TITLE: Field assessment protocol for non-destructive estimation of single- and multi-species cover crop biomass
AUTHORS: Etienne Sutton, Jennifer Blesh

[DESCRIPTION]
This protocol provides step-by-step instructions for estimating cover crop biomass using ground cover photos and height measurements. Compared to traditional destructive sampling techniques, this method is an inexpensive and efficient option for monitoring cover crop biomass with relatively high accuracy. Because the field assessment requires minimal technical expertise and equipment, it is also particularly well suited for citizen science and other applied research contexts aimed at generating large, geographically distributed datasets. For instance, in a two-year study of cover crop performance in the Great Lakes region, the field assessment allowed for collecting cover crop growth data from 257 fields across six states. To validate the accuracy of the field assessment and derive relationships for converting the field assessment output into standard biomass units (e.g., kg ha-1), cover crop biomass was also destructively sampled with a quadrat from 111 of those fields. The R2 values for relationships between the field assessment output and biomass validation samples were 0.84 for cereal rye (N = 55), 0.76 for single-species cover crops more broadly (N = 73), and 0.79 for multi-species mixtures (N = 38). Linear equations were then derived for each of those cover crop subgroups to calculate biomass in kg ha-1. We recommend following a similar validation process when using the field assessment in new regions or contexts. The protocol below is divided into three sections: 1) estimating single-species cover crop biomass; 2) estimating multi-species cover crop biomass; and 3) validation procedures.

[GUIDELINES]
Funding: USDA NIFA Predoctoral Fellowship (GRANT13372495); University of Michigan Libraries, Rackham Program in Public Scholarship, Center for Academic Innovation, and Institute for Global Change Biology at the School for Environment and Sustainability.

Duration: 30-60 minutes


[STEPS]
SECTION: Single-species cover crops: Field survey
1. Walk to a location in the first third of the field with representative cover crop growth, making sure to avoid field edges.
SECTION: Single-species cover crops: Field survey
2. Holding your arm out parallel to the ground at shoulder height, take a photo of the cover crop canopy. The photo should be directly overlooking the cover crop, like an aerial photo (i.e., not at an angle). Make sure to avoid capturing any shadows, shoes, or other objects in the photo. 
   

SECTION: Single-species cover crops: Field survey
3. Record the amount of weed pressure at this location on a scale of 1 to 5.
 

SECTION: Single-species cover crops: Field survey
4. Using a tape measure or other measuring device, collect and record height measurements for three average cover crop plants (in centimeters). 
 

SECTION: Single-species cover crops: Field survey
5. Move to a second location closer to the center of the field with representative cover crop growth.
SECTION: Single-species cover crops: Field survey
6. Take another photo of the cover crop canopy.
SECTION: Single-species cover crops: Field survey
7. Record the amount of weed pressure at this location on a scale of 1 to 5.
SECTION: Single-species cover crops: Field survey
8. Collect and record height measurements for three average cover crop plants (in centimeters).
SECTION: Single-species cover crops: Field survey
9. Walk to a final location in the last third of the field with representative cover crop growth.
SECTION: Single-species cover crops: Field survey
10. Take another photo of the cover crop canopy.
SECTION: Single-species cover crops: Field survey
11. Record the amount of weed pressure at this location on a scale of 1 to 5.
SECTION: Single-species cover crops: Field survey
12. Collect and record height measurements for three average cover crop plants (in centimeters).
SECTION: Single-species cover crops: Data processing
13. 
 
Upload the three cover crop canopy photos per field into Foliage (https://andres-patrignani.github.io/foliage/). Select “Cover crop” from the “Foliage type” drop-down menu.
SECTION: Single-species cover crops: Data processing
14. Calculate average %Cover as the mean of the values that appear in the “Cover” column.
SECTION: Single-species cover crops: Data processing
15. Calculate average height as the mean of all nine height measurements. 
SECTION: Single-species cover crops: Data processing
16. Calculate average weed pressure as the mean of the three weed pressure rankings.
SECTION: Single-species cover crops: Data processing
17. Multiply average %Cover by average height. 
SECTION: Single-species cover crops: Data processing
18. Multiply the resulting number from the previous step by the corresponding weed correction factor listed in the table below: 
  Average weed pressure rankingCorrection factor111 < 20.920.82 < 30.730.63 < 40.540.44 < 50.350.2

SECTION: Multi-species cover crops: Field survey
20. Walk to a location in the first third of the field with representative cover crop growth, making sure to avoid field edges. 
SECTION: Multi-species cover crops: Field survey
21. Holding your arm out parallel to the ground at shoulder height, take a photo of the cover crop canopy. The photo should be directly overlooking the cover crops, like an aerial photo (i.e., not at an angle). Make sure to avoid capturing any shadows, shoes, or other objects in the photo. 
  

SECTION: Multi-species cover crops: Field survey
22. Record the amount of weed pressure at this location on a scale of 1 to 5 .
 

SECTION: Multi-species cover crops: Field survey
23. Visually estimate and record the relative proportion of each species in the mixture (or the four most dominant species if there are more than four) at this location (e.g., SpA = 0.75, SpB = 0.25).
 

SECTION: Multi-species cover crops: Field survey
24. For each of the species in the previous step, record the heights of three average individuals using a tape measure or other measuring device (in centimeters). 
 

SECTION: Multi-species cover crops: Field survey
25. Move to a second location closer to the center of the field with representative cover crop growth.
SECTION: Multi-species cover crops: Field survey
26. Take another photo of the cover crop canopy.
SECTION: Multi-species cover crops: Field survey
27. Record the amount of weed pressure at this location on a scale of 1 to 5.
SECTION: Multi-species cover crops: Field survey
28. Visually estimate and record the relative proportion of each species in the mixture (or the four most dominant species if there are more than four) at this location.
SECTION: Multi-species cover crops: Field survey
29. For each of the species in the previous step, record the heights of three average individuals (in centimeters).
SECTION: Multi-species cover crops: Field survey
30. Walk to a final location in the last third of the field with representative cover crop growth.
SECTION: Multi-species cover crops: Field survey
31. Take another photo of the cover crop canopy.
SECTION: Multi-species cover crops: Field survey
32. Record the amount of weed pressure at this location on a scale of 1 to 5.
SECTION: Multi-species cover crops: Field survey
33. Visually estimate and record the relative proportion of each species in the mixture (or the four most dominant species if there are more than four) at this location.
SECTION: Multi-species cover crops: Field survey
34. For each of the species in the previous step, record the heights of three average individuals (in centimeters).
SECTION: Multi-species cover crops: Data processing
35. 
 
Upload the three cover crop canopy photos per field into Foliage (https://andres-patrignani.github.io/foliage/). Select “Cover crop” from the “Foliage type” drop-down menu.
SECTION: Multi-species cover crops: Data processing
36. Calculate average %Cover as the mean of the values that appear in the “Cover” column.
SECTION: Multi-species cover crops: Data processing
37. Calculate average height for each species (hgt) as the mean of all nine height measurements for that species. 
SECTION: Multi-species cover crops: Data processing
38. Calculate average proportion for each species (prop) as the mean of the three proportion estimates for that species.
SECTION: Multi-species cover crops: Data processing
39. Plug the values from the previous three steps into the following expression:   
%Cover x [(SpA hgt x prop) + (SpB hgt x prop) + (SpC hgt x prop) + (SpD hgt x prop)]
SECTION: Multi-species cover crops: Data processing
40. Calculate average weed pressure as the mean of the three weed pressure rankings.
SECTION: Multi-species cover crops: Data processing
41. Multiply the resulting number from step 39 by the corresponding weed correction factor listed in the table below:
 
  Average weed pressure rankingCorrection factor111 < 20.920.82 < 30.730.63 < 40.540.44 < 50.350.2

SECTION: Multi-species cover crops: Data processing
42. Plug the number calculated in the previous step into the following equation to determine cover crop biomass in kg ha-1:  .
 

SECTION: Validation procedures: Biomass sampling
43. 
 
Walk to a location in the first third of the field with representative cover crop growth, making sure to avoid field edges.
SECTION: Validation procedures: Biomass sampling
44. Toss the quadrat on the ground.
SECTION: Validation procedures: Biomass sampling
45. Clip all aboveground biomass of plants rooted within or directly under the quadrat and place into a paper bag labeled with the field site, species names, and replicate (e.g., rep "a"). Weeds can be grouped into one bag, but all other species should be placed into separate bags.
SECTION: Validation procedures: Biomass sampling
46. Move to a second location closer to the center of the field with representative cover crop growth.
SECTION: Validation procedures: Biomass sampling
47. Toss the quadrat on the ground.
SECTION: Validation procedures: Biomass sampling
48. Collect all aboveground biomass within or underneath the quadrat as in step 45, using new paper bags labeled with the field site, species names, and replicate (e.g., rep "b"). 
SECTION: Validation procedures: Biomass sampling
49. Walk to a final location in the last third of the field with representative cover crop growth.
SECTION: Validation procedures: Biomass sampling
50. Toss the quadrat on the ground.
SECTION: Validation procedures: Biomass sampling
51. Collect all aboveground biomass within or underneath the quadrat as in step 45, using new paper bags labeled with the field site, species names, and replicate (e.g., rep "c").
SECTION: Validation procedures: Biomass sampling
52. Dry all samples at 60 °C for a minimum of 2880 min. 

SECTION: Validation procedures: Biomass sampling
53. Once dried, use a scale to weigh each sample (in grams), making sure to subtract out the bag weight; record weights to two decimal places.
SECTION: Validation procedures: Data processing
54. For each species in each field, calculate the average mass as the mean of the three sample replicates. 
SECTION: Validation procedures: Data processing
55. Calculate biomass in kg ha-1 with the following operation:
  .
SECTION: Validation procedures: Data processing
56. For fields with multiple species, calculate total cover crop biomass by summing the biomass of all species (excluding weeds).
SECTION: Validation procedures: Data processing
57. Using the field assessment output (as calculated in “Single-species cover crops” step 18 or “Multi-species cover crops” step 41 above) as the independent variable and total cover crop biomass as the dependent variable, fit a regression model to the data, with fixed or random effects included as needed (e.g., year, farm). 
SECTION: Validation procedures: Data processing
58. Use the resulting regression equation to calculate cover crop biomass in kg ha-1 for all other fields for which the field assessment was performed. 
SECTION: Validation procedures: Data processing
59. Additional steps for assessing data quality:
SECTION: Validation procedures: Data processing
59.1. For multi-species cover crops, the relative proportion of each species from the biomass validation sampling can be compared to the proportions reported in field assessment to assess the accuracy of the visual estimates.
SECTION: Validation procedures: Data processing
59.2. The weed biomass data from validation sampling can also be used to verify the accuracy of weed pressure rankings in the field assessment. 
SECTION: Single-species cover crops: Data processing
19. If the cover crop is cereal rye, plug the number calculated in the previous step into the following equation to determine cover crop biomass in kg ha-1:  ; if the cover crop is a species other than cereal rye, use this equation:   .