Allometric models for non-destructive leaf area estimation in coffee (Coffea arabica and Coffea canephora)

We aimed to evaluate the currently used allometric models, as well as to propose a reliable and accurate model using non-destructive measurements of leaf length (L) and/or width (W), for estimating the area of leaves of eight field-grown coffee cultivars. For model construction, a total of 1563 leaves were randomly selected from different levels of the tree canopies and encompassed the full spectrum of measurable leaf sizes (0.3–263 cm²) for each genotype. Power models better fit coffee leaf area (LA) than linear models. To validate the model, an independent data set of 388 leaves was used. We demonstrated that the currently used allometric models are biased, underestimating the area of a coffee leaf. We developed a single power model     based on two leaf dimensions [LA = 0.6626 (LW)^1.0116; standard errors: β₀ = 0.0064, β₁ = 0.0019; R² = 0.996] with high precision and accuracy, random dispersion pattern of residuals and also unbiased, irrespective of cultivar and leaf size and shape. Even when the L (but not width) alone was used as the single leaf dimension, the power model developed still predicted with good accuracy the LA but at the expense of some loss of precision, as particularly found for 8% of the leaves sampled with length-to-width ratios below 2.0 or above 3.0.     

Gene introgression into Coffea arabica by way of triploid hybrids (C. arabica x C. canephora)

Interspecific triploid hybrid plants between the tetraploid species Coffea arabica L. and the diploid species C. canephora P. were backcrossed to C. arabica. Although characterised by a low production and an important fruit dropping, all attempted crosses (ie, 6) generated BC(1) progenies. Flow cytometric analysis of the nuclear DNA content revealed that most of the BC1 individuals were nearly tetraploid. Among the male gametes produced by the interspecific triploid hybrids, those presenting a high number of chromosomes appeared strongly favoured. Only pollen mother cells having nearly 22 chromosomes were effective, the others leading to deficient endosperm and fruit dropping. Molecular markers (ie, microsatellite and AFLP) combined with evaluations of morphological characteristics and resistance to leaf rust were applied to verify the occurrence of gene transfer from C. canephora into C. arabica, and to estimate the amount of introgression present in BC(1) individuals. The results reveal a strong deficiency in the C. canephroa alleles indicating a severe counter-selection against the introgression of genetic material from C. canephora into C. arabica by way of triploid hybrids. However, introgressants displaying desirable traits such as a high resistance to leaf rust were obtained. The low level of introgression could be an advantage by facilitating the recovery of the recurrent parent and possibly reducing the number of required backcrosses. On the other hand, this could be a limitation when attempting the transfer of a complex trait or several simply inherited traits.     

Bee pollination and fruit set of Coffea arabica and C. canephora (Rubiaceae)

Self-sterile Coffea canephora and self-fertile C. arabica are important cash crops in many tropical countries. We examined the relative importance of insect, wind, and spontaneous self-pollination and the degree of self-fertility of these two coffee species in 24 agroforestry coffee fields in Indonesia. In both species, open pollination and cross pollination by hand led to the highest fruit set. Wind pollination (including self-pollination) led to 16% lower fruit set than open pollination in C. canephora and to 12.3% lower fruit set in C. arabica. Self-pollinated flowers and unmanipulated controls achieved an extremely low fruit set of 10% or less in the self-sterile species, and of 60% and 48%, respectively in the self-fertile species. These results constitute experimental evidence that cross pollination by bees causes a significant increase in fruit set of not only the self-sterile, but also the self-fertile coffee species. The practical implication is that coffee yield may be improved by managing fields for increased flower visitation by bees.     

Photoinhibition and recovery of photosynthesis in Coffea arabica and C. canephora

Photosynthetic parameters were determined in disks from leaves of C. arabica cv. Red Catuaí and C. canephora cv. Kouillou grown in the field. Kouillou showed a relatively higher irradiance requirement for saturating photosynthesis, lower chlorophyll (Chl) content, and higher Chl a/b ratio than Catuaí. Photoinhibition of photosynthesis under bright irradiance was manifested by decreases in maximum photochemical efficiency (evaluated by the variable to maximum fluorescence ratio, Fv/Fm), as a consequence of an increased initial and a quenched maximum fluorescence. Restoration of Fv/Fm following photoinhibition in low irradiance was faster in Kouillou than in Catuaí. Chloramphenicol both accelerated photoinhibition (mainly in Kouillou) and blocked its recovery for at least 190 min in either cultivar. Photosynthetic oxygen evolution under photoinhibitory conditions was decreased by chloramphenicol; in control leaf disks this decrease was only observed in C. arabica, but with a rapid recovery within 90 min of low irradiance exposure. In both coffee cultivars, the depressed photochemical efficiency of photosystem 2 was not accompanied by a concomitant lowering in oxygen evolution during reversal from photoinhibition.     

Introgression into the allotetraploid coffee ( Coffea arabica L.): segregation and recombination of the C. canephora genome in the tetraploid interspecific hybrid ( C. arabicax C. canephora)

Transfer of desired characters from the diploid relative species such as Coffea canephora into the cultivated allotetraploid coffee species (Coffea arabica L.) is essential to the continued improvement of varieties. Behaviour of the C. canephora genome and its interaction with the C. arabica genome were investigated in tetraploid interspecific hybrids (C. arabica×C. canephora 4x) resulting from a cross between an accession of C. arabica and a tetraploid plant of C. canephora obtained following colchicine treatment. Segregation and co-segregation of restriction fragment length polymorphism (RFLP) and microsatellite loci-markers were studied in two BC₁ populations. These two populations of 28 and 45 individuals, respectively, resulted from the backcross of two tetraploid F₁ plants to C. arabica. The presence in BC₁ plants of specific C. canephora markers was scored for 24 loci (11 RFLP and 13 microsatellites) distributed on at least 7 of the 11 linkage groups identified in C. canephora. At almost all loci analysed, the segregation of C. canephora alleles transmitted by the (C. arabica×C. canephora 4x) hybrids conformed to the expected ratio assuming random chromosome segregation and the absence of selection. The recombination fractions of C. canephora chromosome segments were estimated for seven marker intervals, and compared with the recombination fractions previously observed in C. canephora for the equivalent marker intervals. The recombination frequencies estimated in both plant materials were rather similar, suggesting that recombination in the (C. arabica×C. canephora 4x) hybrid is not significantly restricted by the genetic differentiation between chromosomes belonging to the different genomes. The hybrid (C. arabica×C. canephora 4x) therefore appeared particularly favourable to intergenomic recombination events and gene introgressions. Received: 26 March 2001 / Accepted: 29 June 2001     

Photoinhibition and recovery of photosynthesis in Coffea arabica and C. canephora

Photosynthetic parameters were determined in disks from leaves of C. arabica cv. Red Catuaí and C. canephora cv. Kouillou grown in the field. Kouillou showed a relatively higher irradiance requirement for saturating photosynthesis, lower chlorophyll (Chl) content, and higher Chl a/b ratio than Catuaí. Photoinhibition of photosynthesis under bright irradiance was manifested by decreases in maximum photochemical efficiency (evaluated by the variable to maximum fluorescence ratio, Fv/Fm), as a consequence of an increased initial and a quenched maximum fluorescence. Restoration of Fv/Fm following photoinhibition in low irradiance was faster in Kouillou than in Catuaí. Chloramphenicol both accelerated photoinhibition (mainly in Kouillou) and blocked its recovery for at least 190 min in either cultivar. Photosynthetic oxygen evolution under photoinhibitory conditions was decreased by chloramphenicol; in control leaf disks this decrease was only observed in C. arabica, but with a rapid recovery within 90 min of low irradiance exposure. In both coffee cultivars, the depressed photochemical efficiency of photosystem 2 was not accompanied by a concomitant lowering in oxygen evolution during reversal from photoinhibition.     

Single-locus inheritance in the allotetraploid Coffea arabica L. and interspecific hybrid C. arabica x C. canephora

Molecular cytogenetic analysis has indicated that Coffea arabica is an amphidiploid formed from the hybridization between two closely related diploid progenitor species, C. canephora and C. eugenioides. Our aim was to determine the mode of inheritance in C. arabica and in a tetraploid interspecific hybrid (called arabusta) between C. arabica and C. canephora as revealed by segregation analyses of restriction fragment length polymorphism (RFLP) loci markers. The observed RFLP allele segregations in an F(2) progeny of C. arabica conform to disomic inheritance as expected, with regular bivalent pairing of homologous chromosomes in the F1 hybrid. In contrast, RFLP loci followed tetrasomic inheritance in the arabusta interspecific hybrid, although bivalents have been reported to predominate greatly at meiosis in its hybrid. These results suggest that homologous chromosomes do not pair in C. arabica, not as a consequence of structural differentiation, but because of the functioning of pairing regulating factors. Moreover, the arabusta hybrid seems to offer the possibility of gene exchange between the homologous genomes.     

Identification of novel and conserved microRNAs in Coffea canephora and Coffea arabica

As microRNAs (miRNAs) are important regulators of many biological processes, a series of small RNAomes from plants have been produced in the last decade. However, miRNA data from several groups of plants are still lacking, including some economically important crops. Here microRNAs from Coffea canephora leaves were profiled and 58 unique sequences belonging to 33 families were found, including two novel microRNAs that have never been described before in plants. Some of the microRNA sequences were also identified in Coffea arabica that, together with C. canephora, correspond to the two major sources of coffee production in the world. The targets of almost all miRNAs were also predicted on coffee expressed sequences. This is the first report of novel miRNAs in the genus Coffea, and also the first in the plant order Gentianales. The data obtained establishes the basis for the understanding of the complex miRNA-target network on those two important crops.     

Brevipalpus mites Donnadieu (Prostigmata: Tenuipalpidae) associated with ornamental plants in Distrito Federal, Brazil

Brevipalpus mites colonize a great number of fruit and ornamental plants. Mite species belonging to this genus have been associated with many plant viruses. Citrus leprosis (CiLV) is the most economically important virus transmitted by B. phoenicis mites. It has recently been shown that ornamental plant species can be alternative hosts of this virus. The high volume of trade and frequent movement of live ornamental plants make them efficient pest disseminators. Because of this, it is desirable to expand knowledge of potential pests aiming to guide the adoption of quarantine measures. This work reports ornamental plant hosts of Brevipalpus mites in the Distrito Federal (DF), as well the occurrence of symptoms consistent with Brevipalpus-borne plant viruses in these same hosts. Between July and September of 2005, five surveys were carried out in 14 localities within DF. Leaves and branches of fifty-five ornamental plant species were sampled. The species Pithecellobium avaremotemo Mart. is for the first time reported as a host for B. phoenicis (Geijskes), B. californicus Banks and B. obovatus Donnadieu species. Additionally, seven new species are reported as hosts for Brevipalpus within South America. New hosts are also listed for individual mite species. Typical symptoms of Brevipalpus-borne viruses were observed in Ligustrum sinense Lour., Pelargonium hortorum L.H. Bailey, Hibiscus rosa-sinensis L. and orchids (Dendrobium and Oncidium). The results of this work emphasize the potential role of the ornamental plants as vehicles for dissemination of Brevipalpus mites.     

Inhibition of caffeine biosynthesis in tea (Camellia sinensis) and coffee (Coffea arabica) plants by ribavirin

The effects of ribavirin, an inhibitor of inosine-5'-monophosphate (IMP) dehydrogenase, on [8-(14)C]inosine metabolism in tea leaves, coffee leaves and coffee fruits were investigated. Incorporation of radioactivity from [8-(14)C]inosine into purine alkaloids, such as theobromine and caffeine, guanine residues of RNA, and CO(2) was reduced by ribavirin, while incorporation into nucleotides, including IMP and adenine residues of RNA, was increased. The results indicate that inhibition of IMP dehydrogenase by ribavirin inhibits both caffeine and guanine nucleotide biosynthesis in caffeine-forming plants. The use of IMP dehydrogenase-deficient plants as a potential source of good quality caffeine-deficient tea and coffee plants is discussed.     

N2 Fixation (C2H2 reduction) by epiphylls on coffee,Coffea arabica

Nitrogen fixation (C₂H₂ reduction) by epiphylls on coffee,Coffea arabica, grown in sites with different degrees of shade, was determined. Coffee leaves with nitrogen-fixing epiphylls were found in all sites in approximately equal numbers. Rates of C₂H₂ reduction were similar for all sites and throughout the year, averaging 3.21 nmoles C₂H₂ reduced leaf with epiphylls^–1 day^–1. Apparently, neither rates of activity nor abundance of leaves with nitrogen-fixing epiphylls is related to the degree of shade in a site. No correlation was found between percent epiphyll cover and the presence or magnitude of nitrogen-fixing activity. Calculated annual fixation by epiphylls on coffee was low, ranging from 0.7 g N₂ ha^–1 year^–1 for the shadeless site to 1.4 g N₂ ha^–1 year^–1 for the site withIngajinicuil shade trees. These results suggest that epiphyll fixation is not an important source of nitrogen for the coffee ecosystem studied.     

Lipid transfer proteins in coffee: isolation of Coffea orthologs,Coffea arabica homeologs,expression during coffee fruit development and promoter analysis in transgenic tobacco plants

The aim of the present study was to perform a genomic analysis of non-specific lipid-transfer proteins (nsLTPs) in coffee. Several nsLTPs-encoding cDNA and gene sequences were cloned from Coffea arabica and Coffea canephora species. In this work, their analyses revealed that coffee nsLTPs belong to Type II LTP characterized under their mature forms by a molecular weight of around 7.3 kDa, a basic isoelectric points of 8.5 and the presence of typical CXC pattern, with X being an hydrophobic residue facing towards the hydrophobic cavity. Even if several single nucleotide polymorphisms were identified in these nsLTP-coding sequences, 3D predictions showed that they do not have a significant impact on protein functions. Northern blot and RT-qPCR experiments revealed specific expression of Type II nsLTPs-encoding genes in coffee fruits, mainly during the early development of endosperm of both C. arabica and C. canephora. As part of our search for tissue-specific promoters in coffee, an nsLTP promoter region of around 1.2 kb was isolated. It contained several DNA repeats including boxes identified as essential for grain specific expression in other plants. The whole fragment, and a series of 5′ deletions, were fused to the reporter gene β-glucuronidase (uidA) and analyzed in transgenic Nicotiana tabacum plants. Histochemical and fluorimetric GUS assays showed that the shorter (345 bp) and medium (827 bp) fragments of nsLTP promoter function as grain-specific promoters in transgenic tobacco plants.     

Penicillium brocae, a new species associated with the coffee berry borer in Chiapas, Mexico

Penicillium brocae is a new monoverticillate species isolated from coffee berry borers collected at coffee plantations in Mexico near Cacahoatán, Chiapas, and from borers reared on artificial diets at ECOSUR laboratory facilities in Tapachula, Chiapas. Phenotypically, it is in Penicillium series Implicatum, but because it does not conform to known species we have described it as new. ITS and large subunit rDNA were sequenced and compared to determine the phylogenetic position of this species. It is most closely related to Penicillium adametzii. Penicillium brocae has only been found in association with the coffee berry borer and is one of several fungi that grow in coffee berry borer galleries. Penicillium brocae may provide the exogenous sterols necessary for the coffee berry borer's development and thus be mutualistically associated with the insect.     

Biochemical and genomic analysis of sucrose metabolism during coffee (Coffea arabica) fruit development

Sucrose metabolism and the role of sucrose synthase were investigated in the fruit tissues (pericarp, perisperm, and endosperm) of Coffea arabica during development. Acid invertase, sucrose phosphate synthase, and sucrose synthase activities were monitored and compared with the levels of sucrose and reducing sugars. Among these enzymes, sucrose synthase showed the highest activities during the last stage of endosperm and pericarp development and this activity paralleled closely the accumulation of sucrose in these tissues at this stage. Carbon partitioning in fruits was studied by pulse-chase experiments with (14)C-sugars and revealed high rates of sucrose turnover in perisperm and endosperm tissues. Additional feeding experiments with (14)CO(2) showed that leaf photosynthesis contributed more to seed development than the pericarp in terms of photosynthate supply to the endosperm. Sugar analysis, feeding experiments, and histological studies indicated that the perisperm plays an important role in this downloading process. It was observed that the perisperm presents a transient accumulation of starch which is degraded as the seed develops. Two full-length cDNAs (CaSUS1 and CaSUS2) and the complete gene sequence of the latter were also isolated. They encode sucrose synthase isoforms that are phylogenetically distinct, indicating their involvement in different physiological functions during cherry development. Contrasting expression patterns were observed for CaSUS1 and CaSUS2 in perisperm, endosperm, and pericarp tissues: CaSUS1 mRNAs accumulated mainly during the early development of perisperm and endosperm, as well as during pericarp growing phases, whereas those of CaSUS2 paralleled sucrose synthase activity in the last weeks of pericarp and endosperm development. Taken together, these results indicate that sucrose synthase plays an important role in sugar metabolism during sucrose accumulation in the coffee fruit.     

Inheritance of resistance to coffee wilt disease (Fusarium xylarioides Steyaert) in Robusta coffee (Coffea canephora Pierre) and breeding perspectives

Knowledge on heritability is essential for selecting varieties resistant against coffee wilt disease caused by Fusarium xylarioides, which is currently devastating coffee in East and Central Africa. Variability of the resistance against coffee wilt disease in Coffea canephora and its inheritance were investigated in three experiments corresponding to a clonal trial involving 20 clones, a 10-parent partial diallel progeny, and a half-sib progeny test. There were significant quantitative genetic variations among the clones and progenies, suggesting polygenic control of the resistance. Around 50–65 % tree mortality was the optimal disease level for calculating heritability and genetic gains. General and specific combining abilities calculated within the optimal disease range for partial diallel analysis were significant. Broad-sense heritability for the same analysis and same disease range was moderate (0.329), and corresponding narrow-sense heritability was low (0.112). Broad-sense heritability from clones in the field at the same disease range was also moderate (0.333). Narrow-sense heritability from regression of half-sib progeny means onto parent means in the field and screen house was 0.183 and 0.369, respectively. Selecting tolerant clones for improvement against the disease is possible, and genetic gains are possible by using tolerant parents in breeding programs.