Caldas, Colombia
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The coffee berry borer (CBB) is the most prevalent pest of coffee plantations. Within the Coffea genus, C. arabica is susceptible to CBB and C. liberica shows a lower susceptibility. Two EST libraries were constructed from the total RNA of C. arabica and C. liberica fruits artificially infested with CBBs for 24 h. Using 6000 clones sequenced per library, a unigene database was generated, obtaining 3634 singletons and 1454 contigs. For each contig, the proportion of sequences present in both species was determined and a differential gene expression between the species was detected. C. arabica displayed a higher relative expression of proteins involved in general stress responses, whereas C. liberica showed the induction of a higher number of insect defense proteins. In order to validate the results, quantifications through real-time PCR were done. A hevein-like protein, an isoprene synthase, a salicylic acid carboxyl methyltransferase and a patatin-like protein gene were highly upregulated in C. liberica at 24 and/or 48 h after insect infestation compared to C.arabica. The identification of metabolic pathways induced by this pest insect provides tools to take advantage of the genetic resources available for the control of CBB. © 2012 Taylor and Francis Group, LLC.


Vieira A.,University of Lisbon | Cabral A.,University of Lisbon | Fino J.,University of Lisbon | Azinheira H.G.,University of Lisbon | And 11 more authors.
PLoS ONE | Year: 2016

Colletotrichum kahawae is an emergent fungal pathogen causing severe epidemics of Coffee Berry Disease on Arabica coffee crops in Africa. Currently, the molecular mechanisms underlying the Coffea arabica-C. kahawae interaction are still poorly understood, as well as the differences in pathogen aggressiveness, which makes the development of functional studies for this pathosystem a crucial step. Quantitative real time PCR (qPCR) has been one of the most promising approaches to perform gene expression analyses. However, proper data normalization with suitable reference genes is an absolute requirement. In this study, a set of 8 candidate reference genes were selected based on two different approaches (literature and Illumina RNA-seq datasets) to assess the best normalization factor for qPCR expression analysis of C. kahawae samples. The gene expression stability of candidate reference genes was evaluated for four isolates of C. kahawae bearing different aggressiveness patterns (Ang29, Ang67, Zim12 and Que2), at different stages of fungal development and key time points of the plant-fungus interaction process. Gene expression stability was assessed using the pairwise method incorporated in geNorm and the modelbased method used by NormFinder software. For C. arabica-C. kahawae interaction samples, the best normalization factor included the combination of PP1, Act and ck34620 genes, while for C. kahawae samples the combination of PP1, Act and ck20430 revealed to be the most appropriate choice. These results suggest that RNA-seq analyses can provide alternative sources of reference genes in addition to classical reference genes. The analysis of expression profiles of bifunctional catalase-peroxidase (cat2) and trihydroxynaphthalene reductase (thr1) genes further enabled the validation of the selected reference genes. This study provides, for the first time, the tools required to conduct accurate qPCR studies in C. kahawae considering its aggressiveness pattern, developmental stage and host interaction. © 2016 Vieira et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.


Aguilerag C.,National University of Colombia | Padillah B.E.,Bacteriologa | Florezr C.P.,CENICAFE | Rubiog J.D.,Entomologia | Acunaz J.R.,Entomologia
Revista Colombiana de Entomologia | Year: 2011

Eukaryotic organisms, including insects, have a mechanism for specific gene silencing by short sequences of double-stranded RNA (dsRNA), such a process is known as RNA interference (RNAi). The objective of this research was to follow up this technique to screen candidate genes for the genetic control of the coffee berry borer Hypothenemus hampei. A methodology of feeding by oral drops containing different dsRNAs and dsRNAs' dosages that were placed in the preoral cavity of first instar larvae was used. Then, the larvae were reared on an artificial diet supplemented with the respective dsRNA. Previously, we verified the effectiveness of the dsRNA feeding using two control strategies: a) intake of a fluorescent compound (Calcofluor™) and b) intake of a commercial mixture of protease inhibitors. The genes coding for mannanase, xylanase, cytochrome P450 mono-oxygenase (two genes), ATPase D, α-tubulin and actin, expressed in the coffee berry borer, were selected for silencing. We found that dsRNA of cytochromes P450 produced the greatest effect, causing larval mortality of 64% and 52% respectively, followed by dsRNA of mannanase and xylanase, with a mortality rate of 37% and 33%, respectively, for the highest concentration tested.


Cristancho M.,CENICAFE | Giraldo W.,CENICAFE | Botero D.,CENICAFE | Botero D.,University of Los Andes, Colombia | And 7 more authors.
Advances in Intelligent Systems and Computing | Year: 2014

The coffee rust, caused by the fungus Hemileia vastatrix, is the most serious disease of this crop worldwide. In Colombia the pathogen causes a reduction in production of up to 30% in susceptible varieties of Coffea arabica, if not controlled mild epidemics can occur and complete crop losses in strong epidemics. We applied genomics to study the evolution of the population of this pathogen due to recent outbreaks of the disease in Colombia. Sequencing was performed using 454 and Illumina technology, using DNA and RNA of 8 and 3 isolates of H. vastatrix, respectively. With the software CEGMA we made a first estimate of the genome size of H. vastatrix resulting in an approximate size of 250 Mb. The hybrid assembly was performed with all sequenced genomes given a coverage of 92% with a GC content of 32%. © Springer International Publishing Switzerland 2014.


Moreno Salguero C.A.,National University of Colombia | Bustillo Pardey A.E.,Cenicana | Lopez Nunez J.C.,Cenicafe | Castro Valderrama U.,Cenicana | Ramirez Sanchez G.D.,Cenicana
Revista Colombiana de Entomologia | Year: 2012

The spittlebug, Aeneolamia varia (Hemiptera: Cercopidae) is a limiting pest of sugarcane crops in Venezuela. In Colombia, this species has been recorded for over 40 years in Llanos Orientales, attacking pastures. Only in 2007 this insect was detected in sugarcane crops in the Cauca Valley, where we are looking at biological control agents, particularly entomopathogenic nematodes. In this study, we evaluated the virulence of three entomopathogenic nematode species selected in previous laboratory and greenhouse studies, Steinernema sp.1, Heterorhabditis bacteriophora and Heterorhabditis sp. (Gua 31), and two commercially aviable species, Steinernema sp. and Heterorhabditis sp., on IV instar nymphs of A. varia. The most virulent isolates were Heterorhabditis sp. (Gua 31) causing 42.3% of mortality at a dosage of 5 x 1010 JI/ha, and Heterorhabditis bacteriophora in dosage of 1.5 x 1011 JI/ha causing 76% of mortality on nymphs. This information is intended to give the farmer an effective alternative for the integrated management of A. varia.


Background: Drying helps to prolong the life of the fruit, causing structural changes, nutritional and aromatic level according to the technique and drying the applied temperatures. Convective drying (CD) is the most popular technique for drying different types of food, while drying window reflectance (WR) is a recent technique that uses infrared energy of water, which helps to reduce quality losses dried and heat-sensitive foods such as fruits. Objectives: The objective of this study was to evaluate the effect of drying method; convective drying and reflectance window on the retention of aromas in samples dried “bocadillo banana”. Methods: the convective drying was performed at 50, 60 and 70°C and the WR 70, 80 and 90°C into slices 2 mm thick and 23 mm in diameter “bocadillo banana”. The volatile compounds (VC) were absorbed by the technique of Solid Phase Microextraction (SPME) and aromatic profile was determined by gases chromatography coupled with mass spectrometry (GC-MS), in terms of percent retention of volatile attribute or description of the flavoring and forming reaction. Results: The technique window reflectance retained on average 80% of the volatile compounds of “bocadillo banana”, observing that the treatment at 80°C was better as it retained the highest percentage (91%) of volatile compounds present in the fresh sample. Moreover, in convective drying they were retained 65% on average of the volatiles “bocadillo banana”, presenting higher retention of impact compounds in the treatment at 50°C. Conclusions: Application of the window reflectance technique increased retention of volatile aroma compounds typical of fresh “bocadillo banana” versus hot air drying. © 2016, Universidad de Antioquia. All rights reserved.


Loureiro A.,Institute Investigacao Cientifica Tropical | Nicole M.R.,CIRAD - Agricultural Research for Development | Varzea V.,Institute Investigacao Cientifica Tropical | Moncada P.,Cenicafe | And 2 more authors.
Physiological and Molecular Plant Pathology | Year: 2012

Histological and ultrastructural studies were undertaken to compare Colletotrichum kahawae growth and the sequence of responses it induced in resistant and susceptible coffee genotypes. Coffee resistance was characterized by a restricted fungal growth associated with hypersensitive-like cell death and early accumulation of phenolic compounds, such as flavonoids (cytoplasmic contents) and hydroxycinnamic acid derivatives (cell walls). This accumulation of phenols in the cell walls preceded their lignification and thickening. In the susceptible genotype, a late accumulation of hydroxycinnamic acid derivatives in a number of cell walls and the encasement of some intracellular hyphae were also observed, but these delayed host responses did not prevent fungal growth and sporulation. © 2011 Elsevier Ltd.


PubMed | Cenicafe
Type: Journal Article | Journal: Proceedings of the National Academy of Sciences of the United States of America | Year: 2012

Horizontal gene transfer (HGT) involves the nonsexual transmission of genetic material across species boundaries. Although often detected in prokaryotes, examples of HGT involving animals are relatively rare, and any evolutionary advantage conferred to the recipient is typically obscure. We identified a gene (HhMAN1) from the coffee berry borer beetle, Hypothenemus hampei, a devastating pest of coffee, which shows clear evidence of HGT from bacteria. HhMAN1 encodes a mannanase, representing a class of glycosyl hydrolases that has not previously been reported in insects. Recombinant HhMAN1 protein hydrolyzes coffee berry galactomannan, the major storage polysaccharide in this species and the presumed food of H. hampei. HhMAN1 was found to be widespread in a broad biogeographic survey of H. hampei accessions, indicating that the HGT event occurred before radiation of the insect from West Africa to Asia and South America. However, the gene was not detected in the closely related species H. obscurus (the tropical nut borer or false berry borer), which does not colonize coffee beans. Thus, HGT of HhMAN1 from bacteria represents a likely adaptation to a specific ecological niche and may have been promoted by intensive agricultural practices.

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