Randomised block trial evaluating biological/microbial products claims

1. 1
Trial examining the impact of various organic/biological applications on Poa annua in a mixed
Agrostis/Poa annua green
A number of products are on the Australian market making claims with regard to plant health and
improving root growth. A number of these were trialled to evaluate these.
Initial findings
Experimental protocol
A practice green comprising a mixed Agrostis/Poa annua sward at Bonnie Doon GC was marked out
into 64 x 1m2 plots with a 50mm buffer between each. A randomized block trial was set up
comprising 16 treatments. These were applied as shown opposite with 4 replicates of each including
a control which was untreated.
The trial ran from 6th
February to 16th
April 2018.
Product was applied as per label rate in 400L of water (foliar application) apart from myco-up applied
in 800L/Ha (soil application). All treatments were monthly apart from Product 4 applied every three
weeks
Plots were monitored for turf health using an NDVI (Spectrum TCM 500 Turf Colour metre), together
with GIA Software for root analysis at the cessation of the trial.
A L J N O P P I
F I F M I K O B
B E L H C E J H
D G B J H G A E
M H E L D J L D
C A C A G M C N
P D G F B O F O
N K K I M N K P

2. 2
PRODUCTS: Name Active Ingredient Rate/Ha Rate/1m2
A
B
C Control Control no
treatment
- -
D
E
F
G Product 1 2.8Kg/Ha 0.28g
H Product 2 53g/Ha 0.0053g
I Product 3 20L/Ha 1ml
J
K
L
M Product 4 12.5kg/Ha + 700ml/Ha 1.25g
N
O
P K acetate + Ca
acetate + Mn
acetate + Mg
acetate
30L + 15L + 15L + 15L+15L 3ml + 1.5ml + 1.5ml +
1.5ml + 1.5ml;

3. 3
Results
Results were recorded for all trials and statistical analysis carried. Differences were determined
significant among treatments when the ANOVA f test had a corresponding P value 0.05. In such
cases, a Fisher’s protected LSD test was performed to separate treatments’ differences (Freund and
Wilson 1993)
Root length
Initial root length
Control Product 3 Product 1 Product 4 Acetates +
UAN
Product 2
Sample
1
5.5 9 5.5 7 6.7 6
Sample
2
9.1 8 6.5 7.5 11 7
Sample
3
7.3 9.3 7 5.5 11 7
Sample
4
10 9.8 6 10 7.5 5
Sample
5
8 10 11.2 7
Sample
6
8 7.3 12.5
Mean 7.98 9.03 7.00 8.08 9.05 7.42
ST DEV 1.545855 0.758837 1.767767 2.106577 2.275228926 2.61565798
Statistical analysis (ANOVA Fisher LSD 95% confidence) showed no significant difference between the
root lengths pretreatment)
0
2
4
6
8
10
12
Control Product 3 Product 1 Product 4 Acetates +
UAN
Product 2
Mean maximum root length cm pre treatment

4. 4
Root length at cessation of trial
Control Product
3
Product
1
Product 4 Acetates + UAN Product 2
Sample
1
7.8 11.1 7.7 9.3 8.4 8.2000
Sample
2
8.2 8.9 7.4 7.5 8.4 6.00
Sample
3
8.8 8.8 8.4 9 9.3 8.3000
Sample
4
7.8 10.6 7.9 8.2 7.2 8.6
Mean 8.15 9.85 7.85 8.5 8.325 7.775
ST DEV 0.472582 1.173314 0.420317 0.8124038 0.861684397 1.19547759
Differences were determined significant among treatments when the ANOVA f test had a
corresponding P value 0.05. In such cases, a Fisher’s protected LSD test was performed to separate
treatments’ differences (Freund and Wilson 1993). There were significant differences between the
root lengths post treatment) with Product 3 giving significantly greater root length than the control
and all other treatments.
There was no significant difference between the pre and post treatments.
0
2
4
6
8
10
12
Control Product 3 Product 1 Product 4 Acetates +
UAN
Product 2
Lengthcm
Mean maximum root length cm post treatment

5. 5
NDVI
There was no significant long lasting difference in quality readings throughout the duration of the
trial.
Initial NDVI readings
Control Product
3
Product
2
Product
1
Product
4
Acetates
0.6872 0.6727 0.6762 0.6818 0.6434 0.6779
0.6695 0.6757 0.6864 0.6688 0.6708 0.6827
0.6952 0.6822 0.6758 0.6592 0.7049 0.6978
0.691 0.67 0.6564 0.665091 0.6567 0.6718
Mean 0.685725 0.67515 0.6737 0.668723 0.66895 0.68255
SD 0.011299 0.005245 0.012533 0.009572 0.026449 0.011102
0.61
0.62
0.63
0.64
0.65
0.66
0.67
0.68
0.69
0.7
0.71
Control Product 1 Product 2 Product 3 Product 4 Acetates
Initial pre treatment NDVI

6. 6
Post treatment NDVI readings
The following graphs show the impact of the individual treatments as indicated by the black arrows.
All the treatments gave a ‘’spike’’ in turf quality but this generally was short lasting.
0.6
0.62
0.64
0.66
0.68
0.7
0.72
0.74
12-Feb 19-Feb 26-Feb 5-Mar 12-Mar 19-Mar 26-Mar 2-Apr 9-Apr 16-Apr
Control
0.6
0.62
0.64
0.66
0.68
0.7
0.72
0.74
12-Feb 19-Feb 26-Feb 5-Mar 12-Mar 19-Mar 26-Mar 2-Apr 9-Apr 16-Apr
Product 1

7. 7
0.56
0.58
0.6
0.62
0.64
0.66
0.68
0.7
0.72
0.74
12-Feb 19-Feb 26-Feb 5-Mar 12-Mar 19-Mar 26-Mar 2-Apr 9-Apr 16-Apr
Product 2
0.6
0.62
0.64
0.66
0.68
0.7
0.72
0.74
12-Feb 19-Feb 26-Feb 5-Mar 12-Mar 19-Mar 26-Mar 2-Apr 9-Apr 16-Apr
Product 3

8. 8
0.6
0.62
0.64
0.66
0.68
0.7
0.72
0.74
12-Feb 19-Feb 26-Feb 5-Mar 12-Mar 19-Mar 26-Mar 2-Apr 9-Apr 16-Apr
Product 4
0.6
0.62
0.64
0.66
0.68
0.7
0.72
0.74
12-Feb 19-Feb 26-Feb 5-Mar 12-Mar 19-Mar 26-Mar 2-Apr 9-Apr 16-Apr
Acetates

9. 9
DGCI Analysis
This is a method of quantifying turfgrass color using digital image analysis. Where on a scale of 0–1.0:
1.0 = darkest green
In the case of DGCI SD the lower DGCI SD means more uniform turf color and higher DCGI SD means color is
less uniform
0.25
0.27
0.29
0.31
0.33
0.35
0.37
0.39
0.41
DGCI Mean April
1
DGCI Mean 2
April
DGCI Mean 9
April
DGCI Mean 11
April
DGCI Mean 3
May
DGCI Mean
Control Product 3 Product 2 Product 1 Product 4 Acetates + UAN
0.03
0.035
0.04
0.045
0.05
0.055
0.06
0.065
DGCI SD Mean
April 1
DGCI SD Mean 2
April
DGCI SD Mean 9
April
DGCI SD Mean
11 April
DGCI SD Mean 3
May
DGCI SD Mean
Control Product 3 Product 2 Product 1 Product 4 Acetates + UAN

10. 10
Root Imagery
Root architecture
GIA Analysis
Characterizing root system architecture (RSA) is essential to understanding the development and
function of vascular plants. Identifying RSA-associated genes also represents an underexplored
opportunity for crop improvement. Software tools are needed to accelerate the pace at which
quantitative traits of RSA are estimated from images of root networks.
Testing was carried out using GiA Roots (General Image Analysis of Roots), a semi-automated
software tool designed specifically for the high-throughput analysis of root system images. GiA Roots
includes user-assisted algorithms to distinguish root from background and a fully automated pipeline
that extracts dozens of root system phenotypes. Galkovskyi, T, et al (2012)
Control
Product
1
Product
2
Product
3
Product
4 Acetates
Mean width (um) 23.21 24.25 30.60 27.23 29.27 30.03
SD 2.10 6.34 3.27 2.35 2.51 2.280
GIA
Max Number roots 39.75 51.89 39.9 44.33 52 44.3
Number of Connected
Components 40.16 52.89 45.6 55.75 59.8 51.33
Median Number of roots 27.91 40.77 28.8 34.58 31.7 29.83
Width
Microscopic analysis using a stereo microscope together with a Dino Eye digital microscope running
Dino Eye software revealed significant differences in root width with Product 1, 4 and the acetates
all giving significantly greater root widths than the control.
Product 1 and 4 gave significantly greater numbers of roots compared to the control.
All treatments apart from Product 2 gave a greater number of connected components than the
control.
References
Galkovskyi, T, Mileyko, Y., Bucksch, A., Moore, B., Symonova, O., Price, C.A., Topp, C.N., Iyer-Pascuzzi,
A.S., Zurek, P.R., Fang, S., Harer, J., Benfey, P.N. and Weitz, J.S. (2012) GiA Roots: software for the
high-throughput analysis of plant root system architecture. BMC Plant Biology. 12:116.