Soil Physico-Chemical Properties and Different Altitudes Affect Arbuscular Mycorrhizal Fungi Abundance and Colonization in Cacao Plantations of Cameroon

This study aims to investigate the abundance of AMF according to soil properties and altitudes in different cacao plantations of Cameroon. Physico-chemical analyses were made on soil samples collected from three agro-ecological zones. Soil samples were also used to evaluate directly the AMF abundance following the various altitudes and after trapping by sorghum plant. The results showed that soil properties, AMF spore abundances and colonization fluctuated significantly at different altitudes. The most represented texture was sandy loam. The bimodal zone presented a homogeneous texture (sandy loam) in all its localities. Cacao soil chemical characteristics showed that, the highest nitrogen rate (0.47%;p 0.05, Scott-Knott test) was recorded at Melong in a monomodal zone while Tonga in the Western highlands displayed the lowest rate (0.13%). Soil P concentration was significantly high in monomodal zones (Mbanga and Melong). Soil pH level indicated that the soil from Tonga in the Western highlands was neutral (pH = 6.67), and soils of other localities under study were acidic with the lowest (4.75) pH level recorded at Melong in a monomodal zone. In soil samples, the highest spore density (1.03 spores/g soil) was observed at Ntui in Bimodal zone, while the lowest spore density (0.26 spores/g soil) was observed at Bafang in the Western highlands. Root colonization showed that the sample from Bokito in a bimodal zone displayed the best frequency of mycorrhization (86.11%) while the sample from Bafang in the Western highlands recorded the lowest (27.11%). The PCA analysis highlighted that available phosphorus, pH and altitude all strongly correlated with AMF root colonization ability and can be used as a predictor of AMF colonization ability in cacao rhizosphere.

Characterization of the Sexual Self- and Cross-Compatibility in Genotypes of Cacao

Cacao is a tree crop of high economic importance and the subject of several studies aiming the genetic improvement of the species with the incorporation of characteristics that are agronomically important to create superior genotypes. In this context, this study aimed to characterize 22 genotypes of cacao for sexual compatibility, which is a trait that is strongly associated with yield. Additionally, we identify genotypes incompatible with clone TSH-1188 through controlled intercrosses, due to the great importance of this clone in cocoa breeding. Fifteen genotypes were characterized as self-incompatible and other seven as self-compatible, considering the retention index (20%) of self-pollinated flowers. Nine genotypes were characterized for the first time: CCN-16, COCA-3310, COCA-3370/5, EB-1017, IMC-119, IMC-97, LCTEEN-37A, NA-45, and UF-612. Cacao genotypes showed flower retention index ranging from 0.0% to 55.1% after 15 days of self-pollinations. The number of retained flowers varied after self-pollination of cacao genotypes at 1, 5 and 15 days after pollination. CCN-51, Catongo, CCN-10 and P-4B, were the genotypes with minor losses of flowers after 15 days of pollination, while IMC-119 and TSH-516 had 100% of loss of flowers between 1 and 5 days after pollination. Moreover, the flower retention index 15 days after cross-pollination with TSH-1188 ranged from 0.0% to 87.5% among the tested genotypes. The results of cross-pollinations identified two genotypes that are incompatibles, EB-1017 and IMC-119. This result is important for future allelic studies of incompatibility in cacao, because it might indicate that these genotypes share the same self-incompatibility allele(s). We conclude, therefore, that this study enable the characterization of this working collection of cocoa germplasm for sexual compatibility and this information is important for subsequent crosses between cacao genotypes.

Relationships among Bean Yield Traits in Some Cacao(Theobroma cacao L.)Genotypes

Cacao(Theobroma cacao L.)produces the cocoa bean,a major foreign exchange earner for most West African countries and many smallholders’enterprise.Ample production of cacao is however limited by declining yield among other factors.This study aimed at determining the correlations of the phenotypic traits that were related to the yield of the cacao genotypes.Nine new cacao hybrids were produced from some high-yielding parents in the research farm of Cocoa Research Institute of Nigeria,Ibadan and evaluated from 2012 through 2014 in Owena(7°11′N,5°1′E),Ondo state,Nigeria.Analysis of variance,character correlations and path coefficient analysis were used in the analysis of the relationships among the genotypes.Analysis of variance revealed significant(p≤0.05)variations for number of rows,weight of beans per fruit,fresh weight of one bean,weight of one bean after fermentation,pod value,dry bean length,weight of beans(per fruit)after fermentation and pod index.The study concluded that significant genotypic and phenotypic correlations existed among some of the pairs of the fruit and bean characters with one another and with pod index,suggesting that the contribution of these characters is either positive or negative to growth and yield in the cacao genotype,and that fruit and bean traits are determinants of bean yield in cacao.

Comparison of Antioxidant and Antimicrobial Activities of Acetone and Water Extracts of <i>Theobroma cacao</i>Beans

This present study compared antioxidant and antimicrobial activities of acetone and water extracts of Theobroma cacao beans against Escherichia coli. Total phenolic content (TPC) in both extracts was estimated by the Folin-Denis reagent. The present study showed that the 70% (v/v) acetone extract had a higher extraction yield and TPC (37% and 109 mg TAE g-1 dry weight) than the water extract (33% and 76 mg TAE g-1 dry weight). The antioxidant activities of both extracts were estimated by the DPPH Scavenging Assay. The extract obtained using 70% (v/v) acetone showed higher antioxidant activity (54%) compared to the antioxidant activity obtained using water (34%). Antimicrobial activities of acetone and water extracts from Theobroma cacao were measured against Escherichia coli and were screened by agar well diffusion method and further confirmed with the disc diffusion method. The bacterial growth was measured in Mueller Hinton agar. The extracts inhibited the growth of the Escherichia coli cultured, and the acetone extracts showed antimicrobial capacity comparable or equivalent, as seen in commercial ampicillin.

Effects of Pulp Preconditioning on Total Polyphenols, O-diphenols and Anthocyanin Concentrations during Fermentation and Drying of Cocoa （Theobroma cacao） Beans

Changes in total polyphenols, O-diphenols and anthocyanin concentrations during fermentation and drying of pulp pre-conditioned cocoa （Theobroma cacao） beans were investigated using standard analytical methods. Increasing pod storage, fermentation and drying led to variable reductions in total polyphenols, O-diphenols and anthocyanins content of the beans. The rates of reduction were however more pronounced during fermentation than pod storage and drying. Storage of cocoa pods between 3-7 days with 6 and 7 days of fermentation and drying respectively retained 85%-90% of the total polyphenol and O-diphenols of the cocoa beans. Similarly, anthocyanin content of beans from the 10 days of pod storage decreased by 70% in the sixth day of fermentation. Pod storage decreased the anthocyanin content at all periods of fermentation. These suggest that the post-harvest treatments of pod storage, fermentation and drying all results in variable reductions in polyphenolic content （total polyphenols and O-diphenols） and anthocyanins content of cocoa beans.

Relationships between Cadmium in Tissues of Cacao Trees and Soils in Plantations of Trinidad and Tobago

The primary source of cadmium in cocoa beans has been linked to its direct uptake by the cacao plant from cadmium contaminated soils. This research was conducted to evaluate and interpret significant relationships between cadmium levels in tissues of the cacao plant and soils from cocoa-growing areas in Trinidad and Tobago. Total (HNO3-extractable) concentrations of cadmium in both tissues and soils were determined. The levels of cadmium measured varied in the order: leaves > pods > shells > nibs > soil. Cadmium levels in all the cacao tissues analyzed were significantly (p < 0.05), positively and strongly correlated with each other. Additionally, significant (p < 0.05) positive relationships were also identified between Cd in cacao tissues and corresponding total HNO3-extractable Cd levels in soils. These findings suggest that they can possibly be used as predictive tools for assessing Cd levels in cacao.

Effect of Pectin Lyase Enzyme on Fermentation and Drying of Cocoa (Theobroma cacao L.): An Alternative to Improve Raw Material in the Industry of Chocolate

Cocoa (Theobroma cacao), in all its presentations, is consumed all over the world and is one of the main drivers of the economic in several countries. The world’s Cocoa tendency is focused on developing special beans. This category is subject to postharvest processes of utmost importance such as the fermentation and dry, which are currently carried out with traditional and poorly effective devices, which need to be improved to obtain a high quality product. The aim of this study was to evaluate the influence of the pectin lyase enzyme (E.C.4.2.2.10) on the postharvest cocoa process. We evaluated the enzyme dosage (1.0% and 0.5%) in fermentation and its effect on the variables temperature, acidity and drying time by convection at 60°C. The Pectin lyase activity during fermentation does not cause a significant effect on the variables of temperature and acidity;however, the drying process time required to achieve 7.0% moisture was reduced. The enzyme dosage of 1.0% was the best result, the amount of exudate obtained (115 ml) during fermentation and the best degree of fermentation (77% ± 3.8) were increased and further shows a change in porosity facilitating the scale surface and internal moisture diffusion. The drying rate (Nw) expressed in kgwater/m2 * min was determined based on the empirical model of Newton, where the higher speed was obtained during the falling period. In conclusion, enzyme dosage 1% was the best concentration evaluated because weaken grain husk, which allowed an adequate fermentation,and subsequent time drying reduction until 10.8 h.

The Application of BBCH Scale for Codification and Illustrations of the Floral Stages of Caribbean Fine Cacao Theobroma cacao L.

Cocoa （Theobroma cacao L.） is the major input into the chocolate manufacturing industry, particularly the fine or flavor of the indigenous variety Trinitario, which is the hybrid of Crillo and Forastero. One key problem encountered by researchers is the inability to differentiate the various phenological stages of the cocoa reproductive cycle and to assess or predict the yield. This study sets out to map the various stages of the reproductive biology of cacao and to develop a set of photographic illustrations that appropriately describe the floral biology and development stages of Caribbean free cocoa as a tool for agronomist and farmers. It used a modified version of the codification of the ＂extended BBCH＂ scale （Biologische Bundesantalt, Bundessortenamt and Chemische Industrie, Germany）, which is a numerical system that differentiates between principal, secondary and tertiary growth stages. The phases and all floral stages to fruit development are described and illustrated in a set of photographs, which can now be used for training and referencing purposes. This is the first attempt to undertake and document this study.

Changes in Nib Acidity, Proteolysis and Sugar Concentration as Influenced by Pod Storage and Roasting Conditions of Fermented Cocoa （Theobroma cacao） Beans

Changes in nib acidity, protein and sugar concentration during roasting of pulp pre-conditioned and fermented cocoa （Theobroma cacao） beans were investigated. A 4 - 4 full factorial design with the principal experimental factors as pod storage （0, 3, 7 and l0 d） and roasting time （0, 15, 30 and 45 min） were used. The roasted samples were evaluated for pH, titratable acidity, protein content and sugars concentrations using standard methods. Increasing pod storage caused consistent increases in pH with concomitant decreases in titratable acidity, whereas increasing roasting time caused only marginal and insignificant changes in pH but significantly decreased the titratable acidity. The protein content decreased significantly （P 〈 0.05） with increasing pod storage and roasting time. Reducing sugars increased marginally with increasing pod storage treatments whiles increasing roasting time significantly （P 〈 0.05） decreased the reducing sugars of the beans for all pod storage. The non-reducing sugar and total sugar content of the beans decreased significantly from 3.493 mg/g to 2.641 mg/g and from 9.284 mg/g to 8.891 mg/g, respectively, for pods stored from 0 to 10 days while roasting time caused slight decreases in non-reducing sugars with a considerable decrease in total sugars. Pod storage up to seven days decreased considerable the nib acidity （non-volatile acids）, non-reducing sugars and total sugars while roasting up to 45 min at 120 ℃ caused dramatic decreases in the nib acidity and reducing sugars with only marginal decreases in non-reducing sugars and total sugars.

Chlorocholine Chloride Induces Cacao Reproductive Development Leading to Improved Fruitlets Productivity of Cacao Trees in the Field

Fruitlet abscission （cherelle wilt） is a major factor contributing to lower productivity of cacao plantation in Indonesia. An attempt was made to reduce such cherelle wilt by spraying plant growth retardants （PGRs） on cacao trees at about 10 weeks before flowering session. The treatment was repeated every three weeks. Both the flowering initiation time and the number of flowers on the treated trees were improved significantly. With the most effective composition, flowers were initiated at 24 days after spraying （DAS）, where as the flowering initiation was 11-day later in the controls. At the 35 DAS, the chlorocholine chloride （CCC） 2,000 treatment induced about 20-80 folds more flowers than the controls. Extended observation was made to assess the treatment effect on the fruiting capacity. The treatments were found to improve fruiting ability by increasing the number of fruits set on the trees and shortening the time for fruit setting. At 20 weeks after the first spraying （WAS） there were on average 12.57 fruits per CCC 2,000-treated tree, as opposed to only 4.14 fruits per untreated tree. Analyses of the metabolites content in the flower cushions of the trees indicated that the reproductive growth is significantly correlated with the increased metabolites particularly the reduced sugar. Exogenous addition of sucrose to the PGR treatment gave greater improvement mainly in the fruiting. In conclusion, foliar spray of CCC induces flowering of cacao tress in the field that leads to improve the fruitlets productivity.

Effects of Fermentation and Drying on the Fermentation Index and Cut Test of Pulp Pre-conditioned Ghanaian Cocoa （Theobroma cacao） Beans

Investigations were conducted to establish effects of fermentation and drying on the fermentation index （FI） and cut test of pulp pre-conditioned Ghanaian cocoa beans using a 4 x 3 full factorial experimental design with the principal factors being pod storage （0, 3, 7 and 10 d） and fermentation time （0, 3 and 6 d） to study the changes occurring during the fermentation process. The study also used a 4 x 3 full factorial design with pod storage （0, 3, 7 and 10 d） and drying time （0, 3 and 7 d） being the principal factors investigated to study the changes occurring during the drying process. FI and cut test of the beans were studied during fermentation as well as the drying process. FI of the beans increased significantly with pod storage and fermentation but decreased slightly during drying. FI of the unfermented beans increased slightly from 0.674 for the unstored pods to 0.763 after 10 days of pod storage. The FI of the fermented beans （six days fermentation） also increased from 1.390 for the unstored pods to 1.424 for pods stored for 10 days. It decreased from 1.389 at the start of drying for the unstored pods to 1.105 for pods stored for 10 days at the end of drying （seven days）. FI of all the beans were however, above 1.0 at the end of fermentation and drying for all pod storage treatments. Cut test revealed that storage of pods for 3, 7 and 10 days increased the percentage of brown beans by 66%, 94% and 72%, respectively, by the sixth day of fermentation. Percentage of brown beans decreased to 61%, 76% and 63%, respectively, for pods stored for 3, 7 and 10 d at the end of drying （seven days）. Cocoa pods can be stored for up to 10 days, fermented for six days and dried for seven days with the necessary formation of brown pigments characteristics of well fermented and dried cocoa beans.

Selection of Cocoa Tree (<i>Theobroma cacao</i>Linn) Endophytic Bacteria Solubilizing Tri-Calcium Phosphate, Isolated from Seedlings Grown on Soils of Six Producing Regions of C&ocirc;te d’Ivoire

Phosphorus (P) is an essential macronutrient for the growth and development of cocoa tree (Theobroma cacao Linn). Most of the soils used for cocoa farming in C?te d’Ivoire are low in phosphorus. But cocoa farmers generally have a widespread reluctance to invest in chemicals fertilizers due to high costs and environmental associated risk. Phosphate Solubilizing Bacteria (PSB) are kwon to play an important role in supply of phosphorous to plants in a sustainable manner in P deficient soils. The aim of this research was to screen the endophytic bacteria of cocoa nurseries able to solubilize tri-calcium phosphate. Seedlings of two varieties of cocoa (P7 and NA32) and seedlings of an all-comer, were grown on eighteen (18) samples soils collected in six producing regions of C?te d’Ivoire. A total of 218 endophytic bacteria were isolated and tested on the Pikovskaya’s agar medium, containing Ca3 (PO4) 2. The colonies with clear zone around the microbial growth were suspected as phosphate solubilizing bacteria. Out of 218 bacteria, 90 (41.28%) showed a clear zone around colonies after 7 days of incubation. The Phosphate Solubilization Index (PSI) ranged from 20% to 200%. Bacterial isolated from the soils of Duékoué locality showed the highest mean index of 137.67%. Five PSB (CEBSP5, CEBSP6, CEBSP7, CEBSP8, and CEBSP9) from Duékoué soils and two PSB (CEBSP12 and CEBSP13) from Soubré soils have a PSI ranged from 150% to 200%. Further study in greenhouse and in field condition will confirm the use of these PSB as biofertilizer to increase the available P content in soils, reduce environmental pollution and promote sustainable agriculture.

Correlation and Path Coefficient Analyses of Yield in Cacao(Theobroma cacao L.)

Cacao(Theobroma cacao L.)is an important commodity tree crop which produces the cocoa bean,a major source of income for most West African countries and many smallholder farmers.Declining yield of cacao is a major limitation to cocoa production in Nigeria.This study aimed at determining the correlations of the phenotypic traits that were related in the yield of the cacao genotypes.Nine cacao hybrids produced from some high-yielding parents in the research farm of Cocoa Research Institute of Nigeria,Ibadan,Nigeria were evaluated from 2012 through 2017 in Owena(7°11’N,5°1’E),Ondo state,Nigeria.Character Correlations and Path Coefficient Analysis were used in the description of the performance of the genotypes.The study concluded that significant genotypic and phenotypic correlations existed among many of the pairs of the fruit and bean characters with one another and with pod index,suggesting a complex contribution of these characters either positively or negatively to growth and yield in cacao,and that fruit and bean traits are determinants of yield in cacao.

Inheritance of the Number of Ovules per Ovary and Selection of Cacao Genotypes

We have characterized the number of ovules per ovary (NoOV) in cacao plants originated from crossing the clones CCN 51 and TSH 1188 (segregating progeny) and determined the heritability of this characteristic in order to select plants with higher NoOV in this progeny. The NoOV was calculated as the average of 10 flowers per plant and ranged from 44.8 to 58.6 between the six clones (two parents and four clones belonging to their genealogy). In the progeny (n = 209 plants) the NoOV averaged 54.3 (range 44.1 to 67.8). The NoOV was distributed uniformly among the progeny indicating that this trait is conditioned by polygenes. Its heritability was estimated at 67.7%. The 32 plants with NoOV similar or superior to the genitor CCN 51 (highest amount among clones) were selected for use in the genetic improvement program. We showed that this population is suitable for genetic mapping, molecular marker identification and selection of superior cacao genotypes.

Role of fermentation and microbes in cacao fermentation and their impact on cacao quality

Cocoa seed fermentation involves a well-stabilized succession of microbial processes and the action of enzymes.Microorgan-isms(yeast,lactic acid bacteria,and acetic acid bacteria)play a critical role in cacao fermentation by degrading mucilage pulp by producing a stabilized succession of enzymatic reactions that contribute to its quality.Nowadays,the majority of cacao fermentation practiced in our planet is in the traditional form to produce aroma flavor.The art of using starter culture to substitute naturally occurring microorganisms in cacao fermentation is crucial for the development of the aroma and flavor of cacao for sustainable uniformity and quality improvement and unique cacao flavor development.In this sense,this review presents an overview of cacao fermentation process and its impacts on quality,starter culture and microorganisms involved in cacao fermentation process,cacao bioactive compounds and its health benefits.Moreover,the research perspectives needed and the advance made in terms of fermentation process in order to reduce fermentation period are reviewed.

基于ATR-FTIR及气质联用技术对可可提取物的研究

本研究使用红外光谱技术及GC/MS技术对5种不同类型的可可提取物进行相似度的分析,中国2号可可提取物与英国1号提取物红外光谱相似度达到98.76%,中国1号可可提取物与英国2号可可提取物相似度达到73.43%。5种类型可可提取物共鉴定出48种香气成分,英国1号鉴定出25种,英国2号鉴定出20种,美国1号鉴定出28种,中国1号鉴定出22种,中国2号鉴定出25种。对5种类型可可提取物所鉴定出的香气成分进行主成分分析,可以把中国1号可可提取物与英国2号可可提取物归为一类,把中国2号可可提取物、英国1号可可提取物及美国1号可可提取物归为另一类。通过GC/MS技术结合主成分分析可以对不同类型的可可提取物进行区分,对比传统感官风味评价更具优势,本研究可以为香精产品中原料的替换提供一定的理论依据。

可可果实主要农艺性状相关性及产量因素的通径分析

以79份可可资源为材料,对其果实和种子性状进行相关和通径分析。结果表明:(1)每果种子干重与其他性状的相关系数均表现为正相关,果实经济系数与果重、果壳重、果长、果径围、果壳厚表现出极显著负相关,说明果实越大,就会有更多光合产物分配到果壳中。(2)通径分析结果表明:果壳重对每果种子干重的效应为负,其它性状对每果种子干重均为正向效应;其中单粒重对每果种子干重的贡献最大,单粒重每增加1个单位,每果种子干重会提高0.663 8个单位。每果粒数和果重每增加1个单位,每果种子干重分别提高0.625 4和0.327 9个单位。因此,可可育种过程中要着重关注单粒重、每果粒数和果重等性状。

可可种质资源多样性与创新利用研究进展

就可可种质资源在起源、分布、传播、分类、鉴定及种质创新与利用等方面取得的研究进展进行综述。可可属植物有22种,仅可可(Theobroma cacao)和大花可可(Theobroma grandiflorum)用于栽培。可可种质资源多样性丰富,其分类研究经历了传统形态学划分的Criollo、Forastero和Trinitario遗传类群到现代分子标记划分的Maran^on、Curaray、Criollo、Iquitos、Nanay、Contamana、Amelonado、Purús、Nacional和Guiana遗传类群。目前,可可的生产性品种选育仍以实生选种和芽变选种为主。分子标记的开发、遗传连锁图谱的构建和全基因组测序的完成,为实现可可分子标记辅助育种提供了技术保障。

可可蔗糖磷酸合成酶基因家族进化及组织表达分析

在高等植物中,蔗糖磷酸合成酶（Sucrose phosphate synthase,SPS）是蔗糖合成的限速酶。在多种植物中都发现了SPS基因,而可可中尚未见相关报道。通过分析可可基因组数据库,鉴定出4个SPS候选基因,依次命名为TcSPS1、TcSPS2、TcSPS3和TcSPS4。4个基因的编码区（CDS）长度在3 075~3 228 bp之间,外显子数目为12~14,预测蛋白的平均分子量为118.15 ku,等电点均小于7。进化分析结果表明SPS基因家族分成3个亚族;TcSPS1和TcSPS2属于ClassⅠ亚族,TcSPS3和TcSPS4分别属于ClassⅡ亚族和ClassⅢ亚族。实时荧光定量PCR分析结果表明,TcSPS1与TcSPS2在树皮和果实中高量表达,TcSPS3和TcSPS4主要在叶片中表达。伴随着叶片和花蕾生长发育,各TcSPS基因表达量均呈现出上升的趋势,表明其与主要光合产物--蔗糖的合成或再合成有密切联系,参与可可“源库”器官中光合产物分配。

可可核心种质遗传多样性及果实性状与SSR标记关联分析

利用15对SSR引物,分析具有广泛来源的70份可可资源的遗传多样性。结果表明,15对引物共扩增出76个条带,其中50个多态性条带,占总带数的65.8%。70份可可资源间遗传相似系数（SM）在0.341~0.943之间,平均值为0.625,说明可可资源具有丰富的遗传多样性。在相似系数为0.65水平上,可将70份可可资源分成10类。利用Structure 2.3.2软件分析群体结构,结合可可果实相关性状的表型数据,采用Tassel 2.1的一般线性模型（General linear model,GLM）进行关联分析;结果表明18个位点与果重、果壳重、果长、果径围、果壳厚显著相关（p〈0.05）,各位点对表型变异贡献率为5.5%~13.1%。