Age-dependent determinants of infectious complications profile in children and adults after hematopoietic cell transplantation: lesson from the nationwide study

Incidence and outcome of microbiologically documented bacterial/viral infections and invasive fungal disease (IFD) in children and adults after hematopoietic cell transplantation (HCT) were compared in 650 children and 3200 adults in multicenter cross-sectional nationwide study. Infections were diagnosed in 60.8% children and 35.0% adults, including respectively 69.1% and 63.5% allo-HCT, and 33.1% and 20.8% auto-HCT patients. The incidence of bacterial infections was higher in children (36.0% vs 27.6%; p < 0.0001). Infections with Gram-negative bacteria were more frequent than Gram-positives in adults (64.6% vs 44.8%; p < 0.0001). Outcome of bacterial infections was better in children (95.5% vs 91.4%; p = 0.0011). The IFD incidence (25.3% vs 6.3%; p < 0.0001) and outcome (88.0% vs 74.9%; p < 0.0001) were higher in children. The incidence of viral infections was higher in children after allo-HCT (56.3% vs 29.3%; p < 0.0001), and auto-HCT (6.6% vs 0.8%; p < 0.0001). Outcome of viral infections was better in children (98.6% vs 92.3%; p = 0.0096). Infection-related mortality was 7.8% in children and 18.4% in adults (p < 0.0001). No child after auto-HCT died of infection. Adult age, mismatched transplants, acute leukemia, chronic GVHD, CMV reactivation, infection with Gram-negatives, and duration of infection > 21 days were risk factors for death from infection. In conclusion, pediatric patients have 2.9-fold higher incidence and 2.5-fold better outcome of infections than adults after HCT.


Introduction
Infections are a significant cause of morbidity, mortality, and resource utilization after hematopoietic cell transplantation (HCT) in children and adults. Bacterial infections both after allo-and auto-HCT are known to be associated with high mortality and have become a public health problem of major concern worldwide due to antibiotic resistance. Invasive fungal disease (IFD) remains an important cause of morbidity and mortality after allo-HCT. The incidence of IFD has been reported at 9% after allo-HCT with mortality up to 50% of patients, especially after alternative donor transplantations [1][2][3]. Most studies have reported a high rate of viral infection after allo-HCT but not auto-HCT. High viral infection risk after allo-HCT is likely related to the delayed immune reconstitution after transplantation [4]. Recent EBMT (European Society for Blood and Marrow Transplantation) analysis showed that infections are responsible for 21.6% of deaths after allo-HCT and 11.0% after auto-HCT in all age groups together; however, the risk, types, and outcome of infections varied between age groups [5].
Infections occur in up to 82% of children [6][7][8] and adults [9][10][11] after HCT; however, large multicenter studies on incidence and outcome of bacterial, fungal, and viral infections are lacking. So far also no direct simultaneous comparison was made between children and adults.
In this study, we compared the incidence, type, and outcome of infections in pediatric and adult HCT centers in Poland in multicenter cross-sectional nationwide study. We analyzed also risk factors for the incidence and outcome of infections in 650 children and 3200 adults who received HCT.

Patients and methods
Design of the study All consecutive patients transplanted between 1.01.2012 and 31.12.2015 in 5/5 pediatric, and in 11/ 13, adult HCT Polish centers were included in the retrospective study. Bacterial, fungal, and viral infections were reported biannually by each center and data were analyzed centrally.
Fungal infections The diagnosis of IFD was made according to EORTC/MSG criteria as proven, probable, or possible [15][16][17]. Patients were screened with galactomannan test mainly during neutropenia or on the basis of clinically driven indications. Diagnostics for Pneumocystis jiroveci pneumonia (PjP) was performed in case of clinical indications.
Viral infections Viral infections were classified as episodic (diagnosed on the basis of clinical picture, and supplemented with appropriate tests) or latent (diagnosed at molecular level). The following viruses were detected by PCR analysis: adenovirus (ADV), polyoma BKV, cytomegalovirus (CMV), Epstein-Barr virus (EBV), human herpesvirus 6 (HHV-6), and community-acquired respiratory viruses (CARV) including influenza.
Supportive therapy Uniform, standard anti-infective prophylaxis has been applied for patients undergoing HCT.
Prophylaxis of infections Environmental prophylaxis was applied in all centers according to commonly accepted policy [22]. In children, antibacterial prophylaxis consisted of oral penicillin or second-generation cephalosporin (from day − 10, until neutrophil count > 1 × 10 9 /L or end of immunosuppressive treatment) and oral gentamicin used from the beginning of conditioning until hematological recovery. Children received antifungal prophylaxis with fluconazole; from 2014, posaconazole was used in case of graft versus host disease (GVHD) or in secondary prophylaxis. In children under age of 12 years, the drug was used off-label [17] and administered according to body weight, as shown by Welzen et al [23]. In adults during neutropenia, fluoroquinolones were used for antibacterial prophylaxis and fluconazole in antifungal prophylaxis together with regular screening of serum galactomannan and computed tomography (HRCT/CT) in case of suspected IFD. Both in children and adults, acyclovir was used in prophylaxis of HSV/VZV infection until 1 year post-transplant. Weekly screening for DNA-emia and preemptive treatment were performed for CMVand EBV reactivation. Prevention of PjP included cotrimoxazole after hematopoietic recovery until the end of immunosuppressive treatment. Commercial immunoglobulin preparations were given in case of decreased immunoglobulin concentration during the first month and then monthly until B cell function recovery. Most of children receiving myeloablative conditioning (MAC) were commenced on gut rest from the first 5 days after HCT and received total parenteral nutrition (TPN) until hematopoietic recovery.
Statistical analysis For analysis of incidence, infectious event was defined as the diagnosis of a first specific infectious disorder. Categorical variables were compared with the chisquare test, non-categorical variables were compared with the Mann-Whitney U test. Odds ratio (OR) and confidence intervals (95%CI) were calculated for the difference in occurrence of infections in patients. Cumulative 2-year incidences of bacterial, fungal, or viral infections were calculated using competing risk analysis [25], starting from the day of transplant to the day of the first infection. Death was considered as the competing event. Outcome of infection was regarded as positive in case of survival from infection or negative in case of death from infection. Infection-related mortality (IRM) was defined as any death that occurred in the presence of infection, starting from the day of diagnosis of infection. Death from infection was diagnosed as of bacterial, fungal, or viral cause; however, in many cases of IRM, patients suffered from multiple infections, and clinically the most symptomatic infection was regarded as the primary cause of death. In case of relapse and progression of malignancy, this was regarded as the primary cause of death, regardless of concomitant infection. The Kaplan-Meier method was used to determine IRM, counting from the day of diagnosis of infection. The relationship between the binary outcome, infection incidence, or death from infection, and other variables, regarded as risk factors, were analyzed using multivariate logistic regression: hazard risk (HR) and 95%CI were calculated for each factor. All reported p values are twosided; p < 0.05 was considered as statistically significant.
Infections with Gram-negative bacteria were more frequent than Gram-positive in adults (64.6%), but not in children (44.8%). The difference was highly significant (p < 0.0001; OR = 2.3, 95%CI = 1.8-2.7). The frequency of G-negative   Risk factors In multivariate logistic analysis (Table 5), the risk of infections was higher after allo-HCT than auto-HCT (HR = 1.8; p < 0.001). In allo-HCT patients, the risk was higher in children

Discussion
In this study for the first time ever, simultaneous analysis and comparison of epidemiology and outcome of bacterial, fungal, and viral infections in a large cohorts of children and adults after HCT in a multicenter cross-sectional nationwide study were performed. Both groups largely differed in terms of distribution of primary diseases and their treatment, types of preparative regimens, and types of transplantation. Although both pediatric and adult transplant centers used generally very  similar strategy of anti-infective management [18], some differences between these settings existed, as pediatric centers used many off-label compounds. In this study, we analyzed patients over a period of 4 years, when anti-infective prophylaxis and treatment did not change substantially in both pediatric and adult centers. Bacterial infections occurred mainly during neutropenic, pre-engraftment phase. In adults, Gram-negative bacteria were more often documented, while in children Gram-positive species. The rate of MDR Gram-negative strains was higher in pediatric than in adult centers, while the rate of Gram-positive MDR was comparable in these cohorts. Our results indicate the shift of prevalence from Gram-positive to Gram-negative bacteria in a population of adult hematology patients and increasing incidence of MDR bacteria, especially Gram-   [26,27]. It is debatable, if use of quinolones in adults or oral gentamycin in children have possible negative impact in the selection of resistant gut microbiome [27]. We confirmed that irrespectively to age, transplant performed from alternative donor and prolonged neutropenia were independent risk factors for the development of bacterial infection [28]. The differences in bacterial epidemiology between children and adults resulted in differences in outcome of bacterial infections in these two cohorts, with a higher risk for death related to Gram-negative bacteria. High rate in bacterial infections was found for typical pediatric primary diagnoses like primary immunodeficiencies, neuroblastoma, and Ewing sarcoma; and opposite, in adulthood disease multiple myeloma, the rate was much lower and reached 12.9%. Fungal infections were much more frequently diagnosed in children, regardless of the level of diagnosis; however, it was predominant for possible IFD. It reflects "real-life" pediatric strategy of reducing invasive diagnostics in children. It seems that lower incidence of IFD in adults might result from general strategy of protective environment in transplant setting and the prophylactic use of posaconazole during intensive chemotherapy in AML/MDS, according to ECIL recommendations [29][30][31]. Additionally, modified transplant procedure for pretransplant IFD, such as no-TBI conditioning, RIC, or use of PB as a stem cell source, could have possibly decrease the rate of fungal reactivations [1,32]. Proven IFD were more often diagnosed in children. This was due mainly because of diagnosis of candidemia, as children usually have permanent, while adults rather temporarily central venous catheters. Most of children were also receiving TPN, while it was rather infrequent practice in adults. Relatively high rate of candidemia among proven IFD in children probably contributed to lower IRM than in case of invasive aspergillosis, as reported recently [33].
The incidence of viral infections was higher in children than in adults. This observation can be explained by immature immune system in children, resulting in primary infection or higher rate of reactivation of latent viruses. CMV and EBV were two most often diagnosed viruses in children after allo-HCT. CMV exerts direct and indirect effects in tissues and often plays a role of driver of another infections, including IFD, thus contributing to an increased post-transplant risk of life-threatening complications. With respect to respiratory viral infections, there is no current strategy of routine monitoring of community-acquired respiratory viruses; thus, no firm conclusion can be drawn on this topic from our study.
IRM was higher in adults, what has been evidence-proved for the first time. Additionally, IRM was higher in Gramnegative infections and in patients with acute leukemia. The outcome of infections was better in children both after alloand auto-HCT. In addition to well-defined factors for mortality (acute leukemia, MMUD, GVHD, CMV reactivation), duration of infection > 21 days was associated with an increased risk of death after infection.
The higher infection rate of MDI in children in comparison to adults can be explained by the following factors: (1) much higher rate of auto-HCT in adults resulting in overall lower incidence of infections in adults, especially seen in case of bacterial complications; (2) higher rate of patients with acute leukemia in pediatric cohort, with a well-known high incidence of infectious complications in acute leukemia [8,34]; (3) much higher rate of diagnosis of possible IFD in children being the consequence of the positive results of imaging only; (4) higher incidence of viral infections in children, what can correspond to higher rate of primary infections; and finally, (5) real-life tendency of pediatricians to perform more detailed diagnostic procedures. Due to the same factors, the diagnosis of multiple myeloma was associated with a decreased risk for infection in multivariate analysis. On the other hand, the incidence of infectious complications in this group of patients was similar as presented in recent analyses [35][36][37][38].
The limitation of the study is its retrospective design; however, data were collected periodically. Also no routine screening was performed for viral infections except CMV and EBV. Thus, in most cases of viral infections, the diagnosis was bound to clinical symptoms.
In conclusion, the profile of infections and related deaths largely vary between children and adults. Our study proved age-dependent determinants of pediatric and adulthood profile of infectious complications after HCT: children have higher risk of all types of infections and a better outcome of bacterial infections, while in fungal and viral infections, the IRM was comparable between children and adults. Adult age, MMUD transplants, diagnosis of acute leukemia, chronic GVHD, CMV reactivation, and infection lasting > 21 days are relative risk factors for death from infection after HCT. The potential implication of this comprehensive analysis might be differential infection control and management strategies for children and adults.
institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards. This article does not contain any studies with animals performed by any of the authors.
Informed consent Informed consent was obtained from all individual participants included in the study.
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