Phytosterols are present in Pneumocystis carinii

Abstract

Although originally classified as a protozoan, Pneumocystis carinii is now considered to have fungal characteristics. Drugs typically used for the treatment of fungal infections target ergosterol. Because P. carinii is an important pathogen in AIDS and other immunocompromised patients, knowledge of the sterol content of this organism may be useful as a basis for developing new treatment strategies or for improving diagnosis. P. carinii organisms were harvested from infected rat lungs and were purified by filtration. Control preparations from uninfected animals were identically prepared. Lipids were extracted from the organisms and control preparations and were separated into neutral lipid, glycolipid, and phospholipid fractions by silicic acid chromatography. The neutral lipid fraction was further treated by alkaline hydrolysis and was analyzed by reversed-phase high-pressure liquid chromatography (HPLC), gas chromatography (GC), and GC-mass spectrometry (GC-MS). As shown by HPLC, the neutral lipid fraction from infected rats contained a minimum of six peaks, while in control preparations a single peak with a retention time identical to that of cholesterol was observed. The predominant sterol in these preparations was positively identified by GC-MS as cholesterol and constituted 80 to 90% of the total. The remaining peaks had relative retention times similar to those of phytosterols by both HPLC and GC, and the similarity of these sterols to those derived from plants and fungi was confirmed by MS. Ergosterol, however, was not present. These results provide further evidence for a close phylogenetic relationship between P. carinii and fungi and suggest that these sterols could be used as targets for drug development and for improving diagnosis.Pneumocystis carinii is one of the leading causes of opportunistic infection in AIDS patients (30). Despite the importance of this microorganism as an opportunistic pathogen, much basic information about the parasite is lacking. While originally classified phylogenetically as a protozoan, analyses of P. carinii DNA and mitochondrial and rRNA sequences have generally shown a higher degree of homology to the mitochondrial and rRNA sequences of fungi (yeasts) than to those of protozoa (9,13,27,31,34,35). The exact relationship remains to be determined. Analyses of 5S RNA have suggested a somewhat close phylogenetic relationship to members of the Rhizopoda-Myxomycota-Zygomycota group of "protista fungi." Other studies have suggested similarities to Saccharomyces, Candida, and Neurospora spp. and "red yeast" fungi. In general, the arguments in favor of classifying P. carinii as a protozoan are the amoeboid appearance of the trophic form, susceptibility to antiprotozoal drugs such as pentamidine isethionate or trimethoprim-sulfamethoxazole, insusceptibility to antifungal drugs such as amphotericin B, and the lack of ergosterol in Pneumocystis cell membranes. Arguments in favor of including P. carinii among the fungi include similarities of the sporogenous state to ascospore format...

Comprehensive and definitive structural identities of Pneumocystis carinii sterols

“…The structural assignments of several of these sterol components were made by analyses using TLC, GLC, and GLC-mass spectrometry (MS) in conjunction with chemically synthesized authentic standards. Subsequently, Furlong et al (16) and Urbina et al (17) confirmed the structural assignments for some of the major P. carinii sterol components using comparable analytical methods. However, these workers assigned structures for some sterols containing more than one double bond, which is not possible solely by GLC and GLC-MS analyses.…”

“…We previously examined control lungs from normal untreated rats and corticosteroidtreated uninoculated rats and identified campesterol and ␤-sitosterol among the lung sterols (13). Furlong et al (16) reported that brassicasterol [(24R)-ergosta-5,22-dien-3␤-ol, 24-methylcholesta-5,22-dien-3␤-ol] and stigmasterol (24-ethylcholesta-5,22-3␤-ol) were among the eight major sterols in their P. carinii preparations. They did not report on the sterol composition of control rat lungs to determine whether these sterols could have been scavenged from the host by the organisms in their experimental system.…”

Inhibitors of Sterol Biosynthesis and Amphotericin B Reduce the Viability of Pneumocystis carinii f. sp. carinii

“…In contrast, structural analyses of rat-derived Pneumocystis carinii f. sp. carinii sterols demonstrated that the dominant organism-specific sterols have a double bond at C-7 and an alkyl group consisting of one or two carbons at C-24 of the side chain (10,14,17,25). 24-Alkylsterols with a double bond at C-7 of the sterol nucleus are also found in some plants and fungi (e.g., rust fungi) but are not major components of cellular membranes of vertebrates or of pathogens known to infect the lungs of vertebrates.…”

Heterogeneity of Pneumocystis Sterol Profiles of Samples from Different Sites in the Same Pair of Lungs Suggests Coinfection by Distinct Organism Populations

“…The sterol compositions of trophic forms (24) and mixed life cycle stages (6,15,18) of P. carinii f. sp. carinii do not differ.…”