4.1 Morphological Characteristics of the Musa accessions
Morphological characterization is the first step, according to Buitrago-Bitar et al. (2020), in studying the genetic variability of a population that possesses key features such as colors, shapes, smells, and textures. Although there are morphological, biochemical, and molecular descriptors in bananas, whenever the varietal identification demanded by law is considered, only morphological descriptors have been used to characterize a culture (Lombard et al. 1999; Priolli et al. 2002; Rocha et al. 2002). The difficulty in characterizing the characteristics of many related accessions using only morphological descriptors is high because most of the traits are influenced by the environment and require varied amounts of time since some evaluations were performed at the late stage of development.
In the present study, a greater number of accessions (64%) were recorded below 2 m tall, with only ‘Efolred’ having a pseudostem height range of 2.4–2.6 m. This finding does not correspond to previous reports by Anu et al. (2019) and Vinson et al. (2018), who reported above 3 m height for 80% and 63%, respectively, of Musa spp. The low plant height reported in this work could be attributed to edaphic, climatic, and genetic factors. According to Soares et al. (2012), there was a significant positive correlation between plant height and bunch weight. Taken together, these findings suggest that, in the present study, Efolred produced the highest bunch weight.
The pseudostems of all the accessions were blotched, with SH3436 and PITA 14 being extensively and moderately blotched, respectively (Oselebe et al. 2018). The classification of the petiole margins of Agbagba and SH3436 as clasping was based on the methods of Oselebe et al. (2018).
Three classes of ovule arrangements were present in the accessions studied. This finding does not correspond with the findings of Vilhena et al. (2019), who observed no ovules in their analyzed Musa samples. According to these authors, the bracts of their samples were reflexed as flowers developed. This finding corresponds to the findings of the present study, which reported that 55% of the bracts were retracted and rolled back. In total, 63.64% of the accessions exhibited red, dull purple or yellow outside with pink, dull purple, or yellow inside, similar to the findings of Oselebe et al. (2018).
The bunch shape of a greater number of accessions (36%) was asymmetrical, and the least common shape (9%) was cylindrical. A report by Anu et al. (2019) indicated that the bunch shape of 60% of their studied samples was asymmetrical, with the remaining 40% being cylindrical. The number of fingers per midhand of the bunch ranged from ≤ 12 to ≥ 17 in the present study. However, Vinson et al. (2018) reported means ranging from 11.7 to 17.7 for fingers per hand, while Oselebe et al. (2018) reported a mean number of fingers ranging from 9.00 to 13.00 with a mean fruit length ranging from 14.60 cm to 21.20 cm, which is smaller than the fruit length range reported in the present work (15 cm ≤ fruit length ≥ 31 cm). Soares et al. (2012) reported a positive correlation between fruit length and bunch weight. The high fruit length recorded for ‘Owom’, ‘Efol’, and ‘Numbrantor’ could consequently result in increased bunch weight and hence increased yield.
Based on the morphological descriptors used, the eleven accessions studied were classified into two different ploidy groups, diploid (AB) and triploid (AAB), both of which are hybrids of plantain and banana plants, respectively, with similar scores. This finding contradicts the findings of Buitrago-Bitar et al. (2020), whose work explained the 57 morphological descriptors proposed by IPGRI (1996), which they employed, characterized 12 Musa cultivars into five cultivars with a banana genome (A) and seven cultivars with a plantain genome (B). In comparison, in the work involving morphological and molecular characterization by Cruz-Cárdenas (2017), incomplete separation of the A and B genomes by these markers was reported, although they concluded that both markers are needed to complement each other for clarity. In this work, six score ranges were set for the accessions to be classed, but the accessions were distributed into only two score ranges. This was because the accessions had similar scores, and the majority of the descriptors were influenced by the environment; hence, they were highly unstable. A similar observation was made by Batte et al. (2018), who characterized 11 cultivars using 31 descriptors. The results showed that the cultivars had similar scores, and the descriptors used were not suitable for distinguishing between the cultivars studied. They attributed the result to the high instability shown by the descriptors during scoring. However, Javed et al. (2002) characterized 16 populations of Malaysian wild M. acuminata with the help of 46 morphological characteristics and discovered that the quantitative characteristics were not stable. Due to the instability of quantitative morphological descriptors, Batte et al. (2018) proposed and demonstrated that stable characters should be considered a priori for any cultivar classification. Therefore, a good morphological descriptor should be stable, distinctly identifiable, and heritable across generations. The present results indicated that there was no pure banana or plantain, which could be due to long-term mutations or, according to (Simmonds 1962), could result from inter- and intraspecific hybridization, hence producing hybrids. According to Karamura et al. (1998), diploids AA yielded AAA triploids by meiotic chromosome restitution, while interspecific hybridization between AA types (and perhaps AAA) and M. balbisiana (BB) produced various AAB and ABB types. The morphological ploidy grouping of some accessions was in accordance with that of other works, as described for Agbagba (Pillay et al. 2006). However, for the other accessions, their morphological ploidy grouping did not support previous research, as observed in PITA 14 (Bakry et al. 2009) and Calcutta 4 (Crouch et al. 1999).
4.2 Molecular characteristics of the Musa accessions
Assessing the genetic diversity of these vital crops using informative DNA-based markers can readily facilitate the exploration of Musa accessions for selection and integration into breeding programs. SCoT markers were used for molecular characterization. The utility of SCoT markers has been advocated due to their inherent features, such as reproducibility, the possibility of obtaining codominance during amplification, and accuracy in the investigation of the genetic relatedness and genetic diversity of plants (Talebi et al. 2018). SCoT markers have the advantages of simple operation, low cost, and abundant polymorphisms, and they are more conducive to molecular-assisted breeding (Liu et al. 2017). However, according to Liu et al. (2014), SCoT markers exhibit greater polymorphism than markers such as expressed sequence tag-simple sequence repeats (EST-SSRs). In a study by Igwe et al. (2019), SCoT markers were more useful and informative than directed amplified minisatellite DNA (DAMD) markers when they were used to assess the genetic diversity and classification of accessions of the genus Capsicum. Additionally, they stated that the marker is capable of revealing polymorphisms that might be directly related to gene functions.
In the present study, the 11 SCoT primers used were not amplified from any of the accessions. This finding is similar to that of Ude et al. (2021), who reported that some of the conserved DNA-derived polymorphism (CDDP) primers used on 66 Musa accessions were not amplified. These authors further explained that these primers had a G-C content of less than 60%, thereby confirming that a higher percentage of G-C content is a favorable factor for successful amplification of CDDP primers in plants (Collard and Mackill 2009). However, in the present work, markers with less than 60% G-C content were amplified, while some markers had 100% amplification across the accessions. The lack of amplification could be attributed to issues related to the priming site, such as mutations in the annealing sequence (Colson and Goldstein 1999), null alleles (Chapuis and Estoup 2007), or competitive amplification (Hippolyte et al. 2012).
The majority of the alleles recorded ranged from 0.0909 (SCoT2) to 0.5455 (SCoT35), with an average of 0.2314. This finding showed a very wide range, unlike previous works, which recorded 0.1250 to 0.5000 with a mean of 0.2580 in 70 Musa accessions (Christelova et al. 2011), 0.0460 to 0.4540 with a mean of 0.1710 in 66 Musa accessions (Ude et al. 2021), 0.1700 to 0.5000 with a mean of 0.3000 in 12 Musa accessions (Buitrago-Bitar et al. 2020), 0.1333 to 0.4000 with a mean of 0.2000 using the SCoT marker and 0.1333 to 0.4667 with a mean of 0.2556 using the DAMD marker Capsicum annuum (Igwe et al. 2019). Christelova et al. (2016) reported the highest major allele frequency of 0.5840, which is higher than the value obtained in the present work. Determination of this major allele frequency has provided insight into the variation and common single nucleotide polymorphisms (SNPs) of the Musa accessions of the Ebonyi State University Musa germplasm. Therefore, SCoT35 had the highest occurrence of this SNP across the studied population, while SCoT2 had the least occurrence of this SNP across the germplasm.
In the present work, the number of alleles ranged from 3.0000 to 11.0000, with an average of 8.8182; the gene diversity ranged from 0.5620 to 0.9091, with an average of 0.8460; and the PIC ranged from 0.4762 to 0.9016, with an average of 0.8268. This finding is similar to previous reports on Musa accessions, in which the allele number ranged from 4.0000 to 12.0000, with a mean of 8.0000; genetic diversity ranging from 0.6300 to 0.8800, with a mean of 0.8000; and PIC ranging from 0.5600 to 0.8700, with a mean of 0.7700 (Buitrago-Bitar et al. 2020). The number of alleles ranged from 20.0000 to 51.0000, with a mean of 35.0830; the gene diversity ranged from 0.7820 to 0.9760, with a mean of 0.9240; and the PIC ranged from 0.7680 to 0.9750, with a mean of 0.9180 (Ude et al. 2021), which are far above the values obtained in the present work. An allele number ranging from 5.0000 to 11.0000 was reported by Talebi et al. (2018), and 4.0000 to 18.0000 alleles were reported by Nyine et al. (2017). The richness of the alleles shown by the SCoT markers in this study can be used to indicate genetic diversity and to assess populations purely meant for conservation and breeding purposes (Patil et al. 2013). Considering the mean PIC value of 0.8700, with a range of 0.5300 to 0.9500 (Nyine et al. 2017); a mean of 0.8500, with a range of 0.7600 to 0.9420 (changadeya et al. 2012); and a mean of 0.8270, with a range of 0.6250 to 0.9360 (Christelova et al. 2011), the SCoT markers used in the present work showed high PIC, which implies that they had a high ability to detect polymorphisms and were highly informative and discriminatory.
The maximum values obtained in this study for the effective number of alleles (Ne), Nei's gene diversity (H), Shannon's information index (I), total gene diversity (Ht), gene diversity within the population (Hs), coefficient of gene differentiation (Gst) and estimate of gene flow from Gst or Gcs (Nm) were greater than previously reported values. For example, 1.6440, 0.3619, and 0.5298 were reported as the highest Ne, H, and I, respectively, according to Que et al. (2014), and as maxima of 1.307, 0.269 and 0.178 for Ne, I, and H, respectively, according to Deng et al. (2015), while genetic diversity and relationships among Diospyros germplasms were assessed using SCoT markers. Genetic indices, including Ne, H, and I, are considered crucial in the analysis of genetic diversity in several plants since they measure the degree of genetic diversity of species (Hamilton 2009; Freeland et al. 2011). A report by Cardoso et al. (1998) inferred that H and I are major parameters in genetic diversity studies since H estimates genetic diversity within and between accessions, whereas I estimates genetic diversity within and between populations regardless of the number of accessions accessed. If genetic diversities are found mostly within a population, Ude et al. (2021) explained that fewer populations are required to protect and maintain the overall differences in the accessions or the entire population. However, if genetic diversities are observed mostly between populations, then a greater number of populations should be considered for protection and utilization. Therefore, the accessions studied exhibited a wide genetic gap and high genetic diversity. Since the accessions studied were in one location, a narrow genetic gap was expected, but the opposite could be caused by outcrossing pollination processes and the exchange of genetic materials, as reported for Begonia species involving microsatellite markers (Twyford et al. 2013). Additionally, these interesting accessions with high genetic diversity of neutral markers and alleles could be utilized as suitable candidates for high adaptive variation, fitness, and conservation (Van et al. 2012; Ilves et al. 2013). According to Bilz et al. (2011), conservation efforts related to biodiversity focus on selecting accessions of crops with genetic reservoirs for potential and proven desirable adaptability, precisely under the influence of abiotic and biotic factors.
According to Nei (1978), Gst is classified as high when Gst > 0.1500, medium when it is 0.0500 < GST < 0.1500, and low when its value is < 0.0500. This means that all the accessions studied had high Gst values except for ‘Efolred’ and SH3436, which had intermediate Gst values. A report by Kumar et al. (2014) inferred that the Nm is low when Nm < 1 and moderate (Nm > 1) and extensive (Nm > 4) in nature; hence, all the accessions studied were classified as having moderate NM.
The eleven accessions were grouped into five clusters, with the smallest genetic variation observed between ‘Owom’ and Calcutta 4. A similar finding was given in a previous study involving different marker systems, SSR, AFLP, and RAPD, in which five clusters were detected (Adawy and Atia 2014). However, a different report from that of Ude et al. (2021) generated nine clusters from sixty-six Musa accessions and attributed these results to the nature of the markers and the number of accessions used. By characterizing 23 coconut accessions with the SCoT marker by Rajesh et al. (2015), two clusters were generated. According to the present study, there was no wide genetic distance across the clusters. It has been reported that the further away accessions are from one another, the greater the possibility of acquiring wider genetic diversity, which also indicates their locations within clusters (Skroch and Nienhuis 1995). The grouping of accessions in this study did not agree with the clustering found in the dendrogram. A similar report was given on morphological and molecular characterization of Musa species, in which the resulting genotype groups did not conform to each other (Christelova et al. 2016; Buitrago-Bitar et al. 2020).