Search Results (356)

Adeno-associated virus (AAV)-based massively parallel reporter assays (MPRA) have become an important platform for large-scale functional characterization of regulatory DNA elements. However, plasmids carrying AAV inverted terminal repeats (ITRs) are intrinsically unstable during propagation in Escherichia coli, potentially compromising library integrity before viral packaging. Although ITR-associated recombination has been recognized, the influence of cloning-site position relative to the ITR on plasmid stability has not been systematically investigated. Here, we examined the relationship between cloning-junction proximity to AAV2 ITRs and plasmid recombination using an AAV-MPRA reporter plasmid. We compared four restriction-ligation cloning strategies utilizing restriction sites at defined distances (4–543 bp) from the nearest ITR while preserving ITR integrity, and one strategy in which the ITR itself was disrupted. We observe that plasmid recombination exhibited a pronounced distance dependence. Constructs with ligation junctions located 4, 41, and 182 bp from an intact ITR showed recombination frequencies of 82.5%, 60%, and 20%, respectively, whereas a 0% recombination frequency was detected when the nearest ITR was positioned 543 bp from the cloning junction. In contrast, cleavage within ITR reduced recombination to 15%, demonstrating that preservation of the intact ITR secondary structure is required for efficient recombination. Whole-plasmid sequencing confirmed recurrent large-scale deletions in which the expression cassette and the downstream R-ITR were removed while the L-ITR and plasmid backbone were retained, consistent with preferential processing of the intact L-ITR region. These findings identify cloning-site proximity to an intact AAV ITR as a major determinant of plasmid stability during bacterial propagation and demonstrate that substantial loss of correctly assembled constructs can occur before AAV production. The results have direct implications for the design of AAV-based MPRA libraries and support positioning cloning sites as far as practical from the nearest ITR, together with routine validation of plasmid integrity prior to viral packaging. Full article

Background/Objectives: Healthcare students face an occupational risk of hepatitis B virus (HBV) exposure during clinical training, often many years after receiving their primary vaccination series in infancy or childhood. Progressive waning of anti-hepatitis B surface (anti-HBs) antibody titres, heterogeneous vaccination histories in international student cohorts, high rates of underreported needlestick injuries, and variable occupational health protocols across institutions raise important questions about the adequacy of pre-clinical immunization surveillance in this population. This review aimed to synthesize current evidence on hepatitis B immunization among healthcare students, focusing on occupational exposure risk, seroprotection and immunological memory, clinical management of low or absent anti-HB titres including post-exposure prophylaxis, and the specific challenges posed by multicultural academic settings. Methods: A narrative review was conducted through a systematic literature search of PubMed/MEDLINE and Scopus, covering publications from January 2000 to December 2025, supplemented by key seminal references. After the removal of duplicates and stepwise screening, 49 references were selected for final inclusion. Results: Needlestick and sharps injuries occur at measurable rates during clinical training, with medical students accounting for approximately 30% of occupational exposure events in some series, and with underreporting rates estimated at 19–80% across studies. Pooled seroprotection prevalence at clinical training entry is approximately 73.8% (95% CI 69.1–78.0%), with substantially lower rates among students vaccinated in infancy—as low as 28% at 16–20 years post-vaccination in longitudinal data—compared with adolescence immunization. The majority of students with non-protective titres retain immunological memory, with 90.9% (95% CI 87.7–93.3%) demonstrating an anamnestic response after a single booster dose. Post-exposure prophylaxis pathways depend critically on pre-existing immunological status, reinforcing the operational value of pre-clinical serological screening. International student cohorts present additional complexity due to heterogeneous vaccination schedules, documentation gaps, and variable natural immunity profiles. Conclusions: Systematic pre-clinical serological screening (anti-HBs, HBsAg, and anti-HBc), combined with evidence-based stepwise immunization management and structured educational interventions, is essential to protect healthcare students from occupational HBV exposure. The primary operational goal of screening is the identification of the approximately 5% of true non-responders who require tailored occupational health management. These findings support a proactive, integrated approach to hepatitis B immunization surveillance in healthcare education, aligned with the WHO 2030 viral hepatitis-elimination targets. Full article

Background/Objectives: Nipah virus (NiV) is a highly pathogenic zoonotic agent that causes fatal respiratory and neurological diseases, for which no approved vaccines are currently available. To address this unmet need, we developed and evaluated novel mRNA-LNP vaccine candidates. These vaccines employ an epitope- and structure–guided “immune-focusing” strategy, aiming to maximize the exposure of critical neutralizing epitopes on the viral attachment (G) and fusion (F) glycoproteins. Methods: Head-to-head dimeric antigens (2Gs and 2Fs) were engineered by removing subdominant stalk regions and tandemly duplicating epitope-rich globular head domains. Three LNP-encapsulated candidates-NV1 (encoding 2Gs), NV2 (encoding 2Fs), and NV3 (a 1:1 mixture of NV1 and NV2)-were evaluated in BALB/c mice and Syrian hamsters to systematically assess humoral responses, cytokine secretion, BCR/TCR repertoires, and systemic safety. Results: NV1 and NV3 induced potent binding antibodies and strong neutralizing activity against NiV pseudoviruses. Meanwhile, although NV2 lacked neutralizing activity, it induced superior Th1-biased cellular immunity, accompanied by extensive T-cell clonal proliferation. BCR/TCR sequencing revealed unique adaptive immune characteristics that perfectly align with the distinct immunogenic properties described above. Preliminary safety assessments found no toxicity to vital organs, while active germinal center formation confirmed the success of immune mobilization. Conclusions: An epitope-centered dimer design effectively shapes the unique defense mechanisms of the adaptive immune system. NV3 offers a balanced and synergistic strategy that combines potent humoral and cellular immune defenses, providing a highly promising platform for the development of a Nipah virus (NiV) vaccine. Full article

As false news benchmarks are increasingly being developed from fact-checking sources, this introduces a slight, but under cited, threat to validity: source bias. The two classes are not necessarily similar in style or structure; a classifier could distinguish between fake and real items (both from a fact-checking outlet) based on surface features rather than truth, and obtain scores which do not reflect the ability of the classifier to actually detect the truth. We report this effect for Kazakh–Russian Telegram—a low-resource code-switched environment—and show that a naively engineered corpus yields artificially high performance (up to 0.98 F1) that plummets by about ten percentage points, after the removal of the shortcut. We then introduce a two-stage debiasing protocol: (1) structural cleaning (viral-boilerplate, interrogative openers and verdict labels removal from Tengrinews posts); and (2) a heterogeneous two-source real class constructed from Tengrinews posts and confirmed “True” (Pravda) verdicts on Factcheck.kz. This balanced, cleaned set of 804 posts (402 real/402 fake) is made available to the public. On this debiased benchmark we report honest baselines for six methods: TF-IDF + Logistic Regression, FastText, fine-tuned XLM-RoBERTa, fine-tuned RuBERT, and Llama 3.1 8B with zero- and few-shot prompting. The fine-tuned transformers show good performance (RuBERT 0.8819, XLM-RoBERTa 0.8757 macro F1), whereas classical TF-IDF is competitive (0.8508), and in-context LLM prompting lags behind (zero-shot 0.4522, few-shot 0.6601 macro F1)–quantifying the gap between in-context prompting and few hundred labeled examples. The transformer margin is very small and is calculated from a single split, so we only report the margin as indicative and discuss the evaluation conditions required to confirm the margin. We make a methodological contribution by providing a replicable debiasing recipe and showing that explicit source-bias controls are required to evaluate trustworthy fact-checks in low-resource, fact-check-derived misinformation benchmarks. Data and code are made publicly available so that they can be used to facilitate reproducible research. Full article

Viral diseases are one of the most destructive threats to global agriculture. Among plant viruses, tobacco mosaic virus (TMV) is a highly contagious pathogen infecting numerous economically vital crops. With no curable treatments, the only strategy to mitigate virus spread is early detection and plant removal. The gold standard for TMV detection is reverse transcriptase-polymerase chain reaction (RT-PCR), followed by agarose gel electrophoresis. To reduce TMV diagnosis time and eliminate the requirement for expensive equipment, this study developed and optimized a one-step RT-PCR-coupled cysteamine-functionalized gold nanoparticle assay for the colorimetric determination of the virus. By integrating cDNA synthesis and PCR amplification into a single tube and analyzing the results using cysteamine- functionalized gold nanoparticles (Au@Cys), the diagnosis turnaround time was significantly reduced. Furthermore, this AuNPs-based colorimetric assay enabled straightforward visual inspection with the naked eye, eliminating the need for costly optical devices. Additionally, our regression equation linking RT-PCR product color values, extracted as mean A value based on the CIELAB color space (green-red axis), and viral load allows for the accurate quantification of TMV infection levels in field samples. Our research lays the groundwork for the further development of more cost-effective, quantitative and rapid plant virus diagnosis methods. Full article

Background and Clinical Significance: Foreign body aspiration is an important cause of morbidity and mortality in children younger than three years of age. Although laryngeal foreign bodies are uncommon, they may be life-threatening and are frequently misdiagnosed because of their variable clinical presentation. Eggshell aspiration is exceptionally rare, with only a few cases reported in the literature. We report a case of delayed diagnosis of a glottic eggshell foreign body in a toddler presenting with persistent upper airway symptoms. Case Presentation: A 16-month-old previously healthy girl was referred to our hospital for evaluation of persistent hoarseness and barking cough following a witnessed choking episode while eating boiled egg. The choking episode had occurred 15 days before presentation during an episode of viral upper respiratory tract infection. Initial symptoms were attributed to laryngitis and persisted despite medical treatment. Flexible laryngoscopy revealed a foreign body impacted at the glottic level, while neck radiography demonstrated a radiopaque calcified lesion corresponding to the foreign body. Microlaryngoscopy under deep sedation was performed, and an approximately 10-mm eggshell fragment was successfully removed. A small amount of granulation tissue was observed at the posterior commissure, corresponding to the site of foreign body impaction. The postoperative course was uneventful, and repeat endoscopic examination on postoperative day three demonstrated satisfactory laryngeal healing with regression of the granulation tissue. Conclusions: This case highlights the importance of considering a retained laryngeal foreign body in young children with persistent hoarseness or barking cough following a choking episode, even in the presence of concomitant respiratory infection. Early endoscopic evaluation is essential to avoid diagnostic delay and facilitate prompt treatment. In addition, this report emphasizes the importance of food choking prevention and caregiver education in children younger than three years of age. Full article

Theories on the early steps of the HIV-1 life cycle have been radically revised over the past five years. The long-held assumption that the capsid fully disassembles in the cytoplasm has given way to a more nuanced view: Cytoplasmic disassembly does occur and, in several myeloid systems, is increasingly linked to cytosolic cDNA sensing and to abortive infection. However, a substantial fraction of intact or nearly intact capsid cores instead traverse the nuclear pore complex (NPC) and, upon interacting with the host factor CPSF6, induce liquid–liquid phase separation. This leads to the formation of biomolecular condensates, termed HIV-1 membraneless organelles (HIV-1-MLOs), which subsequently merge with nuclear speckles (NSs). In this Perspective we read these condensates along five interlocking axes. First, the virus drives the host phase separation of cleavage and polyadenylation specificity factor 6 (CPSF6), which quickly fuses with another MLO: the NS composed of the speckle scaffold factors, SON and SRRM2. Second, the resulting condensate behaves as a catalytic site that concentrates the reverse-transcription machinery and thereby promotes integration of the viral DNA into speckle-associated chromatin (SPADs). Third, the same compartment is the final layer of a stratified programme of innate immune evasion, shielding nascent double-stranded DNA from cGAS–STING after cytoplasmic restriction factors and sensors have been outmanoeuvred. Fourth, although demonstrated only in vitro, stable HIV-1-MLOs can maintain the viral RNA genome in the presence of a reverse-transcription inhibitor. Upon removal of the inhibitor, reverse transcription resumes, mirroring, to some extent, the situation in individuals undergoing interruption of antiretroviral therapy and suggesting that these structures may act as a pre-integration reservoir. Fifth, and still largely unexplored, the sanctuary has a pharmacological dimension: anatomical lymphoid compartments, and possibly the condensate itself through selective small-molecule partitioning, may limit antiretroviral drug access. We situate HIV-1-MLOs within the convergent condensate strategies of SARS-CoV-2 and other viruses, and we discuss the clinical, diagnostic, therapeutic, and vaccine implications, including capsid inhibitors as “block-and-expose” tools. Full article

Syndromic multiplex-PCR panels deliver rapid, comprehensive detection of pathogens and resistance determinants in acute infections, but their unrestricted use can generate a high proportion of negative results with substantial economic and stewardship implications. We retrospectively analysed all BioFire FilmArray meningitis/encephalitis (ME), upper-respiratory (RP2.1), pneumonia (PN) and gastrointestinal (GI) determinations performed at the UOSD Microbiology and Virology of P.O. “Vito Fazzi”, ASL Lecce (Apulia, Italy) between 1 July 2024 and 30 April 2026. Repeat determinations from the same patient with the same panel were identified in the laboratory information system and removed before any analysis, so that each determination analysed corresponds to a distinct diagnostic episode. Across 5381 determinations (CNS, n = 761; upper respiratory, n = 2601; lower respiratory, n = 399; gastrointestinal, n = 1620), 59.3% were negative for every target, with a steep appropriateness gradient: 90.0% negativity for the ME panel and 77.1% for the gastrointestinal panel versus 44.2% for the upper-respiratory and 26.6% for the pneumonia panel. CNS positives were predominantly viral (75%), led by enterovirus; Streptococcus pneumoniae was the only consistent bacterial agent. Human rhinovirus/enterovirus dominated the respiratory ecology; influenza A was almost entirely H3 and H1N1pdm09, but 8.1% of influenza-A–positive specimens were equivocal or non-subtypeable and were never referred for sequencing, an avoidable surveillance blind spot for novel/zoonotic (avian) influenza. Pneumonia-panel resistance markers (mecA/C–MREJ, CTX-M, KPC, NDM) clustered in Enterobacterales co-infections. The gastrointestinal panel was dominated by diarrhoeagenic Escherichia coli pathotypes (chiefly EPEC and EAEC) and Clostridioides difficile toxin, its 77% negativity identifying a second over-utilised stream. We argue for CSF-pleocytosis gating, tiered/reflex respiratory algorithms, gastrointestinal-panel gating to community-onset diarrhoea, and mandatory reflex sequencing of unsubtypeable influenza A. Full article

Tomato yellow leaf curl virus (TYLCV) is a major threat to tomato production, creating a need for sensitive, low-cost detection methods that can be applied to early symptomatic or low-viral-load samples. Recombinant antigen configuration may influence serological assay development, although the specific contribution of multiple-cloning-site (MCS)-derived intermediate sequences remains uncertain. In this study, a recombinant Trx-His-coat protein (CP) fusion antigen was produced using an MCS-free direct-fusion construct that retained the Trx-His tag while removing the MCS-derived intermediate sequence, followed by gradient refolding. No direct comparison with linker-containing, tag-cleaved, or tag-free antigen constructs was performed. The purified antigen was used to immunize rabbits and generate a high-titre polyclonal antibody (pAb). The resulting indirect enzyme-linked immunosorbent assay (ELISA) achieved a theoretical limit of detection of 1.8 ng/mL and an estimated pre-dilution equivalent the limit of detection (LOD) of 72 ng/mL after sample dilution. The assay showed favourable tolerance to crude tomato leaf matrices, with spike-recovery rates of 95.45–100.40%. In a preliminary evaluation using a balanced panel of 32 field-collected samples, ELISA absorbance correlated with droplet digital PCR quantification (R² = 0.9819) and plant disease index values (R² = 0.9774). Liquid chromatography–tandem mass spectrometry (LC-MS/MS) peptide mapping and AlphaFold2-based modelling were used only to provide preliminary computational context for antigen interpretation. The assay showed cross-recognition toward Tobacco curly shoot virus (TbCSV), indicating that it should not be considered strictly TYLCV species-specific. Therefore, this assay may support preliminary serological screening under the tested conditions, whereas molecular confirmation remains necessary when species-level identification is required. Full article

Characterizing the mosquito virome is essential for understanding host–microbiota interactions and vector competence, but it can be influenced by sample scale, sequencing strategy, and host depletion. This study evaluated the effect of sampling scale on mosquito virome characterization using a third-generation sequencing (TGS) metagenomics approach based on PacBio HiFi long reads, applied to L4 larvae and adults of Aedes mariae, analyzing single individuals and pools of increasing size before and after host genome removal. The results showed that sequencing yield did not increase with pool size, indicating that the total number of reads is not proportional to the number of individuals. Host genome removal reduced the overall number of reads but altered their composition, increasing the relative proportion of assigned viral reads and reducing unclassified sequences. Despite a similar total read output, virome diversity increased with pool size, with larger pools showing greater taxonomic richness driven by the contribution of each individual. However, the high proportion of unassigned reads suggests the presence of uncharacterized viruses. This methodological workflow was technically feasible for both single-individual and pooled samples. Single-individual analyses may provide complementary information on individual-level virome composition and on low-abundance viral taxa that could be less apparent in pooled samples, whereas pooled samples may facilitate the detection of a broader range of viral taxa and may better capture the shared component of viral diversity within the analyzed population. Full article

Viral infections are the main cause of asthma exacerbation, particularly in children. Impaired interferon induction by the airway epithelium has been linked to increased asthma exacerbation during viral infections. Several studies have suggested that epithelial cells in asthmatic mice induce low levels of type I and type III interferons, which leads to increased viral load. However, emerging evidence suggests that epithelial cells from asthmatic individuals induce delayed interferon responses. This study aimed to assess the ability of asthmatic and healthy epithelial cells to mount interferon-associated responses after exposure to viral stimuli. Variations in gene expression associated with interferon response were analyzed using datasets obtained from the Gene Expression Omnibus (GEO) database. Airway epithelial cells derived from healthy and asthmatic individuals were infected with RNA virus and then subjected to microarray or RNA sequencing. Lung tissues obtained from the animal models (mice and rats) were analyzed using RNA sequencing. Further data analysis was performed using integrated differential expression and pathway analysis (iDEP). Viral infection of airway epithelial cells derived from healthy and asthmatic subjects induces strong expression of interferon- and interferon-related genes. Interferon-stimulated genes (ISGs) were robustly induced in both asthmatic and healthy human epithelial cells after viral infection. However, the induction of virus-induced interferon-related responses was significantly lower in the lung tissues of animals with pre-allergic inflammation. Although previous studies have reported that the antiviral-interferon response is impaired or diminished in asthmatic individuals, our findings suggest that interferon-associated transcriptional responses are preserved in the airway epithelial cells of asthmatics during viral infection. This suggests that asthmatic epithelial cells induce an antiviral immune response, characterized by the induction of interferon-associated genes necessary for viral removal. Future studies should investigate the mechanisms underlying virus-induced asthma exacerbation. Full article

The COVID-19 pandemic highlighted the urgent need to develop effective and broad-spectrum antiviral therapies against coronaviruses. One strategy to address this concern is a combination therapy using repurposed drugs against zoonotic viruses with pandemic potential. We previously demonstrated that the combination of Remdesivir and Ivermectin is highly potent and synergistic in inhibiting the replication of murine hepatitis virus (MHV) in RAW264.7 macrophages. This study investigated the interactions between the drug combination, coronavirus and host by proteomics and RNA sequencing of MHV-infected H2.35 murine liver epithelial cells. Time-of-addition and time-of-removal assays suggested that the drug combination likely affected the synthesis of viral RNA and viral protein. This combination drastically diminished the live virus titer greater than the respective monotherapies in MHV-infected H2.35 cells (by ~4 log₁₀), as well as in SARS-CoV-2-infected VeroE6 cells and human nasal epithelial cells. Proteomic and transcriptomic analyses revealed that viral protein and RNA levels were significantly depressed upon combination treatment. The drug combination exhibited considerable negative effects upon host RNA processes and resulted in the upregulation of host protein processes (e.g., response to unfolded protein; protein insertion into ER membrane). Molecular pathways affected by the combination treatment were markedly distinct from the monotherapies and indicated that Ivermectin enhances Remdesivir by modulating critical host processes to synergistically exert its inhibitory effect on the coronavirus replication cycle. Full article

Effective hand washing takes time and hand sanitizers that contain alcohol have a number of drawbacks, and frequent use of alcohol may cause skin damage. The objective of this study is to formulate nanostructured lipid carrier systems containing chlorhexidine digluconate to be applied topically for hand hygiene, especially for people sensitive to alcohol. A cytotoxicity experiment was conducted to ascertain the safe dosage for each of the three nano-cream formulas (F1, F2 and F3). Following each treatment, the viral titer was assessed using tissue culture infectious dose₅₀ and standard plaque assays. The selected formulation was characterized rheologically. Furthermore, fifteen volunteers of various ages and genders participated in the vivo antimicrobial test of the selected formulation as a hand sanitizer. All of the formulas were found to be safe. Using the disc diffusion method, the three formulations exhibited in vitro antimicrobial effects against different microbes. F1 showed biphasic release, reasonable skin deposition and spherical droplets under a microscope. F1 exhibited a non-Newtonian shear thinning flow behavior. After 30 min, the reduction values for rotavirus and Phix-174 were 21 and 4%, respectively. Additionally, the impact of F1 was assessed on the infectivity of simian rotavirus sa-11 (ds RNA) and Phix-174 (ss DNA) bacteriophage. According to the findings of the in vivo study, the percentage of total bacterial counts that were removed varied from 91 to 100%. Moreover, the range of the removal percentage of total fungi was 95.38 to 100%. In summary, F1 can be used as an economic, safe, and effective hand antiseptic. It can also completely replace alcohol in the market. Full article

Post-acute sequelae of SARS-CoV-2 infection (PASC) extend well beyond the acute respiratory phase, with accumulating virological evidence that SARS-CoV-2 RNA, viral antigens, and proteolytic fragments may persist in cardiovascular and other extrapulmonary tissues, although the extent to which such detection represents replication-competent reservoirs versus residual viral material with uncertain pathological relevance remains under active investigation. Sudden cardiac death (SCD) and fatal pulmonary thromboembolism (PTE) have emerged as forensically and epidemiologically significant outcomes in individuals with prior infection, situated at the intersection of microbiology, public health, and forensic medicine. To synthesize current evidence on the virological mechanisms by which SARS-CoV-2 may contribute to post-acute sudden cardiac death (SCD) and pulmonary thromboembolism (PTE), the population-level epidemiology of these outcomes, and their implications for public health surveillance and forensic practice, we conducted a narrative review of PubMed (MEDLINE), Scopus, and Web of Science Core Collection. The search covered publications from January 2020 to December 2025 and focused on SARS-CoV-2 cellular tropism and tissue persistence, immune-mediated and thromboinflammatory mechanisms, excess cardiovascular and thromboembolic mortality, and autopsy-based pathological findings. After de-duplication of 1837 initially identified records (412 duplicates removed) and screening of 1425 unique records, 78 studies were retained for final synthesis based on virological, epidemiological, and forensic relevance. SARS-CoV-2 enters cardiomyocytes, pericytes, and vascular endothelial cells through ACE2-dependent mechanisms, with cathepsin L compensating for the limited cardiac expression of TMPRSS2. Viral RNA and antigen have been detected in cardiovascular and other extrapulmonary tissues months after symptom onset in selected autopsy series, although persistent detection of viral components does not necessarily indicate ongoing productive infection or direct tissue injury. Endothelial dysfunction, neutrophil extracellular trap (NET) formation, complement activation, and persistent thromboinflammation have been proposed as plausible mechanistic substrates for arrhythmogenic remodelling and thromboembolic events, although definitive causal pathways remain incompletely understood. Population-based studies document persistent excess cardiovascular mortality across multiple jurisdictions, with hazard ratios for pulmonary embolism remaining elevated months after acute infection, particularly in unvaccinated individuals. Autopsy series identify mixed pathological patterns including focal lymphocytic infiltrates, microvascular thrombosis, contraction-band necrosis, and cardiomyocyte vacuolation, although fulminant lymphocytic myocarditis fulfilling Dallas criteria remains uncommon. A microbiology-informed framework uniting tissue-based viral detection, standardized cardiac and pulmonary sampling protocols, and prospective post-mortem registries is needed to better characterize the potential contribution of SARS-CoV-2 to post-acute cardiovascular mortality and to support cause-of-death certification, public health surveillance, and medicolegal practice in the post-pandemic era. Many of the proposed mechanisms remain under active investigation, and definitive causal relationships between viral persistence and adverse cardiovascular outcomes have not yet been conclusively established. Full article

In Africa, the disease burden of diarrheal and respiratory diseases is amplified by limited surveillance capacity, diagnostic limitations, and socioeconomic inequalities. In rapidly urbanizing settings such as Kigali (Rwanda), integrating wastewater-based epidemiology (WBE) into existing surveillance systems offers a promising strategy for generating real-time epidemiological intelligence, identifying community-level hotspots, and addressing gaps in traditional reporting systems. Gastrointestinal and respiratory infections remain major causes of morbidity and mortality globally, particularly in low- and middle-income countries (LMICs), where traditional clinical surveillance systems frequently underreport the true disease burden. This systematic review synthesizes current evidence on the detection of gastrointestinal and respiratory pathogens in wastewater and evaluates the utility of WBE for early warning and public health action. A narrative review approach was used to identify peer-reviewed literature, global health reports, and surveillance studies focusing on the wastewater detection of gastrointestinal and respiratory pathogens. Databases including PubMed, Scopus, and Google Scholar were searched for studies published between 2000 and 2026. The search yielded 1247 records, of which 312 duplicates were removed. After title/abstract screening, 228 full-text articles were retrieved and assessed for eligibility. After a detailed evaluation, 108 studies were excluded for the following reasons: absence of pathogen-specific wastewater data (n = 46), a focus on environmental monitoring without public health relevance (n = 25), insufficient methodological description (n = 21), or other eligibility limitations such as a lack of primary data (n = 16). WBE provides a non-invasive, cost-effective approach for monitoring symptomatic and asymptomatic infections. Challenges involve variability in sampling, environmental factors affecting viral decay, and differences in laboratory workflows. WBE is a powerful complement to traditional infectious disease surveillance, offering early warning capabilities, population-level coverage, and real-time insights into pathogen circulation. Integrating WBE into surveillance programs, especially in LMICs such as Rwanda, can significantly strengthen epidemic preparedness, guide resource allocation, and improve outbreak response. Sustained investment in laboratory capacity, standardized protocols, and multisector collaboration is essential to fully leverage WBE for public health protection. Full article