Infections are the third leading cause of death following kidney transplantation [1], and Pneumocystis pneumonia (PcP) is one of the most severe opportunistic causes. Pneumocystis infects 0.3%–2.6% of kidney transplant recipients (KTR), with a mortality rate reaching 14% in patients admitted to the ICU [2] and an increased risk of transplant loss in surviving patients [3]. The Kidney Disease Improving Global Outcomes (KDIGO) guidelines recommend universal initial PcP prophylaxis with cotrimoxazole for the first 3–6 months after kidney transplantation [4], while the American Society of Transplantation recommends prophylaxis for 6–12 months [5]. However, whether this prophylaxis should be prolonged or resumed in certain high risk situations remains unclear [5]. These recommendations have changed the epidemiology, and now most reports involve late post-transplant recipient PcP [6–9].

Pneumocystis infection elicits T-cell mediated responses including T helper (Th) 1, Th2 and Th17 responses [10], and lymphopenia has been frequently reported as an independent risk factor for PcP [6, 11–13]. Kaminski et al proposed targeted prophylaxis based on simple criteria such as chronic lymphopenia (i.e., < 1,000/μL) [6]. However, the factors that contribute to lymphopenia are not fully understood. Some studies showed that cumulative immunosuppression, corticosteroids pulses, or treated transplant rejection episodes are independent risk factors for PcP, with some conflicting results depending on the cohorts [3, 6, 11–19]. In addition, most study data are collected more than 1 year before the PCP. Identifying clinical and biological biomarkers in the year preceding the PcP could help guide clinicians regarding PcP prophylaxis.

Thus, the main objective of our study was to identify risk factors associated with PcP after kidney transplantation, with a particular focus on events occurring in the year prior PcP. The secondary objective was to study how PcP affects kidney transplant and patient outcomes.

Our case-control study involving KTR yielded 2 main results. First, PcP occurred in 5% of KTR followed in our center and was associated with high related mortality (18%), an increased risk of subsequent chronic rejection, and a return to dialysis. Secondly, several factors were independently associated with PcP, including being treated by immunosuppressive therapy other than anti-rejection drugs, CMV infection in the year before T_PcP, low lymphocyte count, and high creatinine levels. Having a lymphocyte count <1,000/mm³ and/or a CMV infection are two main factors associated with the occurrence of PcP within the year.

PcP is an opportunistic infection that occurs in patients suffering from CD4⁺ T cell response deficiency, which is the case in KTR, who are thus eligible for PcP prophylaxis [4]. In our cohort, 5% developed PcP, which is within the range reported in other cohorts [2, 6, 16, 19]. However, the epidemiology has changed over the last 30 years as a result of updated recommendations and the systematic use of cotrimoxazole, leading to an increase in the proportion of late-onset PcP. It should be noted that PcP occurred in the median time of 40 months, i.e., well after the end of the theoretical prophylaxis recommendation. In this cohort, only half of patients, particularly the most recently included patients, received early prophylaxis with cotrimoxazole. By matching cases and controls on the date of the active transplantation, it is therefore not possible to study the effect of the prophylaxis variable (presence/absence) on the occurrence of PcP. However, the duration of prophylaxis was shorter for cases, suggesting that extending or reinitiating PcP prophylaxis could benefit some patients.

To identify such patients, several associated/risk factors for susceptibility to PcP have been previously identified, but with some discrepancies between studies [3, 6, 11–19]. In addition, events occurring during the year preceding PcP could be informative. As expected, cases were older than controls at the time of PcP, with frailty conferring a higher age-related risk of infection [21]. Cases were also more likely to have a higher creatinine level preceding PcP, supporting the concept of kidney impairment-associated immunosenescence [22]. They were more often considered as having a high immunological risk, raising the possibility of more likely transplant rejection. However, the proportion of acute rejection was similar in cases and controls (14% in each group).

We observed that CMV infection was independently associated with PcP, mainly in the year preceding T_PcP. This association has been reported in several studies [14–16], but not all [6]. In the meta-analysis by Hosseini-Moghaddam et al., CMV infection significantly increased the risk of post-transplant PcP (OR: 3.30, 95% CI: 2.07–5.26). In addition, Lee et al. showed that PcP and CMV co-infection is associated with an increased clinical severity and worse clinical outcomes [23]. The causal link between CMV infection and the occurrence of PcP cannot be asserted, but pathophysiological assumptions can be proposed. First, stronger immunosuppression could be responsible for both opportunistic infections. We observed that cases were more likely to have a low lymphocyte count, as described in other work [6, 12, 13, 18]. The intensity of cumulative immunosuppression remains a difficult variable to quantify. However, we observed no significant difference in the choice of anti-rejection molecules or in the intensity of therapeutic immunosuppression, as assessed by modified Vasudev and TIS scores. Only mTOR inhibitors were more prescribed in cases compared to matched controls, as previously reported [6, 24]. As we discuss above, this association can be explained by the immunosuppressive effect of mTOR inhibitors but without ruling out the possibility of having included mTOR inhibitor-induced interstitial lung disease in some cases [24]. Furthermore, the administration of steroid pulses were not significantly associated with PcP, unlike in the study by Kaminski et al. [6, 13, 25]. However, this result should be interpreted in the light of a low frequency of acute rejection in the cohort. Other immunosuppressive therapies were more frequently prescribed in cases, mainly anti-cancer chemotherapy or anti-C5 therapies, highlighting the role of the cumulative immunosuppressive burden between transplantation and PcP. Secondly, cases were more likely to have impaired renal function, even when adjusted for age. This poorer renal function may reflect the altered terrain in which opportunistic infection occurs more frequently, as the incidence of infections increases linearly as renal function deteriorates [26]. Finally, we observed that for 3⁄4 of patients, PcP occurred within 2 years after CMV infection. CMV infection by itself can induce cellular immunodepression, through mobilization of cellular T immune defenses and secondary immunoparalysis. This hypothesis is reinforced by the results of an in vivo study in mice inoculated with CMV and Pneumocystis, showing that CMV infection induces a decrease in lung cells expressing MHC class II, and in activated T-CD4 lymphocytes in lymphoid organs and the alveolar compartment, associated with a defect in Pneumocystis clearance [27].

Our study confirms that PcP is associated with a poor prognosis in KTR [2, 28, 29], with an attributable mortality rate of 18% and transplant loss in 20% of surviving patients. It is therefore crucial to better understand the risk factors associated with this infection in order to define at risk-situations where anti-Pneumocystis prophylaxis is highly recommended. Global management of PcP involves several nephrotoxic interventions (high dose cotrimoxazole, contrast agent…) and the tapering of immunosuppressive regimen that may further elicit chronic rejection, contribute to the decline in transplant function and precipitate the return to dialysis.

In the end, we identified simple and routine biomarkers (serum creatinine, lymphocyte count) and a frequent opportunistic infectious event (CMV infection) that were associated with the occurrence of PcP. Among cases who received initial anti-Pneumocystis prophylaxis, 22 of 25 (88%) infections could have been prevented if prophylaxis had been restarted or continued in the presence of CMV infection and/or lymphopenia <1,000/mm³. This strategy would have been associated with excess treatment in of 23 out of 79 controls (29%), but is supported by the excellent tolerability of such low doses in real practice, the low cost of the drug, and the good compliance of patients.

The limitations of this study are related to its retrospective and monocentric nature. Some data are missing, even if this number is very limited for most variables. It is possible that over this period of 17 years, unmeasured changes in clinical practice may have influenced the risk to contract PCP, but such difference have been minimized by the controls pairing strategy. We did not provide CD4 and CD8 lymphocytes count since lymphocyte immunophenotyping has only become part of routine follow-up in more recent years and CD4 counts are thus not available for all patients. However, in the study of Kaminsky et al. lymphopenia was identified as the most significantly associated lymphocytic marker of PCP [6]. Patient prognosis could only be partially evaluated and is potentially biased insofar as the matching imposed a follow-up time for controls that was at least equal to that of the index case plus 1 year. Some patients had not received anti-Pneumocystis prophylaxis, but our sensitivity analysis confirmed the same findings in the subgroup of patients who received prophylaxis.

PcP is associated with high mortality and transplant loss in patients who have undergone a kidney transplant. We identified factors that were independently associated with PcP, including immunosuppressive therapy other than anti-rejection drugs, CMV infection in the year before T_PcP, low lymphocyte count and high serum creatinine levels. These risk factors remain unchanged with or without anti-Pneumocystis prophylaxis. Based on these results and previous literature, the occurrence of CMV infection and/or lymphopenia <1,000/mm³ could redefine the indications for continuation or reinitiation of anti-Pneumocystis prophylaxis, which is an inexpensive and well-tolerated treatment.