Journal of Clinical Virology Plus

IntroductionHerpes simplex virus type 1 (HSV-1) is highly prevalent worldwide, making accurate serological testing essential for both clinical diagnosis and epidemiological surveillance. Automated chemiluminescent immunoassays (CLIAs) offer operational advantages over enzyme-linked immunosorbent assays (ELISAs); however, their diagnostic performance relative to Western blot (WB) confirmation in high-prevalence settings remains insufficiently characterized. Hypothesis/Gap StatementThe comparative diagnostic accuracy of CLIA- and ELISA-based assays for HSV-1 IgG detection, when benchmarked against a WB reference standard in endemic populations, remains unclear. AimThis study aimed to evaluate HSV-1 IgG seroprevalence and diagnostic performance of one CLIA and two ELISA platforms using Western blot as the reference method. MethodologyFour hundred archived serum samples from adult male craft and manual workers in Qatar were tested using the Mindray CL-900i CLIA, HerpeSelect ELISA, NovaLisa ELISA, and Euroimmun Western blot. Seroprevalence, diagnostic accuracy, and interassay agreement were assessed using WB as the reference standard, with equivocal and indeterminate results excluded from analysis. ResultsHSV-1 IgG seroprevalence estimates were comparable across assays: HerpeSelect 72.5%, Mindray 70.5%, NovaLisa 66.3%, and Western blot 66.5%, with no statistically significant differences (all p > 0.05). The Mindray CLIA demonstrated the highest diagnostic performance (sensitivity 95.7%, specificity 88.9%, accuracy 93.4%) and strong agreement with Western blot ({kappa} = 0.85). HerpeSelect showed substantial agreement ({kappa} = 0.81), while NovaLisa exhibited lower specificity. ConclusionCLIA- and ELISA-based assays produced comparable HSV-1 seroprevalence estimates in this high-prevalence population; however, diagnostic accuracy varied across platforms. The CLIA platform demonstrated the strongest agreement with Western blot, supporting its use in high-throughput settings, while confirmatory testing remains important to minimize misclassification. Key PointsO_LIWhat is known: HSV-1 serological diagnosis relies mainly on ELISA assays, while automated CLIA platforms are increasingly used in high-throughput laboratories but remain insufficiently evaluated against Western blot confirmation. C_LIO_LIWhat is new: This study provides a large head-to-head comparison of CLIA and ELISA platforms for HSV-1 IgG detection using Western blot as the reference standard in a high-prevalence population. C_LIO_LIClinical implications: Automated CLIA systems demonstrated strong diagnostic accuracy and may represent reliable high-throughput alternatives for HSV-1 serological screening in clinical laboratories. C_LI Impact StatementAccurate serological diagnosis of herpes simplex virus type 1 (HSV-1) is essential for clinical management, epidemiological surveillance, and public health decision-making, particularly in populations where infection is highly prevalent. This study adds to the existing literature by providing a large, head-to-head comparison of automated chemiluminescent immunoassay (CLIA) and enzyme-linked immunosorbent assay (ELISA) platforms for HSV-1 IgG detection, benchmarked against Western blot confirmation in a real-world, high-prevalence setting. By demonstrating that different serological platforms can yield similar population-level seroprevalence estimates yet differ in diagnostic accuracy and specificity, this work highlights the risk of misclassification when confirmatory testing is not considered. The findings are of broad relevance to clinical microbiology laboratories, diagnostic services, and public health surveillance programs that rely on serological assays for HSV-1 screening. The study represents an incremental but important step in refining assay selection and interpretation, supporting more reliable laboratory diagnostics and improved understanding of HSV-1 infection burden in endemic populations. Data Availability StatementThe data that support the findings of this study are available from the corresponding author upon reasonable request.

BackgroundDengue virus infection remains a significant public health concern in India, with changing serotype dynamics influencing disease epidemiology. Understanding local serotype distribution and clinical characteristics is crucial for effective disease management and surveillance. ObjectivesTo determine the prevalence of dengue virus serotypes and analyze their clinical characteristics among NS1-positive patients at a tertiary-care hospital in Gujarat, India. MethodsA cross-sectional study was conducted on NS1-positive dengue patients admitted to AIIMS Rajkot from September 2023 to November 2024. Real-time reverse transcription polymerase chain reaction (RT-PCR) was performed for serotype identification. Clinical and demographic data were collected and analyzed. ResultsNS1-positive patients (70) were confirmed by RT-PCR. DENV-2 was the predominant serotype (53 cases, 75.7%), followed by DENV-1 and DENV-3 (7 cases each, 10.0%), and DENV-4 (2 cases, 2.9%). One co-infection case (DENV-2 + DENV-3) (1.4%) was identified. The mean age was 27.7 {+/-} 14.4 years, with male predominance (58.6%). Young adults (19-35 years) were most affected (45.7%), followed by pediatric patients [≤]18 years (32.9%). Severe dengue occurred in only one case (1.4%), while hospitalization was required in 25 cases (35.7%). All patients presented with fever, chills, headache (50%), rashes (56%), and malaise (56%), being the most common associated symptoms. ConclusionsDENV-2 showed clear predominance in the Rajkot region during the study period, with low rates of severe disease. The significant pediatric and young adult involvement highlights the need for targeted prevention strategies. These findings contribute to the understanding of regional dengue epidemiology and support evidence-based surveillance and control measures.

Background: Although the relaxation of COVID-19 containment measures in China has altered the transmission dynamics of respiratory pathogens, regional data on post-pandemic epidemiological characteristics remain limited.Objective: This study aimed to investigate the pathogen spectrum and epidemiological characteristics of acute respiratory infections (ARIs) in Quzhou City from 2023 to 2024, providing a scientific basis for local prevention and control strategies.Methods: A total of 2,800 respiratory specimens were collected from November 2023 to July 2024, comprising 1,960 influenza-like illness (ILI) cases from outpatient/emergency departments and 840 severe acute respiratory infection (SARI) cases from inpatient departments. All samples were tested for 13 common respiratory pathogens using multiplex fluorescence quantitative PCR. Etiological and epidemiological analyses were performed based on detection results and case information. Results: The overall ARI positivity rate was 59.28% (1,660/2,800), with a male-to-female ratio of 1.07:1 (1,447/1,353). The three most prevalent pathogens were influenza virus (Flu, 23.21%, 650/2,800), Streptococcus pneumoniae (SP, 13.14%, 368/2,800), and adenovirus (ADV, 8.39%, 235/2,800). Single pathogen infections accounted for 73.55% (1,221/1,660) of positive cases, while co-infections with two or more pathogens accounted for 26.45% (439/1,660), yielding an overall co-infection rate of 15.68% (439/2,800). No significant gender difference was observed in detection rates. However, significant differences were found across case types, temporal periods, age groups, and geographic regions (P < 0.01). Children aged [&le;]5 years exhibited the highest positivity rate (78.00%, 378/525), while adults aged [&ge;]65 years showed the lowest (34.53%, 144/417). Among surveillance regions, Kaihua County had the highest positivity rate (72.47%), and Changshan County the lowest (40.55%). Conclusions: Multiple respiratory pathogens and co-infections are prevalent in Quzhou City, with distinct age-specific and seasonal patterns. These findings underscore the need for continuous multi-pathogen surveillance and integrated prevention strategies for influenza and other respiratory infectious diseases in the post-pandemic era.

Background Reactivation of human herpesviruses (HHVs), particularly EBV, is associated with more severe acute SARS-CoV-2 infections and the development of Long COVID (LC). Observations of higher anti-EBV antibody levels in individuals with LC support the idea that chronic reactivation of HHVs could contribute to LC pathology. HHV shedding in saliva has also been previously associated with saliva hormone levels. This study aims to examine the relationship between salivary shedding of HHV DNA and LC symptoms, as well as cortisol, testosterone, and estradiol levels. Methods We enrolled 45 participants with LC, and 45 age-sex-matched controls. Surveys and validated health questionnaires were used to collect demographics, medical history, and symptom profiles. Saliva was self-collected at waking, 15, 30, and 45 minutes, and 8 and 16 hours after waking, across two consecutive days. Salivary cortisol, testosterone and estradiol were measured, and extracted nucleic acid was tested for EBV, HSV 1/2, HCMV and HHV-6 A/B using multiplex qPCR, plus SARS-CoV-2 and RNaseP using RT-qPCR. Findings Detection of salivary EBV and HHV-6 DNA was highest early in the morning. There were no significant differences in salivary cortisol, testosterone, or estradiol, or in EBV or HHV-6 shedding between the LC and control groups. However, salivary HHV-6 DNA levels were positively associated with a greater aggregated LC propensity score, as well as anxiety and depression scores. Interpretation The observed correlation between salivary HHV-6 shedding and symptom severity suggests HHV-6 may contribute to post-acute disease, though mechanisms remain unclear. While our study did not identify a relationship between salivary EBV shedding and LC, EBV may still play a role at earlier time points in the disease course, or in compartments not sampled here. These findings highlight the potential importance of HHV-6 in LC pathophysiology and underscore the need for longitudinal, multi-compartment studies of herpesvirus reactivation in LC.

BackgroundKeratoconus (KC) is a multifactorial disorder with unclear etiology, characterized by localized thinning and a cone-like protrusion of the cornea. The complex etiology of KC exacerbates the lack of an animal model. Previous studies by Tachibana et al. (2002) described an inbred mouse strain (SKC) with a spontaneous, androgen-dependent, cone-like corneal morphology. This study aimed to investigate the corneal phenotypes of SKC mice through an in-depth ophthalmic examination. MethodsMice (n=53) were examined via slit lamp biomicroscopy with fluorescein staining. Spectral-domain optical coherence tomography (SD-OCT) enabled central corneal thickness (CCT) measurement in selected mice (n=26 eyes), and OCT-based pachymetry mapping (n=16 eyes). In vivo corneal confocal microscopy was conducted on eyes to assess cellular morphology (n= 9 eyes). Eyes were collected for histology analysis (n=22). ResultsLesions and epithelial breaks were present in [~]95% of eyes (n=101). Neovascularization, perforation, scarring, and hydrops were seen primarily in males. An opaque, unilateral cone-like morphology was exclusive to males (n=11). Male and female corneas showed no significant difference in CCT, though pachymetry mapping revealed regional thinning patterns in both sexes. Loosened epithelial tight junctions, stromal fibrosis, vascularization, and inflammation of variable severity were identified in both sexes. ConclusionThis study identified previously unreported corneal phenotypes in SKC mice through ophthalmic examination. Unlike previous studies, gross and histological abnormalities were observed in female SKC mice. Our findings suggest a lower penetrance of the cone-like phenotype ([~]20%) than previously reported ([~]33%) and support that the conical phenotype in male mice may be secondary to keratitis.

Objective To evaluate physician knowledge, attitudes, and practices regarding viral exanthem diagnosis and mandatory reporting requirements among practicing physicians in major metropolitan regions of Florida. Study Design An IRB-exempt cross-sectional survey was distributed via REDCap to licensed physicians and residents in family medicine, internal medicine, pediatrics, and infectious disease across Florida. The 19-question survey assessed demographic characteristics, knowledge of viral exanthem diagnosis (measles, rubella, roseola), reporting requirements, physician attitudes, and clinical practices. Knowledge scores were compared by specialty using ANOVA with Tukey post-hoc analysis. Multivariate analysis and linear regression assessed associations between physician confidence and knowledge scores. Results A total of 162 physicians responded, with 146 complete responses included in analysis. Participants included pediatrics (n=74), family medicine (n=48), and internal medicine (n=24). The overall mean knowledge score was 78.5% (SD 20.5). Pediatricians demonstrated the highest scores (82.7%) compared with internal medicine (76.4%) and family medicine (73.3%), with pediatricians scoring significantly higher than family physicians (p=0.04). Differences in vignette-based diagnostic knowledge and mandatory reporting knowledge were not statistically significant across specialties. Roseola was the most commonly diagnosed viral exanthem (66%), followed by measles (30%) and rubella (17%). Most physicians (91.4%) expressed interest in additional training. Conclusions Although overall physician knowledge of viral exanthem diagnosis and reporting was high, clinically meaningful gaps remain, particularly in differentiating similar rash presentations. Pediatricians demonstrated higher knowledge scores than family physicians. Enhanced physician education may improve diagnostic accuracy and public health reporting as vaccination rates decline and outbreaks of vaccine-preventable viral exanthems increase.

Introduction: Hantavirus disease is an emerging and potentially severe zoonosis of global distribution. In Uruguay, it is transmitted by rodents inhabiting peridomestic, suburban, and rural areas. Global incidence is estimated at 150,000 to 200,000 cases per year, with up to 300 annual cases in the Americas. Since 1997, Uruguay's Ministry of Public Health (MPH) has monitored Hantavirus cardiopulmonary syndrome (HCPS), the most common clinical presentation in the region. By 2019, a total of 271 cases had been identified in the country, with an estimated mortality rate of nearly 50%. Objectives: To describe the clinical, epidemiological, and occupational characteristics of patients with Hantavirus disease in Uruguay during the pre-pandemic (2018-2019) and pandemic (2020-2021) periods. Methods: A descriptive, cross-sectional, observational study was conducted, including all serologically confirmed cases of Hantavirus infection reported to the MPH between 2018 and 2021. Clinical and demographic data were extracted from the mandatory reporting form for zoonotic diseases. Incidence and case fatality rates were calculated, and factors associated with fatal outcomes were analyzed. Results: A total of 58 confirmed cases were identified between 2018 and 2021. Most patients were male (62%), with a mean age of 36.5 years (SD 16). A decline in incidence was observed during 2020-2021, with no significant change in case fatality. Direct rodent exposure was the most frequently associated risk factor. Montevideo and Canelones were the most affected departments. Renal and pulmonary involvement were significantly associated with mortality. Conclusion: Hantavirus remains a relevant public health concern in Uruguay. Although a decrease in incidence was observed during the COVID-19 pandemic years, case fatality rates remained high. The findings underscore the need for sustained surveillance and early recognition, particularly in urbanizing regions.

Successive dengue virus (DENV) outbreaks can progressively reshape population immunity influencing disease expression and diagnostic performance. Objectives The aim was to evaluate the impact of secondary infections across sequential outbreaks on clinical severity, serotype dynamics and diagnostic concordance. Methods This retrospective study analyzed 976 febrile-stage samples from three sequential outbreaks in Misiones, Argentina. For serotyping and clinical analyses, 869 viremic samples confirmed by at least one direct method were included (2016: n=512; 2019: n=148; 2024: n=209). Additionally, 318 samples, including 107 non-viremic cases, were used to compare NS1 rapid diagnostic tests (NS1 Ag) and RT-PCR. Viral serotyping and clinical and laboratory markers of disease severity were evaluated. Results Secondary infections increased from 31.05% (2016) to 43.24% (2019) and 53.87% (2024) (p<0.0010). Serotype distribution shifted from DENV-1 predominance in 2016 (95.12%), DENV-1/DENV-4 co-circulation in 2019 (60.71%/39.29%), and DENV-2 predominance in 2024 (97.60%). Secondary infections were associated with more severe disease manifestations, particularly in 2024, with higher hematocrit (p=0.0120) and hemoglobin (p=0.0080), lower white blood cells (p=0.020) and platelet counts (p=0.0030), and elevated AST (p=0.0007) and ALT (p=0.0130). Concordance between NS1 Ag and RT-PCR was lower in secondary infections (k=0.457 vs k=0.759, p=0.0013). Conclusions The rising frequency of secondary infections may affect both clinical severity and diagnostic performance during outbreaks. The clinical impact was more evident in 2024, likely associated with the introduction of a new serotype. These findings highlight the need for optimized surveillance and diagnostic strategies to improve case detection and patient management during epidemics.

Cetacean pathology is a cornerstone for population and marine ecosystem health monitoring, allowing clear differentiation among natural and anthropogenic threats. Previous studies in the Canary Islands reported natural causes of death in 59.4% (1999-2005) and 81% (2006-2012) of stranded cetaceans, versus anthropogenic causes in 33.3% and 19%, respectively. This study aimed to determine the causes of death (CD), pathologic findings, and epidemiological patterns of 316 cetaceans stranded in the Canary Islands between 2013 and 2018. The CDs were classified in pathologic entities (PEs) emphasizing natural versus anthropic origins. Of 316 animals, 224 (70.9%) from 18 species were suitable for pathological investigations. Among natural PEE, natural pathology associated with good nutritional status (NP-GNS) and natural pathology associated with significant loss of nutritional status (NP-LNS) represented 43/224 (19.2%) and 36/224 (16%) cases, respectively. Natural pathology with undetermined nutritional status (NP-UNS) occurred in 19/224 (8.5%) animals. Intra- and interspecific traumatic interactions (ITI) represented 30/224 (13.4%) cases, followed by neonatal/perinatal pathology (NPN) 19/224 (8.5%) and live-stranding stress and/or capture myopathy (LS-CM) 18/224 (8%). Infectious and parasitic diseases predominated in natural PEs. Anthropogenic PEs included interaction with fishing activities (IFA) in 17/224 (7.6%) cases, vessel collisions (VC) in 9/22 (4%) cases, and foreign body-associated pathology (FBAP) in 3/224 (1.3%) animals. Overall, anthropogenic causes accounted for 12.9% of deaths, natural causes for 73.6%, and the CD could not be established in 30/194 (13.4%) cases. This study reaffirms the trends concerning recognized PEs (NP-GNS, NP-LNS, NP-UNS, ITI, NPN, LS-CM, IFA, VC, and FBAP), expands the body of knowledge on cetacean pathology in the Canary Islands, and reports novel findings including mixed infections, clostridiosis in uncommon species, uremic syndrome secondary to urethral nematodiasis, gas embolism in unusual species, epibiont stomatitis, congenital musculo-skeletal malformations, or neoplastic processes. These findings advance understanding of cetacean mortality patterns and support conservation and management strategies.

ObjectivesBenign Prostatic Hyperplasia (BPH) is the non-cancerous enlargement of the prostate accompanied by lower urinary tract symptoms, affecting 50% of men by the age of 501,2. Advanced highly symptomatic BPH exhibits large epithelial glandular nodules with microglandular/atypical adenomatous hyperplasia, but how these features form is unknown3. Our lab has reported that the common parasite Toxoplasma gondii can infect the prostate and induce glandular nodule formation in mice3. The objective of this study is to determine if T. gondii exposure in humans correlates to BPH and nodule formation and if it induces urinary dysfunction concurrent in the mouse model. MethodsWe assessed Toxoplasma exposure by serum ELISA in patients with BPH and non-BPH donor controls, and compared seropositivity rates between the groups. We further assessed the histopathology of these patients for the presence of inflammation and epithelial glandular nodule formation and compared Toxoplasma positive and negative samples. We determined voiding function in Toxoplasma-infected mice between 14 and 60 days of infection with void spot with Void Whizzard software. ResultsMen diagnosed with BPH are more likely to be seropositive for Toxoplasma than age-matched undiagnosed donor controls. In addition, BPH patients that are seropositive for Toxoplasma are more likely to exhibit glandular nodule formation with microglandular / adenomous hyperplasia than seronegative BPH patients. In animal studies, Toxoplasma infection results in abnormal void patterns concurrent with microglandular hyperplasia and nodule formation. ConclusionsThese results suggest that Toxoplasma may be contributing to BPH pathology and lower urinary tract dysfunction in both humans and mice, opening new insights into the development of this important disease. The results also serve to further characterize this model of prostatic hyperplasia and define it as a potential urinary dysfunction model.

BackgroundRespiratory tract infections range from asymptomatic colonisation to an invasive disease. Recent studies suggest that nasopharyngeal microbiota may influence this variability. Emerging evidence points to Dolosigranulum pigrum, a nasopharyngeal commensal, as a potentially protective bacterium. This study aimed to identify variables associated with the presence of D. pigrum in the nasopharynx of children with varying respiratory health statuses. MethodsNasopharyngeal aspirates were collected from children <18 years who were asymptomatic (n=65), had banal viral infection (n=48), or Invasive Pneumococcal Disease (IPD) (n=27). The presence of D. pigrum was defined as >0.1% of total sequences obtained by 16S rRNA gene sequencing. Variables included sex, breastfeeding, delivery mode, S. pneumoniae carriage, respiratory viruses and clinical features. ResultsAmong 140 children (73 males, 67 females), D. pigrum was detected in 79 (56.4%): 44/65 in the healthy group; 26/48 of viral and 9/27 IPD cases. Multivariate analysis revealed significant associations with health status and sex. Healthy children were more likely to carry D. pigrum than IPD cases (44/79 vs. 26/79; p= 0.028). Males were more frequently D. pigrum carriers than females (48/79 vs. 31/79; p= 0.033). ConclusionD. Pigrum was associated with respiratory health, being more prevalent in healthy children, and showed potential sex-related differences.

Background Oropouche virus (OROV) is an emerging vector-borne virus with rapidly expanding geographic range, increasing case counts, and growing evidence of severe outcomes including neuroinvasive disease and vertical transmission. Because OROV infection presents with nonspecific febrile illness that overlaps clinically with other viruses including dengue, zika, and chikungunya, accurate molecular diagnostics are essential for patient care and surveillance. Yet existing assays rely on single genomic targets and are vulnerable to detection failure as the virus evolves and reassorts. Methodology/Principal Findings To support diagnostic capacity, we developed and clinically validated a multiplexed qPCR assay targeting three regions of the OROV S segment, incorporating redundancy to preserve sensitivity across viral diversity while enabling robust clinical interpretation. The multiplex also includes an assay targeting RNaseP as an internal sample control to ensure adequate sample processing. We evaluated assay performance using both historical and contemporary OROV strains and validated the assay on contrived serum, plasma, and cerebrospinal fluid samples, assessing linearity, limit of detection (LOD), accuracy, specificity, precision, and sample stability. The assay met or exceeded all predefined acceptance criteria for clinical testing and achieved an LOD as low as 6 copies per reaction for contemporary outbreak strains. We further implemented a logic-based interpretation matrix that reduced false-positive risk while maintaining sensitivity near the analytical LOD. Conclusions/Significance Our assay sensitively and specifically detects OROV RNA in serum, plasma, and cerebrospinal fluid while incorporating safeguards against viral evolution and reassortment. The assay has been approved for use by CLIA at Nexus Medical Labs in 49 U.S. states, expanding access to timely OROV diagnostics in the United States and providing a durable framework for molecular detection of reassorting, rapidly evolving viruses as OROV continues to spread into new regions.

Background: The global burden of dengue infection has rising, yet limited data exists on its impact in the Caribbean. We describe the incidence and associates of acute kidney injury in adults and children with dengue at a teaching hospital in Jamaica. Methods: A single-centre retrospective cohort study of admissions with laboratory confirmed dengue infection at University Hospital of the West Indies, Mona Jamaica between January 2023 to November 2024. AKI was defined using Kidney Disease Improving Global Outcomes definitions. Patients were included if aged >1year and had at least 2 creatinine values. Clinical, demographic and laboratory data were abstracted by chart review. Summary statistics were used to describe continuous and categorical data, and logistic regression to determine AKI associations. Stratified analysis was performed by age-group (adults-aged [&ge;] 16, and paediatric-aged <16 years). Results: Analyses included 167 persons, 62% (103) were male, mean age was 26.1{+/-}19.5 years. AKI occurred in 25.8%, 65.1% were KDIGO stage 1. AKI incidence was 30.2% and 18.0% among adults and children respectively. There were 3 in-hospital deaths. People with AKI were older 32{+/-}21.4 vs 24 {+/-}18.4 (p=0.021), and had longer duration of stay [6 vs 4 days (p <0.001)]. Male sex [OR 2.09 (95% CI:0.96-4.59), p=0.064], age per year [OR 1.02 (95% CI:1.01-1.04), p=0.015] symptom duration [OR1.11 (CI 0.99-1.24), p = 0.058], admission bilirubin [OR 1.02 (CI: 1.00-1.04), p = 0.022], NLR [OR 1.09 (CI 1.00-1.18), p = 0.037] were associated with AKI. In adults admission potassium was inversely associated with AKI [OR 0.46 (95% CI 0.21-1.01), p 0.056], while in children admission potassium [OR 3.00 (95% CI 0.88-10.6), p 0.088] was associated with AKI. Conclusion: AKI in dengue hospitalizations is higher than most reports at 25.8%. Targeted public health policy on vector control and early symptom recognition may be needed to improve outcomes.

Mpox virus (MPXV) gained increased attention following the declaration of two Public Health Emergencies of International Concern (PHEICs) in 2022 and 2024. The rapid spread of MPXV and the increase in human-to-human transmission highlighted the need for improved diagnostic tools for characterizing infection patterns and transmission dynamics. While PCR is effective for detecting active infections, serological approaches can help identify previous or asymptomatic infections and support retrospective surveillance. However, many serological assays developed during recent outbreaks have not been evaluated in endemic settings such as the Democratic Republic of the Congo (DRC). This study aims to define antigen-specific serological cutoff values to differentiate MPXV-seroreactive individuals from those with other orthopoxvirus (OPXV) exposure or different vaccination histories, specifically for use in the DRC. Here, we analyzed 134 individuals, divided into six distinct cohorts with different exposures. Serum samples were tested using Mesoscale Discovery (MSD) to screen for five MPXV and vaccinia virus (VACV) orthologous antigens: A29L/A27L, A35R/A33R, B6R/B5R, E8L/D8L, and M1R/L1R. Receiver operating characteristic (ROC) analysis identified the best-performing antigens and established seroreactivity cutoff values. A binary composite rule was also evaluated to improve the classification of these results. We identified three MPXV antigens, E8L (cut-off=12.33 AU/mL), A35R (cut-off=5.22 AU/mL), and B6R (cut-off=9.77 AU/mL), that showed the strongest discriminatory performance in the dataset. Collectively, these three antigens form a significant panel that demonstrated clear separation between our mpox survivor cohort and other OPXV-exposed individuals. ImportanceEstablishing antigen-specific serological cutoff values for this assay using unique samples from endemic regions such as the DRC may improve future epidemiological and disease transmission surveillance efforts and contribute to broader efforts to ensure regionally appropriate cutoffs for serological assays.

BackgroundTo date, accessible diagnostic tools to identify whether a patients pneumonia is a bacterial, or viral infection, are not accurate or timely enough to prevent preemptive antibiotic administration. Relying on single biomarkers or clinical presentations has been insufficient. We aimed to incorporate a wide range of novel biomarkers and clinical presentations in a multivariable model and validate its capacity to differentiate cases of bacterial and viral pneumonia. MethodsData from 457 children aged 2-59 months, admitted to Kilifi County Referral Hospital, Kenya, with bacterial (n = 229) and viral (n = 228) infections, were used to develop and validate a predictive multivariable Poisson regression model to differentiate pneumonia etiology. The Receiver Operating Characteristic curve was used to assess biomarker performance and validate the model internally. ResultsSixty-three percent (63%) of the children presented with severe pneumonia. 72% with viral pneumonia had severe pneumonia, compared to 54% with bacterial pneumonia who had severe pneumonia. In crude analyses, chest-wall indrawing, cough, convulsions, crackles, angiotensinogen, and Serpin Family A Member 1 were significantly associated with pneumonia etiology, controlling for age. However, only chest-wall indrawing remained significant in multivariable analyses after controlling for age. The model demonstrated fair, but inadequate, discrimination, with an Area Under the Curve of 0.61. ConclusionAmong the children admitted to hospital with WHO defined pneumonia, a wide range of biomarkers and clinical presentations still failed to distinguish bacterial from viral pneumonia.

Using a commercially available H5 serology assay, we identified a 7.4% (n=5/68) anti-H5 seroreactivity rate among hunters in Northern Canada. All participants reported close contact with wild birds.

Nasopharyngeal (NP) swabs remain the dominant gold standard for respiratory infection diagnostics. While there has been increased use of alternative sample types since the COVID-19 pandemic, guidance on their use for detecting respiratory viruses is not yet definitive, especially for children. In this systematic review and meta-analysis, we aimed to compare the diagnostic accuracy and tolerability of multiple respiratory specimen types for detecting respiratory viruses in pediatric populations. Searches were conducted on July 17, 2025 in MEDLINE, Embase, Web of Science, and Scopus, with screening and data extraction performed in Covidence. English-language primary research articles published since 2000 comparing respiratory virus detection rates in children, using nucleic acid amplification tests between paired respiratory specimens, were included. Risk of bias was assessed using Quality Assessment of Diagnostic Accuracy Studies criteria. We calculated pooled sensitivities and specificities of index specimens: nasopharyngeal aspirates (NPA), mid-turbinate swabs (MT), anterior nasal swabs (ANS), oropharyngeal swabs (OP), and bronchoalveolar lavage fluid (BAL), as compared to the reference, NP swabs, using random-effects modeling, firstly without discrimination by virus. Index specimens were then grouped by sample collection site as nasal, oral, and lower respiratory tract (LRT) specimens for virus-specific analyses. Overall performance and statistical validity were evaluated by hierarchical summary receiver operating characteristic (HSROC) analysis. Data regarding sampling tolerability was also assessed. We screened 2,448 studies and identified 36 publications (total N participants = 10,687) that reported diagnostic test accuracy using paired index-reference data in children. Of these, 18 (total N participants = 4,310) used NP specimens as the reference and were included in the diagnostic test accuracy analysis. Virus-agnostic pooled sensitivity estimates indicated that MT (0.92%) performed most similarly to NP, though sensitivities of ANS (0.79%) and OP (0.70%) were also moderately high for detection of any respiratory virus. BAL sensitivity was the lowest (0.37%). All sample types demonstrated high specificity (0.98%-0.99%). Group estimates and HSROC statistics found that nasal specimens, when grouped, had the highest sensitivity and accuracy for all examined viruses, including for influenza (92%) and RSV (90%). By comparison, oral and LRT specimens performed less well, with more variability, though both showed moderately high sensitivities for RSV (78%, 76%, respectively) and influenza (82%, 80%, respectively), and LRT samples showed high sensitivity for HMPV (82%). Analysis of sample tolerability found that NP swabs consistently ranked as the least comfortable and least preferred, while nasal swabs and saliva both performed well. Datasets for LRT and oral specimens were sparser than for nasal, and this contributed to greater variability, underscoring the need for further diagnostic accuracy studies on alternatives to NP sampling. These data support the viability of nasal and oral alternatives to NP swabs and affirm their application in pediatric care, particularly in outpatient settings. Such alternatives could greatly improve sampling tolerability and increase global access, including in resource-limited settings, to accurate diagnostic methods for respiratory infections.

Background: Infections of the central nervous system (CNS) in children remain a major cause of mortality and long-term disability globally, particularly in low- and middle-income countries (LMICs), where a high proportion of cases lack an identified pathogen. Sporadically, human parvovirus 4 (PARV4) has been detected in a small number of cerebrospinal fluid (CSF) from children with CNS infections, but its pathogenic role is unclear. We investigated the prevalence, clinical impact, and genomic characteristics of PARV4 in children with suspected meningitis. Methods: We retrospectively analyzed CSF samples collected from children with WHO-defined suspected meningitis at the largest pediatric hospital in Bangladesh between 2015-2022. All samples underwent routine diagnostics, including bacterial culture and serological testing. Additional testing for PARV4 and parvovirus B19 was performed using qPCR of samples with >9 white blood cell (WBC)/ul followed by metagenomic sequencing of a subset. Clinical and laboratory data were extracted from patient records. Associations between PARV4 detection and mortality were assessed using logistic regression, adjusting for age, WBC count, and co-infections. Genomic and phylogenetic analyses were conducted on PARV4-positive samples. Findings: Among 2,793 CSF samples with >9 WBC/ul, 526 (18.8%) were PARV4-positive. The median age of PARV4-positive cases was lower than that of PARV4-negative cases (4 vs 7 months, p<0.001). Co-infections were more common among PARV4-positive cases (49.6%) than PARV4-negative cases (16.4%). PARV4 positivity was independently associated with increased in-hospital mortality (adjusted odds ratio 2.09, 95%CI:1.46-2.96; p<0.001). Phylogenetic analysis indicated most strains belonged to genotype 2, with two sequences forming a distinct clade. Interpretation: PARV4 is frequently detected in the CSF of children with suspected meningitis and is associated with increased in-hospital mortality. Its high prevalence, detection early in life, and frequent co-infection with other pathogens highlight the need to investigate PARV4 as an emerging CNS pathogen in LMICs. Funding: Gates Foundation

Syphilis is a sexually transmitted and bloodborne infection caused by Treponema pallidum spp. pallidum. Given the paucity of data on syphilis in Kenyan sex workers and gay, bisexual, and other men who have sex with men (GBMSM), we conducted a retrospective study of syphilis seropositivity in female sex workers (FSW) and GBMSM in Nairobi, Kenya. Seropositivity testing of cryopreserved plasma samples showed that 11.1% (72/647) were positive. Syphilis seropositivity was associated with HIV status, and FSWs were disproportionately represented in the seropositive group (66/72, 92%). Here, we report a higher seropositive rate than in previous studies in Kenya, and ongoing community and surveillance supports are important for addressing the ongoing public health impacts of syphilis.

BackgroundLeptospirosis is a re-emerging zoonosis in French Guiana, with broad and sometimes misleading clinical spectrum. Neurological involvement, referred to as neuroleptospirosis, lacks a consensus definition. Therefore, it is likely underrecognized, raising concerns about missed diagnosis and potentially poor outcomes. This study aimed to characterize the epidemiology, clinical features and outcomes of neuroleptospirosis. Methodology/ Principal findingsA multicenter retrospective study was conducted across all public healthcare facilities of French Guiana (2015-2021). Neuroleptospirosis was defined by the combination of (i) neurological symptoms and (ii) objective evidence of neurological involvement based on cerebrospinal fluid (CSF) analysis and/or neuroimaging. Cases were compared with leptospirosis without neurological involvement. Among 146 consecutive hospital-managed cases of leptospirosis, 18 (12%) met criteria for neuroleptospirosis (incidence 0.88 cases/100,000 inhabitants/year; 95% CI 0.52-1.39). Among them, 78% (14/18) presented a meningeal syndrome, 22% (4/18) an encephalitic syndrome, and 17% (3/18) showed paresthesia. Lumbar puncture was performed in 28/146 (19%) patients, with pleocytosis observed in 18/28 (64%) patients; median CSF leukocyte count was 42 cells/mm3 (range 13-240/mm3), with lymphocytic predominance in 8/13 (62%), a slight protein level (median 0.51 g/l, range 0.32-0.88) and no hypoglycorachia. Brain MRI revealed abnormalities in 2/8 (25%) cases of neuroleptospirosis: one cerebral infarct and one pachymeningitis. Compared with patients without neurological involvement, factors associated with a diagnosis of neuroleptospirosis were an age <30 years (p=0.004, 95% CI 1.72-18), no leukocytosis (p=0.042, 95% CI 1.04-10.39) nor thrombocytopenia (p=0.012, 95% CI 1.39-14.32) during hospitalization. Neuroleptospirosis cases had milder disease: they less often progressed to sepsis (OR 0.28; 95% CI 0.10-0.79); none required intensive care admission nor died. Discussion/ConclusionsThis study provides new insights into the clinical spectrum and outcomes of neuroleptospirosis in French Guiana. Prospective studies incorporating a consensus definition, systematic CSF analysis and microbiological testing are warranted to further characterize the pathophysiology and optimize diagnostic strategies for neuroleptospirosis. Author summaryLeptospirosis is a re-emerging infection in French Guiana, with a broad range of clinical presentations. Yet, its neurological manifestations--referred to as neuroleptospirosis -- remain likely underrecognized, lacking a consensus definition. We conducted a multicenter retrospective study to describe the epidemiological, clinical features and outcomes of neuroleptospirosis. Then, we compared cases with non-neurological leptospirosis. We used a standardized definition of neuroleptospirosis, based on neurological symptoms associated with cerebrospinal fluid analysis and/or neuroimaging. Neuroleptospirosis accounted for one tenth of leptospirosis cases. Clinical presentation was largely dominated by meningitis (three quarters of cases). Laboratory parameters showed lower rates of leukocytosis and renal involvement. CSF profiles revealed moderate lymphocytic pleocytosis, closely mimicking viral infections. Strikingly, compared with non-neurological forms, neuroleptospirosis cases exhibited fewer severity markers and uniformly favorable outcomes, with lower sepsis, no deaths nor need for intensive care admission. Factors associated with a diagnosis of neuroleptospirosis were an age lower than 30, the absence of hyperleukocytosis and thrombocytopenia during hospitalization. Neuroleptospirosis accounted for a non-negligible proportion of leptospirosis cases in French Guiana, exhibiting CSF features mimicking viral meningitis, and carrying excellent outcomes. Increasing awareness and improving recognition of this presentation is essential to reduce underdiagnosis and refine patient management in endemic settings.