GENETIC CHARACTERISATION OF TOLERANCE TO FERROUS IRON TOXICITY IN WEST AFRICAN RICE CULTIVARS
Iron toxicity is one of the main constraints of lowlands rice production in West Africa. The adoption of tolerant varieties constitutes a sustainable approach to address this constraint. The objectives of this study were to: (i) assess the lowland rice production constraints, farmers’ preferred varietal traits and their perceptions of iron toxicity in the Casamance zone (Senegal) through a Participatory Rural Appraisal (PRA) approach; (ii) assess the genetic diversity among rice germplasm from West Africa including landraces from the Casamance area; (iii) identify sources of tolerance to iron toxicity in the rice genotypes; and (iv) identify candidate genes associated with tolerance to iron toxicity. The PRA was conducted in six districts of the Casamance region with data collected through focus group discussion and semi-structured questionnaire. The results indicated that salinity (30%) and iron toxicity (26%) were the main constraints and caused significant yield losses. Most of the farmers observed iron toxicity in their fields during the last five years (95%). Farmers were aware that iron toxicity was associated with the soil characteristics and flooding conditions of their lands. Grain yield (27%) and earliness (20%) were the most preferred traits in addition to tolerance to salinity and iron toxicity. The genetic diversity study with 12 Simple Sequence Repeat (SSR) markers revealed that the entire collection of 480 genotypes including the landraces from Casamance and the collection from AfricaRice which was composed mainly of accessions from the West African countries (57%), presented an intermediate genetic diversity (GD) of 0.63. Mean of the polymorphic information content (PIC) was 0.59. The collection from AfricaRice were more diverse (GD = 0.67) than the landraces (GD = 0.50) collected from 19 villages across six districts of the Casamance zones. Three main genotypic groups where identified corresponding to the Oryza glaberrima and the Oryza sativa rice species and subspecies. Two groups of indica genotypes were identified where one was mainly composed of landraces from Casamance as confirmed by the population structure analysis with 148,934 Single Nucleotide Polymorphic (SNP) resulting from imputation with 0.5% minor allele frequency (MAF). Based on the results from the genetic diversity study, a subset of 282 rice genotypes composed of both rice species were selected for the screening for tolerance to iron toxicity and the genome wide association study (GWAS). The evaluations were conducted in hydroponic solutions with 1500 mg L-1 Fe concentration for ten days and two hot spot fields in Djibelor (Senegal) and Ndeman (The Gambia). Consistency in the genetic variation in response to iron toxicity was also assessed across these different experimental conditions. Differences in the levels of tolerance were observed among the genotypes in the hydroponic screening as well as in the field evaluations. In addition to the classical agro morphological parameters, secondary traits such as chlorophyll content (CC) (SPAD value) and leaf bronzing (LB) were assessed in these experiments. Leaf bronzing had a mean score of 4 in hydroponic screening and varied between 3 and 4 in hot spot fields at Ndeman and Djibelor, respectively. Significant variation was observed among the genotypes for most of the traits recorded in the different experiments. The environment had significant effect on most of the traits. Shoot dry weight and root dry weight were reduced by 59 and 53% respectively in hydroponic conditions. Grain yield and biomass production were reduced by 56% in the field evaluation at Djibelor where the iron toxicity was more severe compared with the non-stress field experiment also implemented at Djibelor. Some stability was found in the genotypic response under the different experiments and consistency was observed in the tolerance of some genotypes. These genotypes comprised accessions from West Africa: BEN11-172, WONKIFON 17, previously identified tolerant variety: IR75887-1-3-WAB1 (ARICA 6) and landraces from Casamance: EHAUB and BANDIOUL MANO. These genotypes presented low iron content (2229 to 7518 mg kg-1) in shoots and high roots iron retention (45769 to 176930 mg kg-1) capacity. The identification of candidate genes associated with tolerance to iron toxicity through GWAS using the 148,934 SNPs was performed mainly on the indica subpopulation which was more representative in terms of diversity and population size (186 genotypes). A total of nine significant SNPs with –log10 P>4 were identified using five traits assessed in hydroponic conditions. For the field conditions, 60 significant SNPs with threshold of –log10 P>4 were detected for eight traits. The new candidate loci identified in hydroponic for SPAD value (CC) were also confirmed in the field conditions. These loci included LOC_Os08g09220 and LOC_Os08g09250, described to be putatively involved in the biosynthesis of OsFBX262- F-box domain and glyoxalase family proteins, reported to be variously regulated when rice seedlings are exposed to oxidative stresses. The locus LOC_Os07g05800 associated with harvest index and the previously reported LOC_Os07g02350, identified for leaf bronzing were reported as putatively involved in the biosynthesis of glutathione S-transferase and casein kinase II subunit alpha-2, respectively. These proteins were suggested as relevant in the tolerance of rice to iron toxicity. Further investigations of these candidate genes including transcriptomic analysis and functional characterization are necessary.