European Journal of Clinical Microbiology & Infectious Diseases

ImportanceCurrent guidelines from the World Health Organization, Centers for Disease Control and Prevention, and Academy of Breastfeeding Medicine recommend discarding all milk remaining in bottles immediately after infant feeding. However, these recommendations lack supporting microbiological evidence from studies of actual infant feeding, imposing substantial financial and emotional burden on the 78 million families worldwide who bottle-feed their infants. ObjectiveTo determine (1) the financial, emotional, and time burden associated with bottle feeding and parental milk disposal practices, and (2) bacterial growth in leftover human milk and formula under different storage conditions. Design(1) Cross-sectional online survey (January 2023-February 2024) and (2) prospective microbiological cohort study. Setting(1) Online survey, (2) infants recruited in Hannover, Germany Participants(1) Survey respondents (n=1056; 99% mothers) and (2) healthy, full-term, bottle-fed infants (n=44; 17 humanmilk, 27 formula) aged 0-12 months. Main Outcomes and MeasuresParental burden scores, milk disposal frequency, and bacterial colony-forming units (CFU)/ml in milk samples before feeding, immediately after feeding, and at 4, 8, and 24 hours post-feeding at 4{degrees}C and 20{degrees}C. ResultsAmong surveyed parents, 46% discarded leftover milk daily, yet 84% reported they would keep milk longer if deemed safe. In microbiological testing, median bacterial burden in humanmilk increased from 4200 CFU/ml (range 300-350,000) pre-feeding to 24,600 CFU/ml (range 1900-29,004,400) post-feeding, but showed no significant further increase at 4 hours (p=0.82) or 8 hours (p=0.64) when stored at either 4{degrees}C or 20{degrees}C. Formula showed similar stability: median CFU/ml increased from 0 (range 0-10,700) to 11,700 (range 1900-630,000) post-feeding, with no significant change at 4 hours (p=0.91) or 8 hours (p=0.73) at either temperature. Significant bacterial growth occurred only after 24 hours at 20{degrees}C (p<0.001). Conclusions and RelevanceBacterial burden in leftover infant milk remained stable below concerning thresholds for 8 hours when refrigerated and 4-8 hours at room temperature, challenging current guidelines that mandate immediate disposal. Evidence-based guideline revision could reduce financial burden and milk waste for families around the globe without compromising infant safety. Key PointsO_ST_ABSQuestionC_ST_ABSHow long is it safe to offer leftover milk in a bottle to an infant that has previously drunk from it? FindingsThe number of bacteria in leftover human milk or formula did not significantly increase from 0 to 8h post-feeding in milk bottles sampled from 44 infants, regardless of whether the milk was kept at room temperature or refrigerated. MeaningLeftover milk may be safely reoffered beyond the limits of the current guidelines.

IntroductionCentral line-associated bloodstream-infection (CLA-BSI) and catheter-related bloodstream infections (CR-BSI) remain a significant concern in pediatric inpatient units. ObjectiveTo analyze a case series of CLA-BSI and CR-BSI in hospitalized pediatric patients in hospitals with rigorous infection prevention measures. Materials and MethodsThis was an analytical, descriptive, and retrospective study conducted in patients aged 0 to 18 years, admitted between August 2023 and March 2025, with a diagnosis of CLA-BSI or CR-BSI in two pediatric hospitals in Rio de Janeiro, Brazil. Variables potentially associated with the occurrence of infection were analyzed. ResultsA total of 86 infections were evaluated, comprising 66 CLA-BSI and 20 CR-BSI. Sixty patients (69.8%) were male, with a mean age of 71.8 months. Sixty-five (83.7%) had previous comorbidities, 63 (73.2%) had a prior hospitalization, and 27 (31.4%) had another invasive device. The mean time from catheter insertion to infection diagnosis was 32.1 days, and the mean time from hospital admission to infection onset was 18.45 days. Gram-negative bacteria were isolated in 40/86 (46.5%) cases. At 30 days post-infection, 61/86 (70.9%) had been discharged, 20/86 (23.3%) remained hospitalized, and 5/86 (5.8%) had died. There was no correlation between the bacterial group and the type of catheter used (p=0.068), nor between infection type (CLA-BSI vs. CR-BSI) and mortality outcome (p=1). ConclusionsCLA-BSI and CR-BSI occurred predominantly in patients with prolonged hospital stays and underlying comorbidities, and were mainly caused by Gram-negative bacteria.

ObjectivesVentilator-associated pneumonia (VAP) is a significant complication in pediatric patients with severe bronchiolitis undergoing mechanical ventilation (MV) in pediatric intensive care units (PICUs). The role of respiratory microbiota in VAP development remains underexplored in this vulnerable population. This study aimed to characterize respiratory microbiota in critically ill children with severe bronchiolitis receiving critical care and identify microbial patterns associated with VAP. MethodsWe conducted a cohort study in paediatric patients with severe bronchiolitis requiring MV at a tertiary PICU in Catalonia, Spain. Epidemiological, clinical and microbiological data were collected. Respiratory microbiota was assessed using 16S rRNA gene sequencing of nasopharyngeal (NP) aspirates and bronchoalveolar lavage (BAL) samples obtained before and during MV. ResultsBaseline NP microbiota differed significantly between VAP and non-VAP groups, with overrepresentation of Moraxella, Enterobacter, and Amniculibacterium genera and underrepresentation of Prevotella in patients who developed VAP. BAL microbiota showed fewer differences, although Enterobacter was more abundant in VAP cases. Random forest models demonstrated strong predictive performance, with the model integrating NP microbiota and clinical parameters achieving the highest accuracy (AUC 0.956). ConclusionsSpecific nasopharyngeal microbial signatures, combined with clinical factors, may serve as risk markers for VAP in mechanically ventilated children, potentially guiding targeted prevention strategies in PICUs.

Background and objectivesCeftazidime-Avibactam (CAZ-AVI) is one of the last options to treat Enterobacteriales and Pseudomonas aeruginosa carbapenem-resistant. We aim to describe the susceptibility profile of bloodstream isolates of Enterobacterales and Pseudomonas aerunosa to ceftazidime-avibactam (CAZ-AVI) among strains resistant to third- and fourth-generation cephalosporins and/or carbapenems. MethodsWe conducted a retrospective descriptive study in two pediatric hospitals of Rio de Janeiro city, Brazil, between January 2023 and February 2025. All blood samples with resistance to third/fourth cephalosporins and/or carbapenem resistance were tested to CAZ-AVI, according to the BRCast methodology. Sensibility of CAZ-AVI and clinical profile of patients and outcomes were described. ResultsWe analyzed 116 blood samples. Of these, 107/116 (92.2%) were resistant to third/fourth-generation cephalosporins with susceptibility to carbapenems, and 9/116 (7.8%) were resistant to both third/fourth-generation cephalosporins and carbapenems. Overall susceptibility to CAZ-AVI was 107/116 (92.2%). The 116 blood samples represented 73 bloodstream infections (BSI) in 66 patients, including 66 single episodes and 7 persistent BSIs. Of the 73 infections, 69(94.5%) were caused by Enterobacterales and 4 (5.5%) by Pseudomonas aeruginosa. Twenty-two (30.1%) infections were detected at hospital admission, and 51 (69.9%) were healthcare-associated infections. Death occurred in 5/73 (6.8%) patients. Length of hospital stay (p=0.01596) were statistically significantly higher in non-survivors compared to survivors. The CAZ-AVI was prescribed for four patients with Enterobacteriales or Pseudomonas aeruginosa infections with clearance from the blood. ConclusionSusceptibility of CAZ-AVI to BSI in children was higher and this antibiotic could be an option to treat carbapenem-resistant infection due to Enterobacteriales and Pseudomonas aeruginosa.

BackgroundRecurrent urinary tract infection (rUTI) is a major diagnostic and management challenge. Dysregulated innate immune responses, including antimicrobial peptides and cytokines, may underlie UTI susceptibility. This study investigates whether urinary concentrations of antimicrobial peptides and cytokines differ in children and adolescents with a history of rUTI and whether they can accurately classify rUTI status. MethodsUrine samples were collected from 42 girls and adolescent females with a history of rUTI and 37 matched healthy controls. Concentrations of antimicrobial peptides (alpha-defensins 1-3, beta-defensin 1, cathelicidin, secretory leukocyte protease inhibitor, lipocalin 2, and ribonuclease 7) and cytokines (interleukin-1 beta, interleukin-6, interleukin-8, and tumor necrosis factor alpha) were quantified using enzyme-linked immunosorbent assays. A logistic regression model with variable selection was developed to classify rUTI participants based on urinary biomarkers and clinical factors. ResultsCompared to controls, participants with rUTI had lower concentrations of beta-defensin 1, cathelicidin, and ribonuclease 7, and higher concentrations of alpha-defensins 1-3, lipocalin 2, and secretory leukocyte protease inhibitor. Cytokine concentrations, including interleukin-1 beta, interleukin-6, interleukin-8, and tumor necrosis factor alpha, were elevated in the rUTI group. The predictive model demonstrated high accuracy, with an area under the receiver operating curve of 0.97 and a prevalence-adjusted area under the precision-recall curve of 0.94. ConclusionsGirls and adolescent females with rUTI exhibit a distinct urinary immune profile characterized by dysregulated antimicrobial peptides and elevated proinflammatory cytokines. A predictive model integrating these biomarkers with clinical features accurately classified rUTI status, supporting their potential utility as diagnostic tools for pediatric UTI.

A multivariant total subclass analysis has been performed for a control cohort (n=15) and a long COVID patient cohort (n=15) measuring the IgG1, IgG2, IgG3, IgG4 and IgE response to the following 14 variants of SARS-CoV-2: Wuhan, Alpha, Delta, BA.1, BA.2, BA.5, EG.5.1, XBB.1.5, BA.2.75, CH.1.1, BA.2.12.1, BQ.1.1, JN.1, and KP.3. Significant differences (p < 0.05 and p < 0.005) between concentrations of IgG subclasses by variant were found in 24% of variants and in mean-normalised distributions. The medians of the mean-normalised distributions were significantly lower for IgG1 (p < 0.05) in long COVID patients compared with controls, and significantly higher (p < 0.005) for levels of IgG3, IgG4 and IgE for long COVID patients. A preliminary diagnostics classification analysis performed by variation of the mean-normalised upper and lower percentiles symmetrically for IgG3 showed a long COVID diagnostic sensitivity of 80%, and specificity of 80% for the 60th percentile threshold of the control cohort. Three types of long COVID can be identified: patients with at least one variant below the threshold (hypo-immune), patients with at least one variant above the threshold (hyperimmune) and patients with IgG3 levels within the reference range. The multivariant subclass spectrum indicates IgG4 and IgE elevations due to potential chronic antigen exposure from persistent virus or autoimmunity and may indicate potential therapeutic interventions.

ObjectivesDiagnosis of community-acquired Legionnaires disease (CALD) relies on microbiological testing. Routine testing in hospitalised CAP patients has low positivity rates. We externally validated a Legionella prediction score, assessed its applicability in routine care, and explored potential updates. MethodsWe analysed data from 196 CALD patients from 20 Swiss hospitals and 196 Legionella-negative CAP controls matched by date of diagnosis ({+/-}14 days; August 2022-March 2024). We assessed the availability of the original score predictors (fever, no/dry cough, hyponatremia, elevated CRP, elevated LDH, low platelet count) in routine care and the original scores discriminative performance. The dataset was split into development and validation cohorts to evaluate whether simplifying modifications improved predictive performance. ResultsThe original score showed 91% (95% CI: 86-96%) sensitivity and 35% (95% CI: 28-42%) specificity at a cut-off [&ge;]2; LDH was infrequently measured, and platelet count was a poor predictor. The simplified SwissLEGIO score (fever >38{degrees}C, sodium <133 mmol/L, CRP >180 mg/L, no/dry cough, prior {beta}-lactam therapy) maintained high sensitivity (88-92%) and showed improved specificity (46-58%) at cut-off [&ge;]2. ConclusionThe SwissLEGIO score is an easy-to-apply screening tool to rule out CALD in hospitalised CAP patients with scores <2 and may reduce testing by 36-52% at a CALD prevalence of 4%.

Mexico has experienced recurrent viral epidemics of substantial intensity, including hyperendemic dengue, COVID-19, and recent reports of avian influenza A (H5N1) infections in birds, which pose an ongoing risk of zoonotic transmission. Mexico was also the location for the earliest detection of the pdmH1N1 virus during the 2009 influenza A pandemic. Under a One Health framework, markets represent a unique opportunity for low-cost virus monitoring at the human-animal interface. Under the hypothesis that these represent sentinel sites for an early virus detection, we implemented a pilot surveillance program at the central market of Merida city, Yucatan, Mexico, considered a regional hotspot for multiple and recent viral outbreaks. Longitudinal sampling was carried out over 11 months at 1-to-6-week intervals from April 2022 to February 2023. We used multi-type surveillance in mosquitoes, live poultry, and wastewater. All samples were screened using RT-qPCR. Positive samples for DENV, SARS-CoV-2 and avian influenza A were further sequenced and analysed under a phylogenetic and epidemiological approach. Through our entomological surveillance, we report the earliest detection of DENV-3 III-B3.2 (genotype III American II lineage, considered a major public health concern in Latin America) in Mexico, overlapping with the resurgence of DENV-3 as the predominant serotype driving the 2023 national epidemic, which showed an increased severity. Through wastewater surveillance, we consistently detect SARS-CoV-2 RNA in wastewater samples, coinciding with the two infection waves officially recorded at a city and state level. Finally, cloacal swabs taken from two juvenile birds at the market suggest that avian influenza A viruses circulated in live poultry sold at the market. These findings show that our market-based surveillance framework is effective for an early detection and monitoring of pathogenic viruses in urban settings, and could complement official epidemiological surveillance in low- and middle-income countries to strengthen early-outbreak warning systems.

BackgroundThere is a need for synthetic peptide-based serologic assays that exploit avidity to replace whole antigens while enabling low-cost diagnostics in resource-limited settings. ObjectiveTo evaluate the diagnostic accuracy of a polymeric peptide-based ELISA leveraging avidity to enhance signal. MethodA 15-member SARS-CoV-2 peptide library corresponding to multiple epitope clusters and proteins was screened by indirect ELISA using pooled sera from RT-PCR-confirmed COVID-19 patients to identify peptides with possible diagnostic utility. The identified lead candidate, S559, possessed terminal cysteine-substitution to allow disulfide polymerization, and the resulting avidity gain was evaluated by comparing the apparent dissociation constant (KDapp) before and after depolymerization with N-acetylcysteine. The performance of an optimized ELISA using S559 was evaluated on 1,222 prospectively collected COVID-19 serum samples and 218 biobanked pre-COVID control serum samples. ResultsPolymeric S559 with a KDapp of 29.26 nM-1was demonstrated to have a 218% avidity gain relative to the completely depolymerized form. At pre-defined thresholds, the optimized S559 ELISA has a sensitivity and specificity of 83.39% (95%CI: 81.18% and 85.43%) and 96.79% (95%CI: 93.50% and 98.70%), respectively. At post hoc thresholds determined by Youden index, sensitivity and specificity reached 95.01 (95% CI: 93.63% - 96.16%) and 100.00% (95% CI: 98.32% - 100.00%), respectively. ConclusionHomomultivalent epitope presentation using polymeric S559 allows a highly specific immunoassay using human sera that may have important value in detecting antibodies, whether for diagnosing infection, confirming vaccination status or conducting surveillance.

The rapid diagnosis of Campylobacter infections is important for the management of infectious gastroenteritis. Although stool culture is considered the gold standard, its sensitivity is limited and it requires prolonged incubation times. We performed a prospective multicenter study at nine healthcare facilities in Japan to evaluate a Campylobacter rapid antigen test using stool specimens between March 2024 and August 2025. Patients with suspected infectious gastroenteritis were consecutively enrolled and tested using QuickNavi-Campylobacter and compared with the FilmArray Gastrointestinal Panel as the reference method. Discordant results were further evaluated by culturing and additional PCR assays. In total, 410 patients were included in the final analysis. The positive, negative, and total concordance rates between QuickNavi-Campylobacter and FilmArray Gastrointestinal Panel were 79%, 99%, and 93%, respectively. The positive concordance rate decreased in specimens collected [&ge;] 6 days after the onset of symptoms (50%). QuickNavi-Campylobacter demonstrated relatively good concordance with the FilmArray Gastrointestinal Panel in a real-world multicenter setting. These results suggest that this rapid antigen test may be particularly useful for the early diagnosis of suspected campylobacteriosis.

BackgroundsFollowing an increase in mpox clade Ib cases in several African countries, the World Health Organization recognized mpox as a public health emergency of international concern, highlighting the need for reliable and accessible diagnostic tools. As several mpox clades are co-circulating in endemic countries, the analytical sensitivity of diagnostic assays should be evaluated for all of them in a comparative manner. MethodsThis study evaluated the analytical sensitivity of three rapid tests (Flowflex Monkeypox virus rapid test from ACON Biotech, Monkeypox antigen rapid test from Assure Tech, and Standard Q Monkeypox Ag Test from SD Biosensor) and two point-of care molecular tests (SD Biosensors M10 and Cepheids Xpert MPOX assays) using serial dilutions of viral stocks from the four clades of Monkeypox virus (MPXV): Ia, Ib, IIa, and IIb. FindingsUpon our analytical comparative benchmarking, the three rapid tests demonstrated comparable analytical sensitivity for all four clades, with a limit of detection of approximately 1,000,000 DNA copies/mL or 1,000 FFU/mL. The two molecular tests demonstrated also comparable analytical sensitivity to an in-house PCR assay for all four clades, detecting concentrations down to 10-100 DNA copies per mL, corresponding to a non-detectable titer of infectious particles. The Xpert assay detected the Clade Ib strain only through its orthopoxvirus target and not its MPXV target, and none of the assays could distinguish between MPXV clades. InterpretationNo differences in sensitivity was observed across MPXV clades neither for Ag-RDTs nor for molecular POCTs. However, the potential of simple, affordable tests, such as Ag-RDTs, is limited by poor sensitivity while the use of highly sensitive POC molecular platforms remains limited by their cost. To date, the lack of accurate, affordable POC MPXV-specific assays with potential to differentiate clades, limits diagnostic capacities as well as our understanding of the virus and its epidemiology. FundingThis work was supported by FIND and internal funds of the Centre for Emerging Viral Diseases. Mpox diagnostic tests were provided by FIND and the World Health Organization (WHO). MOD was supported by Schmidt Science Fellows, in partnership with the Rhodes Trust.

Over-the-counter (OTC) lateral flow tests for respiratory viruses have newly emerged since the beginning of the coronavirus disease 2019 (COVID-19) pandemic and are increasingly available to consumers. While all marketed tests have met standards for Emergency Use Authorization (EUA), De Novo classification or 510(k) clearance, limited data are available to inform consumers of their relative performance. We performed a head-to-head benchtop analytical assessment of OTC tests available for purchase from retailers in the United States in the fall of 2025. Contrived specimen dilution panels were prepared from propagated viral stocks of influenza A H1N1pdm09, influenza A H3N2, influenza B (Victoria lineage), and SARS-CoV-2 JN.1 (Omicron variant) in clinical matrix and applied to the swab provided with each kit. Tests were subsequently performed according to the manufacturers instructions for use and results were interpreted by two readers blinded to the starting test material. Lower limits at which 3 of 3 replicates of test material were detected were within a 4-fold dilution for all four viruses and all eight OTC tests evaluated. We found that at low viral concentrations many OTC tests were interpreted as negative at the start of the stated interpretation window but converted to a positive result by the end of the interpretation window. We conclude that eight OTC tests that are currently readily available to consumers perform similarly in a contrived specimen analytical study, but we recommend that users of OTC lateral flow tests allow for the full incubation time before concluding that a test is truly negative.

ObjectiveUrine cultures are frequently requested at an early stage in primary care and outpatient settings, often without a comprehensive clinical assessment. This practice increases healthcare costs and laboratory workload and may lead to misleading results due to asymptomatic bacteriuria and specimen contamination. This study aimed to evaluate whether routinely reported microscopic urinary leukocyte findings can predict urine culture positivity under real-world clinical conditions. The distribution of isolated microorganisms and the frequency of mixed or contaminated growth were assessed. MethodsThis retrospective, laboratory-based diagnostic accuracy study included all urine samples sent for culture over a one-year period at a tertiary care hospital, provided concurrent microscopic urinalysis was available. No additional clinical exclusion criteria were applied to reflect the routine practice. Leukocyte findings were reported semi-quantitatively and analyzed both categorically and as approximate numerical values. The urine culture results were classified as positive, negative, or mixed/contaminated growth. The diagnostic performance was evaluated using receiver operating characteristic (ROC) curve analysis. ResultsA total of 8,478 urine samples were analyzed in this study. Urine culture positivity was detected in 2,666 (31.4%) samples, whereas 5, 812 (68.6%) showed no growth. Culture positivity increased significantly with higher leukocyte levels (p < 0.001), ranging from 13.1% in the lowest category to 83.1% in samples with abundant leukocytes. ROC analysis demonstrated an acceptable discriminative performance (AUC = 0.747). The Youden index identified an optimal threshold of approximately 5.5 leukocytes per high-power field, with a sensitivity of 60.4% and a specificity of 77.8%. Mixed or contaminated growth was the most common finding among culture-positive samples (43.5%), followed by Escherichia coli (29.5%). ConclusionMicroscopic urinary leukocyte findings were significantly associated with urine culture positivity and demonstrated acceptable predictive performance in real-world clinical practice settings. Although leukocyte microscopy alone is not diagnostic, it may support more selective urine culture ordering, reduce contamination, and contribute to rational diagnosis and antimicrobial management in primary care.

Influenza A, Influenza B, SARS-CoV-2, Respiratory Syncytial Virus and other respiratory pathogens are an ongoing public health concern. The ability to rapidly identify these viruses early in infection is essential for effective treatment and outbreak control. The AMDI Fast PCR Mini Respiratory Panel (MRP) incorporates sample preparation and real-time RT-PCR for detection of Flu A, Flu B, SARS-CoV-2 and RSV from anterior nasal swab (ANS) specimens in less than 10 minutes at the point of care. We established the analytical performance characteristics of the Fast PCR MRP and determined that the limit of detection (LoD) is 250 copies/mL for Flu A and RSV, and 500 copies/mL for Flu B and SARS-CoV-2. In a reproducibility study at 3 clinical sites, there was at least 98.2% positivity for each target for a weak positive (2x LoD) sample, at least 99.6% positivity for a moderate positive (5x LoD) sample and the negative sample returned a negative result for at least 99.3% of the tests. Fast PCR MRP had 100% analytical reactivity to all strains tested (23 Flu A, 6 Flu B, 8 SARS-CoV-2 and 6 RSV) and at least 98% predicted inclusivity from in silico analysis. There was no cross-reactivity to 40 viruses, bacteria and fungi, nor interference for 15 endogenous and exogenous substances in ANS matrix. The Fast PCR MRP delivers excellent analytical performance comparable to high complexity laboratory assays, at the point of care.

ObjectiveThe rapid availability of phenotypic antimicrobial susceptibility results is crucial for the timely detection of multidrug-resistant Gram-negative organisms and for guiding optimized treatment strategies. Recently, novel methods have been introduced that enable direct antimicrobial susceptibility testing (AST) from positive blood cultures. However, their performance has not yet been systematically compared in head-to-head evaluations. This study aimed to assess the analytical performance of two rapid AST approaches--the agar diffusion-based EUCAST rapid AST (RAST) method and the automated QuickMIC system--using a challenging collection of highly resistant Gram-negative organisms. MethodsA total of 101 Gram-negative bacteria (Escherichia coli, n = 24; Klebsiella pneumoniae, n = 22; Acinetobacter baumannii, n = 30; Pseudomonas aeruginosa, n = 25) were spiked into blood cultures and processed according to the respective AST workflows. Broth microdilution (BMD) was performed from pure cultures as the reference method. Time to result (TTR), categorical agreement (CA), and essential agreement (EA) with BMD were evaluated. Boruta analysis was applied to identify genetic determinants associated with AST errors. ResultsOverall TTR for QuickMIC was 3 h 44 min with a CA of 86.2%, an EA of 92.3 % for Enterobacteriaceae and 97.0 % for non-fermenters. Overall CA of RAST ranged from 90.7%-93.7% across reading time points. Overall, very major discrepancy rates were low (QuickMIC n=0.7%, RAST n=0.1%). Presence of NDM-5 and KPC was most frequently associated with errors for QuickMIC and EUCAST RAST, respectively. ConclusionsBoth rapid AST approaches yielded robust results in this diverse and highly resistant bacterial study population, directly from positive blood cultures, with a short turnaround time. These findings underscore the potential of rapid AST methods to facilitate timely optimization of antimicrobial therapy in bloodstream infections, even in the context of extensively drug-resistant pathogens. ImportanceAccurate antimicrobial susceptibility testing (AST) is essential for stewardship and effective therapy, especially as rising antimicrobial resistance increases the risk of empiric treatment failure. Traditional AST methods are limited by slow turnaround times, creating a need for rapid alternatives. This study evaluated the diagnostic accuracy of two rapid AST methods--EUCAST RAST and QuickMIC--using 101 genetically characterized, carbapenem-resistant Enterobacterales, Pseudomonas aeruginosa, and Acinetobacter baumannii tested directly from positive blood cultures. Broth microdilution served as the reference. Both rapid assays provided results within 3.5-6 hours and demonstrated high categorical and essential agreement with few very major discrepancies. Incorrect results were more common in isolates harboring NDM-5 and KPC carbapenemases. Overall, the findings support EUCAST RAST and QuickMIC as reliable tools for challenging resistant pathogens and highlight their potential to enable earlier detection of carbapenem-resistant phenotypes and more timely initiation of appropriate, last-resort antimicrobial therapy.

BackgroundPediatric poisonings are a significant cause of emergency admissions, often linked to accessible household toxins. This study investigates the epidemiological patterns, clinical presentations, and management strategies at a tertiary care hospital in Algiers, Algeria. MethodsA retrospective descriptive analysis was conducted on anonymized pediatric poisonings cases. Variables included age, sex, type of ingestion, intent (accidental vs. voluntary), substances involved, endoscopic findings, hospitalization duration, and management approach. Results59 pediatric poisoning cases were analyzed, which were predominantly accidental and affected children aged 0-6 years. Medication (n=24) and caustic ingestions (n=25) were the most frequent agents. Caustic cases, which primarily affected older toddlers, led to moderate mucosal injury in three patients. Carbon monoxide (n=8) affected a wide age range, including infants and adolescents. Management was mainly supportive; while most medication cases were discharged in two days, caustic cases required hospitalization for up to 14 days. These findings highlight the need for targeted public health interventions. ConclusionPediatric poisonings in this setting are largely preventable. Unsafe storage of medications and caustic agents is the primary contributor. Public health interventions focusing on household safety, caregiver education, and standardized clinical protocols are essential. Key PointsO_ST_ABSWhat is knownC_ST_ABSO_LIPediatric poisonings are common emergency admissions. C_LIO_LIToddlers are the most vulnerable age group. C_LIO_LIUnsafe household storage is a major risk factor. C_LI What is addedO_LIFirst detailed Algerian hospital-based analysis of pediatric intoxications. C_LIO_LICharacterizes the dual burden of caustic and medication poisonings in a North African pediatric setting, with caustic ingestions accounting for the most prolonged hospitalizations and requiring endoscopic evaluation. C_LIO_LIDemonstrates that carbon monoxide poisoning affects a broad age range including infants, highlighting environmental domestic exposure as a distinct and underappreciated risk in this context. C_LI

BackgroundThe influence of genetic and environmental factors, especially during early development, is critical in the pathogenesis of autism. Maternal autoantibodies that recognize specific fetal brain proteins can be strong predictors of autism risk. These antibodies cross the placenta and bind to their target antigens, which play critical roles in neurodevelopment, thereby increasing autism risk. This etiologically defined subtype is now referred to as Maternal Autoantibody-Related Autism (MARA). The newly developed MAR-AutismTM test is an indirect multi-ELISA assay designed to detect specific combinations of these maternal antibodies, which strongly predicts increased autism risk. ObjectiveTranslation of the indirect ELISA assays for the eight relevant antibodies (LDH-A, LDH-B, GDA, STIP1, CRMP1, CRMP2, NSE and YBOX) from an academic laboratory to a clinical development laboratory for optimization and determination of the analytical performance of the individual antibody assays. MethodsFeasibility assays were transferred from the academic laboratory and their performance confirmed prior to optimization of all steps from target protein production to preliminary threshold determination. Validation to rigorous standards was conducted. The ELISAs are qualitative assays using an internal continuous response and a cutoff to define positivity and negativity for each analyte. Analytical performance metrics of linearity, sensitivity, specificity, precision, and stability were determined by standard testing methodologies. ResultsThe optimized ELISAs all performed at acceptable standards for analytical performance. All of the assays except one were demonstrated to be linear upon dilution with buffer and with non-reactive plasma, however, recovery was overestimated with buffer diluent. The precision profile results demonstrated that the Lower Limit of Quantification (LOQ) was greater than the Limit of Detection (LOD) and below the preliminary thresholds determined from a general population cohort distribution. Precision studies showed coefficients of variation less than 15% with two minor exceptions. Common interfering substances, apart from whole human IgG, did not affect assay performance. The microtiter assay plates were stable for at least 6 months without significant drift. ConclusionOverall, the individual antibody assays demonstrated high sensitivity, specificity, and robustness sufficient to enable extension to clinical validation. These assays enable evaluation of specific antibody combinations that were previously reported to strongly and specifically correlate with autism risk, particularly in settings of suspected diagnosis or in families with an older sibling with a confirmed autism diagnosis.

Microbial keratitis is a sight-threatening corneal infection with varying etiological agents, primarily bacteria and fungi. Assessing and contrasting the virulence factors of microorganisms isolated from a non-contact lens-associated keratitis (NCLAK) and contact lens-associated keratitis (CLAK) is the goal of the current investigation. Samples were collected from over 60 patients and analysed using standard microbiological techniques, including culture, Gram staining, KOH mount, biochemical tests, antimicrobial susceptibility testing, and biofilm assays. The results demonstrated that CLAK isolates were predominantly bacterial, especially Pseudomonas aeruginosa, known for strong biofilm production and high multidrug resistance. In contrast, NCLAK showed a higher incidence of fungal infections, particularly Candida albicans. The results highlight the significance of early diagnosis, tailored and improved awareness regarding contact lens hygiene to prevent complications associated with keratitis.

Ziehl-Neelsen (ZN) smear microscopy remains central to tuberculosis (TB) diagnosis and treatment monitoring, yet its sensitivity is limited by incomplete recovery of Mycobacterium tuberculosis during pre-analytical processing. This study evaluated whether modifying centrifugation force and duration improves bacillary recovery and ZN smear performance. Laboratory experiments were conducted using M. tuberculosis H37Rv and pulmonary TB sputum samples. Following NALC-NaOH decontamination, samples were centrifuged at 2,000, 3,000, or 6,000 x g for 40 min, and the effect of centrifugation time at 3,000 x g was assessed by comparing 20 and 40 min using the same specimens. ZN smear grading and positivity were evaluated in triplicate and compared statistically, with significance set at P < 0.05. In H37Rv suspensions, smear grading increased with higher centrifugal force, while smear positivity plateaued at 3,000 x g. In contrast, clinical sputum samples showed progressive increases in both smear grading and positivity with increasing centrifugal force, with statistically significant improvements in positivity (p = 0.0097). Extending centrifugation time at 3,000 x g did not change smear positivity in laboratory suspensions or clinical sputum (P = 0.30). These findings indicate that standard centrifugation conditions are sufficient for laboratory strains, whereas higher centrifugal forces enhance bacillary recovery from clinical sputum, improving ZN smear sensitivity. Optimizing relative centrifugal force during pre-analytical processing may therefore reduce false-negative results and strengthen the diagnostic and treatment-monitoring performance of ZN smear microscopy in routine TB laboratories.

Background and ObjectivesCurrent potassium reference intervals for neonates fail to account for sampling method differences and prematurity-related factors, leading to unnecessary resampling and interventions. We aimed to establish sampling-specific potassium reference intervals for term and preterm neonates using comprehensive electronic health record data. MethodsWe analyzed 195 606 blood gas measurements from 10 290 neonates (2007-2024) at a tertiary neonatal intensive care unit. After ex-cluding values during severely impaired clinical conditions using integrated clinical metadata, we derived reference intervals for venous, arterial, and cap-illary samples. Multivariate analysis identified factors affecting potassium homeostasis. ResultsFrom 55 664 included values, capillary samples showed signif-icantly higher potassium levels than arterial or venous samples. Reference intervals (2.5th-97.5th percentiles) for neonates >7 days: venous [2.6, 5.5] mM, arterial [2.6, 5.9] mM, capillary [3.0, 6.3] mM. Time-matched analysis of 5403 paired samples showed capillary-specific intervals achieved 88% sensitivity and 94% specificity for detecting true hyperkalemia compared to arterial or venous controls. ConclusionCapillary blood gas potassium levels require distinct, higher reference intervals than venous or arterial samples in neonates. Implementa-tion of sampling-specific reference ranges may reduce false-positive results and unnecessary interventions in this vulnerable population. Article SummaryLarge EHR-based study defines sampling-specific neonatal potassium reference intervals; higher capillary ranges reduce false positives and maybe unnecessary interventions. Whats Known on This SubjectExisting neonatal potassium reference intervals often ignore sampling modality and prematurity, contributing to clinical uncertainty and unnecessary repeat testing in neonates. What This Study AddsProvides sampling-specific potassium reference intervals for neonates; capillary samples require distinct, higher ranges, improving discrimination of true hyperkalemia and reducing false positives. Contributors Statement PageRudolf G. Ascherl: Conceptualized and designed the study, coordinated data extraction, perfomed analysis and visualization, drafted and revised the initial manuscript. Benjamin W. Ackermann: Contributed to study design, assisted in revising the manuscript. Matthias Knupfer: Provided clinical oversight, interpreted findings, critically reviewed the manuscript. Equal contribution: Rudolf G. Ascherl and Benjamin W. Ackermann. All authors approved the final manuscript as submitted and agree to be accountable for all aspects of the work.