Use of Wastewater for Mpox Outbreak Surveillance in California

Before April 2022, monkeypox virus infection in humans was seldom reported outside African regions where it is endemic. Currently, cases are occurring worldwide. Transmission, risk factors, clinical presentation, and outcomes of infection are poorly defined.

Orthopoxvirus monkeypox (MPXV) forms two distinct clades: the MPXV Congo Basin clade viruses are endemic in the Congo Basin, human illness typically presents with symptoms similar to discrete, ordinary smallpox and has a case fatality rate of approximately 10% in unvaccinated populations; the MPXV West African clade viruses have been isolated in West Africa and appear to cause a less severe, and less inter-human transmissible disease. Recently, monkeypox outbreaks were reported in US and Sudan caused by MPXV West African and Congo Basin strains respectively. These events demonstrated the ability and trend of the virus to exploit new hosts and emerge globally; it also emphasizes the need for the diagnosis of MPXV, especially the ability to distinguish between Congo Basin and West African monkeypox strains. In this study, three new real-time PCR assays based on TaqMan probe technology were reported: the MPXV West African specific, Congo Basin strain specific and MPXV generic assays. The new assays demonstrated good specificity and sensitivity in the validation study with multiple platforms and various PCR reagent kits, and will improve the rapid detection and differentiation of monkeypox infections from other rash illness.

A number of recent retrospective studies have demonstrated that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA concentrations in wastewater are associated with coronavirus disease 2019 (COVID-19) cases in the corresponding sewersheds. Implementing high-resolution, prospective efforts across multiple plants depends on sensitive measurements that are representative of COVID-19 cases, scalable for high-throughput analysis, and comparable across laboratories. We conducted a prospective study across eight publicly owned treatment works (POTWs). A focus on SARS-CoV-2 RNA in solids enabled us to scale up our measurements with a commercial lab partner. Samples were collected daily, and results were posted to a website within 24 h. SARS-CoV-2 RNA in daily samples correlated with the incidence of COVID-19 cases in the sewersheds; a 1 log 10 increase in SARS-CoV-2 RNA in settled solids corresponds to a 0.58 log 10 (4×) increase in sewershed incidence rate. SARS-CoV-2 RNA signals measured with the commercial laboratory partner were comparable across plants and comparable to measurements conducted in a university laboratory when normalized by pepper mild mottle virus (PMMoV) RNA. Results suggest that SARS-CoV-2 RNA should be detectable in settled solids for COVID-19 incidence rates of >1/100,000 (range, 0.8 to 2.3 cases per 100,000). These sensitive, representative, scalable, and comparable methods will be valuable for future efforts to scale up wastewater-based epidemiology. IMPORTANCE Access to reliable, rapid monitoring data is critical to guide response to an infectious disease outbreak. For pathogens that are shed in feces or urine, monitoring wastewater can provide a cost-effective snapshot of transmission in an entire community via a single sample. In order for a method to be useful for ongoing COVID-19 monitoring, it should be sensitive for detection of low concentrations of SARS-CoV-2, representative of incidence rates in the community, scalable to generate data quickly, and comparable across laboratories. This paper presents a method utilizing wastewater solids to meet these goals, producing measurements of SARS-CoV-2 RNA strongly associated with COVID-19 cases in the sewershed of a publicly owned treatment work. Results, provided within 24 h, can be used to detect incidence rates as low as approximately 1/100,000 cases and can be normalized for comparison across locations generating data using different methods.