused by EV infections is a less common but a more severe disease than aseptic meningitis (18,29,30). Immune-compromised children and adults who are infected with EV may develop chronic meningitis and encephalitis, which may last for years before becoming fatal (16).The early clinical symptoms of meningitis caused by viruses, bacteria, and fungi are quite similar and are difficult to distinguish, but the diagnosis, therapy, and outcome of disease caused by these pathogens vary considerably. A reli...

DNA purified from clinical cerebrospinal fluid and urine specimens by a silica-guanidiniumthiocyanate procedure frequently contained an inhibitor(s) of DNA-processing enzymes which may have been introduced by the purification procedure itself. Inhibition could be relieved by the use of a novel lysis buffer containing alpha-casein. When the novel lysis buffer was used, alpha-casein was bound by the silica particles in the first step of the procedure and eluted together with DNA in the last step, after which it exerted its beneficial effects for DNA-processing enzymes. In the present study we have compared the novel lysis buffer with the previously described lysis buffer with respect to double-stranded DNA yield (which was nearly 100%) and the performance of DNA-processing enzymes.

Adenovirus (Ad) isolates from a large number of human immunodeficiency virus (HIV)-infected individuals were compared serologically and genetically with Ad isolates from immunocompetent patients. Between 1982 and 1994, stool and urine samples from 137 subjects with AIDS hospitalized in The Netherlands yielded 143 Ad strains. Forty additional Ad strains were obtained from 35 HIV-positive patients in Manchester, United Kingdom, in 1992 and 1993. Of these 183 HIV-associated Ad strains, 84% belonged to species D and 3% belonged to species C. These strains were compared with 2,301 Ad strains collected during general diagnostic examinations in The Netherlands from 1973 to 1992. Of the latter strains, 5% belonged to species D and 49% belonged to species C. Two of the Ads isolated from fecal specimens of AIDS patients represent new serotypes: candidate Ad serotype 50 (prototype strain, Wan) of subspecies B1 and candidate Ad serotype 51 (prototype strain, Bom) of species D. The DNA restriction enzyme patterns of strains Wan and Bom differed from the patterns of all established prototypes.