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.

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.

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.

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.

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.

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.

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.

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.

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.

Effects of Agricultural Practices on Soil Organic Nitrogen Fractions in an Inceptisol of a Cocoa Plantation

The effect of agricultural practices on soil organic nitrogen （N） fractions in a cocoa plantation has not been much revealed till now. Despite the fact that soil organic N has been long admitted for its importance to maintain soil fertility. Presented field experiment was conducted in Kaliwining Experimental Station, Indonesian Coffee and Cocoa Research Institute （ICCRI）, Jember, East Java, Indonesia, to investigate the effect of cocoa farm management, namely fertilization, weeding and soil tillage on the content of soil organic N fractions. The design of experiment was arranged in a split-split plot with two levels of weeding as main plots, two levels of soil tillage as subplot and three different fertilizer treatments as sub-sub plot. The analysis of soil N including total N and soil organic N fractions, namely, total hydrolized N, ammonium N, amino sugar N and amino acid N, were performed. The result showed that the effect of fertilization treatment was significant to the content of total N, ammonium N, amino sugar N and amino acid N. No-tillage treatment resulted in total N and amino sugar-N content increasing by 8% and 24%, respectively, over tillage treatment. Slashing treatment caused increase of the total N by 3% from herbicide treatment, whilst decrease of ammonium N and amino sugar N by 7% and 24%, respectively.

Charcoal Production in a Cocoa-Farming Area： Plant Species Used and Their Sources

The authors investigated the plant species used for charcoal production and their sources and the willingness of the producers to plant charcoal-making species. Furthermore, the authors indirectly determined the involvement of farmers in charcoal production in Atwima Mponua District of Ghana. A total of 120 producers were interviewed and simple ranking was used for the analysis. Twenty-three plant species were used with the first, second, third and fourth which were Celtis mildbraediL Cylicodiscus gabunensis, Margaritoria discoides and Amphimas pterocarpoides respectively. However, the Amphimas pterocarpoides was not among the traditional charcoal-producing species. The wood used came from seven different sources. None of the charcoal producers had a plantation of charcoal-producing species. Only 13 % of the producers were willing to plant, 87% of them who were not willing to plant indicated that they would leave charcoal production if the resource got finished. The 120 charcoal producers were made up roughly of 56% males, and 44% females and in which 69.2% of them were farmers.

施用生物有机肥对可可苗期生长及土壤酶活性的影响

通过温室盆栽试验研究施用不同生物有机肥对可可苗期生长及土壤酶活性的影响。结果表明:施用生物有机肥BIOa和BIOb的处理可可植株干重,总根长、根系直径、根系表面积和根系总体积显著高于对照(单施化肥)及施用牛粪的处理;施用生物有机肥BIOa的处理可可叶片净光合速率、蒸腾速率和叶绿素含量与对照相比分别增加了169%、270%和28%;施用BIOa的处理土壤酸性磷酸酶、脲酶和蔗糖酶活性与对照相比显著提高了98.7%、92.2%和83.2%。综上所述,施用生物有机肥BIO可显著促进可可地上部植株和根系生长,同时也可显著提高相关土壤酶活性。

可可泛素结合酶基因家族的生物信息学初步分析

泛素结合酶（E2s）促进底物泛素化或者与E3s链接,是靶蛋白泛素化的关键酶,在泛素-蛋白酶体途径中起重要作用。利用可可（Theobroma cacao L.）全基因组测序数据,共鉴定出45个E2s基因家族成员,包括39个UBC和6个UEV基因。通过生物信息学方法,对可可E2s家族的基本理化性质、基因结构、二级结构预测、亚细胞定位、进化关系等方面进行初步分析。结果表明：可可E2s基因CDS长度在441（Tc UEV3）～3 651 bp（Tc UBC7）之间,对应的编码蛋白氨基酸数目在146（Tc UEV3）～1 216 aa（Tc UBC7）之间,编码蛋白分子量在16.39～134.71 ku之间。E2s基因外显子在1～11之间,多数基因外显子数目在5～7之间。E2s基因在10条染色体上均有分布,1号染色体为7个,数量最多;6号染色体2个基因,数量最少。可可E2s蛋白大多为不稳定蛋白,且均为亲水性蛋白,其二级结构以α-螺旋和无规则卷曲为主要构成元件。大多数可可E2s蛋白定位在细胞核,少数定位在内质网或者细胞质。进化树分析表明：可可E2s蛋白被分为20个亚家族,包括16个UBC亚家族和4个UEV亚家族,第Ⅵ亚家族E2s数目最多为9个;E2s家族蛋白在物种进化过程中具有高度保守性。

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%。

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。

椰子可可间作下种植密度对作物产量及经济效益的影响

研究椰子可可间作条件下,不同种植密度可可对椰子和可可产量及经济效益的影响。试验共设7个处理:在椰园间作不同密度的可可2.0 m×2.0 m(A)、3.0 m×2.0 m(B)和3.0 m×3.0 m(C),以单作椰子6.0 m×6.0 m(CN)和单作可可2.0 m×2.0 m(CA)、3.0 m×2.0 m(CB)和3.0 m×3.0 m(CC)为对照。结果表明:间作不同密度的可可均显著提高椰子叶片全氮、全磷和全钾等养分含量;间作园土壤有机质、全氮、有效磷、速效钾等养分含量均显著高于单作椰园;间作可可使椰子产量显著提高,间作可可产量高于单作可可,但差异不显著;处理C的产投比和土地当量比较高。在海南椰园,采用3.0 m×3.0 m的密度间种可可有利于经济效益提高,并起到节本增效的作用。

基于MaxEnt模型的可可潜在适宜分布研究

根据野外采样和文献查阅,系统整理了可可的地理分布记录,并利用MaxEnt生态位模型和ArcGIS软件对可可的潜在适宜分布范围进行预测。结果表明:北美洲南部、中南美洲北部、非洲西部、亚洲东南部以及太平洋美拉尼西亚群岛地区均是可可的潜在适宜分布区域。其中,中国海南、台湾南部、云南西双版纳、广东雷州半岛也属于可可的适生范围。经ROC(Receiver operating characteristic)曲线分析法验证,MaxEnt模型的AUC(Area under curve)值为0.977,表明预测结果具有较高的可信度。各环境变量重要性的Jacknife检验表明,极端最低温度、年降雨量、年温度变化范围、最暖季降雨量对可可的潜在分布影响最大。