Background: Witches’ broom disease (WBD) of cacao (Theobroma cacaoL.), caused byMoniliophthora perniciosa, is the most important limiting factor for the cacao production in Brazil. Hence, the development of cacao genotypes with durable resistance is the key challenge for control the disease. Proteomic methods are often used to study the interactions between hosts and pathogens, therefore helping classical plant breeding projects on the development of resistant genotypes. The present study compared the proteomic alterations between two cacao genotypes standard for WBD resistance and susceptibility, in response toM. perniciosainfection at 72 h and 45 days post-inoculation; respectively the very early stages of the biotrophic and necrotrophic stages of the cacao xM. perniciosainteraction.Results: A total of 554 proteins were identified, being 246 in the susceptible Catongo and 308 in the resistant TSH1188 genotypes. The identified proteins were involved mainly in metabolism, energy, defense and oxidative stress. The resistant genotype showed more expressed proteins with more variability associated with stress and defense, while the susceptible genotype exhibited more repressed proteins. Among these proteins, stand out pathogenesis related proteins (PRs), oxidative stress regulation related proteins, and trypsin inhibitors. Interaction networks were predicted, and a complex protein-protein interaction was observed. Some proteins showed a high number of interactions, suggesting that those proteins may function as cross-talkers between these biological functions.Conclusions: We present the first study reporting the proteomic alterations of resistant and susceptible genotypes in theT. cacaoxM. perniciosapathosystem. The important altered proteins identified in the present study are related to key biologic functions in resistance, such as oxidative stress, especially in the resistant genotype TSH1188, that showed a strong mechanism of detoxification. Also, the positive regulation of defense and stress proteins were more evident in this genotype. Proteins with significant roles against fungal plant pathogens, such as chitinases, trypsin inhibitors and PR 5 were also identified, and they may be good resistance markers. Finally, important biological functions, such as stress and defense, photosynthesis, oxidative stress and carbohydrate metabolism were differentially impacted withM. perniciosainfection in each genotype.

Six months-old seminal plants of 36 cacao genotypes grown under greenhouse conditions were subjected to two soil water regimes (control and drought) to assess, the effects of water deficit on growth, chemical composition and oxidative stress. In the control, soil moisture was maintained near field capacity with leaf water potentials (ΨWL) ranging from −0.1 to −0.5 MPa. In the drought treatment, the soil moisture was reduced gradually by withholding additional water until ΨWL reached values of between −2.0 to −2.5 MPa. The tolerant genotypes PS-1319, MO-20 and MA-15 recorded significant increases in guaiacol peroxidase activity reflecting a more efficient antioxidant metabolism. In relation to drought tolerance, the most important variables in the distinguishing contrasting groups were: total leaf area per plant; leaf, stem and total dry biomass; relative growth rate; plant shoot biomass and leaf content of N, Ca, and Mg. From the results of these analyses, six genotypes were selected with contrasting characteristics for tolerance to soil water deficit [CC-40, C. SUL-4 and SIC-2 (non-tolerant) and MA-15, MO-20, and PA-13 (tolerant)] for further assessment of the expression of genes NCED5, PP2C, psbA and psbO to water deficit. Increased expression of NCED5, PP2C, psbA and psbO genes were found for non-tolerant genotypes, while in the majority of tolerant genotypes there was repression of these genes, with the exception of PA-13 that showed an increased expression of psbA. Mutivariate analysis showed that growth variables, leaf and total dry biomass, relative growth rate as well as Mg content of the leaves were the most important factor in the classification of the genotypes as tolerant, moderately tolerant and sensitive to water deficit. Therefore these variables are reliable plant traits in the selection of plants tolerant to drought.

Flooding is common in lowlands and areas with high rainfall or excessive irrigation. A major effect of flooding is the deprivation of O₂ in the root zone, which affects several biochemical and morphophysiological plant processes. The objective of this study was to elucidate biochemical and physiological characteristics associated with tolerance to O₂ deficiency in two clonal cacao genotypes. The experiment was conducted in a greenhouse with two contrasting clones differing in flood tolerance: TSA-792 (tolerant) and TSH-774 (susceptible). Leaf gas exchange, chlorophyll (Chl) fluorescence, chemical composition and oxidative stress were assessed during 40 d for control and flooded plants. Flooding induced a decrease in net photosynthesis, stomatal conductance and transpiration of both genotypes. In flood conditions, the flood-susceptible clone showed changes in chlorophyll fluorescence, reductions in chlorophyll content and increased activity of peroxidase and polyphenol oxidase. Flooding also caused changes in macro- and micronutrients, total soluble sugars and starch concentrations in different plant organs of both genotypes. Response curves for the relationship between photosynthetically active radiation (PAR) and net photosynthetic rate (P _N) for flooded plants were similar for both genotypes. In flood conditions, the flood-susceptible clone exhibited (1) nonstomatal limitations to photosynthesis since decreased in maximum potential quantum yield of PSII (F_v/F_m) values indicated possible damage to the PSII light-harvesting complex; (2) oxidative stress; (3) increased leaf chlorosis; and (4) a reduction in root carbohydrate levels. These stresses resulted in death of several plants after 30 d of flooding.