Sequence Polymorphisms in the<i>Pneumocystis carinii</i>Cytochrome<i>b</i>Gene and Their Association with Atovaquone Prophylaxis Failure

Walker, Daniel J.; Wakefield, Ann E.; Dohn, Michael N.; Miller, R F; Baughman, Robert P.; Hossler, Paul A.; Bartlett, Marilyn S.; Smith, James W.; Kazanjian, Powel; Meshnick, Steven R.

doi:10.1086/314509

Cited by 126 publications

(89 citation statements)

References 38 publications

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“…carinii sp. f. hominis has been suggested by the identification of isolates with polymorphisms in the gene encoding dihydropteroate synthase (DHPS), the target of sulphonamide drugs and in the gene encoding cytochrome b, the target of atovaquone [41]. The use of OP samples for sequence typing at these gene targets will greatly facilitate studies on the identification of drug resistance in l?…”

Section: Discussionmentioning

confidence: 99%

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

Tsolaki

Miller

Wakefield

1999

Journal of Medical Microbiology

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A nested PCR, amplifying a portion of the gene encoding the mitochondria1 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 R carinii pneumonia (PCP) to monitor response to anti-R carinii treatment. Genotyping of R 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 l? 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.

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Section: Discussionmentioning

confidence: 99%

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

Tsolaki

Miller

Wakefield

1999

Journal of Medical Microbiology

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“…Because atovaquone is a valuable alternative drug for malaria, PcP, and toxoplasmosis, the development of drug-resistant populations is of serious concern (35,42,58,73). Increased cytochrome b mutations were detected in human-derived P. jirovecii from AIDS patients with prior exposure to atovaquone (35).…”

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“…This study showed that multiple mutations around the binding sites can lead to atovaquone resistance by decreasing the affinity for the drug by reducing the volume of the binding pocket. Atovaquone resistance may be more complex (73), as evidence is growing for multiple functions of CoQ (5,20,40,51,54,55,61,70) and its presence at cellular sites other than the mitochondrion.…”

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Ubiquinone Synthesis in Mitochondrial and Microsomal Subcellular Fractions of Pneumocystis spp.: Differential Sensitivities to Atovaquone

et al. 2005

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The lung pathogen Pneumocystis spp. is the causative agent of a type of pneumonia that can be fatal in people with defective immune systems, such as AIDS patients. Atovaquone, an analog of ubiquinone (coenzyme Q [CoQ]), inhibits mitochondrial electron transport and is effective in clearing mild to moderate cases of the infection. Purified rat-derived intact Pneumocystis carinii cells synthesize de novo four CoQ homologs, CoQ 7 , CoQ 8 , CoQ 9 , and CoQ 10 , as demonstrated by the incorporation of radiolabeled precursors of both the benzoquinone ring and the polyprenyl chain. A central step in CoQ biosynthesis is the condensation of p-hydroxybenzoic acid (PHBA) with a long-chain polyprenyl diphosphate molecule. In the present study, CoQ biosynthesis was evaluated by the incorporation of PHBA into completed CoQ molecules using P. carinii cell-free preparations. CoQ synthesis in whole-cell homogenates was not affected by the respiratory inhibitors antimycin A and dicyclohexylcarbodiimide but was diminished by atovaquone. Thus, atovaquone has inhibitory activity on both electron transport and CoQ synthesis in this pathogen. Furthermore, both the mitochondrial and microsomal fractions were shown to synthesize de novo all four P. carinii CoQ homologs. Interestingly, atovaquone inhibited microsomal CoQ synthesis, whereas it had no effect on mitochondrial CoQ synthesis. This is the first pathogenic eukaryotic microorganism in which biosynthesis of CoQ molecules from the initial PHBA:polyprenyl transferase reaction has been unambiguously shown to occur in two distinct compartments of the same cell.

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“…Basically, most fungi, e.g., Aspergillus nidulans, Neurospora crassa, Saccharomyces cerevisiae, and Schizosaccharomyces pombe, contain an intron(s) in their cyt b gene sequences (28). cyt b gene sequences of Pneumocystis carinii and C. glabrata do not contain introns (11,45). Among the 23 species of Trichosporon studied, 11 species had an intron(s) in the region sequenced.…”

Section: Discussionmentioning

confidence: 99%

Molecular Phylogenetics of the Genus Trichosporon Inferred from Mitochondrial Cytochrome b Gene Sequences

et al. 2005

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Trichosporon Behrend is a member of the basidiomycetous yeasts. The species of the genus are causative agents of the superficial to deep-seated mycosis known as trichosporonosis. Generally, it is found that trichosporonosis is caused by six species: Trichosporon asahii, Trichosporon asteroides, Trichosporon cutaneum, Trichosporon inkin, Trichosporon mucoides, and Trichosporon ovoides (16,19,36,37). It is also found that the major causative agents of trichosporonosis differ in causing infection. T. asahii and T. mucoides are involved in deep-seated infection. T. asteroides and T. cutaneum are associated with superficial infection. T. ovoides and T. inkin are involved in white piedra of the head and genital area, respectively. Trichosporon pullulans is not a major causative agent of trichosporonosis and is rarely isolated from fungemia patients (25,26). Recent studies indicate that the number of patients with illness caused by a Trichosporon species has been increasing (21,22,42,43,52,56). Deep-seated trichosporonosis has a high mortality rate, and the prognosis for patients is very poor. Trichosporon species are also responsible for summer-type hypersensitivity pneumonitis (1, 2, 34, 55).The genus Trichosporon was revised using a combination of physiological and molecular criteria (14, 15). Subsequently, Sugita et al. (35,36,38) contributed significantly to the classification and taxonomic position of the genus Trichosporon. Recently, the number of species of the genus Trichosporon has expanded from 19 (17) to 23 (13). All of these classifications are based on the analysis of different regions of the rRNA genes, such as 5S rRNA genes, 26S rRNA genes, 18S rRNA genes, the D1/D2 region of large-subunit rRNA genes, or the internal transcribed spacer region of rRNA genes. There is a need for further evidence based on different DNA sequences to establish phylogenetic relationships (27). It was therefore of interest to find the relationships among various species of Trichosporon by analyzing mitochondrial (mt) DNA rather than rRNA genes.Mitochondrial genes are an attractive marker for inferring the phylogeny of closely related species because of the rapid evolution of the mitochondrial genome, the lack of recombination, and the strict maternal inheritance (29). Restriction fragment length polymorphism analysis of mt DNA has been shown to be useful for estimating the relationships among fungi (18,24,44). The mt cytochrome b (cyt b) gene has been used to study the evolution and phylogenetic relationships of many animals, including birds, mammals, and fish (3,9,10,12,20,23), but the sequence of this gene had been determined for only six species of fungi before our study (46). We have reported that the mt cyt b gene is useful for the identification, classification, and phylogenetic analysis of fungi (4-7, 46-49, 53, 54).We have previously shown that the cyt b gene phylogeny of basidiomycetous yeasts correlates with the cell wall biochemistry and septal ultrastructure (5). However, only two species of Trichosporon, T. aquat...

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Sequence Polymorphisms in thePneumocystis cariniiCytochromebGene and Their Association with Atovaquone Prophylaxis Failure

Cited by 126 publications

References 38 publications

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

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

Ubiquinone Synthesis in Mitochondrial and Microsomal Subcellular Fractions of Pneumocystis spp.: Differential Sensitivities to Atovaquone

Molecular Phylogenetics of the Genus Trichosporon Inferred from Mitochondrial Cytochrome b Gene Sequences

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