Carriage of Pneumocystis carinii by immunosuppressed patients and molecular typing of the organisms.

Abstract

Pneumocystis carinii causes severe pneumonia in immunocompromised patients, mainly those infected with HIV. The disease is now believed to be frequently caused by de-novo infection from exogenous sources, which, however, remain unknown. The detection of P. carinii DNA in air samples collected in rural areas suggests the existence of an environmental reservoir. Recently, authors have reported the detection of P. carinii in patients without established P. carinii pneumonia (PCP), raising the question of the existence of carriers who could constitute a reservoir of the pathogen. These patients were most often immunocompromised individuals with AIDS, but a few were receiving an immunosuppressing therapy or were immunocompetent with pulmonary disease. However, it is often unclear from those reports whether the patients received anti-PCP prophylaxis or treatment before or after the collection of the specimen, raising the question of an early infection rather than true carriage. As far as immunosuppressed HIV-negative patients are concerned, three studies suggest the carriage of P. carinii in a few patients who clearly did not receive anti-PCP drugs [1–3]. However, in those studies, it was not reported whether immunosuppression was modified or withheld, a fact that could account for the absence of symptoms during the follow-up. Also, follow-up was documented in only one study (0.2–16 months [3]). Three well-characterized cases are reported here, which strongly suggest that carriage in immunosuppressed HIV-negative patients can occur. In addition, molecular typing of the carried micro-organisms was performed for the first time. A collection of bronchoalveolar lavage (BAL) specimens from 91 patients was examined blindly for the presence of P. carinii using the polymerase chain reaction (PCR) and two staining methods (Gomori silver staining cysts and Giemsa staining trophozoites). Among the 91 patients, 64 were HIV negative and 27 were HIV positive. PCR and microscopy were concordant for the vast majority of the patients: 80 were negative for both techniques and eight smear-positive PCP cases were also positive by PCR (two HIV negative and six HIV positive). However, in three patients, P. carinii was detected only using PCR upon two independent DNA extractions. A low number of organisms probably explains the detection only by PCR. Contamination is largely unlikely because: (i) the amplifications of four different P. carinii genomic regions were positive; and (ii) the same results were obtained for the two DNA extractions using the typing method based on the detection of the polymorphisms in the four genomic regions by the single-strand conformation polymorphism technique [4]. Patients 1 and 2 were infected with a single P. carinii type and patient 3 was probably infected with two types (Table 1). All types identified in the three patients have been observed many times in symptomatic PCP patients with AIDS [5].Table 1: Demographic and clinical characteristics of HIV negative carriers of Pneumocystis carinii and molecular typing of the organisms. The demographic and clinical characteristics of these three cases are given in Table 1. All were HIV-negative patients receiving chronic immunosuppressive treatment for polymyositis or graft, a treatment which remained unchanged before and after BAL collection. Despite the absence of anti-PCP prophylaxis and treatment, none of the patients developed established PCP during the entire follow-up. All had an alternative diagnosis for their pulmonary infiltrates, for which they were treated successfully. Patient 1 presented with a second episode of pulmonary infiltrates after one month and had a second BAL. This specimen was again positive for P. carinii, but only by PCR. It contained the same P. carinii type as the first specimen. The diagnosis was the same as for the first episode and infiltrates resolved in the absence of PCP treatment or a change in immunosuppressive therapy. For two patients, the PCR positive for P. carinii coincides with an episode of cytomegalovirus (CMV) infection, a known predisposition for PCP [6], raising the question of CMV predisposing or unmasking P. carinii carriage. There was no epidemiological or typing evidence that these carriers acquired the organism from other PCP patients present in the hospital. In conclusion, these observations suggest that the three patients carried P. carinii. Carriage can last at least one month confirming another similar observation [2]. The presence of three carriers among the specimens investigated suggests that, as previously reported [3], carriers may not be infrequent. There is no evidence that the types colonizing carriers of P. carinii are different from those causing PCP, suggesting that carriers could be a potential reservoir of P. carinii for other immunosuppressed patients, including those with AIDS. Acknowledgement The authors would like to thank Arlette Cruchon for excellent technical assistance. Philippe M. Hausera Dominique S. Blanca Jacques Billeb Aimable Nahimanaa Patrick Franciolia