Gene arrays at Pneumocystis carinii telomeres.

In the fungus Pneumocystis carinii, at least three gene families (PRT1, MSR, and MSG) have the potential to generate high-frequency antigenic variation, which is likely to be a strategy by which this parasitic fungus is able to prolong its survival in the rat lung. Members of these gene families are clustered at chromosome termini, a location that fosters recombination, which has been implicated in selective expression of MSG genes. To gain insight into the architecture, evolution, and regulation of these gene clusters, six telomeric segments of the genome were sequenced. Each of the segments began with one or more unique genes, after which were members of different gene families, arranged in a head-to-tail array. The three-gene repeat PRT1-MSR-MSG was common, suggesting that duplications of these repeats have contributed to expansion of all three families. However, members of a gene family in an array were no more similar to one another than to members in other arrays, indicating rapid divergence after duplication. The intergenic spacers were more conserved than the genes and contained sequence motifs also present in subtelomeres, which in other species have been implicated in gene expression and recombination. Long mononucleotide tracts were present in some MSR genes. These unstable sequences can be expected to suffer frequent frameshift mutations, providing P. carinii with another mechanism to generate antigen variation.

Isolates of Pneumocystis jirovecii from Harare show high genotypic similarity to isolates from London at the superoxide dismutase locus.

Pneumocystis jirovecii is the cause of Pneumocystis pneumonia (PCP) in humans. Isolates of P. jirovecii obtained from patients in Harare, Zimbabwe were genotyped at the superoxide dismutase locus. High genotypic similarity to isolates of P. jirovecii obtained from patients in London, UK was observed. These data provide additional support for the hypothesis that P. jirovecii is genetically indistinguishable in isolates from geographically diverse locations.

Genotypes of Pneumocystis jiroveci isolates obtained in Harare, Zimbabwe, and London, United Kingdom.

Isolates of Pneumocystis jiroveci from sulfa-exposed and nonexposed patients from London, United Kingdom, and Harare, Zimbabwe, were genotyped. At the dihydropteroate synthase (DHPS) locus, there was evidence of selection pressure from sulfa drug exposure, and reversal of DHPS genotype ratios occurred when selection pressure was absent or was removed.

Limited asymptomatic carriage of Pneumocystis jiroveci in human immunodeficiency virus-infected patients.

Forty-seven bronchoalveolar lavage fluid samples from 16 human immunodeficiency virus (HIV)-infected patients were used to test the latency model of Pneumocystis infection in the human host. Identification of DNA sequence polymorphisms at 4 independent loci were used to genotype Pneumocystis jiroveci from the 35 samples that contained detectable P. jiroveci DNA. Eighteen of those 35 samples came from patients who did not have Pneumocystis pneumonia (PCP) and had confirmed alternative diagnoses. Seven patients had asymptomatic carriage of P. jiroveci over periods of < or = 9.5 months after an episode of PCP, and in all 7 cases, a change in genotype from that in the original episode of PCP was observed. The absence of P. jiroveci DNA in one-fourth of the 47 samples and the observed changes in genotype during asymptomatic carriage do not support the latency model of infection. Asymptomatic carriage in HIV-infected patients may play a role in transmission of P. jiroveci and may even supply a reservoir for future infections.

A new name (Pneumocystis jiroveci) for Pneumocystis from humans.

The disease known as Pneumocystis carinii pneumonia (PCP) is a major cause of illness and death in persons with impaired immune systems. While the genus Pneumocystis has been known to science for nearly a century, understanding of its members remained rudimentary until DNA analysis showed its extensive diversity. Pneumocystis organisms from different host species have very different DNA sequences, indicating multiple species. In recognition of its genetic and functional distinctness, the organism that causes human PCP is now named Pneumocystis jiroveci Frenkel 1999. Changing the organism's name does not preclude the use of the acronym PCP because it can be read "Pneumocystis pneumonia." DNA sequence variation exists among samples of P. jiroveci, a feature that allows reexamination of the relationships between host and pathogen. Instead of lifelong latency, transient colonization may be the rule.

Pneumocystis carinii.

Pneumocystis carinii is an atypical fungus that causes pneumonia in immunocompromised individuals. P. carinii comprises a heterogeneous group of organisms that have been isolated from a wide range of mammalian host species. P. carinii infection is host species specific, the P. carinii organisms that infect humans have only been found in humans. This review discusses the application of molecular techniques to the study of the biology and epidemiology of P. carinii infection. It addresses the use of DNA amplification for the detection and diagnosis of P. carinii pneumonia. Studies investigating the reservoir of infectious P. carinii organisms, the routes of transmission of the infection, and the emergence of drug resistant strains of P. carinii are also discussed.

Probable mother-to-infant transmission of Pneumocystis carinii f. sp. hominis infection.

A mother and her 4.5-week-old infant had Pneumocystis carinii pneumonia contemporaneously. Genotyping of P. carinii f. sp. hominis DNA at three independent loci showed the same genotype in samples from mother and infant. These data suggest transmission of P. carinii organisms from the mother to her infant.

Oropharyngeal samples for genotyping and monitoring response to treatment in AIDS patients with Pneumocystis carinii pneumonia.

A nested PCR, amplifying a portion of the gene encoding the mitochondrial large subunit ribosomal RNA (mt LSU rRNA) of Pneumocystis carinii sp. f. hominis was applied to oropharyngeal samples obtained on repeated occasions from 12 HIV-infected patients with P. carinii pneumonia (PCP) to monitor response to anti-P. carinii treatment. Genotyping of P. carinii sp. f. hominis was also performed on paired samples of oropharyngeal and broncho-alveolar lavage samples before the start of treatment, and on oropharyngeal samples during the course of treatment, by analysis of sequence variation at the internal transcribed spacer (ITS) regions of the nuclear rRNA operon. When a simple dilutional method was used, a reduction in the amount of amplification product was observed in samples from all patients during the course of treatment. In eight of the 12 patients, a single ITS sequence type was found in the oropharyngeal samples and also in the paired broncho-alveolar lavage sample. A mixed infection was identified in the samples from three patients. In eight patients, the ITS sequence types identified in the oropharyngeal sample were the same as in the broncho-alveolar lavage sample. Nested PCR amplifying the mt LSU rRNA on oropharyngeal samples provides a non-invasive method of monitoring response to treatment of PCP. ITS sequence typing of P. carinii sp. f. hominis from oropharyngeal samples appears to be a reliable alternative to broncho-alveolar lavage samples and provides a non-invasive tool for further epidemiological studies.

A Pneumocystis carinii multi-gene family with homology to subtilisin-like serine proteases.

Copies of multi-gene family, named PRT1 (protease 1), encoding a subtilisin-like serine protease were cloned from the opportunistic fungal pathogen Pneumocystis carinii. Comparison of the nucleotide sequence of a genomic clone and a cDNA clone of PRT1 from P. carinii f. sp. carinii revealed the presence of seven short introns. Several different domains were predicted from the deduced amino acid sequence: an N-terminal hydrophobic signal sequence, a pro-domain, a subtilisin-like catalytic domain, a P-domain (essential for proteolytic activity), a proline-rich domain, a serine/threonine-rich domain and a C-terminal hydrophobic domain. The catalytic domain showed high homology to other eukaryotic subtilisin-like serine proteases and possessed the three essential residues of the catalytic active site. Karyotypic analysis showed that PRT1 was a multi-gene family, copies of which were present on all but one of the P. carinii f. sp. carinii chromosomes. The different copies of the PRT1 genes showed nucleotide sequence heterogeneity, the highest level of divergence being in the proline-rich domain, which varied in both length and composition. Some copies of PRT1 were contiguous with genes encoding the P. carinii major surface glycoprotein.