Development of a yeast assay for rapid screening of inhibitors of human-derived Pneumocystis carinii dihydrofolate reductase.

Human-derived Pneumocystis carinii dihydrofolate reductase (DHFR) was expressed in a Saccharomyces cerevisiae strain whose growth depends on complementation by this enzyme. We utilized a quantitative assay to measure the sensitivity of this yeast strain to DHFR inhibitors. This assay should be useful for identifying new inhibitors of human-derived P. carinii DHFR.

Alteration in expression of the rat mitochondrial ATPase 6 gene during Pneumocystis carinii infection.

BACKGROUND: Pneumocystis carinii causes pneumonia in immunocompromised patients with a high morbidity and mortality rate, but the interaction between this organism and the host cell is not well understood. The purpose of this research was to study the response of host cells to P. carinii infection on a molecular level. RESULTS: The technique of mRNA differential display was used to detect genes whose expression may be affected by P. carinii infection. The nucleotide sequence of one differentially displayed DNA fragment was found to be identical to that of the rat mitochondrial ATPase 6 gene, which is a subunit of the F0F1-ATP synthase complex. A four-fold increase in expression of this gene was verified by Northern blot analysis of total RNA extracted from P. carinii-infected rat lung versus that from mock-infected rat lung. Localization of the cells containing ATPase 6 mRNA was accomplished by in situ hybridization. In sections of non-infected rat lung, these cells were found lining the distal parts of the respiratory tree and in apical areas of the alveoli. Histological location of these cells suggested that they were Clara cells and type II pneumocytes. This hypothesis was confirmed by co-localizing the mRNAs for ATPase 6 and surfactant protein B (SP-B) to the same cells by two-color fluorescent in situ hybridization. CONCLUSIONS: The ATPase 6 gene is over expressed during P. carinii infection, and type II pneumocytes and Clara cells are the cell types responsible for this over-expression.

Are cytochrome b gene mutations the only cause of atovaquone resistance in Pneumocystis?

There is evidence that exposure of the opportunistic pathogen Pneumocystis to atovaquone enhances the development of resistance to the drug. Atovaquone is a structural analog of ubiquinone, which binds to the mitochondrial cytochrome bc(1) complex and inhibits electron transport. Like the parasites Plasmodium and Toxoplasma, atovaquone resistance can result from mutations in the cytochrome b gene of Pneumocystis. However, atovaquone resistance cannot be explained by cytochrome b gene mutations in all cases. The discovery that atovaquone also inhibits biosynthesis of ubiquinone in P. carinii may unfold other mechanisms by which drug resistance develops.

Chronic Pneumocystis carinii infection of the liver. A case report and review of the literature.

Pneumocystis carinii infection of the liver is being reported with increasing frequency in patients with acquired immune deficiency syndrome (AIDS). The clinical picture typically resembles hepatitis. We report such an occurrence in a patient with persistent elevation of alkaline phosphatase and gamma-glutamyl transpeptidase with relatively normal transaminases who was found to have P. carinii on antemortem liver biopsy. The differential diagnosis of abnormal alkaline phosphatase and gamma-glutamyl transpeptidase in patients with AIDS should include P. carinii.