Recently, wastewater-based epidemiology (WBE) research has experienced a strong impetus during the Coronavirus disease 2019 (COVID-19) pandemic. However, a few technical issues related to surveillance strategies, such as standardized procedures ranging from sampling to testing protocols, need to be resolved in preparation for future infectious disease outbreaks. This review highlights the study characteristics, potential use of WBE and overview of methods, as well as methods utilized to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) including its variant in wastewater. A literature search was performed electronically in PubMed and Scopus according to PRISMA guidelines for relevant peer-reviewed articles published between January 2020 and March 2022. The search identified 588 articles, out of which 221 fulfilled the necessary criteria and are discussed in this review. Most global WBE studies were conducted in North America (n = 75, 34 %), followed by Europe (n = 68, 30.8 %), and Asia (n = 43, 19.5 %). The review also showed that most of the application of WBE observed were to correlate SARS-CoV-2 ribonucleic acid (RNA) trends in sewage with epidemiological data (n = 90, 40.7 %). The techniques that were often used globally for sample collection, concentration, preferred matrix recovery control and various sample types were also discussed. Overall, this review provided a framework for researchers specializing in WBE to apply strategic approaches to their research questions in achieving better functional insights. In addition, areas that needed more in-depth analysis, data collection, and ideas for new initiatives were identified.

The Coronavirus disease 2019 (COVID-19) pandemic has caused significant global threats, as Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is primarily transmitted through airborne droplets and bioaerosols. Healthcare workers are particularly at high risk, yet there is limited research on the presence of SARS-CoV-2 in bioaerosols within healthcare facilities in Malaysia. This study aimed to determine the presence and viability of SARS-CoV-2 and its variants of concern in the air and ventilation systems of designated COVID-19 facilities from December 2021 to February 2022. Samples were collected from two hospitals and one quarantine centre (QC), including medical wards, intensive care units, emergency departments, and QC halls. Air samples were obtained using air samplers, while surface samples were taken from return air grilles. SARS-CoV-2 ribonucleic acid (RNA) and its variants were detected using reverse transcription droplet digital polymerase chain reaction (RT-ddPCR) and PCR-based genotyping, respectively. Results showed that Hospital A had a higher rate (24.6%) of positive samples than Hospital B (8.8%). Surface samples had a higher positivity rate (50.0%) compared to air samples (8.3%). The detected variants included delta (34.7%), a mixture of delta and omicron (8.7%), non-variant of concern (non-VOC) (8.7%), and omicron (4.3%). This study emphasizes the need for strict airborne infection control measures for healthcare workers.

Background: Wastewater-based surveillance (WBS) has emerged as a valuable tool for population-level monitoring of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) transmission, yet the interplay between sampling strategies and disease prevalence in shaping detection performance remains ambiguous. We investigated how grab and composite sampling influence SARS-CoV-2 ribonucleic acid (RNA) detection dynamics and predictive lag times across high- and low-prevalence communities in Selangor, Malaysia. Methods: A 28-week longitudinal study was conducted in Selangor, Malaysia, comparing grab and composite wastewater sampling in communities with high and low Coronavirus disease 2019 (COVID-19) prevalence. SARS-CoV-2 RNA in 348 samples was quantified using digital Reverse Transcription Polymerase Chain Reaction (RT-dPCR), and viral lineages were characterized by Nanopore sequencing. Detection sensitivity and lead times relative to reported cases were evaluated. Results: In low-prevalence settings, grab sampling showed higher detection sensitivity than composite sampling (92.0% vs. 70.0%), whereas both methods achieved similarly high detection in high-prevalence areas (>97.0%). Lag-time analysis indicated that grab sampling in high-prevalence settings was significantly associated with case trends at potential two-week lead (p = 0.024), while composite sampling in low-prevalence settings showed the strongest association at a potential one-week lead (p = 0.0022). Overall, lag structures varied by both sampling strategy and prevalence context. Both sampling approaches captured the replacement of Omicron sublineages (XBB.1.5, XBB.1.9.1, XBB.1.16) and identified additional circulating variants, including EG.5, that were not captured in the available clinical sequencing dataset during the same period. Conclusions: These findings reveal that local transmission intensity is associated with the utility of different sampling designs. Context-specific optimization of WBS sampling strategies enhances sensitivity, reduces detection lag, and strengthens early warning and genomic-tracking capacity in public health surveillance frameworks.

Dengue fever, with a high proportion of asymptomatic infections, poses a major global public health challenge that traditional surveillance systems frequently underestimate. Wastewater-based epidemiology (WBE) has emerged as a promising approach to monitoring infectious diseases beyond enteric viruses. Dengue virus is shed in urine, feces, and saliva, providing a biological basis for wastewater detection alongside clinical surveillance. This systematic review and meta-analysis synthesize current evidence on dengue virus (DENV) detection in wastewater and evaluate methodological factors influencing detection success in WBE. A systematic literature search using selected databases and predetermined keywords, followed by eligibility screening, resulted in ten studies being included, covering community surveillance and experimental trials. DENV ribonucleic acids (RNA) were most consistently detected and enriched in wastewater solids, indicating this matrix as the most reliable for surveillance. Among concentration methods, ultrafiltration achieved the highest viral recovery efficiency, while reverse transcription digital polymerase chain reaction (RT-dPCR) demonstrated superior sensitivity and precision compared to those of reverse transcription quantitative polymerase chain reaction (RT-qPCR), particularly at low viral concentrations. Storage at −80 °C was critical for preserving RNA integrity. The meta-analysis yielded a pooled DENV positivity rate of 24% (95% CI: 20–28%) after exclusion of outliers. Overall, solid-phase analysis combined with RT-dPCR represents the most sensitive methodological approach across the included studies. Harmonized protocols are needed to support future translation of dengue WBE into community surveillance as current evidence mainly demonstrates methodological feasibility and provides a technical foundation for future public health integration. Therefore, further longitudinal and multi-site validation is required to establish its broader applicability for dengue surveillance.

Dengue Virus (DENV) circulates as four antigenically distinct serotypes whose dominance fluctuates over time in many endemic regions, a phenomenon known as serotype shift that is frequently associated with large outbreaks and increased disease severity. This review, through a synthesis of epidemiological, virological, immunological, entomological, and environmental evidence, observes that serotype shift likely arises from the interaction of multiple determinants rather than solely from viral evolution, with population immunity playing a central role. The accumulation of serotype-specific herd immunity, together with short-lived cross-protection and Antibody-Dependent Enhancement (ADE), reshapes population susceptibility and creates ecological space for heterologous serotypes with higher transmission potential. The synthesis of global dengue studies indicates that these immune dynamics interact with viral genetic diversity, vector competence, climate variability, and human factors such as demography, socioeconomic status, population density and mobility to drive cyclical and sometimes abrupt changes in serotype dominance. Notably, the review indicates that serotype changes often precede or coincide with more clinical severity and patterns of outbreaks, with direct implications for the process of forecasting outbreaks, vaccine performance, and preparedness to respond with appropriate health measures. On the whole, this review confirms the opinion that the change of dengue serotype occurrence becomes a consequence of interconnected biological and ecological processes involved in the transmission of dengue serotype shifts in hyperendemic areas.