No abstract

The suitability of a 16S rRNA-based mycoplasma group-specific PCR for the detection of mycoplasma contamination in cell cultures was investigated. A total of 104 cell cultures were tested by using microbiological culture, DNA fluorochrome staining, DNA-rRNA hybridization, and PCR techniques. A comparison of the results obtained with these techniques revealed agreement for 95 cell cultures. Discrepant results, which were interpreted as false negative or false positive on the basis of a comparison with the results obtained with other methods, were observed with nine cell cultures. The microbiological culture technique produced false-negative results for four cell cultures. The hybridization technique produced false-negative results for two cell cultures, and for one of these cell cultures the DNA staining technique also produced a false-negative result. The PCR may have produced false-positive results for one cell culture. Ambiguous results were obtained with the remaining two cell cultures. Furthermore, the presence of contaminating bacteria interfered with the interpretation of the DNA staining results for 16 cell cultures. For the same reason the hybridization signals of nine cell cultures could not be interpreted. Our results demonstrate the drawbacks of each of the detection methods and the suitability of the PCR for the detection of mycoplasmas in cell cultures.

The use of a 16S rRNA based polymerase chain reaction (PCR) for the detection of Mycoplasma pneumoniae infection was investigated. Sputum samples from 34 patients with respiratory illness and evidence of pneumonia as judged by chest X-ray were analyzed by PCR and microbiological culture. Throat swabs from 14 healthy individuals were used as controls. For serology, an enzyme immunoassay for the detection of immunoglobulin M antibodies and a complement fixation assay were performed. Evidence of Mycoplasma pneumoniae infection was obtained in ten patients (29%), eight of whom were found positive by both PCR and serology. Two of the sputum samples from these eight patients were negative by culture. Of the remaining two patients positive for Mycoplasma pneumoniae, one was positive by PCR and culture but negative by serology, and one was found positive by serology but negative by PCR and culture. Thirteen of the 14 controls were negative by both PCR and serology. One control, however, was negative by serology but positive by PCR, which was probably due to asymptomatic carriage of Mycoplasma pneumoniae. The results of this study indicate the suitability of the PCR for the detection of Mycoplasma pneumoniae in clinical samples as well as its potential value as an additional tool for the diagnosis of infection.

Recently, an rRNA-based polymerase chain reaction (PCR) has been developed for the detection of murine mycoplasmas at both the genus and species level (F.

The risk of cross-colonization and subsequent infection by Pseudomonas aeruginosa in holiday camps for cystic fibrosis patients was studied in 91 children by culturing sputum at their arrival, at their departure, 2 months later, and at regular intervals thereafter. The isolated strains were subjected to serotyping, phage typing, pyocin typing, and genotyping by random amplified polymorphic DNA fingerprinting-PCR. It was concluded from random amplified polymorphic DNA fingerprinting-PCR typing that the Pseudomonas flora was not constant in most children. Some children harbored one genotype, whereas some harbored two or more different genotypes simultaneously. Most culture-positive children easily acquired a strain of another genotype which replaced the former one or coexisted with the original one. The incidence of sputum conversion was 7.7% in previously negative children; the incidence of permanent colonization and infection was 1.9%. This risk was comparable with that observed in the community. We conclude that the risk of cross-infection is trivial compared with the obvious joy and social benefit derived from a holiday camp.