A diagnostic gap to fill: Development of molecular tools to distinguish the cyst-forming nematode species Heterodera carotae and Heterodera cruciferae

The carrot cyst nematode, Heterodera carotae , is an important pest causing qualitative and quantitative production losses across several countries worldwide. In culture crops, Heterodera carotae and Heterodera cruciferae , can be found in mixture and are virtually unrecognizable from a morphological point of view. However, only H. carotae is able to develop on carrot. To monitor carrot production and set up suitable H . carotae control plans, simple and reliable diagnostic methods to detect and identify these two species are needed. In this study, we developed and successfully tested two real-time PCR protocols (SYBR Green singleplex and TaqMan multiplex real-time PCRs) against both species: H . carotae and H . cruciferae. Using two specific sets of primers, we managed, whatever the methods, to distinguish the two species. Moreover, we showed a higher specificity using the TaqMan assays but a better sensitivity with the SYBR Green assays. In this study, we are proposing two useful and reliable diagnostic tools to enable farmers and scientists to rapidly detect two morphologically and genetically closely related species and supervise agricultural management. • The cyst nematode H. carotae and H. cruciferae occur in mixture in carrot fields • H. carotae and H. cruciferae are indistinguishable using the available molecular tools • Two real-time PCRs were developped to distinguish H. carotae from H. cruciferae • The two real-time PCRs were complementary in terms of sensitivity and specificity • Microsatellite loci are usefull markers to identify species-specific polymorphisms