Inheritance of Resistance to Bacterial Halo Blight and Bacterial Leaf Spot in Arabica Coffee.

Developing coffee cultivars resistant to multiple diseases by combining resistance genes is a top priority in breeding programs. To create cultivars resistant to diseases and nematodes, we transferred genes for resistance to bacterial infections caused by Pseudomonas coronafaciens pv. garcae, which causes bacterial halo blight (BHB), and P. amygdali pv. tabaci, which causes bacterial leaf spots (BLS), into Arabica coffee. Genetic analyses were conducted on breeding populations to estimate the number and function of genes that confer resistance to BHB and BLS. In total, 2,109 plants in the F2 generation and reciprocal backcrosses were inoculated with P. coronafaciens pv. garcae, while 1,996 plants were inoculated with P. amygdali pv. tabaci. Results showed that resistance to both pathogens had a heritability of 0.99, and the segregations of resistance indicated that each disease was controlled by a single dominant gene. The analyses also revealed that the resistance genes for BHB and BLS were linked, with an average distance of 10.75 cM between them on the same chromosome.

The Rhizosphere Microbiomes of Five Species of Coffee Trees.

Coffee is one of the most important commodities in the global market. Of the 130 species of Coffea, only Coffea arabica and Coffea canephora are actually cultivated on a large scale. Despite the economic and social importance of coffee, little research has been done on the coffee tree microbiome. To assess the structure and function of the rhizosphere microbiome, we performed a deep shotgun metagenomic sequencing of the rhizospheres of five different species, C. arabica, C. canephora, Coffea stenophylla, Coffea racemosa, and Coffea liberica. Our findings indicated that C. arabica and C. stenophylla have different microbiomes, while no differences were detected between the other Coffea species. The core rhizosphere microbiome comprises genera such as Streptomyces, Mycobacterium, Bradyrhizobium, Burkholderia, Sphingomonas, Penicillium, Trichoderma, and Rhizophagus, several of which are potential plant-beneficial microbes. Streptomyces and mycorrhizal fungi dominate the microbial communities. The concentration of sucrose in the rhizosphere seems to influence fungal communities, and the concentration of caffeine/theobromine has little effect on the microbiome. We also detected a possible relationship between drought tolerance in Coffea and known growth-promoting microorganisms. The results provide important information to guide future studies of the coffee tree microbiome to improve plant production and health. IMPORTANCE The microbiome has been identified as a fundamental factor for the maintenance of plant health, helping plants to fight diseases and the deleterious effects of abiotic stresses. Despite this, in-depth studies of the microbiome have been limited to a few species, generally with a short life cycle, and perennial species have mostly been neglected. The coffee tree microbiome, on the other hand, has gained interest in recent years as Coffea trees are perennial tropical species of enormous importance, especially for developing countries. A better understanding of the microorganisms associated with coffee trees can help to mitigate the deleterious effects of climate change on the crop, improving plant health and making the system more sustainable.

A metabolomic platform to identify and quantify polyphenols in coffee and related species using liquid chromatography mass spectrometry.

Introduction: Products of plant secondary metabolism, such as phenolic compounds, flavonoids, alkaloids, and hormones, play an important role in plant growth, development, stress resistance. The plant family Rubiaceae is extremely diverse and abundant in Central America and contains several economically important genera, e.g. Coffea and other medicinal plants. These are known for the production of bioactive polyphenols (e.g. caffeine and quinine), which have had major impacts on human society. The overall goal of this study was to develop a high-throughput workflow to identify and quantify plant polyphenols. Methods: First, a method was optimized to extract over 40 families of phytochemicals. Then, a high-throughput metabolomic platform has been developed to identify and quantify 184 polyphenols in 15 min. Results: The current metabolomics study of secondary metabolites was conducted on leaves from one commercial coffee variety and two wild species that also belong to the Rubiaceae family. Global profiling was performed using liquid chromatography high-resolution time-of-flight mass spectrometry. Features whose abundance was significantly different between coffee species were discriminated using statistical analysis and annotated using spectral databases. The identified features were validated by commercially available standards using our newly developed liquid chromatography tandem mass spectrometry method. Discussion: Caffeine, trigonelline and theobromine were highly abundant in coffee leaves, as expected. Interestingly, wild Rubiaceae leaves had a higher diversity of phytochemicals in comparison to commercial coffee: defense-related molecules, such as phenylpropanoids (e.g., cinnamic acid), the terpenoid gibberellic acid, and the monolignol sinapaldehyde were found more abundantly in wild Rubiaceae leaves.

Unsaponifiable matter from oil of green coffee beans: cosmetic properties and safety evaluation.

CONTEXT: Unsaponifiable matter (UM), a fraction of green coffee oil (GCO) contains functional compounds responsible for desirable cosmetic properties such as UV-B absorption. OBJECTIVES: To evaluate oil content and sun protection factor (SPF) variability of the two most important species of coffee and, the toxic and cytotoxic effects, as well as cosmetic properties, including antioxidant and antimicrobial activities of UM obtained from green Coffea arabica seed oil. MATERIALS AND METHODS: The safety and potential cosmetic properties of UM extracted from green coffee oil (GCO) were evaluated by the brine shrimp viability and the MTT cytotoxicity assays. The SPF and antioxidant activity were evaluated using in vitro methods. RESULTS: Relevant cytotoxicity was found against keratinocytes for concentrations ≥25 µg/mL and in the brine shrimp assay (LC50 24 µg/mL). Antimicrobial and antioxidant activities (IC50 1448 µg/mL) were low in UM but SPF was 10 times higher than in GCO. CONCLUSION: UM is a novel potential UV-B absorbent but its use as a cosmetic ingredient should be better considered due to the considerable cytotoxicity shown in the experimental conditions described.

Large-scale analysis of differential gene expression in coffee genotypes resistant and susceptible to leaf miner-toward the identification of candidate genes for marker assisted-selection.

BACKGROUND: A successful development of herbivorous insects into plant tissues depends on coordination of metabolic processes. Plants have evolved complex mechanisms to recognize such attacks, and to trigger a defense response. To understand the transcriptional basis of this response, we compare gene expression profiles of two coffee genotypes, susceptible and resistant to leaf miner (Leucoptera coffella). A total of 22000 EST sequences from the Coffee Genome Database were selected for a microarray analysis. Fluorescence probes were synthesized using mRNA from the infested and non-infested coffee plants. Array hybridization, scanning and data normalization were performed using Nimble Scan® e ArrayStar® platforms. Genes with foldchange values +/-2 were considered differentially expressed. A validation of 18 differentially expressed genes was performed in infected plants using qRT-PCR approach. RESULTS: The microarray analysis indicated that resistant plants differ in gene expression profile. We identified relevant transcriptional changes in defense strategies before insect attack. Expression changes (>2.00-fold) were found in resistant plants for 2137 genes (1266 up-regulated and 873 down-regulated). Up-regulated genes include those responsible for defense mechanisms, hypersensitive response and genes involved with cellular function and maintenance. Also, our analyses indicated that differential expression profiles between resistant and susceptible genotypes are observed in the absence of leaf-miner, indicating that defense is already build up in resistant plants, as a priming mechanism. Validation of selected genes pointed to four selected genes as suitable candidates for markers in assisted-selection of novel cultivars. CONCLUSIONS: Our results show evidences that coffee defense responses against leaf-miner attack are balanced with other cellular functions. Also analyses suggest a major metabolic reconfiguration that highlights the complexity of this response.

Molecular characterization of a miraculin-like gene differentially expressed during coffee development and coffee leaf miner infestation.

The characterization of a coffee gene encoding a protein similar to miraculin-like proteins, which are members of the plant Kunitz serine trypsin inhibitor (STI) family of proteinase inhibitors (PIs), is described. PIs are important proteins in plant defence against insects and in the regulation of proteolysis during plant development. This gene has high identity with the Richadella dulcifica taste-modifying protein miraculin and with the tomato protein LeMir; and was named as CoMir (Coffea miraculin). Structural protein modelling indicated that CoMir had structural similarities with the Kunitz STI proteins, but suggested specific folding structures. CoMir was up-regulated after coffee leaf miner (Leucoptera coffella) oviposition in resistant plants of a progeny derived from crosses between C. racemosa (resistant) and C. arabica (susceptible). Interestingly, this gene was down-regulated during coffee leaf miner herbivory in susceptible plants. CoMir expression was up-regulated after abscisic acid application and wounding stress and was prominent during the early stages of flower and fruit development. In situ hybridization revealed that CoMir transcripts accumulated in the anther tissues that display programmed cell death (tapetum, endothecium and stomium) and in the metaxylem vessels of the petals, stigma and leaves. In addition, the recombinant protein CoMir shows inhibitory activity against trypsin. According to the present results CoMir may act in proteolytic regulation during coffee development and in the defence against L. coffeella. The similarity of CoMir with other Kunitz STI proteins and the role of CoMir in plant development and plant stress are discussed.

Phenol contents, oxidase activities, and the resistance of coffee to the leaf miner Leucoptera coffeella.

We examined the role of phenolic compounds, and the enzymes peroxidase and polyphenol oxidase, in the expression of resistance of coffee plants to Leucoptera coffeella (Lepidoptera: Lyonetiidae). The concentrations of total soluble phenols and chlorogenic acid (5-caffeoylquinic acid), and the activities of the oxidative enzymes peroxidase (POD) and polyphenol oxidase (PPO), were estimated in leaves of Coffea arabica, C. racemosa, and progenies of crosses between these species, which have different levels of resistance, before and after attack by this insect. The results indicate that phenols do not play a central role in resistance to the coffee leaf miner. Differences were detected between the parental species in terms of total soluble phenol concentrations and activities of the oxidative enzymes. However, resistant and susceptible hybrid plants did not differ in any of these characteristics. Significant induction of chlorogenic acid and PPO was only found in C. racemosa, the parental donator of the resistance genes against L. coffeella. High-performance liquid chromatography (HPLC) analysis also showed qualitative similarity between hybrids and the susceptible C. arabica. These results suggest that the phenolic content and activities of POD and PPO in response to the attack by the leaf miner may not be a strong evidence of their participation in direct defensive mechanisms.