TcTI, a Kunitz-type trypsin inhibitor from cocoa associated with defense against pathogens.

Protease inhibitors (PIs) are important biotechnological tools of interest in agriculture. Usually they are the first proteins to be activated in plant-induced resistance against pathogens. Therefore, the aim of this study was to characterize a Theobroma cacao trypsin inhibitor called TcTI. The ORF has 740 bp encoding a protein with 219 amino acids, molecular weight of approximately 23 kDa. rTcTI was expressed in the soluble fraction of Escherichia coli strain Rosetta [DE3]. The purified His-Tag rTcTI showed inhibitory activity against commercial porcine trypsin. The kinetic model demonstrated that rTcTI is a competitive inhibitor, with a Ki value of 4.08 × 10-7 mol L-1. The thermostability analysis of rTcTI showed that 100% inhibitory activity was retained up to 60 °C and that at 70-80 °C, inhibitory activity remained above 50%. Circular dichroism analysis indicated that the protein is rich in loop structures and ß-conformations. Furthermore, in vivo assays against Helicoverpa armigera larvae were also performed with rTcTI in 0.1 mg mL-1 spray solutions on leaf surfaces, which reduced larval growth by 70% compared to the control treatment. Trials with cocoa plants infected with Mp showed a greater accumulation of TcTI in resistant varieties of T. cacao, so this regulation may be associated with different isoforms of TcTI. This inhibitor has biochemical characteristics suitable for biotechnological applications as well as in resistance studies of T. cacao and other crops.

Effects of habitat management on different feeding guilds of herbivorous insects in cacao agroforestry systems / Efecto del manejo de hábitat en diferentes gremios alimentarios de insectos herbívoros en sistemas agroforestales de cacao

AbstractHuman pressure on natural habitats increases the importance of agroforests for biodiversity conservation. The objective of this study was to evaluate the role of cacao traditional cultivation system (CTCS) on the conservation of the herbivorous insect community when compared with a monodominant rubber agroforest, a type of agricultural system for cacao cultivation. The insects were sampled in three habitats in Southeastern Bahia, Brazil: native forests, CTCS and rubber agroforests. In each habitat, 18 plots of 10 m2 were established, and the structural measures were collected and herbivorous insects were sampled with a Malaise/window trap. The diversity of folivorous decreased with the simplification of vegetation structure, but species composition was similar among habitats. In addition to a decrease in the availability of resources in monodominant rubber agroforests, the latex present in these systems have limited the occurrence of species that cannot circumvent latex toxicity. The diversity of sap-sucking insects was similar among habitats, but species composition was similar only in the CTCS and native forest, and it was different in the rubber agroforest. We observed turnover and a higher frequency of individuals of the family Psyllidae in the rubber agroforest. The biology and behavior of Psyllids and absence of natural enemies enable their diversity to increase when they are adapted to a new host. We observed a shift in the composition of xylophagous insects in the rubber agroforest compared to that in other habitats. Moreover, this agroforest has low species richness, but high individual abundance. Latex extraction is likely an important additional source of volatile compounds discharged into the environment, and it increases the attraction and recruitment of coleoborers to these sites. We concluded that CTCS has an herbivorous insect community with a structure similar to the community found in native forests of the region, and they present a more interesting conservation strategy when compared to rubber agroforests. We also emphasized the potential risk of local pest outbreaks in rubber agroforests for both the rubber and associated cacao trees. Rev. Biol. Trop. 64 (2): 763-777. Epub 2016 June 01.

ResumenLa presión humana sobre los hábitats naturales aumenta la importancia de los sistemas agroforestales para la conservación de la biodiversidad. El objetivo de este estudio fue evaluar el papel del sistema tradicional de cultivo de cacao en la conservación de la comunidad de insectos herbívoros en comparación con el agrobosque monodominante de caucho, un tipo de sistema agrícola para el cultivo de cacao. Los insectos fueron muestreados en tres hábitats en el sureste de Bahia, Brasil: bosque nativo, sistema tradicional de cultivo de cacao y agrobosque de caucho. En cada hábitat, se establecieron 18 parcelas de 10 m2, se tomaron medidas estructurales y se muestrearon los insectos herbívoros con una trampa tipo Malaise. La diversidad de folívoros disminuyó con la simplificación de la estructura de la vegetación, pero la composición de especies fue similar entre hábitats. Además se presentó una disminución de la disponibilidad de recursos en agrobosques de caucho, el látex presente en estos sistemas han limitado la aparición de especies que no pueden evadir la toxicidad de látex. La diversidad de insectos chupadores de savia fue similar entre los hábitats, pero la composición de especies fue similar sólo en el sistema tradicional de cultivo de cacao y el bosque nativo, y diferente en el agrobosque de caucho. Hemos observado que hay una mayor frecuencia de individuos de la familia Psyllidae en el agrobosque de caucho. La biología y el comportamiento de los psílidos y ausencia de enemigos naturales permiten que su diversidad aumente cuando se adaptan a un nuevo huésped. Hemos observado un cambio en la composición de los insectos xilófagos en el agrobosque de caucho en comparación con la de otros hábitats. Por otra parte, este agrobosque tiene una baja riqueza de especies, pero gran abundancia individual. La extracción de látex probablemente es una fuente adicional importante de compuestos volátiles que son vertidos en el ambiente, y aumenta la atracción y el reclutamiento de curculiónidos en estos sitios. Hemos llegado a la conclusión de que el sistema tradicional de cultivo de cacao tiene una comunidad de insectos herbívoros con una estructura similar a la comunidad que se encuentra en los bosques nativos de la región, y presenta una estrategia de conservación más interesante si se compara con los sistemas agroforestales de caucho. También hicimos hincapié en el riesgo potencial de aparición de plagas locales en sistemas agroforestales de caucho, tanto para el caucho como para los árboles de cacao asociados.

Theobroma cacao L. pathogenesis-related gene tandem array members show diverse expression dynamics in response to pathogen colonization.

BACKGROUND: The pathogenesis-related (PR) group of proteins are operationally defined as polypeptides that increase in concentration in plant tissues upon contact with a pathogen. To date, 17 classes of highly divergent proteins have been described that act through multiple mechanisms of pathogen resistance. Characterizing these families in cacao, an economically important tree crop, and comparing the families to those in other species, is an important step in understanding cacao's immune response. RESULTS: Using publically available resources, all members of the 17 recognized pathogenesis-related gene families in the genome of Theobroma cacao were identified and annotated resulting in a set of ~350 members in both published cacao genomes. Approximately 50 % of these genes are organized in tandem arrays scattered throughout the genome. This feature was observed in five additional plant taxa (three dicots and two monocots), suggesting that tandem duplication has played an important role in the evolution of the PR genes in higher plants. Expression profiling captured the dynamics and complexity of PR genes expression at basal levels and after induction by two cacao pathogens (the oomycete, Phytophthora palmivora, and the fungus, Colletotrichum theobromicola), identifying specific genes within families that are more responsive to pathogen challenge. Subsequent qRT-PCR validated the induction of several PR-1, PR-3, PR-4, and PR-10 family members, with greater than 1000 fold induction detected for specific genes. CONCLUSIONS: We describe candidate genes that are likely to be involved in cacao's defense against Phytophthora and Colletotrichum infection and could be potentially useful for marker-assisted selection for breeding of disease resistant cacao varieties. The data presented here, along with existing cacao-omics resources, will enable targeted functional genetic screening of defense genes likely to play critical functions in cacao's defense against its pathogens.

Effects of habitat management on different feeding guilds of herbivorous insects in cacao agroforestry systems.

Human pressure on natural habitats increases the importance of agroforests for biodiversity conservation. The objective of this study was to evaluate the role of cacao traditional cultivation system (CTCS) on the conservation of the herbivorous insect community when compared with a monodominant rubber agroforest, a type of agricultural system for cacao cultivation. The insects were sampled in three habitats in Southeastern Bahia, Brazil: native forests, CTCS and rubber agroforests. In each habitat, 18 plots of 10 m2 were established, and the structural measures were collected and herbivorous insects were sampled with a Malaise/window trap. The diversity of folivorous decreased with the simplification of vegetation structure, but species composition was similar among habitats. In addition to a decrease in the availability of resources in monodominant rubber agroforests, the latex present in these systems have limited the occurrence of species that cannot circumvent latex toxicity. The diversity of sap-sucking insects was similar among habitats, but species composition was similar only in the CTCS and native forest, and it was different in the rubber agroforest. We observed turnover and a higher frequency of individuals of the family Psyllidae in the rubber agroforest. The biology and behavior of Psyllids and absence of natural enemies enable their diversity to increase when they are adapted to a new host. We observed a shift in the composition of xylophagous insects in the rubber agroforest compared to that in other habitats. Moreover, this agroforest has low species richness, but high individual abundance. Latex extraction is likely an important additional source of volatile compounds discharged into the environment, and it increases the attraction and recruitment of coleoborers to these sites. We concluded that CTCS has an herbivorous insect community with a structure similar to the community found in native forests of the region, and they present a more interesting conservation strategy when compared to rubber agroforests. We also emphasized the potential risk of local pest outbreaks in rubber agroforests for both the rubber and associated cacao trees.

Tree spatial structure, host composition and resource availability influence mirid density or black pod prevalence in cacao agroforests in Cameroon.

Combining crop plants with other plant species in agro-ecosystems is one way to enhance ecological pest and disease regulation mechanisms. Resource availability and microclimatic variation mechanisms affect processes related to pest and pathogen life cycles. These mechanisms are supported both by empirical research and by epidemiological models, yet their relative importance in a real complex agro-ecosystem is still not known. Our aim was thus to assess the independent effects and the relative importance of different variables related to resource availability and microclimatic variation that explain pest and disease occurrence at the plot scale in real complex agro-ecosystems. The study was conducted in cacao (Theobroma cacao) agroforests in Cameroon, where cocoa production is mainly impacted by the mirid bug, Sahlbergella singularis, and black pod disease, caused by Phytophthora megakarya. Vegetation composition and spatial structure, resource availability and pest and disease occurrence were characterized in 20 real agroforest plots. Hierarchical partitioning was used to identify the causal variables that explain mirid density and black pod prevalence. The results of this study show that cacao agroforests can be differentiated on the basis of vegetation composition and spatial structure. This original approach revealed that mirid density decreased when a minimum number of randomly distributed forest trees were present compared with the aggregated distribution of forest trees, or when forest tree density was low. Moreover, a decrease in mirid density was also related to decreased availability of sensitive tissue, independently of the effect of forest tree structure. Contrary to expectations, black pod prevalence decreased with increasing cacao tree abundance. By revealing the effects of vegetation composition and spatial structure on mirids and black pod, this study opens new perspectives for the joint agro-ecological management of cacao pests and diseases at the plot scale, through the optimization of the spatial structure and composition of the vegetation.

Shade tree diversity, cocoa pest damage, yield compensating inputs and farmers' net returns in West Africa.

Cocoa agroforests can significantly support biodiversity, yet intensification of farming practices is degrading agroforestry habitats and compromising ecosystem services such as biological pest control. Effective conservation strategies depend on the type of relationship between agricultural matrix, biodiversity and ecosystem services, but to date the shape of this relationship is unknown. We linked shade index calculated from eight vegetation variables, with insect pests and beneficial insects (ants, wasps and spiders) in 20 cocoa agroforests differing in woody and herbaceous vegetation diversity. We measured herbivory and predatory rates, and quantified resulting increases in cocoa yield and net returns. We found that number of spider webs and wasp nests significantly decreased with increasing density of exotic shade tree species. Greater species richness of native shade tree species was associated with a higher number of wasp nests and spider webs while species richness of understory plants did not have a strong impact on these beneficial species. Species richness of ants, wasp nests and spider webs peaked at higher levels of plant species richness. The number of herbivore species (mirid bugs and cocoa pod borers) and the rate of herbivory on cocoa pods decreased with increasing shade index. Shade index was negatively related to yield, with yield significantly higher at shade and herb covers<50%. However, higher inputs in the cocoa farms do not necessarily result in a higher net return. In conclusion, our study shows the importance of a diverse shade canopy in reducing damage caused by cocoa pests. It also highlights the importance of conservation initiatives in tropical agroforestry landscapes.

True bug (Heteroptera) impact on cocoa fruit mortality and productivity.

The real impact of true bug damage on cocoa pods has never been assessed precisely. We conducted a 2-yr study on 1,080 cocoa trees on 36 farms in Cameroon to assess the contribution of true bugs to fruit mortality and production loss. The cocoa fruiting cycle, fruit mortality, and damage caused by true bugs as well as other pests and diseases were monitored on a weekly basis. True bug damage also was described on 2,500 ripe pods per year. Pod weight, bean number, and bean weight were measured and compared for different degrees and types of damage on the ripe pods. Our results showed that true bugs were the main external cause of young fruit abortion. They reduced the abundance of young fruit by up to 10%. In contrast, although one-third of the ripe pods sampled had true bug lesions, only 4% were moderately to heavily damaged. The mean weight of ripe pods was reduced by 12% when there was medium to heavy damage. While the mean weight of wet beans was reduced significantly (by 3-10%), the number of beans per pod was not changed by damage. Despite the reduction in mean weight, the overall weight of beans for the pods sampled was reduced by <2%. Therefore, our study confirmed the common assumption that the economic impact of true bug damage on mature pods is negligible on cocoa farms in Cameroon. However, true bugs have a significant impact on young fruit mortality.

Sexual dimorphism of pupae and adults of the cocoa pod borer, Conopomorpha cramerella.

This paper describes the main distinguishing characteristics of female and male pupae and adults of cocoa pod borer, Conopomorpha cramerella (Snellen) (Lepidoptera: Gracillariidae). Two pairs of tubercles present on the sterna of segments IX and X of the female pupae are useful in differentiating female from male pupae. The female genital opening is located anterior to the first pair of tubercles and forms a plateau in which the center has a light brown longitudinal depression that indicates the female genital opening. The male genital opening is a conspicuous, brown, longitudinal slit located between the two pairs of tubercles. The sex of the adult moth can be determined by examining the ventrocaudal segments of the abdomen. The last segment of the female abdomen is white, compressed laterally and at the tip, and the hairy anal papillae can be seen. In the male, the ventrocaudal end of the abdomen is black and robust. This information will be useful for laboratory and field diagnosis and while working on sex ratios of this important pest of cocoa.

Field evaluation of synthetic sex pheromone traps for the cocoa mirid Sahlbergella singularis (Hemiptera: Miridae).

BACKGROUND: Field trapping experiments were conducted to evaluate the attractiveness of five different pheromone blends as well as the impact of trap design on attractiveness towards cocoa mirids, Sahlbergella singularis Hagl. and Distantiella theobroma (Dist.), in Cameroon. RESULTS: A total of 361 adults of S. singularis (359 males and two females) were caught. The highest numbers of mirids were found in traps with pheromone blends that combined a monoester and a diester, compared with traps with the diester or the monoester individually and control traps with no pheromone. Rectangular traps caught significantly more mirids compared with delta traps. The mean number of 5.1 mirids trap⁻¹ year⁻¹ caught in rectangular traps was significantly higher compared with the 1.8 mirids trap⁻¹ year⁻¹ for the delta traps. CONCLUSION: The data revealed that rectangular traps containing pheromone blends combining both the monoester and the diester have a good potential to lure and trap adult males of S. singularis on cocoa farms. The pheromone blends used were specific for S. singularis, and the use of pheromone traps appears to be a promising strategy for incorporation into integrated pest management strategies for the monitoring or even the control of S. singularis in cocoa plantations.

Endophytic fungal diversity in Theobroma cacao (cacao) and T. grandiflorum (cupuaçu) trees and their potential for growth promotion and biocontrol of black-pod disease.

The endophytic niches of plants are a rich source of microbes that can directly and indirectly promote plant protection, growth and development. The diversity of culturable endophytic fungi from stems and branches of Theobroma cacao (cacao) and Theobroma grandiflorum (cupuaçu) trees growing in the Amazon region of Brazil was assessed. The collection of fungal endophytic isolates obtained was applied in field experiments to evaluate their potential as biocontrol agents against Phytophthora palmivora, the causal agent of the black-pod rot disease of cacao, one of the most important pathogens in cocoa-producing regions worldwide. The isolated endophytic fungi from 60 traditional, farmer-planted, healthy cacao and 10 cupuaçu plants were cultured in PDA under conditions inducing sporulation. Isolates were classified based upon the morphological characteristics of their cultures and reproductive structures. Spore suspensions from a total of 103 isolates that could be classified at least up to genus level were tested against P. palmivora in pods attached to cacao trees in the field. Results indicated that ∼70% of isolates showed biocontrol effects to a certain extent, suggesting that culturable endophytic fungal biodiversity in this system is of a mostly mutualistic type of interaction with the host. Eight isolates from genera Trichoderma (reference isolate), Pestalotiopsis, Curvularia, Tolypocladium and Fusarium showed the highest level of activity against the pathogen, and were further characterized. All demonstrated their endophytic nature by colonizing axenic cacao plantlets, and confirmed their biocontrol activity on attached pods trials by showing significant decrease in disease severity in relation to the positive control. None, however, showed detectable growth-promotion effects. Aspects related to endophytic biodiversity and host-pathogen-endophyte interactions in the environment of this study were discussed on the context of developing sustainable strategies for biological control of black-pod rot of cacao.

Cocoa Pod Borer (Conopomorpha cramerella Snellen) in Papua New Guinea: biosecurity models for New Ireland and the autonomous region of Bougainville.

Cocoa Pod Borer (Conopomorpha cramerella Snellen) (CPB) is an important pest of cocoa. Following its emergence as a pest in East New Britain, Papua New Guinea, in 2006, it was considered relevant to assess its potential spread to other cocoa growing regions. Its likelihood of introduction to the islands of Bougainville and New Ireland from East New Britain Province, Papua New Guinea, was modeled using Monte Carlo simulation. This dispersal model was based around different scenarios, identifying trends rather than explicitly attempting to encapsulate true values. The model suggested that CPB is far more likely to establish on New Ireland than on Bougainville. More important, incertitude resulting from incomplete knowledge of the amount and frequency of cocoa transported between islands had a significant effect on model outputs. Quarantine and agriculture officials will be able to refine these parameter values, and then use the relevant scenarios from those presented here as a guide to develop quarantine procedures. In addition, a contingency model was employed to estimate the optimal sampling effort to use following an incursion of CPB into Bougainville or New Ireland and the seemingly successful implementation of an initial eradication program. The model suggests that at a 1% infestation level, sampling should continue for 2.5-2.7 years (90% CI) after claiming eradication, and this estimate changed little for higher infestation levels. Through modeling variations in sampling intensity, the model also suggested that determining the full spread of CPB is more important than increased sampling within one region.

[Scolitids (Coleoptera: Scolytidae) associated to the cocoa agroecosystem in Tabasco, Mexico]. / Escolítidos (Coleoptera: Scolytidae) asociados al agroecosistema cacao en Tabasco, México.

The Scolytidae fauna associated to the cacao agroecosystem in Tabasco, Mexico was studied during 2007. Adult insects were captured by using ethanol and light traps, and by direct collecting in their host plants. 3,192 specimens of 34 species belonging to 18 genera of Scolytidae were collected. Twenty-two species are new records for the State of Tabasco. The genera with more species were Xyleborus (6) and Hypothenemus (6). We conclude that the Scolytidae fauna associated to the cacao agroecosystem captured with the three trap systems are similar through the year of study. We also conclude that although the ethanol traps were more efficient, obtaining the highest species diversity, the use of other methods is important.

Escolítidos (Coleoptera: Scolytidae) asociados al agroecosistema cacao en Tabasco, México / Scolitids (Coleoptera: Scolytidae) associated to the cocoa agroecosystem in Tabasco, Mexico

El objetivo de este trabajo fue conocer la fauna de escolítidos asociada al agroecosistema cacao durante el año 2007 en Tabasco, México; para ello se utilizaron trampas de alcohol etílico, trampa de luz y captura sobre sus plantas huéspedes como métodos de recolecta. Se registraron 3,192 especimenes de 34 especies pertenecientes a 18 géneros de escolítidos. Veintidós especies son registros nuevos para el Estado de Tabasco. Los géneros con mayor riqueza de especies fueron Xyleborus (6) e Hypothenemus (6). Se concluye que la fauna de escolítidos capturadas con los tres métodos de recolecta presentó una fluctuación poblacional similar a través del año de estudio. Además concluimos que aunque la trampa de alcohol etílico fue el método de captura más eficiente para la obtención de la mayor diversidad de especies es importante la utilización de los otros métodos que complementan y enriquecen la información obtenida.

The Scolytidae fauna associated to the cacao agroecosystem in Tabasco, Mexico was studied during 2007. Adult insects were captured by using ethanol and light traps, and by direct collecting in their host plants. 3,192 specimens of 34 species belonging to 18 genera of Scolytidae were collected. Twenty-two species are new records for the State of Tabasco. The genera with more species were Xyleborus (6) and Hypothenemus (6). We conclude that the Scolytidae fauna associated to the cacao agroecosystem captured with the three trap systems are similar through the year of study. We also conclude that although the ethanol traps were more efficient, obtaining the highest species diversity, the use of other methods is important.

Activity evaluation of cocoa pod borer sex pheromone in cacao fields.

The previously identified female sex pheromone of cocoa pod borer, Conopomorpha cramerella, was re-evaluated for its attractive activity in different field conditions. It was found that lures containing 100-mug of synthetic sex pheromone blend, (E,Z,Z)- and (E,E,Z)-4,6,10-hexadecatrienyl acetates, and the corresponding alcohols in a ratio of 40:60:4:6 in a polyethylene vial attracted male C. cramerella moths in Sabah and peninsular Malaysia and in Sumatra and Sulawesi, Indonesia, suggesting that the same pheromone strain existed in a wide stretch of the Indo-Malayan archipelago. Of the three kinds of trap designs tested, the Delta traps were more effective than Pherocon V scale traps. Male captures were not significantly different among traps baited with 100-, 300-, or 1,000-mug doses of sex pheromone. A release rate study of pheromone formulation conducted in the laboratory showed that volatile active ingredients were desorbed from polyethylene vials following first-order kinetics, which indicates a satisfactory "half-life time" of a 100-mug loading is approximately 6 wk under laboratory conditions. A satisfactory attractiveness of the lure with a 100-mug loading was approximately 1-2 mo in the fields.

Effect of genotype of cocoa (Theobroma cacao) on attractiveness to the mirid Sahlbergella singularis (Hemiptera: Miridae) in the laboratory.

BACKGROUND: Mirids are a major constraint to cocoa growing in Africa. Cocoa breeding for tolerance/resistance could be effective in an integrated pest management system. Attractiveness is one aspect of tolerance/resistance, and decreasing the attractiveness of cocoa trees should be a good way of reducing damage. RESULTS: Small-scale laboratory tests were carried out in Cameroon to assess differences in the attractiveness to mirids of eleven cocoa genotypes. The genotypes were ranked according to their attractiveness score and a distance from a mean value. An analysis of variance was performed and revealed significant differences between cocoa genotypes (F=3.15, P<0.001). The cocoa genotype groupings revealed three major categories, with BE10 and SNK413 proving to be the most attractive. In contrast, five genotypes, IMC60, the Catongo Trinitario genotypes, Playa Alta2 (from Venezuela), SIC5 and SNK614, proved to be less attractive than the mean. Four genotypes (PA107, SNK619, UPA134 and T60/887) displayed similar attractiveness to the mean. CONCLUSION: The circular microtest offered the advantage of not needing a reference cocoa genotype. The least attractive clones, such as IMC60, were also the most tolerant in the field. Comparisons with the results of other studies are proposed.

Análisis de variabilidad genética en Moniliophthora roreri con AP-PCR y RAPD en Antioquia, Colombia / Analysis of genetic variability in Moniliophthora roreri with AP-PCR and RAPD in Antioquia, Colombia

Moniliophthora roreri es el agente causante de la moniliasis del cacao, la enfermedad más severa en las plantaciones de cacao en el departamento de Antioquia, Colombia. Los marcadores moleculares RAPD (Random Amplyfied Polymorphism of DNA) y AP-PCR (Arbitraly Primed Polymerase Chain Reaction) fueron usados para estudiar la variabilidad genética de 170 aislamientos de M.roreri colectados en doce municipios de Antioquia. El análisis dividió la población en seis grupos,el grupo G1 fue el más grande y contenía el 95 por cien de los aislamientos con una alta similitud genética (coeficiente de similitud de 0,7 a 1), mientras los otros cinco grupos contenían solo aislamientos de Apartadó y Dabeiba con una similitud genética moderadamente baja (coeficiente de similitud entre 0,45 a 0,55). El análisis de componentes principales mostró una alta similitud genética entre la población excepto entre los aislamientos de Apartadó y Dabeiba, que registraron los más altos niveles de variabilidad genética con valores altos del índice de Shannon y el porcentaje de loci polimórficos, mientras los otros aislamientos registraron una baja variabilidad genética. Los valores de diversidad y diferenciación genética en la población muestran una introducción reciente de M.roreri en las plantaciones de cacao de Antioquia, y una reproducción predominantemente clonal en la población. De acuerdo con Amova, la mayoría de la variación genética se encontró dentro de los municipios (75,68 por cien) con solo un 5,94 por cien presente entre las subregiones.

Cocoa pod (Theobroma caco)--a potential breeding habit of Aedes albopictus in dengue-sensitive Kerala State, India.

Aedes (Stegomyia) albopictus (Skuse), a potential dengue vector, particularly in the foot-hill areas of the Western Ghat region of Kerala State has been found breeding for the first time in cocoa pods (Theobroma cacao) in India. Breeding in cocoa pods is considered an opportunistic adaptive behavior of the mosquito to maintain density in the absence of common breeding sites, especially latex-collecting cups in rubber plantations, during the monsoon periods. Vector breeding in cocoa pods was observed up to 8 m from the ground, and in both hanging and grounded pods, whose endocarp had been devoured by rodents. Ae. albopictus breeding in cocoa plant is discussed in relation to accentuating disease transmission risk.

Activity of Bacillus thuringiensis toxins against cocoa pod borer larvae.

Twelve Cry proteins from Bacillus thuringiensis Berliner were tested in bioassays on cacao plantations in Indonesia for activity against the larvae of cocoa pod borer (Conopomorpha cramerella (Snellen)), an insect pest of the cacao tree. Through the damage caused by their feeding, the larvae of cocoa pod borer cause the pods of the cocoa tree to ripen prematurely. They are difficult to control with conventional measures. Preliminary assays identified five toxins that were more active than others. In two subsequent bioassays the activity of selected toxins was determined more accurately. Three Cryl proteins with relatively little homology were all found to be toxic, opening perspectives for controlling cocoa pod borer by expression of Cry proteins in transgenic plants.

Creation of BAC genomic resources for cocoa ( Theobroma cacao L.) for physical mapping of RGA containing BAC clones.

We have constructed and validated the first cocoa ( Theobroma cacao L.) BAC library, with the aim of developing molecular resources to study the structure and evolution of the genome of this perennial crop. This library contains 36,864 clones with an average insert size of 120 kb, representing approximately ten haploid genome equivalents. It was constructed from the genotype Scavina-6 (Sca-6), a Forastero clone highly resistant to cocoa pathogens and a parent of existing mapping populations. Validation of the BAC library was carried out with a set of 13 genetically-anchored single copy and one duplicated markers. An average of nine BAC clones per probe was identified, giving an initial experimental estimation of the genome coverage represented in the library. Screening of the library with a set of resistance gene analogues (RGAs), previously mapped in cocoa and co-localizing with QTL for resistance to Phytophthora traits, confirmed at the physical level the tight clustering of RGAs in the cocoa genome and provided the first insights into the relationships between genetic and physical distances in the cocoa genome. This library represents an available BAC resource for structural genomic studies or map-based cloning of genes corresponding to important QTLs for agronomic traits such as resistance genes to major cocoa pathogens like Phytophthora spp ( palmivora and megakarya), Crinipellis perniciosa and Moniliophthora roreri.