How to start a WES program

Pull together a multidisciplinary team to plan and implement a WES program, which includes:

  • sample collection
  • sample processing and analysis
  • data interpretation and sharing
  • data application and use

The following steps are relevant to many contexts, but some steps might only be applicable in certain situations, and additional steps may be required.

A photo of a sign that says "TOILETS" next to painted images of a man and a woman in colorful clothing. The sign is hanging from chains from a roof outside a building and there are trees in the background.
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A WES program needs team members from each of the three primary practice domains:

  1. Sanitation practitioners are typically responsible for collecting, storing, and transporting samples of wastewater, fecal sludge, or surface water that contains human feces. The sampling team may include wastewater treatment plant operators, field technicians, environmental engineers, environmental scientists, or other representatives of a wastewater utility or organization involved in fecal sludge management.
  2. Laboratory scientists analyze the samples to identify and/or quantify trends/levels of specific targets. They should be trained in environmental microbiology and use appropriate equipment, validated analytical methods, and quality assurance controls to ensure data reliability.
  3. Public health practitioners may include biostatisticians, epidemiologists, public health generalists, healthcare providers, and administrators from local or regional health departments, ministries of health, hospitals, and other institutions who identify targets of interest, process and analyze the wastewater data, alongside complementary public health data, and determine whether any action is required.

Team members for a WES program will come from many different disciplines across these practice domains. For a new WES program, individuals from these domains should be identified to fill key program roles. Representatives from NGOs, private sector, and research institutions may also play a role in the WES program, particularly if there is a lack of capacity within one of the three primary practice domains.

The program team should then define the goals of the WES program and the public health action(s) that will be prompted by surveillance data. This may include:

  • identifying the target(s) of interest (e.g. pathogens, ARGs) for which the WES program will be testing;
  • identifying the purpose of the testing, such as detecting the presence of targets in a community or monitoring trends of those targets;
  • identifying the public health actions that can be taken in response to positive detections, increasing trends, or decreasing trends;
  • documenting the planned public health actions or criteria for determining public health actions.

Sample collection can vary greatly depending on the circumstances. A sampling plan is typically used to document where to sample, how often to sample, what to sample, how to sample, and how to safely collect, store, and transport samples (CDC, 2023).

Select your sampling site(s) to cover a certain portion of the population, provide data on high-risk communities, or provide data for communities where clinical testing is unavailable or underutilized. Define the population represented by each sample:

  • Wastewater surveillance: Defining the sewer catchment area upstream of the sampling point requires knowledge of the sewer network and sewer connections. Sewer maps can aid in defining sewer catchment areas, delineating which houses, institutions, or other buildings may feed into the sewer upstream of the collection point.
  • Non-sewered sanitation surveillance: Defining the population that uses non-sewered communal sanitation facilities can be a challenge. Focus on sampling from communal facilities, rather than household-level facilities, because samples from communal facilities represent a larger population. Facility user surveys conducted over a few days are time-consuming but can provide an idea of where users live. Key stakeholder interviews are less resource-intensive and may clarify generally where users are coming from, though they will not provide as much detail as user surveys.
  • Environmental surveillance: Defining the population represented by a surface water sample requires topographical information for the area. Areas with informal housing and unsafely managed sanitation located within the drainage basin of the sampling point are likely to contribute to the environmental surveillance sample.

Other considerations include the accessibility, safety, and security of the location.

Sampling frequency should be chosen for each target of interest. For example, a testing program for SARS-CoV-2 might choose to take samples two to three days per week to identify trends in the data more quickly and accurately, whereas a polio surveillance program might rely on monthly testing to detect the presence or absence of poliovirus.

The three main types of samples are passive samples, grab samples, and composite samples.

Sample collection method

Description

Pros

Cons

Passive sample

Collected by deploying a device into the wastewater or surface water of interest for a specific amount of time (Schang et al., 2021)

The sampler contains adsorbent material, such as cotton gauze, an electronegative membrane, or other material that interacts with the sample, which can be contained in a housing or tied directly to a string

The sample in the adsorbent material is processed after the passive sampler is removed from the environment

Inexpensive

Low-theft value

Minimal maintenance requirements

Does not require electricity

Less labor-intensive with no specialized skills required

Results difficult to quantify, due to unknown volume flowing through sampler, though quantification may be estimated with normalization to a fecal indicator

The string anchoring the passive sampler can break, and the sampler can be lost

Grab sample

A specific volume of wastewater, surface water, or sludge is collected at a single point in time

Samples represent a single moment in time

Can be collected rapidly using simple equipment

Inexpensive

Does not require a power source

Results are quantifiable

Highly influenced by fluctuations in flow and composition

Likely to be less representative than composite samples, particularly when deployed in smaller systems

Composite sample

Collected by pooling multiple grab samples at a specified frequency over a set time period (CDC, 2023)

Often facilitated by an automatic sampler

Samples can be time proportional (i.e. sub-samples are collected at equal time intervals during the sample period) or flow proportional (i.e. sub-samples are collected at equal flow intervals, regardless of the time between sub-samples during the sample period). Flow proportional samples require a flow meter.

Results are quantifiable

Samples are considered more representative of community fecal contributions than single grab samples (see above)

Sampling is usually automated

Equipment is expensive

Requires a power source (electricity or battery)

Equipment footprint can be large

Equipment may be susceptible to theft

Equipment maintenance can be challenging and expensive; parts and service may be hard to acquire in some locations

  • Liquid wastewater collected from within a sewer collection system or at the influent of a treatment plant.
  • Sludge, such as primary sludge from a treatment plant or fecal sludge from a communal pit latrine, septic tank, or other containment chamber within a non-sewered system.
  • Liquid samples from fecally-impacted surface waters, including open drains or rivers.

Samples should be collected, stored, and transported in a manner that protects workers and prevents sample degradation. Those collecting samples should follow occupational safety practices that may include engineering and administrative controls, handwashing, specific safe work practices, and the use of personal protective equipment (PPE) normally required when handling untreated wastewater and fecal sludge. For composite and grab samples, the volumes collected should allow for at least two replicates of sample to be processed, allowing for a back-up sample, an archived sample, or a replicate if needed (APHL, 2022b). Any required metadata, such as date and time of collection, location, and sample type, should be recorded when the sample is taken (CDC, 2023b) (NOAA, n.d.). Metadata for environmental and non-sewered samples will vary from wastewater samples.

Selected metadata

All samples: date and time of collection, sample location including GPS coordinates, sample type (composite, grab, passive), sample matrix (wastewater, primary sludge, septic tank, pit latrine, dry pit latrine, surface water body, drainageway), time between collection and arrival at laboratory, temperature of sample at collection and at arrival, institution type if sample represents a single institution or building, estimated number of people represented by the sample, general site conditions

Wastewater samples: flowrate, sampling frequency for composite samples, pretreatment conditions if any

Non-sewered samples: point on the sanitation service chain where sample was collected (i.e. containment, conveyance, treatment), time elapsed since last emptying, addition of chemicals or additives

Environmental samples: recent rainfall/weather data, presence of trash (rubbish, litter), general observations/conditions

Passive samples: time of placement and time of removal of the sampler, type of passive sampler and adsorbent material used

If refrigeration of samples is required for the target(s) of interest, samples should be refrigerated as soon as possible during the collection process at temperatures no higher than 4 °C (39 °F) and then transported to the testing location as quickly as possible (same day or next-day) with cold packs. Many US jurisdictions designate wastewater samples as “UN3373, Biological substances, Category B” for shipment. Category B substances are infectious substances, including those transported for diagnostic or investigational purposes (such as for wastewater surveillance), that are “not in a form generally capable of causing permanent disability or life-threatening or fatal disease in otherwise healthy humans or animals when exposure to it occurs” (PHMSA, 2020).  APHL’s Packing and Shipping Guidance for Biological Substances, Category B Specimens (APHL, 2022a) offers instructions on how to package Category B substances for shipment. If possible, samples should be processed within 24 hours of collection to prevent sample degradation and ensure data is actionable. Remaining samples should be frozen at -70°C for archiving, avoiding more than one freeze-thaw cycle if future analysis is anticipated.

A laboratory analysis protocol should lay out each of the steps for processing and testing the samples and laboratory controls, the required personnel, space and equipment needs, and the health and safety protocols to be observed (APHL, 2022). The required laboratory analysis steps may vary depending on the target of interest, the sample type, and the surveillance program goals. For example, analytical methods to test for SARS-CoV-2 in wastewater using polymerase chain reaction (PCR) usually includes inactivation, concentration, extraction, and quantification steps, but these steps may be different for other targets of interest. Several laboratory methodologies and technologies may exist to process and test samples and can be selected based on cost, supply chain availability, personnel requirements, and other factors. Take measures to ensure that the testing results are accurate and repeatable, including using quality assurance (QA) control procedures to evaluate data quality.

Document the results of the testing in a reliable data management system that ideally includes protected online storage and access. Use sample IDs to link all metadata and QA data with sampling data. After testing is complete, all samples, reagents, and contaminated material must be decontaminated and/or disposed in accordance with all applicable regulations. Document all steps in the analytical workflow, including processing times.

Rapid, easy-to-use testing technologies, like the GeneXpert and mgLAMP may exist for certain targets (Daigle et al., 2022; Zhu et al., 2022) and could be considered, particularly in remote settings, emergency contexts, or locations without access to a capable laboratory.

 

Log and process the data in a way that facilitates data analysis and sharing, while maintaining data privacy. Protect the privacy of small communities and vulnerable populations by implementing controls, such as limiting public data sharing to populations above a certain size threshold, avoiding public data sharing for selected vulnerable populations, and not disclosing the exact location of sampling sites. The data processing and sharing protocol should identify the individuals responsible for processing and sharing the data, as well as those individuals and groups who should receive the data and in what format. Choose and document the data sharing frequency for different data user groups (e.g. internal team members, decision makers, general public).

Make sure that all equipment and supplies required for sampling, testing, data processing, and data sharing are available when needed to avoid delays. You may need to order supplies many months in advance due to global procurement chain challenges. Create a plan for reordering consumables, considering availability, expiration dates, and shipping times, as well as other site-specific constraints. The plan should identify the suppliers for all equipment and supplies. Take note of service schedules for equipment, and identify service providers for equipment maintenance and repair.

Once the planning steps are complete and the sampling, testing, and data protocols are in place, the WES program can begin. It can be helpful to identify a start-up period to allow for troubleshooting, follow-up training, and streamlining of data processing and data sharing procedures. Protocols should be reviewed on a regular basis, with procedures adjusted as necessary.