Description of the phenological, vegetative and yield traits of the accessions per habitat
The process of data management included the computation of mean squares for the assessed phenological, vegetative and yield traits of the accessions with the sampling habitats considered as the treatment factor. The error mean squares served in the multiple comparison of means reported in Table 1.
Regarding the phenological traits, the accessions from the habitat of Zh have the longest period from seeding to first male (102.39 d) and first female (108.14 d) flower appearances, and the longest period from seeding to physiological maturity (153.95 d). For those traits, the accessions from Tiassale and Soubre are not significantly different from those of Zh. And, accessions from Tiassale and Zh have the longest periods from seeding to 50% flowering. On the other hand, accessions from Korho, Ferke, Bondu and Burki develop their first male and female flowers and attain 50% flowering in a very short period. They also reach physiological maturity faster. Accessions from Korho, however, have the longest period from seeding to 50% emergence (6.07 d) and accessions from Bondu have the longest period from first female flower appearance to physiological maturity (53.04 d).
For the vegetative traits, accessions from Tiassale and Soubre have the largest girth size (4.43 cm and 4.63 cm, respectively). Accessions from Tiassale have the longest (24.98 cm) and widest (19.94 cm) leaves, the longest male (16.2 cm) and female (4.03 cm) peduncles and the longest petioles (34.94 cm). The measures for those organs on accessions from Soubre rank second to those of Tiassale. On the other hand, accessions from Korho, Ferke, Bondu and Burki are characterized by smaller girth size, smaller leaves, smaller petioles and smaller peduncles of male and female flowers. But the accessions from Bondu are the tallest (586.91 cm) followed by the accessions from Ferke (489.20 cm). And the accessions from Zh are the shortest (417.38 cm).
For the flowering and yield traits, accessions from Tiassale and Soubre show the largest numbers of male (27.33 units and 22.58 units, respectively) and female (5.22 units and 6.05 units, respectively) flowers per plant, largest numbers of fruits per plant (2.78 units and 2.53 units, respectively) and largest measures of all fruit-related traits. Their seeds are very large, but in small numbers. In contrast, accessions from Korho, Ferke, Bondu and Burki have the smallest numbers of male and female flowers per plant, the smallest numbers of fruits per plant and the smallest measures of fruit-related traits. They have large numbers of seeds, but their seeds are smaller, except the seeds of the accessions from Burki. Refer to Table 1 for more detailed information.
Variability of the phenological, vegetative and yield traits
Table 2 shows the spread of the phenological and morphological traits of the assessed accessions of C. moschata. All the evaluated traits showed very wide ranges of distribution of the observations. Some conspicuously wide ranges of traits include number of days to 50% flowering (DTF) that goes from 52 to 152 d, plant height with a minimum of 48 cm and a maximum of 1510 cm, diameter of the fruit that is between 5.8 cm and 35 cm, weight of the fruit that varies between 150 g and 10,930 g and number of seeds per fruit that spreads in the interval from 32 units per fruit to 729 units per fruit. Excluding the number of days to 50% emergence (DTE), all the other assessed traits have remarkably wide ranges of phenotypic expressions (Table 2). All the traits but DTE, DTF, days from first female flower appearance to fruit maturity, fruit length and length of the dry seed, had outliers. The number of outliers ranged from 1 to 67. Except the outliers observed with the width of the dry seed, all the outliers were above 1.5*IQR + Q3 where IQR is the inter-quartile range and Q3 is the third quartile. The presence of outliers is indicative of the richness and large variability of the population of accessions. The outliers are exceptional performances that fall outside the normal distribution of the observations. They are a stock of unusual traits that can be used in a crop improvement program when beneficial. For example, the observed outliers for diameter of the fruit, weight of the fruit or thickness of the pulp can be used in a breeding program for the improvement of fruit yield. Similarly, outliers for beneficial traits related to the seed can be used to improve C. moschata crop for seed yield. Besides, the computed mean squares (data not reported) showed highly significant variations between accessions for the assessed traits. They all yielded p-values less than 0.01, providing additional support to the evidence of large variability among the accessions of C. moschata of Cote d’Ivoire. The computed standard deviation, and median absolute deviation for each trait are additional evidence. We should note that in most cases, the mean squares associated to year (data not reported) were not significant, indicating the relative stability of the assessed traits.
The components of variance, the quantitative genetic differentiation, the overall mean, and the coefficients of variation are reported in Table 3. The lme4 package37 used in the determination of the components of variance, does not provide p-values in the analysis of mixed or random models. The reported quantities in Table 3 are not accompanied with tests of significance. It is worth mentioning that the respective units of measure of the assessed traits are squared for the variances and the evaluated estimates will be reported without the units of measure. The phenotypic variance ((sigma_{p}^{2})) is partitioned into variance between morphotypes or genotypic variance ((sigma_{g}^{2})), and within morphotypes or residual variance ((sigma_{e}^{2})). For the class of phenological traits, considerable genotypic variances were observed with days to 50% flowering (266.21) and days to first male flower appearance (254.40), compared with their respective residual variances (148.13 and 199.50). Regarding the class of vegetative traits, only the peduncle length of male flowers had a genotypic variance (9.22) greater than its residual variance (8.86). In the class of flowering and yield traits, 8 of the 15 traits assessed showed large genotypic variances in comparison with their respective residual variances. They are number of female flowers per plant ((sigma_{g}^{2}) = 3.02 versus (sigma_{e}^{2}) = 2.36), length of the fruit ((sigma_{g}^{2}) = 53.96 versus (sigma_{e}^{2}) = 48.97), diameter of the fruit ((sigma_{g}^{2}) = 37.17 versus (sigma_{e}^{2}) = 16.76), volume of the fruit ((sigma_{g}^{2}) = 10,713,468 versus (sigma_{e}^{2}) = 3,904,590), weight of the fruit ((sigma_{g}^{2}) = 5,413,819 versus (sigma_{e}^{2}) = 1,420,187), diameter of the cavity enclosing the seed ((sigma_{g}^{2}) = 19.12 versus (sigma_{e}^{2}) = 7.75), thickness of the fruit pulp ((sigma_{g}^{2}) = 1.11 versus (sigma_{e}^{2}) = 0.94) and weight of the fruit pulp ((sigma_{g}^{2}) = 5,979,212 versus (sigma_{e}^{2}) = 1,088,750). For a trait to have a lager genotypic variance than the residual variance is synonymous to a relative ease of improvement of the crop for that trait through a breeding program.
The coefficient of variation (CV) is another statistic that measures variation. It is actually the dispersion of a trait per unit measure of its mean, which can be used to compare variations of traits with different measurement units or different scales. As a rule-of-thumb, a coefficient of variation greater than 20% is indicative of large variation for the trait. The phenotypic coefficient of variation is considerably high for 25 of the 28 assessed traits. Only the number of days from seeding to physiological maturity, the first and second longest axes of the dry seed show coefficients of variation less than 20%. Traits with very large phenotypic coefficients of variation include the peduncle length of female flowers ((CV_{p}) = 93.98%), weight of the pulp ((CV_{p}) = 92.96%), volume of the fruit ((CV_{p}) = 89.17%), weight of the fruit ((CV_{p}) = 78.30%) and number of female flowers per plant ((CV_{p}) = 65.81%). With respect to the residual coefficients of variation, only the number of days from seeding to 50% emergence and number of days from first female flower appearance to physiological maturity have residual coefficients of variation greater than 20%, among the phenological traits. All the vegetative traits have residual coefficients of variation greater than 20%, and show a near-perfect linear relation (r = 0.98; p < 0.001) with the phenotypic coefficients of variation. From that observation, we may conclude that the variations in the phenotypic expressions of the vegetative traits are largely due to the variations within morphotypes. For the flowering and yield traits, all the residual coefficients of variation are greater than 20%, except the first and second longest axes of the seed. The genotypic coefficient of variation is less than 20% for all the phenological and vegetative traits except the peduncle length of male flowers with a coefficient of variation of 33.01%. On the other hand, 13 of the 15 flowering and yield traits have genotypic coefficient of variation greater than 20%. Among them, are the weight of the pulp with a genotypic coefficient of variation of 90.26%, the volume of the fruit with a genotypic coefficient of variation of 84.20% and the weight of the fruit with a genotypic coefficient of variation of 75.12%. Besides, for the flowering and yield traits, the genotypic coefficients of variation are highly correlated (r = 0.99, p < 0.001) with the phenotypic coefficients of variation in a near-perfect linear trend ((b = 0.97;p < 0.001)). That finding forms the basis to infer that most of the variations in the phenotypic expressions of the flowering and yield traits are largely caused by genotypic variability, without dismissing the contribution from the variability within morphotype.
The quantitative genetic differentiation, termed (Q_{ST})38, is broadly the ratio of genotypic variance to phenotypic variance. It is closely related to the estimator of heritability. It scales between 0 and 1. It is well suited to the genetic analysis of morphological traits. In this study, the computed estimates of (Q_{ST}) take values between 0.01 and 0.73. A value of (Q_{ST}) = 0.28 is considered moderate quantitative genetic differentiation38. And it is easy to see that a value of (Q_{ST} = tfrac{1}{3}) implies that the between-morphotype variance is equal to the within-morphotype variance for a morphological or phenological trait. And a (Q_{ST}) = 0.5 means the between-morphotype variance is twice the within-morphotype variance and can be qualified as a considerably large estimate of genetic differentiation. Based on our estimates of genetic differentiation, we may affirm that moderate to considerably large differentiation has occurred for several phenological, vegetative and yield traits and the differentiation is particularly high for fruit-related traits such as diameter of the fruit ((Q_{ST}) = 0.53), diameter of the cavity enclosing the seeds ((Q_{ST}) = 0.55), volume of the fruit ((Q_{ST}) = 0.58), weight of the fruit ((Q_{ST}) = 0.66), and weight of the pulp ((Q_{ST}) = 0.73). The observed morphological differences of the accessions in the plots of the experimental trials led to the attempt to regroup the morphotypes of C. moschata in clusters with unsupervised methods. The results are given in the section below.
Segmentation of the accessions and identification of ecotypes
The clustering was first performed with base R39. The NbClust package40 determined 3 clusters based on the majority rule. The hclust object was then used with the ape package41 to create the circular phylogenetic tree of Fig. 1. The cluster validation was verified with the fpc package42. The tree regrouped morphotypes according to their phenological and morphological similarities in three clusters that also reflect the 3 geographical zones labeled forest, mountain, and savannah. The 3 zones are characterized by distinct bioclimatic parameters (seasons, rainfall, temperature and humidity, see Table 4). Accessions from the same geographical zone were similar and accessions from different zones were distant. In general the forest region is characterized by two rainy seasons that alternate with two dry seasons, an accumulated annual rainfall of 1200 mm, an average daily temperature of 27 °C and a relative humidity of 70%. The mountain region has one very long rainy season with an accumulated annual rainfall of 1400 mm and a short dry season, an average daily temperature of 27 °C and a relative humidity of 69%. The savannah region has a long dry season followed by a short rainy season with an accumulated annual rainfall of 900 mm, an elevated daily temperature of 29 °C, and an elevated relative humidity of 80%. The forest region includes the morphotypes of Tiassale and Soubre, the mountain region has the morphotypes of the habitat of Zh and the savannah region has the morphotypes of the habitats of Bondu, Ferke, Korho, and Burki. The K-means algorithm was also used to cluster the accessions and the results similarly showed that the accessions within a cluster were from the same geographical zone as defined above (data not shown). The three regions showed large genotypic diversity among the accessions of C moschata. Figure 2 gives a picture of the diversity of the accessions with the dissimilarity measures between and within geographic regions representing the main growing areas of C. moschata. The diversity is presented by the quartiles, the minimum and the maximum rank of the dissimilarities within a region. The width of the box is determined by the number of morphotypes considered in the drawing of the boxplot and is not related to the genotypic richness of the accessions in a region. The median dissimilarity within the forest region is ranked approximately 190000th with a total number of 220,000 dissimilarity points in the population. The forest region also presents some outliers which are accessions of C. moschata that are morphologically or phenologically distinct from the commonly observed morphology and phenology of C. moschata in Cote d’Ivoire. The forest region has the largest genotypic diversity. The genotypic diversity in the other two regions is also considerably large with median dissimilarity ranking about 85000th and 90000th, respectively for the mountain and the savannah regions. The minimum and maximum dissimilarity ranks of the mountain region are about the same as the minimum and the maximum dissimilarity ranks of the distribution of accessions between regions.
Circular phylogenetic tree of the 34 morphotypes of C. moschata grown in Cote d’Ivoire. The three colors define the grouping of the morphotypes according to their morphological and phenological similarities.
Rank order of dissimilarities between and within geographic regions of differentiation of the accessions of C. moschata of Cote d’Ivoire.
A principal components analysis (Fig. 3) separated the morphotypes in distinct clusters. The morphotypes from the forest region form a distant cluster in the lower left of the two-dimensional representation of the first two principal components. The morphotypes of the mountain region form another cluster at the upper-right of the biplot. And the morphotypes from the savannah are grouped at the lower-right. The vectors indicate the traits that most characterize the morphotypes of a given region. It appears that the length of the vector is an indication of the degree of significance of the trait in the differentiation of the accessions. There is no evidence that the characters number of day to 50% emergence, weight of dry seeds and weight of fresh seeds exerted any genotypic differentiation. However, girth size and all fruit-related traits such as number of fruits per plant, diameter of the fruit, thickness of the pulp, weight of the fruit, diameter of the cavity enclosing the seeds, and volume of the fruit set the morphotypes of the forest region apart. The number of seeds per fruit and the phenological traits including the number of days from seeding to first female and male flower appearances, number of days to 50% flowering and number of days to physiological maturity are strong characteristics of the morphotypes from the mountain region. The morphotypes of the savannah region diverged morphologically with longer plant height, and accelerated phenology with shorter vegetative and reproductive phases. The morphological and phenological divergence of the accessions from the three regions is reflective of the ecosystems where they are thriving, to the point that the accessions from a region may be considered a separate variety or ecotype. All three regions showed high diversity of C. moschata with a Shanon-Weaver diversity index ranging between 1.39 for the forest region, to 1.95 for the savannah region. The Shanon-Weaver index is an indicator of the richness in term of number of different genotypes of C. moschata in a region, and evenness meaning that the different genotypes are represented in fairly equal proportion43,44,45. The Simpson index is an indicator of evenness. It scales between 0 and 1. The Simpson indices are 0.69 for the forest region, 0.77 for the mountain region and 0.80 for the savannah region (Fig. 3). The computation of the two indices takes into account the sample size. And the lower Shannon–Weaver and Simpson indices for the forest region could be due to smaller sample size compared to the other regions.
Separation of the morphotypes from the forest, mountain, and savannah regions and vectors of morphological and phenological traits obtained from the principal components analysis along with the diversity indices in each region.
Source: Ecology - nature.com
