Frédéric Breton

Possible sources of genetic resistance in oil palm (Elaeis guineensis Jacq.) to basal stem rot caused by Ganoderma boninense - prospects for future breeding.

Oil palm estates in southeast Asia suffer from substantial losses due to basal stem rot caused by Ganoderma boninense. Field observations have been carried out in North Sumatra, Indonesia, on a series of planting materials of known origin. Differences in susceptibility to the disease have been detected within the two Elaeis species, guineensis and oleifera. Within Elaeis guineensis, material of Deli origin is highly susceptible compared to material of African origin. It is also possible to detect differences in reaction between parents and between crosses within a given origin. The variability of resistance to basal stem rot within the same cross is also illustrated by the diverse responses of clones derived from palms of the same origin. The prospects opened up by these results are discussed, and the importance of performing an early selection test is highlighted.

Scopoletin production and degradation in relation to resistance of Hevea brasiliensis to Corynespora cassiicola

Summary Inoculation of leaves of resistant and susceptible Hevea brasiliensis clones with Corynespora cassiicola induced foliar necrosis and biosynthesis of scopoletin (Scp), considered as a Hevea phytoalexin. Foliar symptoms appeared, as soon as 24 h after infection, and precede Scp accumulation in inoculum droplets (peak: 48 h after inoculation). Scp concentration was 5-fold higher in the susceptible than in the resistant clone 48 h after infection. Nevertheless, a fungitoxic effect of Scp on spore germination and on mycelium growth was shown in bioassays, but the efficient concentrations were greater than those found with another foliar pathogenic fungus of rubber tree such as Microcyclus ulei and Colletotrichum gloeosporioides. Bioassay showed that the low Scp fungitoxicity to C. cassiicola could also be related to the ability of fungus to detoxify Sep. Isoelectric focusing analysis of peroxidase activity in Hevea infected leaflets has shown an increase in acidic and basic isoperoxidases that are able to use Scp as substrate. The rapid increase of Scp-oxidase activity (as soon as 16 h after inoculation) was higher in the resistant than in the susceptible clone. In vitro testing of Scp peroxidation products has shown a toxic effect on C. cassiicola (conidia germination and mycelium growth), but not significantly higher than Scp. It appears that the level of Sep accumulation was a balance between its synthesis and its degradation by the pathogen or/and by foliar Scp-peroxidases. In conclusion, these results lead to the fact that Scp cannot be considered as a major defense mechanism of Hevea towards C. cassiicola disease.

Unfolding and refolding of active apple polyphenol oxidase

For the first time, unfolding (6 M guanidine) and refolding of partially proteolysed purified polyphenol oxidase (PPOr) was achieved, with 88% of activity recovered. Optimal refolding conditions consisted in stepwise dialysis of guanidine treated extracts, the dialysis buffers containing 1 M (NH4)2SO4 and 100 microM CuSO4. However, CuSO4 had limited effect on the recovering of PPOr activity, whereas (NH4)2SO4 was essential. Concerning the PPO tertiary structure, denaturing conditions (combinations of boiling and reducing agent) used on SDS-PAGE have shown (i) a compact tertiary structure and (ii) the presence of disulfide bonds in PPOr, accounting for the shift between 27 and 41 kDa, and 41 and 42 kDa, respectively. Resistance to proteolytic cleavage was used to study the conformational changes induced by the denaturing treatments. Folded PPOr was resistant to further proteolysis whereas unfolded PPO was totally digested, indicating the role of tertiary structure of PPOr in the resistance to proteases.

Identification and development of new polymorphic microsatellite markers using genome assembly for Ganoderma boninense, causal agent of oil palm basal stem rot disease

Ganoderma boninense is a telluric lignicolous basidiomycete and the causal agent of basal stem rot, one of the most devastating diseases of the oil palm (Elaeis guineensis). While the fight against G. boninense is of major concern in Southeast Asia, little information is available regarding the genetic diversity and evolutionary history of the fungus. In this context, the development of an informative molecular marker set to characterize the diversity of G. boninense is a key step towards understanding the biology of this pathogen. A G. boninense draft genome sequence of 63 Mbp, assembled using 454 and Illumina sequencing technology, was used to identify and develop a set of microsatellite markers (simple sequence repeats, SSRs). A total of 2487 SSRs were identified, for which 145 SSR primer pairs were designed. These SSRs are characterized by di- to hexanucleotide motifs with 5 to 34 repetitions. Ninety-seven SSR loci were successfully amplified on an initial small set of G. boninense isolates from Indonesia. A collection of 107 isolates from several regions in Southeast Asia were screened to characterize each locus for allele number, polymorphism information criterion and the presence or absence of null alleles at each locus. These results allowed us to propose an effective set of 17 SSRs for studying genetic diversity within G. boninense.

Les composés phénoliques et la résistance des plantes aux agents pathogènes

Summary Plant defense responses against microorganisms require many mechanisms as constitutive phenolic compounds and phytoalexins. These secondary metabolic products can be accumulated and involved in plant defence. Their activity was related to antimicrobial properties, involvement in cell wall reinforcement, modulation and induction of plant responses. Clonage of most genes coding for enzymes of phenolic metabolism give many means for genetic manipulations aiming to improve the effectiveness of plant defences. Recent results were very interesting but do not yet allow to define the real level of the involvement of phenolic compounds in plant disease resistance.

About Ganoderma boninense in oil palm plantations of Sumatra and peninsular Malaysia: Ancient population expansion, extensive gene flow and large scale dispersion ability

Wood rot fungi form one of the main classes of phytopathogenic fungus. The group includes many species, but has remained poorly studied. Many species belonging to the Ganoderma genus are well known for causing decay in a wide range of tree species around the world. Ganoderma boninense, causal agent of oil palm basal stem rot, is responsible for considerable yield losses in Southeast Asian oil palm plantations. In a large-scale sampling operation, 357 sporophores were collected from oil palm plantations spread over peninsular Malaysia and Sumatra and genotyped using 11 SSR markers. The genotyping of these samples made it possible to investigate the population structure and demographic history of G. boninense across the oldest known area of interaction between oil palm and G. boninense. Results show that G. boninense possesses a high degree of genetic diversity and no detectable genetic structure at the scale of Sumatra and peninsular Malaysia. The fact that few duplicate genotypes were found in several studies including this one supports the hypothesis of spore dispersal in the spread of G. boninense. Meanwhile, spatial autocorrelation analysis shows that G. boninense is able to disperse across both short and long distances. These results bring new insight into mechanisms by which G. boninense spreads in oil palm plantations. Finally, the use of approximate Bayesian computation (ABC) modelling indicates that G. boninense has undergone a demographic expansion in the past, probably before the oil palm was introduced into Southeast Asia.