Odile Huchette, research engineer Christèle Bellamy, research technician Didier Perrin and Eric Vade, research assistants G&H February 2003, Liverpool WP3 / cultivation, P4 P4 : persons involved COOPD'OR R&D Influence of environmental and genetic factors on the alliin content of garlic bulbs

G&H February 2003, Liverpool WP3 / cultivation, P4

–Greenhouse experiment in various S regimes(P4) Sampling of fresh material during the cultivation treatment for CSO analysis ( P5) Freeze drying of samples (P4) for total sulfur analysis (P2) –Micro-propagation of 4 genotypes for new experiments in vitro (P4) Experiment III (October 2002 => April 2003) : Differential display of clones exposed in vitro to various light conditions (intensity + quality) and S nutrition treatments (0 S in macro-nutrients, normal and excess) Experiment IV (January 2003 => July 2003) : Differential display of clones exposed in vitro to various mineral nutrition treatments (S combined with N) COOPD'OR R&D G&H February 2003, Liverpool WP3 / cultivation, P4 Year 3 : what was done

Greenhouse conditions 2001 PRI G&H February 2003, Liverpool WP3 / cultivation, P4 Plantation in boxes with washed river sand Mineral nutrition with drop irrigation Greenhouse conditions 2002

G&H February 2003, Liverpool WP3 / cultivation, P4 Greenhouse experiment 2001 COOPD'OR R&D

3 varieties : PRI, MOR, MES 3 Sulphur nutrition treatments: 0 (in the macro-elements) = Treatment S- 2 meq (normal) = Treatment TS 8 meq = Treament S++ G&H February 2003, Liverpool WP3 / cultivation, P4 Greenhouse experiment 2002 COOPD'OR R&D Weekly observations :Growing of the leaves and bulbs Measurements at harvest : - Weight of the bulbs - alliin content (P5) - total sulfur (P2) in progress

G&H February 2003, Liverpool WP3 / cultivation, P4 No differences during the growing season between the 3 treatments Bulb growing = diameter of the bulb / diameter of the neck 01/03 : Plantation 17/06 : Harvest of MES 10-11/07 : Harvest of MOR 16-17/07 : Harvest of PRI

G&H February 2003, Liverpool WP3 / cultivation, P4 Results at harvest COOPD'OR R&D  influence of the variety  no significative influence of the S treatment, except for MES

–Alliin contents in bulbs are 3 to 4 times higher in 2002 compared to 2001 –Printanor appeared in 2002 as the richest variety in alliin, contrary to the previous results –Garlic plants are not affected by an absence of sulfur in the macro-nutrients absorption of S atmospheric to be confirmed in vitro in 2003 –Garlic plants are not significatively affected by the sulfur nutrition alone, except for one variety tested influence of the nitrogen to be confirmed in vitro and in greenhouse in 2003 COOPD'OR R&D G&H February 2003, Liverpool WP3 / cultivation, P4 Conclusion of greenhouse experiments

COOPD'OR R&D 4 genotypes exposed to various environmental factors... –quality of the light –sucrose concentration of the medium –temperature –cold induction period... and various mineral nutritions –sulfur G&H February 2003, Liverpool WP3 / cultivation, P4 In vitro experiments 2001/02

Among all the conditions tested, the better to induce more alliin in garlic bulbs : Sucrose in the medium : 60g/l [ g/l tested] Cold induction period (2 to 3 months) Environmental factor : F+i light Sulphur in the medium : 6 Meq SO 3 [ meq tested] G&H February 2003, Liverpool WP3 / cultivation, P4 Main results of in vitro experiments COOPD'OR R&D To be confirmed in 2003 : Influence of the intensity of the light Effect of higher dosis of sulphur (9 meq) Effect of nitrogen combined to sulphur Role of S atmospheric

G&H February 2003, Liverpool WP3 / cultivation, P4 Year 4 : what is planed COOPD'OR R&D

G&H February 2003, Liverpool WP3 / cultivation, P4 In vitro experiment III (in macro-elements) Gradient inSO4 meq 22-24°C - P = 16 hrs - F 1month 22-24°C – P=16 hrs F 950 lux 3mediums (S- TS S++) 3 varieties : PRI + MES + MORl Cold induction3°C - P = 10 hrs - F 22-24°C – P=16 hrs F+i lux 22-24°C – P=16 hrs F 2000 lux 2months Effect of various light conditions (intensity + quality) and S nutrition

G&H February 2003, Liverpool WP3 / cultivation, P4 In vitro experiment IV SO Gradient inSO4 and nitrogen meq (in macro-elements) 22-24°C - P = 16 hrs - F 1month 9 9 medias varieties : PRI + MES + MORl + MORd Cold induction 3°C - P = 10 hrs - F 22-24°C – P=16 hrs F+i lux 2months Nitrogen N 3N N 2N 3N N/2 N 2N 3N Effect of S nutrition combined to N nutrition

Observations : - stage of the plant (1= growing / 2=beginning of bulbification / 3=maturation / 4=mature bulb) - number of green leaves - height of the leaves Measurements at harvest : - weight, size and observation of the bulbs - height of the leaves - alliin content (P5) G&H February 2003, Liverpool WP3 / cultivation, P4 In vitro experiments 2003 COOPD'OR R&D

PRI F °C P = 16h S- TS S++ G&H February 2003, Liverpool WP3 / cultivation, P4 In vitro experiment III-2003  Observation of the 03 February COOPD'OR R&D

3 varieties and 3 treatments : -S ++ N = excess of sulfur, N -S ++ 2N = excess of sulfur, 2xN -S ++ 3N = excess of sulfur, 3xN (I) Effect of N combined to an excess of S (II) Cloves from experiment 2002 Measurements : - growing of the leaves (I-II) and bulbs (I) - weight and CSO content of the bulbs (P5) at harvest (I-II) G&H February 2003, Liverpool WP3 / cultivation, P4 Greenhouse experiment 2003

G&H February 2003, Liverpool WP3 / cultivation, P4 Effect of virus contamination on AlCSOs in garlic bulbs COOPD'OR R&D

G&H February 2003, Liverpool WP3 / cultivation, P4 Deliverables P4 DP 2  Chinese garlic powder to be analysed DP 6  Garlic powder issued from various S-fertilisation in field condition DP 14  Chemical distinction between garlic accessions when cultivated in various sulphur nutrition regimes and under distinct temperatures in vitro DP 15  Physiological (growth traits) and chemical distinction (CSO) between accessions when cultivated in various sulphur nutrition regimes over two years DP 19  Chemical distinction (CSO) between accessions when cultivated in various sulphur (greenhouse) and mineral nutrition regimes (in vitro) DP 22 Paper on the interaction genotype x environment on the sulphur content of garlic plants grown in vitro, in greenhouse and in field conditions DP 30 Paper on the effect of combined sulphur, nitrogen and selenium nutrition on garlic growth, flavour precursor content and on biological potential of garlic for disease prevention DP 34 Chemical distinction (CSO) between garlic accessions when cultivated in various sulphur regimes (greenhouse) and mineral nutrition regimes (in vitro)

2 varieties : Morasol, Printanor 3 treatments : S- : no sulphur in the macro-nutrients TS : 2 meq S in the fertilising solution S++ : 8 meq S in the fertilising solution G&H February 2003, Liverpool WP3 / cultivation, P4 garlic powder available from the experiment 2002 in greenhouse COOPD'OR R&D