In this study we found that PCP was a severe complication following liver transplantion; the outbreak of PCP in liver transplant recipients at our center was not caused by nosocomial transmission according to the analysis of their clinical, epidemiological, and molecular characteristics though PCP patients, colonized patients, and HCW with close contact to PCP patients, were a potential source of P. jirovecii transmission; moreover, a decrease in absolute CD19 + B-cell counts might play an important role in the development of PCP in liver transplant recipients.
Previously, PCP was most frequently associated with HIV patients; in contrast, its incidence has been decreasing recently (1, 2). Growing evidence has shown an increase in PCP incidence and mortality in solid liver transplant recipients during the last decades, which has contributed to the increasing number of PCP outbreaks (4). An outbreak cluster of PCP in immunocompromised patients has been repeatedly reported to be caused by nosocomial transmission via four proposed routes, including i) directly contacting PCP patients; ii) contacting the same HCW who had close contact with PCP patients; iii) contacting the same colonized patients who might or might not have close contact with previous PCP patients; iv) airborne transmission of environmental aerosols contaminated with P. jirovecii from PCP patients (7, 8, 14–16).
Therefore, we conducted the first systematical research to investigate all the potential sources of transmission. From the transmission map, there was no spatiotemporal possibility for these PCP patients to meet each other before diagnosis either during hospitalization or at outpatient clinic, which led to the rejection of this route. Then, air sample detection was all negative. There were two main reasons that accounted for this phenomena: the traditional passive sedimentation method which was less sensitive compared with an air sampler, and laminar airflow rooms which made it difficult to collect P. jirovecii. Nevertheless, it was still impossible for PCP patients to get infected from each other through airborne transmission route combining transmission map analysis. Finally, colonized HCW and colonized patients were found to have higher occurrence of P. jirovecii when compared with controls. There was a possibility that they got infected from PCP patients or vice versa. Additionally, we had to admit that PCR on swabs was less sensitive than lower respiratory samples, which may result in possible false-negative results. Invasive investigations could improve active screening of P. jirovecii infected patients in case of high suspicion.
Furthermore, positive P. jirovecii samples were genotyped. Apart from sequence typing failure in some samples or loci, the results were still sufficient to differentiate the P. jirovecii strains. To our surprise, PCP patients showed completely different P. jirovecii strains, which were partially distinct from those detected in unrelated controls from other departments because of incomplete sequencing. This meant PCP patients were infected from different sources so it was reasonable to infer that HCW and transplant recipients might get infected from them when they had close contact. Moreover, we found an unreported mutation at the CYB gene (CYB10/CYB11) only in one PCP patient (22, 23).
Finally, risk factors for the development of PCP were analyzed. In our study, we identified decreased CD19 + B-cell counts to be the only risk factors. Nevertheless, in contrast to published studies that have reported several predictors such as age, CMV, acute rejection and decreased CD4 + T-cell counts, no such increased risk of PCP was observed in our study (9–11). Iriart X et al reported PCP occurred in elderly patients ≥ 65 (9). In our analysis, as age-matched patients were used as a control group we could not confirm age as a relevant risk factor. Nevertheless, the mean age of the cluster was 55 years old with only two patients ≥ 65. The presence of CMV viremia was associated with PCP. Basically, CMV infection may simply reflect the degree of immunosuppression. Rostved el. al. described a high rate of CMV co-infection but could not detect increased rates of CMV prior to diagnosis of PCP (24). Acute rejection implies the activation of the immune system while PCP occurred in immunocompromised patients. Therefore, acute rejection usually precedes PCP. It is the anti-rejection treatment that suppresses the immune system leading to lymphocyte deficiency, especially decreased CD4 + T-cell counts (25–27). In essence, acute rejection is a reflection of decreased CD4 + T-cell counts. Allograft dysfunction could increase the risk of PCP (28). In liver transplant recipients, immunosuppressive therapy consisted of induction and maintenance, which were all targeted at CD4 + T-cells. Decreased CD4 + T-cell counts is a clearly defined risk factor for PCP in HIV patients, while no clear threshold could be defined for CD4 + T-cell counts in liver transplant recipients (29).
A growing number of studies have stressed the key role CD19 + B-cells have played in the immune system. Rong et al found Pneumocystis burden in B cell-deficient mice progressively increased. Moreover, the clearance of Pneumocystis was delayed in B cell-activating factor receptor-deficient mice, which had few B cells and Pneumocystis-specific IgG and IgM antibodies (30). Hernandez-Novoa B et al reported CD40L-KO mice are highly susceptible to developing severe Pneumocystis pneumonia due to a decrease in CD19 + B-cell compared with immunocompetent mice (31). Therefore, the CD19 + B-cells exhibited an antifungal effect against P. jirovecii. In conjunction with these data, our study advocates the assumption that a decreased number of CD19 + B-cell in the blood may be the risk factor for PCP.
The main limitation of this study is that it represents the experience of a single center with a small number of patients. Future studies, preferably multi-center controlled clinical trials, are needed to further validate our initial report.