[Clinical features and genetic analysis of three patients with Immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome due to variants of FOXP3 gene].

OBJECTIVE: To analyze the clinical characteristics of three patients with Immune dysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome. METHODS: Three patients with IPEX syndrome diagnosed at the Children's Hospital of Fudan University from January 24, 2013 to July 29, 2019 were selected as the study subjects. Their clinical features, laboratory investigations and results of genetic testing were summarized. Treatment and prognosis were also explored. RESULTS: All of the three children had developed the disorder during infancy. One child had initial features including diabetes and diabetic ketoacidosis, whilst the other two had initiated by diarrhea. All patients had gastrointestinal involvement, and one was diagnosed as very early onset inflammatory bowel disease by colonoscopy and biopsy. Two children also had endocrine glands involvement. One child had manifested type 1 diabetes and positivity for thyroglobulin and thyroid peroxidase antibodies, though his thyroid function had remained normal. Another one had hypothyroidism and was treated by levothyroxine. Genetic testing revealed that all children had harbored missense variants of the FOXP3 gene, including c.1222G>A (p.V408M), c.767T>C (p.M256T) and c.1021A>G (p.T341A). The clinical symptoms of one patient were alleviated following allogeneic hematopoietic stem cell transplantation. One patient was stable after treatment with infliximab plus insulin, and one child had died of refractory septic shock and multiple organ dysfunction syndrome at 3 months old. CONCLUSION: FOXP3 gene variant-associated IPEX syndrome may have very early onset and diverse clinical manifestations. For male patients with infantile onset chronic diarrhea, multiple endocrine or multiple system involvement, genetic testing is recommended, which may facilitate early diagnosis, treatment and genetic counseling.

Diagnosis of mixed infection and a primary immunodeficiency disease using next-generation sequencing: a case report.

Major Histocompatibility Complex Class II (MHC II) deficiency is a rare primary immunodeficiency disorder (PID) with autosomal recessive inheritance pattern. The outcome is almost fatal owing to delayed diagnosis and lacking of effective therapy. Therefore, prompt diagnosis, timely and effective treatment are critical. Here, we report a 117-day-old boy with diarrhea, cough, cyanosis and tachypnea who was failed to be cured by empiric antimicrobial therapy initially and progressed to severe pneumonia and respiratory failure. The patient was admitted to the pediatric intensive care unit (PICU) immediately and underwent a series of tests. Blood examination revealed elevated levels of inflammatory markers and cytomegalovirus DNA. Imaging findings showed signs of severe infection of lungs. Finally, the diagnosis was obtained mainly through next-generation sequencing (NGS). We found out what pathogenic microorganism he was infected via repeated conventional detection methods and metagenomic next-generation sequencing (mNGS) of sputum and bronchoalveolar lavage fluid (BALF). And his whole exome sequencing (WES) examination suggested that CIITA gene was heterozygous mutation, a kind of MHC II deficiency diseases. After aggressive respiratory support and repeated adjustment of antimicrobial regimens, the patient was weaned from ventilator on the 56th day of admission and transferred to the immunology ward on the 60th day. The patient was successful discharged after hospitalizing for 91 days, taking antimicrobials orally to prevent infections post-discharge and waiting for stem cell transplantation. This case highlights the potential importance of NGS in providing better diagnostic testing for unexplained infection and illness. Furthermore, pathogens would be identified more accurately if conventional detection techniques were combined with mNGS.

Case report: The clinical utility of metagenomic next-generation sequencing in mucormycosis diagnosis caused by fatal Lichtheimia ramosa infection in pediatric neuroblastoma.

Lichtheimia ramosa (L. ramosa) is an opportunistic fungal pathogen of the order Mucorales that may result in a rare but serious mucormycosis infection. Mucormycosis could be angioinvasive, causing thrombosis and necrosis in the nose, brain, digestive tract, and respiratory tract. The infection is highly lethal, especially in immunocompromised hosts, and the incidence has been on the rise. However, due to its relatively low incidence in pediatric population and the challenges with diagnosis, the awareness and management experience for pediatric mucormycosis are extremely limited, which might lead to poor outcomes. In this study, we comprehensively reviewed the course of a fatal rhinocerebral mucormycosis case in a pediatric neuroblastoma patient receiving chemotherapy. Due to a lack of awareness of the infection, the standard care of amphotericin B treatment was delayed and not administered until the identification of L. ramosa by metagenomic next-generation sequencing (mNGS)-based pan-pathogen detection of the patient's peripheral blood sample. We also reviewed the literature on L. ramosa infection cases reported worldwide between 2010 and 2022, with an analysis of clinical manifestation, prognosis, and epidemiological data. Our study not only highlighted the clinical value of comprehensive mNGS in rapid pathogen detection but also raised awareness of recognizing lethal fungal infection early in immunocompromised hosts including pediatric cancer patients.

Metagenomic next-generation sequencing of bronchoalveolar lavage fluid from children with severe pneumonia in pediatric intensive care unit.

Background: Severe pneumonia due to lower respiratory tract infections (LRTIs) is a significant cause of morbidity and mortality in children. Noninfectious respiratory syndromes resembling LRTIs can complicate the diagnosis and may also make targeted therapy difficult because of the difficulty of identifying LRTI pathogens. In the present study, a highly sensitive metagenomic next-generation sequencing (mNGS) approach was used to characterize the microbiome of bronchoalveolar lavage fluid (BALF) in children with severe lower pneumonia and identify pathogenic microorganisms that may cause severe pneumonia. The purpose of this study was to use mNGS to explore the potential microbiomes of children with severe pneumonia in a PICU. Methods: We enrolled patients meeting diagnostic criteria for severe pneumonia admitted at PICU of the Children's Hospital of Fudan University, China, from February 2018 to February 2020. In total, 126 BALF samples were collected, and mNGS was performed at the DNA and/or RNA level. The pathogenic microorganisms in BALF were identified and correlated with serological inflammatory indicators, lymphocyte subtypes, and clinical symptoms. Results: mNGS of BALF identified potentially pathogenic bacteria in children with severe pneumonia in the PICU. An increased BALF bacterial diversity index was positively correlated with serum inflammatory indicators and lymphocyte subtypes. Children with severe pneumonia in the PICU had the potential for coinfection with viruses including Epstein-Barr virus, Cytomegalovirus, and Human betaherpesvirus 6B, the abundance of which was positively correlated with immunodeficiency and pneumonia severity, suggesting that the virus may be reactivated in children in the PICU. There was also the potential for coinfection with fungal pathogens including Pneumocystis jirovecii and Aspergillus fumigatus in children with severe pneumonia in the PICU, and an increase in potentially pathogenic eukaryotic diversity in BALF was positively associated with the occurrence of death and sepsis. Conclusions: mNGS can be used for clinical microbiological testing of BALF samples from children in the PICU. Bacterial combined with viral or fungal infections may be present in the BALF of patients with severe pneumonia in the PICU. Viral or fungal infections are associated with greater disease severity and death.

Heparin-binding protein as a biomarker of severe sepsis in the pediatric intensive care unit: A multicenter, prospective study.

OBJECTIVES: The aim of this study is to assess Heparin-binding protein (HBP) as a diagnostic and prognostic biomarker of severe sepsis in the pediatric intensive care unit (PICU). METHODS: A multicenter, prospective study was conducted among children with sepsis in nine PICUs in China from October 2019 to June 2021. Plasma levels of HBP, procalcitonin (PCT), C-reactive protein (CRP), lactate, and white blood cell (WBC) count were determined at enrollment and 72 h after enrollment. RESULTS: Of 355 included patients, 132 patients were diagnosed with non-severe sepsis (referred to as sepsis), 223 patients had severe sepsis. Patients with severe sepsis had significantly elevated levels of HBP compared with sepsis (median 170.5 vs. 74.1 ng/mL, P < 0.001). Adding HBP to a diagnostic model with PCT and lactate could significantly improve the diagnostic capability for severe sepsis. The plasma levels of HBP correlated positively with the number of dysfunctional organs. After adjusting for confounding factors, the declined levels of HBP at 72 h had a significant association with decreased in-hospital mortality (adjusted odds ratio (aOR) 0.242, P < 0.001). The levels of HBP showed weak positive correlations with PCT, CRP, WBC, and no correlation to lactate. CONCLUSIONS: HBP at enrollment can be an independent indicator for severe sepsis and the dynamic changes at 72 h can be a predictor for in-hospital mortality in PICU.

The feasibility, safety, and efficacy of Paxlovid treatment in SARS-CoV-2-infected children aged 6-14 years: a cohort study.

Background: Paxlovid is recognized as an effective medication in preventing the progression of coronavirus disease of 2019 (COVID-19) to severe form in adults; however, its efficacy has remained unknown in pediatric cases. This study aimed to analyze the feasibility, safety, and efficacy of Paxlovid treatment in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-infected children aged 6-14 years. Methods: We conducted a cohort study based on prospectively collected clinical data. We recruited 5 pediatric cases with underlying diseases treated with Paxlovid from 7 April 2022 to 26 May 2022 and 30 age-matched patients with underlying diseases who were not treated with Paxlovid as controls. The safety and efficacy of Paxlovid were primarily assessed by inter-group comparisons. Results: Of the 5 Paxlovid-treated cases, including 1 male and 4 females, 3 and 2 cases were mildly and moderately ill, respectively. The underlying diseases included congenital heart defects, cerebral palsy, Down syndrome, and leukemia. Only 1 patient had received 1 dose of an inactivated SARS-CoV-2 vaccine. Paxlovid was initiated within 5 days after the onset of symptoms in all cases. Comedications were used in 2 cases. In the safety analyses, after Paxlovid initiation, 1 patient had transient diarrhea, and 1 patient had transiently elevated liver enzymes [alanine transaminase (ALT), 125 U/L; aspartate transaminase (AST), 83 U/L; normal range, <40 U/L]. In the efficacy analyses, all 5 Paxlovid-treated cases recovered, with the respective viral shedding times of 11, 4, 10, 9, and 9 days. Compared with age-matched controls, the viral shedding times were not significantly different between groups. Conclusions: Based on the current small sample size study, Paxlovid is a feasible option for treating SARS-CoV-2-infected children aged 6-14 years with underlying diseases. However, the safety and efficacy of Paxlovid warrant further large-scale studies.

Prognostic death factors in secondary hemophagocytic lymphohistiocytosis children with multiple organ dysfunction syndrome receiving continuous renal replacement therapy: A multicenter prospective nested case-control study.

INTRODUCTION: Multiple organ dysfunction syndrome (MODS) with secondary hemophagocytic lymphohistiocytosis (SHLH) causes significant mortality. We aimed to identify the predictor factors for death in pediatric patients with SHLH-associated MODS receiving continuous renal replacement therapy (CRRT). METHODS: This multicentered nested case-control study was conducted from 2016 to 2020. The characteristics were compared between survivors and non-survivors. Logistic regression was applied to identify the risk factors for death. The cutoff values were assessed by receiver operating characteristics curves. RESULTS: Fifty two patients were enrolled in this study. Interleukin-6 level (p = 0.018) and the number of organ dysfunction (p = 0.047) were independent risk factors for death. The cutoff value of 13.12 pg/ml interleukin-6 and three organs dysfunction at CRRT initiation presented a high sensitivity and specificity. CONCLUSION: The number of organ dysfunction and interleukin-6 at CRRT initiation are independent risk factors for death in pediatric patients with SHLH-associated MODS.

Diagnosing Balamuthia mandrillaris encephalitis via next-generation sequencing in a 13-year-old girl.

Balamuthia amoebic encephalitis has a subacute-to-chronic course and is almost invariably fatal owing to delayed diagnosis and a lack of effective therapy. Here, we report a 13-year-old girl with cutaneous lesions and multifocal granulomatous encephalitis. The patient underwent a series of tests and was suspected as having tuberculosis. She was treated with various empiric therapies without improvement. She was finally correctly diagnosed via next-generation sequencing of the cerebrospinal fluid. The patient deteriorated rapidly and died 2 months after being diagnosed with Balamuthia mandrillaris encephalitis. This study highlights the important clinical significance of next-generation sequencing, which provides better diagnostic testing for unexplained paediatric encephalitis, especially that caused by rare or emerging pathogens.

Neuroprotective effects of pinocembrin on ischemia/reperfusion-induced brain injury by inhibiting autophagy.

BACKGROUND: Cerebral ischemia/reperfusion (I/R) injury is a common pathological process after cardiac arrest, shock and acute cerebral infarction recanalization, which causes serious injury in brain function. Pinocembrin (Pino), a natural flavonoid at the highest concentration in propolis, exhibited a variety of biological effects, including antitumor, antimicrobial and anti-inflammatory activities. However, the effects of Pino on brain injured after I/R and the mechanisms of its neuroprotective effects remain elusive. METHODS: In the present study, we used I/R model rats underwent transient cerebral ischemia inducing by four-vessel occlusion and reperfusion. Pino alone or in combination with autophagy inducer rapamycin (RAPA) was administered to I/R rats. The behavior and cognitive function were evaluated by open field test and Morris water maze test. HE staining was used to determine the survival of hippocampus CA1 pyramidal cells. Three key proteins of autophagy, LC3, Beclin1 and p62, were detected by Western blot. RESULTS: Our results showed that Pino could significantly reduce the damage of hippocampus CA1 pyramidal neurons and alleviate the impairments of behavior and cognitive function in I/R rats. Pino also decreased the expression of LC3II and Beclin1 and increased the level of p62 in hippocampus CA1 of I/R rats. In addition, Pino also decreased RAPA-induced neuronal damage and excessive activation of autophagy in I/R rats. CONCLUSIONS: Taken together, these results suggested that Pino could protect the brain injury induced by I/R and the potential mechanisms might attribute to inhibition of autophagy activity.