1. Response of near isogenic sorghum lines, differing at the P locus for plant color, to grain mold and head smut fungi Deanna Funnell-Harris

2. Jeff Pedersen Plant Geneticist John Toy Agronomist Pedersen, J. F., Toy, J. J. (2001) Germination, emergence and yield of 20 plant color, seed- color near-isogenic lines of grain sorghum. Crop Science, 41, 107-110.

3. PurpleTan Red “White” There were no significant differences in mean lesion lengths when tan or purple plants (with red or white grain) were inoculated with Fusarium thapsinum or Alternaria sp. When assessing grain grown on purple or tan plants, results indicated that purple plant color may protect grain from Fusarium pathogens. Hypothesis: Purple plant color protects grain against panicle diseases. Funnell, D. L., Pedersen, J. F. (2006) Association of plant color and pericarp color with colonization of grain by members of Fusarium and Alternaria in near-isogenic sorghum lines. Plant Disease, 90, 411-418

4. Grain with white pericarps grown on tan plants has a neutral flavor, appealing appearance and higher digestibility. Grain grown on purple or red plants results in an off- color. Bach Knudsen et al., 1988 Dykes et al., 2005 Rooney & Awika, 2005

5. Louis Prom Scott Sattler Jeff Pedersen

6. Phenylalanine Cinnamic acid p-Coumaric acid p-Coumaryl Coenzyme A … P?P? Snyder and Nicholson. 1990. Science 248: 1637-1639 Lo and Nicholson. 1998. Plant Physiol. 116: 979-989 Vermeriss and Nicolson. 2008. Phenolic Compound Biochemistry 3-Deoxyanthocyanidins Anthocyanidins Anthocyanins Naringenin Lignin biosynthesis Flavonoid biosynthesis

7. Sorghum pigment genetics Plant color-two loci P and Q -P-purple plant color -p-tan plant color -Q-purplish-black -P is epistatic over Q Pericarp color-two loci R and Y -RRYY-red -rrYY-yellow -RRyy, rryy-white Plant color lines – Purple-PPQQ – Tan-ppQQ – Grain-RRyy PurpleTan White grain Rooney. 2000. Pages 261 – 307 In Sorghum: Origin, History, Technology and Production Zanta et al. 1994. Journal of Heredity, 85, 23 - 29

8. Experiment Ten near-isogenic lines, 5 producing the purple/red pigments (“purple”), 5 not (“tan”) all with white grain. Planted in 4 replica plots 2 (Lincoln and Ithaca, NE) or 3 (Corpus Christi, TX) years. Collected grain, assessed for grain mold fungi. At CC assessed for incidence of head smut symptoms and signs. Funnell-Harris, D. L., Prom, L. K., Sattler, S. E. and Pedersen, J. F. 2013. Ann. Appl. Biol., 163, 91-101

9. Grain mold fungi Alternaria A. alternata A. tenuissima Fusarium G. fujikuroi Other Fusarium Cochliobolus spp. Curvularia lunata Cu. sorghina Bipolaris sorghicola Kindly provided by S. Navi Little and Magill. 2003. Physiol. Mol. Plant Pathol. 63: 271-279. Funnell-Harris et al. 2010. Phytopathology 100: 671-681 Funnell-Harris et al. 2013. Can. J. Microbiol. 59: 87-96

10. Mean numbers and standard errors of Alternaria spp. colonies per 100 grains, by selection on DCPA or DRBC media, obtained from grain grown on purple or tan sorghum plants at three locations. y Fungal groupMedium z LocationPlant color IthacaLincolnCorpus ChristiPurpleTan Total Alternaria spp. DCPA51.7 ± 14.351.5 ± 14.246.2 ± 14.450.7 ± 8.749.1 ± 8.8 P = 0.951P = 0.749 DRBC53.9 ± 14.953.1 ± 14.845.9 ± 14.951.4 ± 8.850.5 ± 8.8 P = 0.920P = 0.786 A. alternataDCPA35.1 ± 13.038.9 ± 13.034.0 ± 13.137.8 ± 7.734.2 ± 7.7 P = 0.963P = 0.317 DRBC37.9 ± 13.740.7 ± 13.733.1 ± 13.836.8 ± 8.037.6 ± 8.0 P = 0.927P = 0.777 y Least squares means were compared within location or plant color, for medium selected upon z Grains were screened for fungal growth by plating onto the media, dichloran chloramphenicol peptone agar (DCPA) and dichloran, rose bengal, chloramphenical (DRBC) agar. Numbers of Alternaria isolation per 100 grains: no significant differences Funnell-Harris, D. L., Prom, L. K., Sattler, S. E. and Pedersen, J. F. 2013. Ann. Appl. Biol., 163, 91-101

11. Location effects and medium × plant color interactions on colonization of grain by Cochliobolus spp. Mean numbers and standard errors of Cochliobolus spp. colonies per 100 grains, as determined by selection on DCPA or DRBC media, obtained from grain grown on purple or tan sorghum plants at three locations. y Medium z LocationPlant color IthacaLincolnCorpus ChristiPurpleTan DCPA0.3 ± 0.3 3.2 ± 1.10.5 ± 0.52.0 ± 0.5 P = 0.058P = 0.052 DRBC0.3 ± 0.81.5 ± 0.84.3 ± 0.82.9 ± 0.81.2 ± 0.5 P = 0.010P = 0.063 y Least squares means were compared within location or plant color, for medium selected upon. z Grains were screened for fungal growth by plating onto the media, dichloran chloramphenicol peptone agar (DCPA) and dicholoran, rose bengal, chloramphenical agar (DRBC). Funnell-Harris, D. L., Prom, L. K., Sattler, S. E. and Pedersen, J. F. 2013. Ann. Appl. Biol., 163, 91-101 Funnell-Harris, D. L., Prom, L. K., and Pedersen, J. F. 2013. Can. J. Microbiol., 59, 87-96

12. Location effects on colonization of grain by Fusarium species but no effects of plant color Mean numbers and standard errors of Fusarium spp. per 100 grains, by selection on DCPA, DRBC and PCNB media, obtained from grain grown on purple or tan sorghum plants at three locations w. Medium x LocationPlant color IthacaLincolnCorpus ChristiPurpleTan DCPA1.3 ± 1.21.3 ± 1.05.7 ± 1.83.4 ± 1.02.1 ± 1.0 P = 0.204P = 0.272 DRBC3.7 ± 2.71.1 ± 2.66.0 ± 2.93.9 ± 1.83.3 ± 1.7 P = 0.721P = 0.237 PCNB0.3 ± 0.30.7 ± 0.43.3 ± 0.91.8 ± 0.51.0 ± 0.5 P = 0.015P = 0.253 w Least squares means were compared within location or plant color, for the fungal group and medium. x Grains were screened for fungal growth by plating onto the media, dichloran chloramphenicol peptone agar (DCPA), dicholoran, rose bengal, chloramphenical agar (DRBC) and pentachloronitrobenzene agar (PCNB). Funnell-Harris, D. L., Prom, L. K., Sattler, S. E. and Pedersen, J. F. 2013. Ann. Appl. Biol., 163, 91-101

13. Two pathogenic species were detected in grain grown at all three locations. Known Fusarium sorghum pathogens isolated from grain grown on purple or tan plants, at three locations, identified using a molecular identification technique. LocationSpecies y Purple plant colorTan plant color % similarityn n Ithaca, NEF. proliferatum9911001 F. thapsinum993 4 Lincoln, NEF. andiyazi1001Not tested F. proliferatum99-1002 F. thapsinum993 Corpus Christi, TXF. proliferatum9911001 F. thapsinum997 4 F. verticillioides1001n/a0 y The 5’ region of the translation elongation factor gene (TEF) was amplified and sequenced and compared with sequences of type isolates in the FUSARIUM-ID database (http://isolate.fusariumdb.org/index.php).http://isolate.fusariumdb.org/index.php F. thapsinum Funnell-Harris, D. L., Prom, L. K., Sattler, S. E. and Pedersen, J. F. 2013. Ann. Appl. Biol., 163, 91-101 Base changes in TEF F. proliferatum

14. Conclusion-grain mold Grain from purple plants was not more resistant to the three grain mold fungal genera than grain grown on tan plants.

15. Sorghum head smut (Sporisorium reilianum) Frederiksen. 2002. Pages 18-20 In “Compendium of Sorghum Diseases”

16. At Corpus Christi, purple plants had greater head smut incidence than tan plants Mean incidence (percent plants affected) of head smut in plants near-isogenic for plant color Plant colorPercent head smut Purple14.8* Tan6.8 SE for means are 2.4. Means are significantly different at P < 0.001.

17. Phenylalanine Cinnamic acid p-Coumaric acid p-Coumaryl Coenzyme A Lignin biosynthesis … P?P? Snyder and Nicholson. 1990. Science 248: 1637-1639 Lo and Nicholson. 1998. Plant Physiol. 116: 979-989 Vermeriss and Nicolson. 2008. Phenolic Compound Biochemistry Pedersen and Toy. 2001. Crop Sci. 41: 107 - 110 Cui et al. 1996. Physiol. Mol. Plant Pathol. 49: 187-199. Halkier and Moller. 1989. Plant Phys. 90: 473-479. Phenylalanine ammonia lyase Chalcone synthase ? 3-Deoxyanthocyanidins Anthocyanidins Anthocyanins Naringenin ? Flavonoid biosynthesis

18. Conclusions The near-isogenic purple and tan lines with white grain are potentially valuable to investigate other defenses of sorghum besides the well-characterized 3-deoxyanthocyanidin phytoalexins. The tan lines would be ideal to develop food- grade sorghum hybrids, especially in combination with lines having race-specific resistance to sorghum head smut.

19. Deanna Funnell-Harris Plant pathologist Scott Sattler Plant molecular biologist USDA-ARS Sorghum project, Lincoln, NE Melinda Yerka Plant geneticist

20. National Institute of Food & Agriculture, grant 2011-67009- 30026