Objectives and Study

T-cell mediated immunity (CMI) is crucial in Cytomegalovirus (CMV) control. We present the interim results of a single-centre prospective study assessing the capacity of CMV-CMI assays to predict children’s failure to spontaneously control CMV after liver transplantation (LT).

Methods

LT recipients (0-17 yo) were included and followed until 200 days after LT. Preemptive therapy (PET) with ganciclovir was initiated if i) ≥ 100.000 CMV-DNA IU/mL whole blood (wb) or ii) ≥ 50.000 IU/mL CMV-DNA wb rapidly increasing in HR patients. CMV-CMI was measured through QuantiFERON CMV (cut-off > 0.2 IU/mL), and T-SPOT against CMV pp65/IE-1 antigens (cut-off > 20 SFU). Donor (D)-to-recipient (R) serological risk was classified as high (HR: CMV-IgG D+/R-), intermediate (MR: D_any/R+) and low (LR: D-/R-).

Results

22 children were included (11 females, median age 1.5 years, IQR: 0.8-7).

Six remained uninfected; 9 (7 MR, 2 HR) spontaneously controlled the infection (group SC: median CMV-DNA 1130 IU/mL, IQR 410-2773); 7 (1 MR, 6 HR) needed PET (group PET: median CMV-DNA 85594 IU/mL, IQR 50903-130503).

CMV-CMI was higher in the SC than in the PET group according to IE-1 T-SPOT (18.5 ± 26 vs 2.5 ± 5 SFU at +2 weeks; 20.5 ± 34 vs 3.9 ± 4.5 SFU at +4 weeks; P = 0.025), and QuantiFERON CMV (0.5 ± 0.7 vs 0.06 ± 0.17 IU/mL at +2 weeks; 3.9 ± 6 vs 0.7 ± 1 IU/mL at +4 weeks; P = 0.034) but not pp65 T-SPOT (P = 0.196).

A ratio between the CMV-CMI/viremia·10^-2 at +2 weeks discriminated SC from PET group with AUCs of 0.75 (P = 0.11), 0.93 (P = 0.067), and 0.82 (P = 0.04) for IE-1 and pp65 T-SPOTs, and QuantiFERON CMV, respectively.

Conclusions

CMV-CMI assays are promising in predicting the need for PET in LT children.

This study is funded by the ESPGHAN Research Grant 2019.

Objectives and Study

The risk of infection after paediatric liver transplantation is increased both in the short and long term. Although hepatitis A virus (HAV) infection in children is often asymptomatic and self-limiting, some case studies describe severe courses resulting in acute liver failure. HAV vaccination is a simple, less invasive and cost-effective method that has led to incidence reduction. Only few data exist describing vaccination response in such immunocompromised patients. In this retrospective, single-centre study, we investigated HAV vaccination response in HAV-naïve children after liver transplantation (LT) and describe long-term immunity over five years.

Methods

Vaccination status is reviewed at annual check-ups and vaccinations may be carried out again from the first check-up (12 months) after LT. Hepatitis A immunity is determined at each annual check-up by analysing anti-HAV-IgG using automated chemiluminescent microparticle immunoassay.

Results

458 LT recipients were analysed between September 2004 and June 2021. In total, data on 153 HAV-naïve children (of these 71 female, 46.4%) with a median annual follow-up of 7.6 years (IQR: 4.6 – 11.2) after LT was available. More than 47% of patients were diagnosed with biliary atresia (BA), followed by hepatic malignancy (11.8%) and acute liver failure (11.1%). Median age at time of transplant was 1.0 years (0.6 – 2.5). 59 children received immunisation with two HAV vaccine doses. At follow-up, 88.1% were anti-HAV-IgG positive, decreasing to 74.4% five years later (p=0.08). In contrast, 26 patients received one HAV vaccination, of which 65.4% had detectable anti-HAV-IgG. After 5 years, this decreased to 50.0% (p=0.31).

Conclusions

More than 88% and 65% of liver-transplanted children show seroconversion after HAV vaccination with one and two doses, respectively. Seropositivity rates 5 years post-vaccination are slightly lower. In contrast, studies in healthy children 10 years post-vaccination show better immunogenicity with seroconversion of 72% and 96%, respectively, indicating less immunogenicity following paediatric LT.

Objectives and Study

Gut microbiata dysbiosis has been reported in adults with liver cirrhosis. Data on gut microbiota (GM) in children with end-stage liver disease (ESLD) are scarce. We investigated GM in children before and up to 3 months after liver transplantation (pLT) aiming to delineate GM compositions associated with disease severity.

Methods

21 fecal samples of 19 children (9f, age 3.1(0.4-15.7) years) with ESLD (53% biliary atresia) awaiting pLT were analyzed using 16S rRNA gene sequencing. Samples after pLT were obtained in n=11 children at 1-4, 12&24 weeks after pLT. Results were compared to n=26 age-matched healthy controls (12f, age 3.6(0.1-15.9) years).

Results

ESLD samples showed lower GM α-diversity than controls (Shannon-Index 2.9±1.0 vs. 3.8±0.8,p<0.01). On the Phylum level, ESLD samples contained higher proportions of Proteobacteria and reduced levels of Firmicutes (22.1±28% vs. 2.6±4% and 50.1±23% vs. 67.7±16%, p<0.001 respectively). On the Family level, this translated into higher proportions of Enterobacteriaceae (20.4±29% vs. 1.4±3.5%, p<0.01) and reduced levels of Lachnospiraceae (17.9±13% vs. 35.1±12%), Peptostreptococcaceae (0.6±0.9% vs. 1.9±1.6%) and Ruminococcacea (5.1±6% vs. 21.2±9%, p<0.001). UniFrac analysis reflected differences between patients and controls and suggested an impact of disease severity on GM composition (Fig1a). Emergency hospitalisation for decompensated ascites (HOSP) and Rifampicin-treatment for refractory pruritus (RIFA) as markers of disease severity (n=6 each) were associated with reduced α-diversity compared to non-HOSP or non-RIFA ESLD-samples (2.0±1.0 vs. 3.2±0.8/2.7±0.5 vs. 3.4±0.7, p=0.01 respectively) and were independent predictors of α-diversity in multiple-regression-analysis (F(3,16)=7.7, p<0.01, R²=0.59). After a post-pLT drop, α-diversity returned to pre-pLT values after 3 months, but retained the dysbiotic GM composition seen pre-pLT (Fig1b).

Conclusions

Deviations in GM composition in end-stage liver disease are associated with occurrence of complications. GM dysbiosis did not normalize up to 6 months after pLT. Windows of opportunity for therapeutic modulation of GM need to be defined in order to improve patient outcome.

Objectives and Study

To present our experience and impact of the optimization of liver grafts by prioritizing the pediatric recipient, promoting bipartition (Split).

Methods

We reviewed our pediatric liver transplant series focusing on split grafts. We compared the historical series (1994-2019) with the last 3 years (2019-2022) and analyzed its impact on living related donor liver transplantation (LRDLT).

Results

There were 26 bipartitions in period 2 (26.8%, only one urgent, 60% in situ) compared to 52 in period 1 (6.6%, 71% urgent). The main indication was biliary atresia (58%), with a median of 12.3 months (range 0.9-89) and 8.1 kg (range 3.1-21.9). In 18 (73%) we kept the left and proper hepatic artery, leaving the right hepatic artery, common hepatic artery, and celiac trunk for the right graft (with reconstruction). Several technical difficulties were managed: double suprahepatic anastomosis (1), venous grafts from the graft surface (2) cava substitution with a jugular graft (1), pericardium dissection (2), venous grafts between superior mesenteric and portal veins (7), donor iliac artery grafts (8), and double bile duct (7). Eleven required delayed abdominal wall closure.

The transplant was 100% successful (12/52 grafts had been lost in period 1, 23%); a biliary fistula and an arterial thrombosis were resolved surgically. To our knowledge, all right liver grafts were successfully implanted, except for two that required retransplantation and one which was not finally used. The LRDLT/ Split ratio decreased significantly (p<0.05): 36:1 between 2016-2019 vs 8:26 between 2019-2022. The biliary complications rate also decreased in the new Split era compared with the LRDLT group.

Conclusions

Bipartition has quadrupled in our pediatric liver transplant program, with excellent results and a reduction in the need for LRDLT, ceasing to be just an emergency option. The in-situ technique and an adequate distribution of the artery are advantages that benefit both teams.

Objectives and Study

Epstein-Barr virus (EBV) infection is associated with post-transplant lymphoproliferative disorders (PTLD) after liver transplantation (LT). EBV-specific T cells play an important role in the infection. We aimed to determine the response of these cells in paediatric LT recipients.

Methods

Peripheral immune cells and EBV viral load were evaluated before LT and periodically monitored until 48 weeks after LT. Significant EBV viremia (SEV) was defined as EBV viral load >2,000 IU/mL. EBV-specific T cells and reactive natural killer cells were determined by using flow cytometry techniques and EBV antigens, including BZLF1, Epstein–Barr nuclear antigen 1 (EBNA1), and latent membrane protein 2 (LMP2). All patients underwent the same immunosuppressive protocol.

Results

Among 40 LT recipients, seven children (17.5%) were diagnosed with SEV with the median (IQR) onset of 29 (19, 33) weeks after LT, and 2/7 children later had PTLD. The children with SEV had lower CD4+ T cells and natural killer T (NKT) cells before LT than children without SEV (P=.04, both). LMP2-reactive NKT cells in the children with SEV were also higher at 4 weeks after LT (P<.01). Eighteen seronegative children (45%) received liver grafts from EBV-seropositive donors (D+/R-), and five of them subsequently had primary EBV infection (PEI). Among the D+/R- serostatus, CD8+ T cells were higher in the children with PEI, while BZLF1-specific CD8+ T cells were lower at 2 weeks after LT compared to children without PEI (P<.01, both). EBNA1-reactive NKT cells were also lower at 12 weeks after LT in children with PEI (P=.01).

Conclusions

The relatively lower EBV-specific T cells, particularly BZLF1-specific CD8+ T cells, was associated with PEI after LT. Some reactive NKT cells were associated with PEI and SEV. These findings may lead to further studies concerning the prediction and prevention of EBV infection in paediatric LT recipients.