TITLE: W-1 WATER FIELD SAMPLING
AUTHORS: REDI-NET Consortium

[DESCRIPTION]
This protocol describes water field sampling.

[GUIDELINES]
OBJECTIVE

To outline steps to properly collect water samples and evaluate the risk of zoonotic disease transmission by the detection of pathogens from environmental DNA (eDNA).

SUMMARY/SCOPE

The overarching aim of the REDI-NET is to develop a collaborative laboratory network between domestic and international partnering institutions to address disease surveillance needs in order to effectively detect, predict and contain potentially emergent zoonosis. This SOP provides guidance on the Protocol for the correct sampling of water at CONUS/OCONUS sites to evaluate and predict the risk of zoonotic diseases transmission.

RESPONSIBLE PERSON

Principal Investigator, Study Coordinator, Entomology Component Lead, Managers
 

MAINTENANCE OF EQUIPMENT

BEFORE EACH COLLECTION

Clean water collection devices.
Freeze and clean ice-packs.
Clean cool-boxes.
Charge all equipment(e.g., GPS unit, tablets). Make sure the tablet has enough free-space for field sampling pictures.

AFTER EACH COLLECTION

Clean all equipment thoroughly between sampling sites, including boots, cooler box (inside and outside), etc. Store sterile equipment separate from used equipment and samples.

QUALITY CONTROL

This SOP is reviewed by the applicable supervisor annually or as required in order to maintain its relevance.



 

 

[STEPS]
SECTION: SAMPLING TEAMS
1. 
 
Field sampling of eDNA water samples involves two people. One person serves as the ‘sampler’ and the other person serves as a ‘helper’. The helper can look up details in these instructions when needed, keep track of samples, handle objects that are contamination risks, serve as a second set of eyes for potential contamination, and ensure safety of the sampler in potentially hazardous field conditions.
SECTION: SAMPLING SITE SELECTION
2. Find two suitable sampling locations within the water body representing different ecological systems, if possible.
SECTION: SAMPLING SITE SELECTION
3. At each sampling location, soil samples should be taken from two points, one at the edge of the waterhole and the second one meter in from the edge. Sediment collection at each sampling location should be done in triplicate.
SECTION: SAMPLING SITE SELECTION
4. Because ‘the edge’ of the water body may change due to precipitation changes, the edge should be defined at each month's sampling.
SECTION: SAMPLING SITE SELECTION
5. If the water body has a "drop off" (embankment of more than 20 cm), the soil and water samples should be taken where the water "meets" the soil. Aim to collect at locations where finer sediment is present, if possible.
SECTION: SAMPLE COLLECTION
6. 
 
At each sampling location, the helper opens the bag containing the vials to have one open and ready for the sampler. The helper will open and close the bags containing the collection materials. This is to prevent potential contamination of samples.
SECTION: SAMPLE COLLECTION
7. The Sampler takes the appropriately labeled water dipper (for example, SL1-edge would be the dipper head for sampling location 1 at the edge) from the bag and opens the corresponding Whirl-Pak® Bag which can be placed in a bucket for easier manipulation. 
 

SECTION: SAMPLE COLLECTION
8. Check the depth of the water. If the depth of the water is greater than 1 m, the water column should be gently mixed using the water dipper without disturbing bottom sediment.
 

SECTION: SAMPLE COLLECTION
9. Once the water column is homogenized, collect water filling the dipper, and empty the sample into the Whirl-Pak® Bag until the 250 mL sample is collected (3/4 of a 532 mL Whirl-Pak® Bag).
 

SECTION: SAMPLE COLLECTION
10. Once the water is collected, measure pH of the sample by emerging litmus paper.
SECTION: SAMPLE COLLECTION
11. Close the Whirl-Pak® Bag and dry it using paper towels.
SECTION: SAMPLE COLLECTION
12. Label the Whirl-Pak® Bag with a Sharpie or water-proof marker.
SECTION: SAMPLE COLLECTION
13. Place the Whirl-Pak® Bags into the cooler box filled with ice for storage.
SECTION: SAMPLE COLLECTION
14. Proceed with REDI-NET SOP S-1 Soil Collection at the same sampling location.
SECTION: SAMPLE COLLECTION
15. Repeat the water and soil sampling 3 times per sampling location point.
SECTION: SAMPLE COLLECTION
16. Repeat steps 8-15 at the one meter mark at the same sampling location.
SECTION: SAMPLE COLLECTION
17. Clean boots thoroughly between sampling location sites.
SECTION: SAMPLE COLLECTION
18. Repeat steps 7-17 at the second sampling location until all the samples are collected.
SECTION: SAMPLE COLLECTION
19. When all samples for a collection site are taken, bring the samples back to the laboratory facility for further processing according to REDI-NET SOP W-2 Water Processing (filtration should be done within 1440 min).
SECTION: WATER STORAGE
20. Keep freshly collected water in labeled whirl-pak bags at 4 °C cold chain (cooler box) until return to laboratory. Water samples can be stored at 4 °C for up to 1 week.
SECTION: WATER STORAGE
21. Once at the laboratory, it is optimal to filter the water samples and freeze the filter disk in separate, labeled ziplock bags as soon as possible. If water samples can not be processed on the same day of collection, whole sample bags should be frozen at -80 °C. If -80 °C storage is unavailable, water bags/ filter disks should be stored at -20 °C until processing. Water samples can be stored at -20 °C for up to 1 month and at -80 °C for longer term storage.