Transcription of Biotic Stress Associated Genes in White Clover (Trifolium repens L.) Differs in Response to Cyst and Root-Knot Nematode Infection.

The transcription of four members of the Kunitz proteinase inhibitor (KPI) gene family of white clover (Trifolium repens L.), designated as Tr-KPI1, Tr-KPI2, Tr-KPI4 and Tr-KPI5, was investigated at both local infection (roots) and systemic (leaf tissue) sites in white clover in response to infection with the clover root knot nematode (CRKN) Meloidogyne trifoliophila and the clover cyst nematode (CCN) Heterodera trifolii. Invasion by the CRKN resulted in a significant decrease in transcript abundance of Tr-KPI4 locally at both 4 days post-infection (dpi) and at 8 dpi, and an increase in transcription of Tr-KPI1 systemically at 8 dpi. In contrast, an increase in transcript abundance of all four Tr-KPI genes locally at 4 and 8 dpi, and an increase of Tr-KPI1, Tr-KPI2, and Tr-KPI5 at 8 dpi systemically was observed in response to infection with the CCN. Challenge of a resistant (R) genotype and a susceptible (S) genotype of white clover with the CCN revealed a significant increase in transcript abundance of all four Tr-KPI genes locally in the R genotype, while an increase in abundance of only Tr-KPI1, Tr-KPI2, and Tr-KPI5 was observed in the S genotype, and only at 4 dpi. The transcript abundance of a member of the1-AMINOCYCLOPROPANE-1-CARBOXYLATE (ACC) SYNTHASE gene family from white clover (Tr-ACS1) was significantly down-regulated locally in response to CRKN infection at 4 and 8 dpi and at 4 dpi, systemically, while abundance increased locally and systemically at 8 dpi in response to CCN challenge. Conversely, the abundance of the jasmonic acid (JA) signalling gene, CORONATINE-INSENSITIVE PROTEIN 1 from white clover (Tr-COI1) increased significantly at 8 dpi locally in response to CRKN infection, but decreased at 8 dpi in response to CCN infection. The significance of this differential regulation of transcription is discussed with respect to differences in infection strategy of the two nematode species.

Resistance Reactions to Meloidogyne trifoliophila in Trifolium repens and T. semipilosum.

The predominant root-knot nematode in New Zealand pastures is Meloidogyne trifoliophila, and a recurrent selection program in Trifolium repens has developed resistance to this species. No data are available, however, on the mechanisms of resistance in T. repens or resistant genotypes of T. semipilosum. The development of M. trifoliophila in roots of T. repens and T. semipilosum was examined weekly after a 2-day inoculation with eggs. More second-stage juveniles (J2) were found in two resistant genotypes of T. repens than in two susceptible ones 1 week after inoculation. J2 did not develop further in resistant genotypes, but in susceptible plants development proceeded to the adult stage, visible at 4 weeks after inoculation. The mode of action of resistance to M. trifoliophila in T. repens and in T. semipilosum was compared after a 24-hour inoculation with J2. Numbers of J2 per root tip ranged from 0 to 12 with a median of one for each species. At 24 hours after inoculation (HAI), similar numbers of J2 were seen in the cortex oriented toward the root tip in both resistant and susceptible genotypes of both plant species. At 48 HAI, accumulations of J2 were seen in the meristem in both resistant and susceptible genotypes of both plant species. At 72 HAI, differences in nematode responses were evident between resistant and susceptible genotypes of both plant species; in susceptible roots, J2 heads were embedded in the developing stele. At this time, a browning reaction in resistant genotypes of both plant species indicated a hypersensitive response, and differences in the reaction were recorded between T. repens and T. semipilosum. More study is needed to determine whether the resistance reaction in T. semipilosum is suitable for introgression or insertion into T. repens.