Detection of SARS-CoV-2 RNA in wastewater from an enclosed college campus serves as an early warning surveillance system.

SARS-CoV-2, the causative agent of Covid-19, is shed from infected persons in respiratory droplets, feces, and urine. Using quantitative PCR (qPCR), our group hypothesized that we could detect SARS-CoV-2 in wastewater samples collected on a university campus prior to the detection of the virus in individuals on campus. Wastewater samples were collected 3 times a week from 5 locations on the main campus of the University of North Carolina Wilmington (UNCW) from July 24, 2020 to December 21, 2020. Post-collection, total RNA was extracted and SARS-CoV-2 RNA in the samples was detected by qPCR. SARS-CoV-2 signal was detected on campus beginning on August 19 as classes began and the signal increased in both intensity and breadth as the Fall semester progressed. A comparison of two RNA extraction methods from wastewater showed that SARS-CoV-2 was detected more frequently on filter samples versus the direct extracts. Aligning our wastewater data with the reported SARS-CoV-2 cases on the campus Covid-19 dashboard showed the virus signal was routinely detected in the wastewater prior to clusters of individual cases being reported. These data support the testing of wastewater for the presence of SARS-CoV-2 and may be used as part of a surveillance program for detecting the virus in a community prior to an outbreak occurring and could ultimately be incorporated with other SARS-CoV-2 metrics to better inform public health enabling a quick response to contain or mitigating spread of the virus.

Investigation of polyethylene terephthalate (PET) drinking bottles as marine reservoirs for fecal bacteria and phytoplankton.

Polyethylene terephthalate (PET) is frequently used in the food and beverage industry and therefore contributes greatly to plastic marine debris. The fecal pollution indicator bacteria Enterococcus is used for marine water contamination assessments and is regularly found in storm water discharge. In order to examine if PET drinking bottles act as refuges for Enterococcus, a study was conducted within euhaline tidal waters of Wrightsville Beach, NC, USA via the deployment of bottle floats positioned nearby two stormwater outfall pipes. Bottles were retrieved weekly to assess the accumulation of fecal bacteria and phytoplankton. Each bottle was analyzed for the presence of Enterococcus on plastic surfaces and within water inside the bottle. Abundance of Enterococcus and planktonic chlorophyll α was found to be significantly greater in association with PET bottles versus the surrounding waters. Bottles were observed to act as reservoirs for both Enterococcus and phytoplankton with concentrations well above the state, federal, and WHO standards.

Waste nutrients from U.S. animal feeding operations: Regulations are inconsistent across states and inadequately assess nutrient export risk.

Livestock production in the United States has been transformed over the past several decades, largely as a result of widespread development of industrial-scale mass production facilities, termed Animal Feeding Operations (AFOs). These facilities generate massive amounts of animal waste that can concentrate in small areas. Animal wastes from AFOs have led to high levels of nutrients and other pollutants in nearby surface waters, as well as groundwater. The environmental problems associated with these disposal practices have led to federal and state modifications to the rules and regulations governing waste practices. We summarize the federal guidelines for AFO nutrient management, focusing on swine, and compare the regulations of four AFO-rich states in different regions of the USA. Furthermore, we discuss inconsistencies among regulations and regulatory gaps, and identify issues with waste nutrient management practices that lead to environmental degradation in watersheds hosting AFOs. Finally, we address these shortcomings and the need to implement policy updates that would alleviate some of these environmental and human concerns.

Inflow and infiltration in coastal wastewater collection systems: effects of rainfall, temperature, and sea level.

Wastewater collection and treatment systems are vital to public health, economic growth, and environmental quality, but do not receive as much consideration for upgrades and improvements as other forms of public infrastructure. Extraneous inputs to a wastewater collection system are caused by rainfall and submergence ("Inflow and infiltration," I&I) and other factors, such as variation in sea level in coastal settings. These factors all pose risks for system degradation, sanitary system overflows (SSOs), and water quality impairment, but remain poorly quantified. Multiple regression analyses of total flows through 19 wastewater collection systems in coastal North Carolina, all using gravity collection systems, over a 2-year period (2010-2011) demonstrated statistically significant effects of rainfall, temperature, and sea level as drivers of extraneous flows. Rainfall effects were significant for 18/19 (95%) of these systems. Temperature effects were also significant for 18/19 (95%) of these systems. Sea level effects, primarily driven by spring-neap tidal oscillations, were significant for 11/19 (58%). Further, single factor regression analyses of the effects of temperature and sea level on system flows demonstrated significant effects for 16/19 (84%) and 18/19 (95%) systems, respectively. These collective results demonstrate the potential vulnerability of coastal wastewater collection and treatment systems to breaches in system integrity that allow extraneous flows, primarily through groundwater elevation, to drive further infrastructure degradation and environmental pollution. PRACTITIONER POINTS: Heavy rainfalls drive statistically significant inflow and infiltration (I&I) in over 90% of central wastewater systems in coastal North Carolina. Temperature, most likely as effects of seasonal variation in groundwater levels, also had a significant effect on I&I in over 90% of these systems. Sea level, expressed as daily high-high tide, drove significant effects on flow through over 90% of these systems.

Micro-zooplankton grazing as a means of fecal bacteria removal in stormwater BMPs.

A priority for environmental managers is control of stormwater runoff pollution, especially fecal microbial pollution. This research was designed to determine if fecal bacterial grazing by micro-zooplankton is a significant control on fecal bacteria in aquatic best management practices (BMPs); if grazing differs between a wet detention pond and a constructed wetland; and if environmental factors enhance grazing. Both 3-day grazing tests and 24-h dilution assays were used to determine grazing differences between the two types of BMP. Micro-zooplankton grazing was a stronger bacteria removal mechanism in stormwater wetlands rich in aquatic vegetation compared to a standard wet detention pond, although grazing was important in detention ponds as well. Our experiments indicated that the majority of grazers that fed on fecal bacteria were <20 µm in size. Grazing rates were positively correlated with fecal coliform abundance and increased water temperatures. Enumeration of grazers demonstrated that protozoans were significantly more abundant among wetland vegetation than in open water, and open wetland waters contained more flagellates and dinoflagellates than open wet detention pond waters. Grazing on fecal bacteria in BMPs is enhanced by aquatic vegetation, and grazing in aquatic BMPs in warmer climates should be greater than in cooler climates.

Rainfall effects on inflow and infiltration in wastewater treatment systems in a coastal plain region.

Aging wastewater collection and treatment systems have not received as much attention as other forms of infrastructure, even though they are vital to public health, economic growth, and environmental quality. Inflow and infiltration (I&I) are among potentially widespread problems facing central sewage collection and treatment systems, posing risks of sanitary system overflows (SSOs), system degradation, and water quality impairment, but remain poorly quantified. Whole-system analyses of I&I were conducted by regression analyses of system flow responses to rainfall and temperature for 93 wastewater treatment plants in 23 counties in eastern North Carolina, USA, a coastal plain region with high water tables and generally higher rainfalls than the continental interior. Statistically significant flow responses to rainfall were found in 92% of these systems, with 2-year average I&I values exceeding 10% of rainless system flow in over 40% of them. The effects of rainfall, which can be intense in this coastal region, have region-wide implications for sewer system performance and environmental management. The positive association between rainfall and excessive I&I parallels the effects of storm water runoff on water quality, in that excessive I&I can also drive SSOs, thus confounding water quality protection efforts.

Multiple modes of water quality impairment by fecal contamination in a rapidly developing coastal area: southwest Brunswick County, North Carolina.

Fecal contamination of surface waters is a significant problem, particularly in rapidly developing coastal watersheds. Data from a water quality monitoring program in southwest Brunswick County, North Carolina, gathered in support of a regional wastewater and stormwater management program were used to examine likely modes and sources of fecal contamination. Sampling was conducted at 42 locations at 3-4-week intervals between 1996 and 2003, including streams, ponds, and estuarine waters in a variety of land use settings. Expected fecal sources included human wastewater systems (on-site and central), stormwater runoff, and direct deposition by animals. Fecal coliform levels were positively associated with rainfall measures, but frequent high fecal coliform concentrations at times of no rain indicated other modes of contamination as well. Fecal coliform levels were also positively associated with silicate levels, a groundwater source signal, indicating that flux of fecal-contaminated groundwater was a mode of contamination, potentially elevating FC levels in impacted waters independent of stormwater runoff. Fecal contamination by failing septic or sewer systems at many locations was significant and in addition to effects of stormwater runoff. Rainfall was also linked to fecal contamination by central sewage treatment system failures. These results highlight the importance of considering multiple modes of water pollution and different ways in which human activities cause water quality degradation. Management of water quality in coastal regions must therefore recognize diverse drivers of fecal contamination to surface waters.

Stimulation of fecal bacteria in ambient waters by experimental inputs of organic and inorganic phosphorus.

Fecal microbial pollution of recreational and shellfishing waters is a major human health and economic issue. Microbial pollution sourced from stormwater runoff is especially widespread, and strongly associated with urbanization. However, non-point source nutrient pollution is also problematic, and may come from sources different from fecal-derived pollution (i.e. fertilization of farm fields, lawns and gardens, and ornamental urban areas). Fecal bacteria require nutrients; thus the impact of such nutrient loading on survival and abundance of fecal coliform bacteria in ambient waters was experimentally investigated in a constructed wetland in coastal North Carolina, USA. A series of nutrient-addition bioassays testing impacts of inorganic and organic nitrogen and phosphorus demonstrated that additions of neither organic nor inorganic nitrogen stimulated fecal coliform bacteria. However, phosphorus additions provided significant stimulation of fecal coliform growth at times; on other occasions such additions did not. Dilution bioassays combined with nutrient additions were subsequently devised to assess potential impacts of microzooplankton grazing on the target fecal bacteria populations. Results demonstrated grazing to be a significant bacterial reduction factor in 63% of tests, potentially obscuring nutrient effects. Thus, combining dilution experiments with nutrient addition bioassays yielded simultaneous information on microzooplankton grazing rates on fecal bacteria, fecal bacterial growth rates, and nutrient limitation. Overall, when tested against a non-amended control, additions of either organic or inorganic phosphorus significantly stimulated fecal coliform bacterial growth on 50% of occasions tested, with organic phosphorus generally providing greater stimulation. The finding of significant phosphorus stimulation of fecal bacteria indicates that extraneous nutrient loading can, at times, augment the impacts of fecal microbial pollution of shellfishing and human contact waters.

Impacts of a raw sewage spill on water and sediment quality in an urbanized estuary.

A sewer main serving a large municipal wastewater system ruptured, discharging approximately 3,000,000 gallons (11,355,000 L) of raw human sewage into a multi-branched tidal creek estuary along the US East Coast. The biochemical oxygen demand caused severe hypoxia in the system, causing a large fish kill. The sewage load led to high fecal coliform bacteria concentrations in the creek (maximum of 270,000 CFU 100ml(-1)), which declined in an approximate logarithmic manner over the first few days. The spill caused elevated sediment fecal coliform bacteria and enterococcus counts that declined much more gradually than water column counts. Persistence of relatively high concentrations of fecal indicator bacteria in sediments for several weeks after the spill suggests that sediment sampling should be included in response to major sewage spills. The high concentration of nutrients in the spilled sewage led to several algal blooms. However, nutrient concentrations in the water column declined rapidly, demonstrating the value of conserving marshes because of their pollutant filtration function.