Oakes Irrigation Research Site
Carrington Research Extension Center * North Dakota State University
P.O. Box 531, Oakes, ND 58474-0531, Phone: (701) 742-2744, FAX: (701) 742-2700, E-mail:													Kelly.c.Cooper@ndsu.edu
Field Evaluation of Oilseed Sunflowers for Resistance to Sclerotinia Head Rot
M. Wunsch, B. Kraft, M. Schaefer and S. Kallis, K. Cooper, L. Besemann and H. Eslinger
Methods
General Agronomics: The study was on a Maddock sandy loam soil type.  The previous crop was onion and the tillage operation consisted of disking once in the fall and spring cultivations (May 2, May 27 and June 1) to remove small weeds and to smooth the seedbed.  Authority Elite (sulfentrazone, 0.7 lb ai/gal, and s-metolachlor, 6.3 lb ai/gal; FMC Corp., Philadelphia, PA) was applied pre-emergence at 22 fl oz/acre on June 3.
Experimental design: A completely randomized block design with four replicates. The seeded plot size was 5 feet (center to center) by 25 feet long. The harvested plot size was five feet (center to center) by approximately twenty-one feet long. There were two rows per plot and the row spacing was 30 inches. Guard plots (10 feet wide) were established along all perimeters of the trial.
Planting details: Sunflowers were planted on June 2 using a Kinze 2000 planter with cone units. The seeding rate was 3.83 seeds/linear foot of row = 60,000 seeds/acre. The final plant population was 1 plant every 10 inches of row = 21,000 plants/acre. The final population was established by manual thinning the sunflowers during early vegetative growth.
Disease establishment: Inoculations were conducted over multiple days such that every head was inoculated twice - once at approximately R5.4 to R5.6 (40 to 60% of the disk flowers blooming or already bloomed) and once at approximately R5.7 to R5.9 (70 to 90% of the disk flowers blooming or already bloomed).  To ensure that every plant was inoculated twice, plants were given a dot of spray paint on an upper leaf at each inoculation; once a plant received two dots of spray paint, the plant received no additional inoculations.  Inoculations were conducted on August 3, 5, 6, 9, 10, and 12.
Inoculations:  At each inoculation, 15,000 spores were applied to the front of each head (each head received a total of 30,000 spores over two inoculations).
Irrigation:  Overhead irrigation was applied through micro-sprinkler misting systems.  From August 3 (when the first inoculations commenced) through August 17, irrigation was applied, with water delivered 2 or 3 minutes every 30 or 60 minutes as needed to keep the front of sunflower heads moist to the touch.
Parts of this trial experienced severe lodging caused by several strong wind storms that occurred in August and September.
Data collection: (disease assessments, harvest) was conducted only in plots that suffered less than 30% lodging.
Disease assessments:  Conducted shortly before harvest on September 19-21 at the late R8 to early R9 growth stage, with every plant was assessed for the percent of the head exhibiting Sclerotinia head rot.  Plants exhibiting damage from sunflower midge were excluded from the analysis; otherwise, all plants in each row were evaluated. Strong storms in August and early September caused significant lodging in parts of the trial.  Plots were only assessed for disease if fewer than 30% of plants within the plot were lodged, and disease assessments were not taken on any lodged plants.
Statistical analysis: Data were evaluated with analysis of variance except those data sets for which multiple plots included missing data (Sclerotinia heat rot severity, shattering incidence and severity; for these data sets, any plots in which no disease was observed were recorded as missing data). Assumptions of ANOVA: (1) The assumption of constant variance was assessed with Levene's test for homogeneity of variances and visually confirmed by plotting residuals against predicted values. (2) The assumption of normality was assessed with the Shapiro-Wilk test and visually confirmed with a normal probability plot. (3) The assumption of additivity of main-factor effects across replicates (no replicate-by-treatment interaction) was evaluated with Tukey's test for nonadditivity.The R9 Sclerotinia head rot incidence and severity index data and % seed over 25/64 sieve for hybrid ‘12GCF05’ exhibited a moderate deviation from normality due to outliers; a systematic transformation could not be found that addressed the problem, and the untransformed data were analyzed. For data that violated model assumptions, a systematic natural-log transformation [LN(x+1) for data sets with values less than 1, otherwise LN(x)] was applied. Assessment of whether results differed by inoculation timing and/or hybrid and whether there was an interaction between inoculation timing and hybrid: Analyses were conducted with replicate, main factor, main-factor by replicate interaction, sub-factor, and sub-factor by main-factor interaction in the model, with F-tests for replicate and the main factor (hybrid) utilizing replicate-by-hybrid interaction for the error term.    Assessment of inoculation timing: The impact of inoculation timing treatments was evaluated separately for each hybrid; where no significant (P < 0.05) hybrid x inoculation treatment interaction occurred, it was also evaluated in the combined data across hybrids, with data from each hybrid  considered a separate replicate (for a total of 16 replicates). Analyses were conducted with replicate and treatment as main factor effects. Single-degree-of-freedom contrasts were performed for all pairwise comparisons of isolates; to control the Type I error rate at the level of the experiment. Analyses were implemented in PROC UNIVARIATE and PROC GLM of SAS (version 9.3; SAS Institute, Cary, NC).
Assessments of shattered head tissue:  Conducted immediately before harvest on September 26 at the R9 growth stage, with every plant exhibiting Sclerotinia head rot assessed for the percent of the head that was shattered (diseased head tissue often shatters and falls to the ground prior to harvest).  Strong storms in August and early September caused significant lodging in parts of the trial.  Plots were only assessed for disease and shattering if fewer than 30% of plants within the plot were lodged, and disease and shattering assessments were not taken on any lodged plants.
Harvest and seed yield and quality assessment: Sunflower heads were manually clipped at the R9 growth stage on September 26, placed in a drier, and passed through the combine on October 5.  Strong storms in August and early September caused significant lodging in parts of the trial, and plots were only harvested if fewer than 30% of plants within the plot were lodged.
Sclerotia in harvested grain:  Percent sclerotia by weight in the harvested grain prior to cleaning the grain.  From each plot, sclerotia were manually removed from a 120-gram subsample of harvested grain.
Sclerotia in cleaned grain:  Percent sclerotia by weight that could not be mechanically removed on the basis of size.  Harvested grain was cleaned on a mechanical shaker equipped with an upper sieve with 18/64 or 20/64 round holes, depending on the variety, and a lower sieve with 6 x 3/4 slotted holes.   From each plot, sclerotia were manually removed from the cleaned grain from an average 441-gram (maximum, 651-gram; minimum, 126-gram) subsample.
Seed quality – Dehulled seed and seeds per pound:  Percent of seeds that were dehulled.   Dehulled seeds within a subsample of 500 seeds were manually counted from each plot.  Seed weight of hulled seeds.  Five hundred seeds were counted, dehulled seeds were counted and manually removed, and the number and weight of the remaining hulled seeds was used to calculate seeds per pound from each plot.
* The results suggest that, to be effective, fungicides will probably need to be applied at bloom initiation when conditions favorable to disease occur.
Test weight and yield:  After the grain was mechanically cleaned, all remaining sclerotia were manually removed from the samples utilized to evaluate the test weight.  After the grain was mechanically cleaned, the percent contamination of the cleaned grain with sclerotia was assessed on the cleaned grain, and plot-level yields were proportionally adjusted such that reported yields excluded contamination with sclerotia.
Statistical analysis: Data were evaluated with analysis of variance.  Assumptions of ANOVA: (1) The assumption of constant variance was assessed with Levene's test for homogeneity of variances and visually confirmed by plotting residuals against predicted values.  (2) The assumption of normality was assessed the Shapiro-Wilk test and visually confirmed with a normal probability plot.   (3) The assumption of additivity of main-factor effects across replicates (no replicate-by-treatment interaction) was evaluated with Tukey's test for nonadditivity. To meet model assumptions, a systematic natural-log transformation was applied to the dehulled seed data.  All other data met model assumptions.
Assessment of treatment differences:  Analyses were conducted with replicate and treatment as main factor effects.  Single-degree-of-freedom contrasts were performed for all pairwise comparisons of isolates; to control the Type I error rate at the level of the experiment, the Tukey multiple comparison procedure was employed.  Analyses were implemented in PROC UNIVARIATE and PROC GLM of SAS (version 9.4; SAS Institute, Cary, NC).
FUNDED BY THE USDA NATIONAL SCLEROTINIA INITIATIVE
HYBRIDS FOR WHICH DATA WERE COLLECTED FROM (MINIMUM-3 REPLICATES):  Plots exhibiting severe lodging (>30% lodged) were not assessed.
SUMMARY TABLE - oilseed sunflowers, Oakes (2016).
				Sclerotinia Head Rot						Shattering - diseased heads						Weighted Sclerotinia		Adjusted Sclerotinia		Sclerotia in		Sclerotia in					Seeds per pound
				Incidence		Severity		Sev. Index		Incidence		Severity		Sev. Index		head rot index		head rot index		harvested		cleaned		Dehulled					Test weight		Yield
Entry	Type	Company	Entry	Sept. 19-21 (R8-R9 growth stage)						Sept. 25 (R9 growth stage)						Sept. 25 (R9)		Sept. 25 (R9)		grain		grain		seed		Oil	--------------------  10% moisture  --------------------
Number	Commercial / experimental		Name	%		%		%		%		%		%		0 to 100		0 to 100		% by weight		% by weight		%		%	seeds		bu/ac		lbs/ac
1	Resistant Check	Croplan	343 DMR HO	29	ab*	100	a*	29	ab*	65	bcd*	78	bc*	50	b-e*	22	abc*	17	a*	2	a*	1	a*	0	a*‡	-	7831	abc*	34.4	a*	1652	a*
4	Experimental	Syngenta	NX64189	63	cd	100	a	63	cd	37	ab	64	abc	23	abc	39	cd	57	cd	10	bc	6	bc	10	de	-	8593	a-d	30.3	d	1245	ab
6	Experimental	Syngenta	NX54267	14	a	100	a	14	a	55	bcd	80	bc	43	bcd	10	a	8	a	2	a	1	a	2	abc	-	9285	cd	32.1	bcd	1721	a
7	Commercial	Syngenta	SY7919	23	ab	96	a	22	ab	43	abc	78	abc	35	a-d	15	abc	16	a	2	a	1	ab	4	a-d	-	10000	de	31.5	cd	1948	a
8	Experimental	NuSeed	EXP6561	63	cd	100	a	63	cd	64	bcd	86	c	55	cde	49	de	44	bc	10	bc	6	bc	6	bcd	-	9640	cde	33.6	abc	1080	ab
9	Experimental	NuSeed	EXP8962	44	bc	100	a	44	bc	65	bcd	90	c	57	cde	37	bcd	23	ab	4	a	2	ab	2	abc	-	9149	bcd	32.9	a-d	1956	a
14	Experimental	NuSeed	EXP1157	32	ab	100	a	32	ab	77	cd	91	c	71	de	30	a-d	7	a	4	a	1	ab	1	ab	-	7425	ab	34.4	ab	1302	ab
16	Experimental	NuSeed	EXP2570	76	de	100	a	75	de	12	a	40	a	5	a	39	cd	75	de	13	c	8	cd	16	e	-	10894	ef	25.9	e	1188	ab
19	Experimental	NuSeed	EXP2548	89	e	99	a	89	e	66	bcd	80	bc	52	cde	68	e	81	e	21	d	11	d	5	cde	-	7302	a	25.5	e	806	b
20	Experimental	NuSeed	EXP6938	82	de	100	a	82	de	38	abc	71	abc	27	abc	52	de	78	de	15	c	8	cd	5	bcd	-	7406	a	25.7	e	1130	ab
23	Commercial	NuSeed	COBALT II	72	cde	100	a	72	cde	92	d	91	c	83	e	66	e	31	ab	6	ab	4	ab	3	a-d	-	9743	de	34.1	ab	628	b
24	Commercial	NuSeed	FALCON	20	ab	95	a	20	ab	26	ab	63	ab	15	ab	11	ab	18	ab	3	a	1	ab	2	a-d	-	11731	f	34.9	a	1499	ab
			F:	23.56		1.88		24.06		8.81		5.24		10.63		16.25		31.29		27.63		18.60		8.72			20.16		55.71		5.54
			P>F:	< 0.0001		0.0851		< 0.0001		< 0.0001		0.0002		< 0.0001		< 0.0001		< 0.0001		< 0.0001		< 0.0001		< 0.0001			< 0.0001		< 0.0001		0.0002
			CV:	19.6		2.2		19.4		27.5		20.1		31.8		24.7		23.7		27.3		35.5		31.6			6.6		2.9		25.1
HYBRIDS THAT WERE EXCLUDED FROM THE ANALYSIS DUE TO SEVERE LODGING:
HYBRIDS FOR WHICH DATA WERE COLLECTED FROM (MINIMUM-3 REPLICATES):  Plots exhibiting severe lodging (>30% lodged) were not assessed.
SUMMARY TABLE - oilseed sunflowers, Oakes (2016).
				Sclerotinia Head Rot						Shattering - diseased heads						Weighted Sclerotinia		Adjusted Sclerotinia		Sclerotia in		Sclerotia in
				Incidence		Severity		Sev. Index		Incidence		Severity		Sev. Index		head rot index		head rot index		harvested		cleaned		Dehulled			Seeds/pound		Test weight		Yield
Entry	Type	Company	Entry	Sept. 19-21 (R8-R9 growth stage)						Sept. 25 (R9 growth stage)						Sept. 25 (R9)		Sept. 25 (R9)		grain		grain		seed		Oil	--------------------  10% moisture  --------------------
Number	Commercial / experimental		Name	%		%		%		%		%		%		0 to 100		0 to 100		% by weight		% by weight		%		%	seeds		bu/ac		lbs/ac
2	Susceptible Check	Croplan	305 DMR NS	----------   NO DATA (all replicates were severely lodged)   ---------												----------------------------------------------   NO DATA (all replicates were severely lodged)   ---------------------------------------------------
3	Experimental	Syngenta	NX64288	67		100		67		18		100		18		39		62		8		4		5		0	8817		33		1276
5	Experimental	Syngenta	NX64290	88		100		88		63		84		53		64		77		15		11		7		0	11446		28		592
10	Experimental	NuSeed	EXP3712	17		100		17		63		75		47		13		10		4		1		1		0	10776		30		1931
11	Experimental	NuSeed	EXP6912	12		100		12		100		80		80		11		3		0		1		0		0	10319		34		1688
12	Experimental	NuSeed	EXP0757	82		100		82		96		93		89		78		33		45		25		20		0	9935		No Data		189
13	Experimental	NuSeed	EXP0857	31		100		31		40		83		34		21		23		7		5		4		0	10540		31		1142
15	Experimental	NuSeed	EXP5457	----------   NO DATA (all replicates were severely lodged)   ---------												----------------------------------------------   NO DATA (all replicates were severely lodged)   ---------------------------------------------------
17	Experimental	NuSeed	EXP2577	68		100		68		55		75		42		48		56		16		12		13		0	12162		28		575
18	Experimental	NuSeed	EXP3331	76		100		76		56		68		38		53		66		16		10		6		0	10500		27		775
21	Experimental	NuSeed	EXP8912	----------   NO DATA (all replicates were severely lodged)   ---------												----------------------------------------------   NO DATA (all replicates were severely lodged)   ---------------------------------------------------
22	Commercial	NuSeed	CAMARO II	44		100		44		91		96		88		40		15		4		2		1		0	9329		35		1103
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