1. Standard Operating Procedure WWSOP02000 Effective Date: 04/01/2013 KENTUCKY WATERSHED WATCH STREAM GRAB SAMPLE COLLECTION

2. This presentation is based on the “Kentucky Watershed Watch Stream Grab Sample Collection Standard Operating Procedure WWSOP02000” and is available from wwky.org Grab Sample SOP

3. Stream safetyStream safety Sample siteSample site location location Pre-samplingPre-sampling information information Collection proceduresCollection procedures PreservationPreservation Stream Grab Sample CollectionProcedures RecordkeepingRecordkeeping TransportationTransportation ResultsResults

4. Site Coordinates and Photos GPS (decimal degrees – 5 decimal places) GPS (decimal degrees – 5 decimal places) OR Digital map (KGS WW Site - http://kgs.uky.edu/kgsmap/krww/viewer.asp http://kgs.uky.edu/kgsmap/krww/viewer.asp or Google Earth) Take photos upstream and downstream of sampling point – with recognizable landmarks; should be taken annually during Summer sampling event Take photos upstream and downstream of sampling point – with recognizable landmarks; should be taken annually during Summer sampling event Label photos: “Site#upstream” and “Site#downstream” Label photos: “Site#upstream” and “Site#downstream” Submit information to Water Watch office Email: OllieTheOtter@ky.gov Submit information to Water Watch office Email: OllieTheOtter@ky.gov Phone: 1-800-928-0045 Sample Site Location

5. You will receive materials from program coordinators or by mail You will receive materials from program coordinators or by mail Read instructions upon receipt! (Critical info on sample times and delivery included) Read instructions upon receipt! (Critical info on sample times and delivery included) Inspect all sample and preservative containers upon receipt for damage or breakage Inspect all sample and preservative containers upon receipt for damage or breakage You will receive for sampling: InstructionsInstructions Transport informationTransport information Chain of Custody RecordsChain of Custody Records Gloves (disposable)Gloves (disposable) Sample containersSample containers Preservative (if required)Preservative (if required) Pre-sampling Information

6. Sample containers vary for the job at hand Pre-sampling Information

7. Watershed Watch Chain of Custody Record The sampling containers are pre-marked with a unique Site Number The Site Number should match the number on the “Chain of Custody Record” Pre-sampling Information

8. I received a bottle with a site number I don’t recognize! Check the documents in your packet, you may have been randomly assigned to collect a blank or a duplicate Pre-sampling information

9. Blank - A sample bottle filled while at the stream site with distilled water Serves as a check for possible contamination during sample handling or processing at the labServes as a check for possible contamination during sample handling or processing at the lab DO NOT indicate that the sample is a blank – the lab should not knowDO NOT indicate that the sample is a blank – the lab should not know Pre-sampling Information

10. Duplicate - A sample bottle filled with stream water at your site at the same time as your regular sample Serves as a check for variability among samples that should yield approximately the same lab resultsServes as a check for variability among samples that should yield approximately the same lab results DO NOT indicate that the sample is a duplicate – the lab should not knowDO NOT indicate that the sample is a duplicate – the lab should not know Pre-sampling Information

11. Proper attire for handling samples Gloves (disposable)Gloves (disposable) Eye protectionEye protection Pre-sampling Information

12. Basicequipmentneeded Collection Procedures

13. Selecting your sample collection location Collection Procedures On your first visit to your site, select an area of the stream where the flow is concentrated and where you can completely submerge and fill the bottle without touching the bottom NOTE: It is important that you collect your sample at the same location each time

14. Concentrated Flow Location Collection Procedures

15. SampleCollectionLocation High Gradient Collection Procedures

16. SampleCollectionLocation Low Gradient Collection Procedures

17. Grab Sample Collection Enter stream at a downstream location Enter stream at a downstream location Walk upstream to avoid disturbing bottom Walk upstream to avoid disturbing bottom Collect sample at the same location each time in Collect sample at the same location each time in the middle of your concentrated flow location the middle of your concentrated flow location and at mid-depth of stream and at mid-depth of stream Collection Procedures

18. Grab Sample Collection Collection Procedures 0 = Dry 1 = Ponded 2 = Low 3 = Normal 4 = Bank Full 5 = FLOOD! Do not sample during these flow conditions Flow Rate

19. Put on disposable gloves Put on disposable gloves DO NOT RINSE BOTTLE DO NOT RINSE BOTTLE Point mouth of bottle directly over surface of water, dip Point mouth of bottle directly over surface of water, dip and scoop bottle upstream beneath water surface, and and scoop bottle upstream beneath water surface, and through water column in one swift motion to fill above through water column in one swift motion to fill above line marked on bottle line marked on bottle If preservative is in bottle, take caution to angle mouth so If preservative is in bottle, take caution to angle mouth so as not to spill any while collecting sample as not to spill any while collecting sample Cap container, leaving at least a small airspace at top to Cap container, leaving at least a small airspace at top to allow lab mixing allow lab mixing Place in wet ice immediately Place in wet ice immediately ForE. coli For E. coli grab sample collection: Collection Procedures

20. Put on disposable gloves Put on disposable gloves Rinse bottle with stream water three times Rinse bottle with stream water three times (fill bottle ¼ full of stream water, cap, shake, and discard downstream) Point mouth of bottle upstream (against the flow), Point mouth of bottle upstream (against the flow), submerge entire bottle to mid-depth of stream, submerge entire bottle to mid-depth of stream, scoop and fill scoop and fill Rinse cap in stream and cap bottle Rinse cap in stream and cap bottle Add preservative (if necessary) and place in wet ice Add preservative (if necessary) and place in wet ice Collection Procedures For grab sample collection other thanE. coli: For grab sample collection other than E. coli:

21. What to do if you get a chemical on you For emergency first aid in the field, flush For emergency first aid in the field, flush affected area with large amounts of water affected area with large amounts of water and seek medical attention immediately and seek medical attention immediately Follow the first aid procedures found on Follow the first aid procedures found on the Material Safety Data Sheets (MSDS) the Material Safety Data Sheets (MSDS) Equipment Chemical Safety

22. Samples must be kept immersed in wet ice Samples must be kept immersed in wet ice (but not frozen) until received by laboratory Some samples may require other preservatives Some samples may require other preservatives (check your sample event packet) Preservation

23. Like containers, preservation methods vary (You don’t have to memorize these) Preservation

24. Preservation Sample Holding Time: length of time a preserved sample can be stored length of time a preserved sample can be stored after collection and before it is analyzed without after collection and before it is analyzed without significantly affecting the results significantly affecting the results from the minute you collect a sample, the from the minute you collect a sample, the clock starts ticking on the holding time clock starts ticking on the holding time varies for different types of analysis varies for different types of analysis established by the US Environmental established by the US Environmental Protection Agency Protection Agency

25. E. coli = 6 hours maximum Other than E. coli = Follow holding time instructions received in your sampling event packet Preservation Sample Holding Times

26. document and record thedocument and record the transfer of the samples transfer of the samples from the stream to the from the stream to the laboratory laboratory function as a fieldfunction as a field measurement form measurement form provide a place for fieldprovide a place for field observations observationsRecordkeeping The Chain of Custody Record (COC) serves to: Watershed Watch Chain of Custody Record

27. Site #Site # Location descriptionLocation description Sample collector nameSample collector name and contact info Correct any preprinted Correct any preprinted information if necessary information if necessaryRecordkeeping Chain of Custody Record prepopulated information prepopulated information

28. Watershed Watch Chain of Custody Record Date and Time of sample collectionDate and Time of sample collection Field Observations and MeasurementsField Observations and Measurements Date and time of sampleDate and time of sample Relinquished/Received Relinquished/Received must be identical Recordkeeping Volunteer entries on the Chain of Custody Cross through any mistakes and initial – do not erase

29. Transportation 1. Make delivery arrangements before the sampling event before the sampling event 2. Take your sample and COC to the designated drop-off to the designated drop-off location location 3. Double-check your recordkeeping paperwork recordkeeping paperwork 4. Make sure to sign your COC Sample Delivery

30. Runner LAB Sampler Drop Off Sampler Sampler Sampler Sampler 6 hrs (E. coli)Collection 7:00-10:00 AM Runners 9:00-1:00 PM Transportation

31. Results Professional labs analyze samples

32. Results Bacteria Analysis (Older Method) Colony Forming Units (CFU) Lab processes your stream sample and applies it to a special plate Lab processes your stream sample and applies it to a special plate Plate is kept warm for a specified period of time Plate is kept warm for a specified period of time Colonies of microscopic bacteria that have formed are counted Colonies of microscopic bacteria that have formed are counted Results are recorded in CFU Results are recorded in CFU

33. Results E. coli Analysis (Newer Method) Add Colilert to sample and dissolve Pour sample into Quanti-Tray Seal tray and keep warm for 24 hours Count fluorescent wells and convert

34. RESULTS Results will be posted on the Kentucky Geological Survey website after the basin data manager and KY Watershed Watch have reviewed for errors http://kgs.uky.edu/kgsmap/krww/viewer.asp

35. Interpretation E. coli Enlarged photo of E. coli bacteria shortened form of Escherichia coli shortened form of Escherichia coli microscopic bacteria that live in the microscopic bacteria that live in the intestines of warm-blooded animals intestines of warm-blooded animals and are passed in fecal material and are passed in fecal material used as an indicator of the possible used as an indicator of the possible presence of pathogens that may cause presence of pathogens that may cause infection or illness infection or illness

36. Interpretation Sources of E. coli in Streams in Streams

37. Interpretation Surface Water Quality Standards for E. coli Swimming Standard of 240 cfu/100 ml (single grab sample) Swimming Standard of 240 cfu/100 ml (single grab sample) Standards apply during Recreation Season from May 1 to October 31 Standards apply during Recreation Season from May 1 to October 31

38. InterpretationNutrients generally refers to forms of generally refers to forms of nitrogen (N) and phosphorus (P) nitrogen (N) and phosphorus (P) excess nutrients in streams can excess nutrients in streams can cause overgrowth of algae and cause overgrowth of algae and plants, which disrupts healthy plants, which disrupts healthy natural processes natural processes this can lead to large decreases this can lead to large decreases in dissolved oxygen, a condition in dissolved oxygen, a condition called eutrophication called eutrophication

39. InterpretationNutrients Excess of nutrients (N & P) Abundant growth of algae Algae die Decomposition of algae by bacteria consumes oxygen Low dissolved oxygen levels can lead to fish kills EUTROPHICATION

40. Interpretation Sources of Nutrients in Streams in Streams

41. Interpretation Surface Water Quality Standards forNutrients Kentucky does not have numeric surface water standards in regulation for all nutrients of concern (including Kentucky does not have numeric surface water standards in regulation for all nutrients of concern (including Total Nitrogen and Total Phosphorus) Surface water standards in Kentucky regulations include: Surface water standards in Kentucky regulations include: Ammonia (un-ionized)0.05 mg/l (parts per million) Ammonia (un-ionized)0.05 mg/l (parts per million) Nitrate (NO 3 )10 mg/l Nitrate (NO 3 )10 mg/l Sulfate (SO 4 )250 mg/l Sulfate (SO 4 )250 mg/l

42. InterpretationMetals elemental substances that are good elemental substances that are good conductors of heat and electricity conductors of heat and electricity naturally occurring in rock and soil, and naturally occurring in rock and soil, and can be weathered and carried to can be weathered and carried to streams when exposed streams when exposed can be harmful to humans or aquatic life can be harmful to humans or aquatic life at low levels over a long time (chronic) at low levels over a long time (chronic) or at higher levels for a short time or at higher levels for a short time (acute) (acute) Heavy Metal

43. Interpretation Sources of Metals in Streams in Streams

44. Interpretation Surface Water Quality Standards for Metals for Metals Aluminum Aluminum Antimony Antimony Arsenic Arsenic Barium Barium Beryllium Beryllium Boron Boron Cadmium Cadmium Calcium Calcium Chromium Chromium Cobalt Cobalt Copper Copper Gold Gold Iron Iron Lead Lead Lithium Lithium Magnesium Magnesium Manganese Manganese Nickel Nickel Phosphorus Phosphorus Potassium Potassium Selenium Selenium Silicon Silicon Silver Silver Sodium Sodium Strontium Strontium Sulfur Sulfur Thallium Thallium Tin Tin Vanadium Vanadium Zinc Zinc Underlined metals are ones for which Kentucky has numeric water quality standards, which can be found in 401 KAR 10:031 Section 6.

45. InterpretationMetals A fish consumption advisory is in effect statewide All Kentucky waters are under advisory for mercury. Women of childbearing age and children 6 years of age or younger should eat no more than one meal per week of freshwater fish. Adult men and other women are not included in the consumption notice. All Kentucky waters are under advisory for mercury. Women of childbearing age and children 6 years of age or younger should eat no more than one meal per week of freshwater fish. Adult men and other women are not included in the consumption notice. This is not an emergency as organic mercury can occur This is not an emergency as organic mercury can occur naturally in the environment and does not affect swimmers, skiers or boaters. Fish can accumulate these low levels of mercury by eating plankton and other small aquatic creatures.

46. InterpretationPesticides chemicals used to kill undesirable plants chemicals used to kill undesirable plants or insects or insects improper timing of application and improper timing of application and overuse can lead to runoff into streams overuse can lead to runoff into streams can be harmful to humans or aquatic life can be harmful to humans or aquatic life at low levels over a long time (chronic) at low levels over a long time (chronic) or at higher levels for a short time or at higher levels for a short time (acute) (acute)

47. Interpretation Sources of Pesticides in Streams in Streams

48. Interpretation Surface Water Quality Standards forPesticides Kentucky does not have numeric water quality standards for all pesticides (including atrazine) Kentucky does not have numeric water quality standards for all pesticides (including atrazine) Surface water standards in Kentucky regulations include: Surface water standards in Kentucky regulations include: 2,4-D0.1 mg/l (parts per million) 2,4-D0.1 mg/l (parts per million)

49. (CFU)

52. At the creek Practice grab sample collection