Objective·To evaluate the diagnostic performance and application value of a domestically developed microfluidic technology-based nucleic acid detection system for gastrointestinal pathogens.Methods·A total of 534 fecal samples were collected from Shanghai Children's Medical Center, Shanghai Jiao Tong University School of Medicine, between April 5th, 2024 and October 6th, 2025. Based on the purpose of testing, the samples were divided into three independent cohorts. Cohort A included 15 samples confirmed positive for Clostridium difficile by the Xpert Clostridium difficile assay (Xpert C. difficile), which were used to evaluate the performance of the gastrointestinal pathogen detection panel (high definition gastrointestinal panel, HD GI panel) for detecting Clostridium difficile. Cohort B included 167 samples from patients requiring comprehensive gastrointestinal pathogen screening and was used to compare the multi-pathogen detection performance of the HD GI panel with the FilmArray gastrointestinal panel (FilmArray GI panel). Cohort C consisted of 352 fecal samples from children with clinically suspected gastrointestinal infections who were admitted to the pediatric intensive care unit (PICU), and was used to analyze the epidemiological characteristics of gastrointestinal pathogens in critically ill children and the clinical application value of the HD GI panel. Sociodemographic and baseline clinical data of children in Cohort C were collected. The Kappa test assessed the agreement between the two detection methods. The McNemar test was used to compare the differences in positive detection rates between the two methods. Pearson correlation analysis was used to evaluate the correlation CT values obtained by the two methods. Using the results of the FilmArray GI panel as the reference standard, the sensitivity, specificity, positive predictive value, and negative predictive value of the HD GI panel, along with their 95% confidence intervals, were calculated. Multivariate Logistic regression analysis was used to identify factors affecting the detection accuracy.Results·The detection results for Clostridium difficile obtained with the HD GI panel and the Xpert C. difficile assay were consistent, and the CT values showed a moderate positive correlation (r=0.594, P=0.019). The overall compliance between the HD GI panel and the FilmArray GI panel was 92.22%, and the Kappa test indicated good consistency between the two methods (κ=0.823, P<0.001). Compared with the FilmArray GI panel, the HD GI panel achieved a sensitivity of 88.89%, a specificity of 93.81%, a positive predictive value of 87.27%, and a negative predictive value of 94.64%. Among critically ill children, the overall positivity rate detected by the HD GI panel was 39.77%. Norovirus GⅡ, rotavirus A, and adenovirus were the predominant viral pathogens, whereas enteroaggregative Escherichia coli (EAEC), Clostridium difficile, and Salmonella were the main bacterial pathogens. Co-infections accounted for 30.00%. Compared with clinical diagnosis, the HD GI panel demonstrated a sensitivity and a negative predictive value of 100.00%, with a diagnostic accuracy of 93.60%. The Kappa test showed a high level of agreement between the two methods (κ=0.860, P<0.001). Multivariate Logistic regression analysis further revealed that the presence of underlying diseases was an independent factor associated with detection accuracy (OR=0.051, 95%CI 0.014‒0.141, P<0.001).Conclusion·The HD GI panel demonstrates diagnostic performance comparable to that of mainstream platforms for gastrointestinal pathogen detection, offering advantages such as simple operation, rapid turnaround, and a pathogen spectrum consistent with the characteristics of pediatric gastrointestinal infections in China. It is well suited for use in primary hospitals and high-throughput testing settings.

We aimed to evaluate whether the FILMARRAY® Pneumonia Panel (FAPP) added to standard care could effectively guide antibiotic prescriptions in hospitalized pediatric patients with infectious pneumonia. In this single-center, open-label, parallel-group randomized controlled clinical trial, we enrolled hospitalized children (28 days to 18 years old) diagnosed with infectious pneumonia. Participants were randomized to standard care only (control) or to added FAPP (intervention group). The primary outcome was the proportion of patients with antibiotic change within 72 h. Between December 8, 2021, and February 16, 2023, we enrolled 315 patients (157 in the intervention group and 158 in the control group). Pathogens were detected in 86.6 https://www.chictr.org.cn/showproj.html?proj=128024 .

Inflammatory bowel disease (IBD) is a chronic relapsing and remitting disorder in which loss of intrinsic enteric neurons (iENs) has been documented. However, the contribution of gut microbiota to the loss of iENs in IBD remains poorly defined. Here, we identify an IBD-enriched intestinal pathogen, Clostridium symbiosum (C. symbiosum), which exacerbates iEN loss and colitis. Mechanistically, C. symbiosum-derived succinate, emerging as a central mediator, drives macrophage glycolysis via the H3K79succ/HK2 axis, thereby sustaining IL-1β secretion, which, in turn, promotes neuronal-specific NLRP3 inflammasome activation and consequent neuronal loss. We further demonstrated that preventing iEN loss effectively improves outcomes in C. symbiosum-exacerbated colitis. Importantly, we identified phiCS-1, an endolysin from C. symbiosum-specific bacteriophages, which efficiently lyses C. symbiosum and markedly attenuates C. symbiosum-mediated iEN loss and colitis. Together, our study provides insights into the intricate interplay between gut microbiota and immune-neuron crosstalk, offering avenues for targeted therapeutic interventions in IBD.

Dopaminylation, the covalent attachment of dopamine to the side chain of glutamine in proteins, represents a newly characterized class of posttranslational modifications. Because of the limited identification of substrates, the functions and molecular mechanisms associated with dopaminylation remain largely uncharacterized. Using an alkyne-functionalized dopamine probe, we developed a method for selectively enriching dopaminylated proteins in whole-cell systems. This approach provided a comprehensive resource of 4,133 dopamine-enriched protein candidates and peptide-level analysis with acid-cleavable tags identified 1,181 putative dopaminylated proteins, including histone H4 dopaminylation at Q27 (H4Q27dop), which we further validated. Functionally, H4Q27dop acts as a transcriptional repressor in a neuroblastoma model, where it blocks CEBPD binding at the CCND1 promoter, leading to transcriptional downregulation of CCND1 and subsequent suppression of cell proliferation. Our findings provide both a valuable resource of dopaminylated substrate proteins and a distinct mechanistic insight into how dopamine regulates neuroblastoma cell growth.

Elevated lactate levels are a hallmark of severe infections and are associated with poor outcomes in sepsis patients, but the underlying mechanisms remain poorly understood. Recent findings have shown that lactate can covalently modify histones (e.g., histone lactylation) in macrophages, acting as a critical epigenetic regulator of inflammatory response. Here, we demonstrate that histone lactylation also occurs in neutrophils—the first immune cells mobilized during acute inflammation—and is functionally important for their activation. Using both DMSO-differentiated HL-60 (dHL-60) cells and primary neutrophils, we found that LPS stimulation significantly increased intracellular lactate levels and histone lactylation, particularly at the H4K8 site. These changes enhanced cytokine release, ROS production, and chemotaxis. Lactate further amplified these effects, while inhibition of glycolysis or p300 suppressed them. Multi-omics analyses revealed substantial enrichment of H4K8la at the promoter region of WTAP, a key m6A methyltransferase component, promoting its expression via CEBP/β recruitment. WTAP knockdown significantly reduced m6A modifications of TLR2 mRNA and impaired its stability. Both WTAP knockdown and TLR2 inhibition markedly dampened the inflammatory responses. Importantly, this glycolysis-H4K8la-WTAP-TLR2 axis was further validated in LPS-induced septic mice and pediatric sepsis patients, highlighting its clinical relevance. In summary, our findings uncover a novel lactate-driven epigenetic—post-transcriptional regulatory circuit that amplifies neutrophil inflammatory responses, expanding the regulatory framework of innate immunity and providing potential therapeutic targets for hyperinflammation.

Staphylococcal aureus ( S. aureus ) infection can lead to a wide range of diseases such as sepsis and pneumonia. Staphylococcal superantigen-like (SSL) proteins, expressed by all known S. aureus strains, are shown to be involved in immune evasion during S. aureus infection. Here, we show that SSL10, an SSL family protein, exhibits potent cytotoxicity against human cells (HEK293T and HUVEC) by inducing necroptosis upon binding to its receptor TNFR1 on the cell membrane. After binding, two distinct signaling pathways are activated downstream of TNFR1 in a RIPK3-dependent manner, i.e., the RIPK1-RIPK3-MLKL and RIPK3-CaMKII-mitochondrial permeability transition pore (mPTP) pathways. Knockout of ssl10 in S. aureus significantly reduces cytotoxicity of the culture supernatants of S. aureus , indicating that SSL10 is involved in extracellular cytotoxicity during infection. We determined the crystal structure of SSL10 at 1.9 Å resolution and identified a positively charged surface of SSL10 responsible for TNFR1 binding and cytotoxic activity. This study thus provides the description of cytotoxicity through induction of necroptosis by the SSL10 protein, and a potential target for clinical treatment of S. aureus -associated diseases.

Respiratory tract infections (RTIs) are a significant global health concern ranging from common colds to severe respiratory illnesses, particularly in children among underdeveloped regions. Despite the development and deployment of multiple diagnostic technologies, PCR and nucleic acid amplification remain central due to their high sensitivity and specificity. Economical and rapid multiplexed Point-Of-Care Testing (POCT) solutions are crucial for enabling clinicians to swiftly identify infection causes, thereby facilitating targeted therapy, reducing antibiotic use and drug resistance, and monitoring community-wide infection and resistance trends. At Shanghai Children’s Medical Center (SCMC), we employed both multiplexed POCT (FlashDx RP1.1 for 13 common pathogens, primarily targeting upper RTIs) and metagenomic Next-Generation Sequencing (mNGS/tNGS) by VisionMedicals to screen patients presenting with acute RTI symptoms. Over 50,000 outpatient and emergency room samples were analyzed using nasal pharyngeal swabs (POCT) between 2023 and 2024, in addition to more than 600 inpatient samples were evaluated using sputum/BALF samples (NGS) in late 2023, after the pandemic period. The POCT platform showed a 68.5% overall positivity rate and a 9% rate of multiple infections. The most common pathogens were HRV/HEV (25%), Adenovirus (19%) and Mycoplasma pneumoniae (MP) (13%), this is also a dynamic and varied infection spectrum reflective of the complex post-pandemic pathogen landscape. mNGS/tNGS analysis revealed an increase in infections caused by Streptococcus pneumoniae, Staphylococcus aureus and E. coli, particularly during the MP outbreak, where the drug resistance rate surged from 45% to 85%, highlighting the critical role of these technologies in guiding treatments. The integration of multiplexed POCT and NGS tests into Respiratory Syndromic Testing has proven to be pivotal. Our findings demonstrate a shift in the infection spectrum post-pandemic, emphasizing the value of these diagnostic tools in improving patient management, reducing antibiotic misuse, and ultimately decreasing healthcare costs.

Neuroblastoma(NB)is the most common extracranial solid tumor in children.Despite treatment advances,the survival rates of high-risk NB patients remain low.This highlights the urgent need for a deeper understanding of the molecular mechanisms driving NB progression to support the development of new therapeutic strategies.In this study,we demonstrated that the reduced levels of DNAJC12,a protein involved in metabolic regulation,are associated with poor prognosis in NB patients.Our data indicate that low DNAJC12 expression activates glycolysis in NB cells,leading to increased lactic acid production and histone H4 lysine 5 lactylation(H4K5la).Elevated H4K5la upregulates the transcription of COL1A1,a gene implicated in cell metastasis.Immunohistochemistry staining of NB patient samples confirmed that high H4K5la levels correlate with poor clinical outcomes.Furthermore,we showed that inhibiting glycolysis,reducing H4K5la,or targeting COL1A1 can mitigate the invasive behavior of NB cells.These findings reveal a critical link between metabolic reprogramming and epigenetic modifications in the context of NB progression,suggesting that H4K5la could serve as a novel diagnostic and prognostic marker,and shed light on identifying new therapeutic targets within metabolic pathways for the treatment of this aggressive pediatric cancer.

Background Diarrhea is a common complication of hematopoietic stem cell transplantation (HSCT) and is associated with substantial morbidity, but its etiology is often unknown. Etiologies of diarrhea in this population include infectious causes, chemotherapy- or medication-induced mucosal injury and graft-versus-host disease (GVHD). Distinguishing these potential causes of diarrhea is challenging since diarrheal symptoms are often multifactorial, and the etiologies often overlap in transplant patients. The objectives of this study were to evaluate whether the FilmArray gastrointestinal (GI) panel would increase diagnostic yield and the degree to which pre-transplantation colonization predicts post-transplantation infection. Methods From November 2019 to February 2021, a total of 158 patients undergoing HSCT were prospectively included in the study. Stool specimens were obtained from all HSCT recipients prior to conditioning therapy, 28 ± 7 days after transplantation and at any new episode of diarrhea. All stool samples were tested by the FilmArray GI panel and other clinical microbiological assays. Results The primary cause of post-transplantation diarrhea was infection (57/84, 67.86%), followed by medication (38/84, 45.24%) and GVHD (21/84, 25.00%). Ninety-five of 158 patients were colonized with at least one gastrointestinal pathogen before conditioning therapy, and the incidence of infectious diarrhea was significantly higher in colonized patients (47/95, 49.47%) than in non-colonized patients (10/63, 15.87%) ( P < 0.001). Fourteen of 19 (73.68%) patients who were initially colonized with norovirus pre-transplantation developed a post-transplantation norovirus infection. Twenty-four of 62 (38.71%) patients colonized with Clostridium difficile developed a diarrheal infection. In addition, FilmArray GI panel testing improved the diagnostic yield by almost twofold in our study (55/92, 59.78% vs. 30/92, 32.61%). Conclusions Our data show that more than half of pediatric patients who were admitted for HSCT were colonized with various gastrointestinal pathogens, and more than one-third of these pathogens were associated with post-transplantation diarrhea. In addition, the FilmArray GI panel can increase the detection rate of diarrheal pathogens in pediatric HSCT patients, but the panel needs to be optimized for pathogen species, and further studies assessing its clinical impact and cost-effectiveness in this specific patient population are also needed. Graphical abstract

BackgroundThe correlation between the gut microbiota and airway inflammation in childhood allergic rhinitis (AR), particularly concerning allergen exposure, remains insufficiently explored. This study aimed to link gut microbiota changes with house dust mite (HDM)-specific IgE responses in pediatric AR.MethodsUsing metagenomic shotgun sequencing, we compared the fecal microbiota of 60 children with HDM-AR to 48 healthy controls (HC), analyzing the link to IgE reactions. We examined the effects of oral Escherichia (E.) fergusonii treatment in mice sensitized with ovalbumin and HDM on allergic symptoms, mucosal cell infiltration, Th1/Th2/Tregs balance in the spleen, serum cytokine levels, and E. fergusonii presence in feces.ResultsChildren with HDM-AR have a less diverse gut microbiome and lower levels of E. fergusonii compared to controls, with a negative correlation between E. fergusonii abundance and HDM-specific IgE levels. In mice sensitized with OVA and HDM, oral administration of E. fergusonii improved allergic symptoms, reduced nasal eosinophils/mast cells infiltration and adjusted Th cell populations towards a non-allergic profile in splenic lymphocytes with exception of IFN-gamma change in serum.ConclusionThese findings underline the potential of targeting gut microbiota, particularly E. fergusonii, in managing childhood HDM-AR, suggesting a promising approach for future interventions.ImpactThe composition and distribution of gut microbiota in children with HDM-AR are significant changed.The abundance of genus is decreased in HDM-AR children.HDM-specific IgE levels are strongly negatively associated with abundance.Oral administration of effectively suppresses allergic responses in murine model.These findings offer novel insights into the diagnosis and treatment of HDM-AR, which suggested that holds promise as a potential therapeutic avenue for managing HDM-AR.

Dopaminylation, the covalent attachment of dopamine to the side chain of glutamine (Gln, Q) in proteins, has been identified as a class of posttranslational modification. Due to the limited number of substrates, the functions and underlying molecular mechanisms of dopaminylation are not fully characterized. Utilizing an alkyne-functionalized dopamine probe, we have developed a method to selectively enrich dopaminylated proteins in the whole-cell context. We identified 4,133 proteins potentially modified with dopamine and validated the modification of histone H4 glutamine 27 by dopamine (H4Q27dop). H4Q27dop mainly exerts transcriptional inhibition function in neuroblastoma cells, and can inhibit cell proliferation through downregulating cyclin D1 gene CCND1 transcription, a classical well-known proliferation booster. Our study provides a valuable resource of putative substrate proteins modified with dopamine and reveals a novel mechanism of dopamine regulating cell growth in a neuroblastoma cancer model.

IntroductionAllergen-specific immunotherapy (AIT) induces long-term immune tolerance to allergens and is effective for treating allergic rhinitis (AR). However, the impact of sublingual immunotherapy (SLIT) on gut microbiota from AR patients and its correlation with treatment efficacy remains unclear.MethodsIn the present study, we enrolled 24 AR patients sensitized to Dermatophagoides farinae (Der-f) and 6 healthy donors (HD). All AR patients received SLIT treatment using standardized Der-f drops. Stool samples were collected from AR patients before treatment, and 1- and 3-months post-treatment, as well as from HD, for metagenomic sequencing analysis.ResultsAR patients had significantly lower richness and diversity in gut microbiota compared to HD, with notable alterations in composition and function. Besides, three months post-SLIT treatment, significant changes in gut microbiota composition at the genus and species levels were observed in AR patients. Streptococcus parasanguinis_B and Streptococcus parasanguinis, which were significantly lower in AR patients compared to HD, increased notably after three months of treatment. LEfSe analysis identified these species as markers distinguishing HD from AR patients and AR patients pre- from post-SLIT treatment. Furthermore, changes in the relative abundance of S. parasanguinis_B were negatively correlated with changes in VAS scores but positively correlated with changes in RCAT scores, suggesting a positive correlation with effective SLIT treatment. DiscussionSLIT treatment significantly alters the gut microbiota of AR patients, with S. parasanguinis_B potentially linked to its effectiveness. This study offers insights into SLIT mechanisms and suggests that specific strains may serve as biomarkers for predicting SLIT efficacy and as modulators for improving SLIT efficacy.

BackgroundMolecular diagnostic technology is the foundation of precision medicine, which has the advantages of good specificity, high sensitivity, strong targeting, rapiddiagnosis, etc. It has a wide range of applications in the field of pediatrics. However, molecular diagnostic technology is characterized by complicated experimental operation, high difficulty in data analysis and interpretation, in consistent standards among laboratories, and difficult standardization of technical processes. Enhancing the application value of molecular diagnostic technology in pediatrics and promoting the high-quality development of the discipline requires careful consideration by relevant management and professionals.MethodsThis study firstly outlines the development history of molecular diagnostic technology. Then, it analyzes the application of molecular diagnostic technology in the field of pediatrics. Finally, it explores the countermeasures for the management of molecular diagnostic laboratories.ResultsThis study highlights the importance of molecular diagnostic technology in providing information and decision-making basis for disease prevention, prediction,diagnosis, treatment and regression. It has a wide range of applications in the molecular diagnosis of pediatric hereditary diseases, malignant tumors and infectious diseases. In addition, the countermeasures for the management of molecular diagnostic laboratories are proposed from the five aspects of laboratory, personnel team construction, standardized management,multidisciplinary cross-discipline, research and translation, safety managementand ethical supervision, and management upgrading and modernization.ConclusionsMolecular diagnostic technology, as the basis of precision medicine, has become one of the important frontier fields in the development of contemporary pediatric medicine. Enhanced laboratory capacity in molecular diagnostic techniques can improve outcomes in the prevention, prediction, diagnosis, treatment, prognosis and research of pediatricdiseases, and lay the groundwork for child healthcare.

OBJECTIVE:Rare variants of CCNK (cyclin K) give rise to a syndrome with intellectual disability. The purpose of this study was to describe the genotype-phenotype spectrum of CCNK-related syndrome and the underlying molecular mechanisms of pathogenesis.METHODS:We identified a number of de novo CCNK variants in unrelated patients. We generated patient-induced pluripotent stem cells (iPSCs) and neural progenitor cells (NPCs) as disease models. In addition, we constructed NPC-specific Ccnk knockout (KO) mice and performed molecular and morphological analyses.RESULTS:We identified 2 new patients harboring CCNK missense variants and followed-up 3 previous reported patients, which constitute the largest patient population analysis of the disease. We demonstrate that both the patient-derived NPC models and the Ccnk KO mouse displayed deficient NPC proliferation and enhanced apoptotic cell death. RNA sequencing analyses of these NPC models uncovered transcriptomic signatures unique to CCNK-related syndrome, revealing significant changes in genes, including WNT5A, critical for progenitor proliferation and cell death. Further, to confirm WNT5A's role, we conducted rescue experiments using NPC and mouse models. We found that a Wnt5a inhibitor significantly increased proliferation and reduced apoptosis in NPCs derived from patients with CCNK-related syndrome and NPCs in the developing cortex of Ccnk KO mice.INTERPRETATION:We discussed the genotype-phenotype relationship of CCNK-related syndrome. Importantly, we demonstrated that CCNK plays critical roles in NPC proliferation and NPC apoptosis in vivo and in vitro. Together, our study highlights that Wnt5a may serve as a promising therapeutic target for the disease intervention. ANN NEUROL 2023;94:1136-1154.

Background Studies on mNGS application in pediatric oncology patients, who are at high risk of infection, are quite limited. Methods From March 2020 to June 2022, a total of 224 blood samples from 195 pediatric oncology patients who were suspected as bloodstream infections were enrolled in this study. Their clinical and laboratory data were retrospectively reviewed, and the diagnostic performance of mNGS was assessed. Results Compared to the reference tests, mNGS showed significantly higher sensitivity (89.8% vs 32.5%, P < 0.001) and clinical agreement (76.3% vs 51.3%, P < 0.001) in detecting potential pathogens and distinguishing BSI from non-BSI. Especially, mNGS had an outstanding performance for virus detection, contributing to 100% clinical diagnosed virus. Samples from patients with neutropenia showed higher incidence of bacterial infections ( P = 0.035). The most identified bacteria were Escherichia coli , and the overall infections by gram-negative bacteria were significantly more prevalent than those by gram-positive ones (90% vs 10%, P < 0.001). Overall, mNGS had an impact on the antimicrobial regimens’ usage in 54.3% of the samples in this study. Conclusions mNGS has the advantage of rapid and effective pathogen diagnosis in pediatric oncology patients with suspected BSI, especially for virus. Impact Compared with reference tests, mNGS showed significantly higher sensitivity and clinical agreement in detecting potential pathogens and distinguishing bloodstream infections (BSI) from non-BSI. mNGS is particularly prominent in clinical diagnosed virus detection. The incidence of bacterial infection was higher in patients with neutropenia, and the overall infection rate of Gram-negative bacteria was significantly higher than that of Gram-positive bacteria. mNGS affects the antimicrobial regimens’ usage in more than half of patients.

Objective: To evaluate the risk of major infections in children with newly diagnosed childhood-onset systemic lupus erythematosus Methods: Predictors of major infections were identified by the multivariable logistic regression. Major infection free was defined as no major infection events within 6 months after the diagnosis of cSLE. The Kaplan-Meier survival plot was performed. A prediction model for major infection events was established and examined by receiver operating characteristic (ROC) curve analysis. Results: A total of 98 eligible patients were recorded in the medical charts. Sixty-three documented events of major infections were found in 60 (61.2%) cSLE patients. Furthermore, 90.5% (57/63) of infection events occurred within the first 6 months after the diagnosis of cSLE. The high SLEDAI (SLEDAI >10), lupus nephritis and lymphocyte count <0.8x109/L were predictors for major infections. The CALL score (Children with high disease activity [SLEDAI >10], lymphopenia, and LN) was defined by the number of predictors. Patients were then categorized into two groups: low-risk (score 0-1) and high-risk (score 2-3). Patients in the high-risk group had higher rates of the major infection occurrence than those in the low-risk group during the 6 months after the diagnosis of the cSLE (P<0.001) (HR:14.10, 95% CI 8.43 to 23.59). The ROC curve analysis indicated that the CALL score was effective both in the whole cSLE cohort [area under the curve (AUC) = 0.89, 95% CI: 0.81-0.97] and in the subgroup of lung infections (n = 35) (AUC = 0.79, 95% CI: 0.57-0.99). Conclusion: High disease activity, LN and lymphopenia were predictors for major infections in newly diagnosed cSLE patients. Specific predictors help identify the cSLE patients with the high risk of major infections. The CALL score could be a useful tool to stratify cSLE patients in practice.

With adult patients, the measurement of [TIMP-2]*[IGFBP7] can predict the risk of moderate to severe AKI within 12 h of testing. In pediatrics, however, the performance of [TIMP-2]*[IGFBP7] as a predictor of AKI was less studied and yet to be widely utilized in clinical practice. This study was conducted to validate the utility of [TIMP-2]*[IGFBP7] as an earlier biomarker for AKI prediction in Chinese infants and small children. We measured urinary [TIMP-2]*[IGFBP7] using NEPHROCHECK® at eight perioperative time points in 230 patients undergoing complex cardiac surgery and evaluated the performance of [TIMP-2]*[IGFBP7] for predicting severe AKI within 72 h of surgery. A total of 50 (22%) of 230 developed AKI stages 2–3 within 72 h after CPB initiation. In the AKI stage 2–3 patients, two patterns of serum creatinine (SCr) elevations were observed. The patients with only a transient increase in SCr within 24 h (< 24 h, early AKI 2–3) did not experience a worse outcome than patients in AKI stage 0–1. AKI stage 2–3 patients with SCr elevation after 24 h (24–72 h, late AKI 2–3), as well as AKI dialysis patients (together designated severe AKI), did experience worse outcomes. Compared to AKI stages 0–1, significant elevations of [TIMP-2]*[IGFBP7] values were observed in severe AKI patients at hours T2, T4, T12, and T24 following CPB initiation. The AUC for predicting severe AKI with [TIMP-2]*[IGFBP7] at T2 (AUC = 0.76) and maximum T2/T24 (AUC = 0.80) are higher than other time points. The addition of the NEPHROCHECK® test to the postoperative parameters improved the risk assessment of severe AKI. Multiple AKI phenotypes (early versus late AKI) were identified after pediatric complex cardiac surgery according to SCr-based AKI definition. Urinary [TIMP-2]*[IGFBP7] predicts late severe AKI (but not early AKI) as early as 2 h following CPB initiation. A higher resolution version of the Graphical abstract is available as Supplementary information

BACKGROUND:Metagenomic next-generation sequencing (mNGS) has the potential to become a complementary, if not essential, test in some clinical settings. However, the clinical application of mNGS in a large population of children with various types of infectious diseases (IDs) has not been previously evaluated.METHODS:From April 2019 to April 2021, 640 samples were collected at a single pediatric hospital and classified as ID [479 (74.8%)], non-ID [NID; 156 (24.4%)], and unknown cases [5 (0.8%)], according to the final clinical diagnosis. We compared the diagnostic performance in pathogen detection between mNGS and standard reference tests.RESULTS:According to final clinical diagnosis, the sensitivity and specificity of mNGS were 75.0% (95% CI: 70.8%-79.2%) and 59.0% (95% CI: 51.3%-66.7%), respectively. For distinguishing ID from NID, the sensitivity of mNGS was approximately 45.0% higher than that of standard tests (75.0% vs 30.0%; P < 0.001). For fungal detection, mNGS showed positive results in 93.0% of cases, compared to 43.7% for standard tests (P < 0.001). Diagnostic information was increased in respiratory system samples through the addition of meta-transcriptomic sequencing. Further analysis also showed that the read counts in sequencing data were highly correlated with clinical diagnosis, regardless of whether infection was by single or multiple pathogens (Kendall's tau b = 0.484, P < 0.001).CONCLUSIONS:For pediatric patients in critical condition with suspected infection, mNGS tests can provide valuable diagnostic information to resolve negative or inconclusive routine test results, differentiate ID from NID cases, and facilitate accurate and effective clinical therapeutic decision-making.