Development of a new in vivo protocol through soil inoculation to investigate sugar beet resistance towards Ditylenchus dipsaci penetration

The stem nematode,Ditylenchus dipsaci, causes severe damage in sugar beet. To date, nematode inoculation throughthe leaf axil has been used as the standard method to investigateD. dipsaciinteraction with sugar beet underin vivoconditions. Toget as close as possible to field conditions, we established a new screening mechanism to perform soil inoculation. The most suitableinoculation time point, inoculum level and positioning on sugar beet, as well as rearing process on carrots, were determined. At a15:8°C day:night temperature regime, penetration rates ofD. dipsaciwere at maximum following soil inoculation at plant emergence.Up to 115 nematodes penetrated sugar beet seedlings 22 days post-planting with an inoculum level of 1000 nematodes into the soil atplant emergence.Ditylenchus dipsacipenetration rate was higher in plants with soil inoculation than with inoculation on to the leaf axil.High soil moisture increased nematode migration into seedlings whenD. dipsaciinoculation was carried out in four holes 1 cm fromthe plant base. Rearing the nematodes for 35 days at 20°C on carrot discs resulted in an infective inoculum containing up to 50% eggs.We recommend a soil inoculation of 1000 freshly extracted nematodes per pot at plant emergence. The nematode suspension has to bepreviously reared for 35 days on carrot discs to obtain activeD. dipsaciinoculum. This system will allow for the selection of suitablesugar beet genotypes that suppress nematode penetration, in support of breeding for resistance againstD. dipsaci.