Musa Germplasm Research Articles

Genome composition and genetic diversity of Musa germplasm from China revealed by PCR-RFLP and SSR markers

Banana ( Musa spp.) is one of the most important crops in the world. In this study, 216 banana accessions, 184 from the National Banana Germplasm Collection of China (NBGCC) and 32 from the International Network for the Improvement of Banana and Plantain (INIBAP), were used to determine the genome composition of banana plants in these collections and to estimate their genetic diversity. The genome composition was examined using PCR-RFLP markers. The molecular data for all but one accession (ITC 1231) from INIBAP were in agreement with the initial records based on phenotypic characteristics. Microsatellite (SSR) markers were used to investigate the genetic variability and relationships among these banana accessions. Ten of the 47 primer pairs tested consistently produced reproducible and discrete fragments. We identified a total of 92 alleles, ranging from 5 to 15 per locus. The genetic similarity between the accessions ranged between 0.1 and 1, when estimated using Jaccard's coefficient. The UPGMA method based on genetic similarities, grouped the NBGCC accessions according to those containing the ‘A’ and ‘B’ genomes. However, this analysis could not separate all the accessions, especially the somatic mutations, using the primers in this study. These data indicated that limited genetic variation exists within these accessions and the collections of NBGCC should include a much wider range of banana plant material.

Molecular phylogeny of banana cultivars from Thailand based on HAT-RAPD markers

Musa acuminata Colla (AA genomes) and Musa balbisiana Colla (BB genomes) are the wild progenitors of the cultivated banana, they are highly variable in Thailand. The genetic system is relatively unknown and complicated due to interspecific hybridization, heterozygosity and polyploidy, which are common in most clones. These factors make identification of closely related banana cultivars difficult, especially when sterile. The high annealing temperature-random amplified polymorphic DNA (RAPD) technique was used to estimate the genetic relationship between 22 selected banana cultivars, utilizing 14 random primers. Phylogenetic relationship was determined by unweighted pair group method with arithmetical averages cluster analysis. The dendrogram constructed from the similarity data showed that all the 22 cultivars analysed were closely related with a narrow genetic base. There were sufficient RAPD polymorphisms that were collectively useful in distinguishing the cultivars. The dendrogram grouped all the AA, BB, AAA, AAB and ABB genomes into a major cluster. Several subgroups are recognized within the major clade. As expected, Ensete glauca Roxb. (Musaceae) and Strelitzia reginae Banks (Strelitziaceae) were clearly differentiated from the analysed edible bananas. Our study showed that RAPD markers are sufficiently abundant to classify and readily dissect genetic differences between the closely related Musa germplasm and provide a basis for the selection of parents for improvement of this germplasm.

Ploidy investigation of bananas ( Musa spp.) from the National Banana Germplasm Collection at Rubona–Rwanda by flow cytometry

Bananas ( Musa spp.) are important staple and income-generating crops for millions of people in the highlands of East and Central Africa. This region is considered as a secondary center of banana diversity. The determination of ploidy level in different germplasms is important for better management of genetic diversity and is required before starting concerted breeding programmes. The use of reliable and high-resolution methods such as flow cytometry is recommended. In our study, flow cytometry was used on 89 frozen leaf accessions from the National Banana Germplasm collection at Rubona–Rwanda. Our results indicate that the ploidy level of 65 Highland banana clones analysed, is triploid, as was previously reported using morphological characteristics. However, the clones ‘Pomme’, ‘Kamaramasenge’, ‘Gisubi kayinja’, ‘Gisubi kagongo’, and ‘Dibis’ previously classified in National Banana Germplasm collection at Rubona as diploid, diploid, diploid, triploid, and tetraploid, respectively, were found to be triploid, triploid, triploid, diploid and triploid. These results should therefore, have great significance in banana germplasm management, for breeding programmes as well as for the Musa Germplasm Information System (MGIS).

Ploidy and genome composition of Musa germplasm at the International Institute of Tropical Agriculture (IITA)

Musa spp (bananas and plantains) constitute a hybrid-polyploid complex and are classified according to different genome compositions such as AA, BB, AB, AAA, AAB, ABB, AAAA, ABBB, AAAB and AABB. Knowledge of ploidy and exact genome compositions of the parental material is essential for Musa breeding. This study determined the ploidy levels and genome composition of the Musa germplasm collection, constituting over 300 accessions, at the International Institute of Tropical Agriculture in Nigeria and Uganda. Flow cytometric analysis of nuclear DNA content was used to estimate ploidy levels, while genome composition was ascertained with RAPD markers that are specific for the A and B genomes of Musa. It was determined that at least 8% of the plants in the germplasm collection were miss-classified in terms of ploidy and/or genome composition. The cultivars 'Pisang awak', 'Foulah 4' and 'Nzizi', previously classified as triploids, were found to be tetraploids by flow cytometry and conventional root tip chromosome counts. Similarly, cultivars that were previously classified as diploids including ‘Too’, and ‘Toowoolee’ were found to be triploids in our analysis. Ploidy and genome classification in Musa was generally determined from morphological characteristics. While our study showed that such a system is not always reliable, it was interesting to find that none of the plantains in the germplasm collection were miss-classified with regards to both ploidy and genome composition.

Genetic diversity of Musa diploid and triploid accessions from the Brazilian banana breeding program estimated by microsatellite markers

Banana (Musa spp.) is one of the most consumed fruits worldwide. Production is based mainly on triploid cultivars, and most genetic improvement programs aim to generate tetraploid hybrids obtained from the crossing of established triploid cultivars with a diploid parent genotype, improved or wild, exhibiting the trait of interest, normally resistance to biotic factors. Microsatellites were used to investigate the genetic variability and relationships between 58 Musa genotypes, including 49 diploids and nine triploid cultivars maintained at the Musa germplasm collection of the Brazilian dessert banana breeding program. Thirty-three primer pairs developed for banana were tested, and nine amplified reproducible and discrete fragments, producing a total of 115 alleles. The average number of alleles amplified per primer was 12.8, ranging from 10 to 15. The diploid genotypes presented the largest genetic variability, demonstrated by the large number of alleles detected, and the low similarity between the clones. The phenetic analysis clustered the triploid cultivars in a separated group, with the exception of the ‘Nanica’ and ‘Gros Michel’ cultivars, which showed high similarity with the diploid cultivar ‘Mambee Thu’. It was not possible to separate the wild diploid genotypes from the cultivated ones, indicating a common origin of these genotypes. A high proportion of duplicated alleles and/or loci was observed for diploid and triploid genotypes. The information gathered about the similarity between diploid and triploid accessions will help to define potential crosses to maximize the recovery of the typical fruit qualities required in Brazil (AAB, Pome and Silk dessert banana).

In vitro screening for resistance to Radopholus similis in Musa spp.

The burrowing nematode, Radopholus similis, is the most important nematode pest of banana and plantain worldwide, causing severe yield losses in commercial and local consumption cultivars. Chemical control is currently the most-used method to manage the nematodes, although nematicides are dangerous, toxic and expensive. Therefore, nematode control through genetic improvement is widely encouraged. Protocols have been set up to detect sources of resistance in the existing Musa germplasm, but the screening under greenhouse and field conditions is laborious and time consuming. In vitro screening could facilitate this process by picking up sources of resistance at an early phase. In this study, a procedure for in vitro screening for the resistance to R. similis was set up by determining the optimal incubation time and the optimal nematode inoculum. In a second step, the optimized procedure was confirmed by screening a selection of Musa varieties with a known host plant response. Except for ‘Yangambi Km5’, their host response under in vitro conditions corresponded to their host response under greenhouse or field conditions. The susceptible status of ‘Grande Naine’, ‘Gros Michel’ and ‘Cachaco’ was confirmed, as well as the resistant status of ‘Pisang Jari Buaya’ and ‘SH-3142’.

Determination of the Optimum Reaction Conditions for the PCR in Kenyan Musa Germplasm

Determination of the Optimum Reaction Conditions for the PCR in Kenyan Musa Germplasm

Screening of Vietnamese Musa germplasm for resistance to root knot and root lesion nematodes in the greenhouse

Twenty-six Vietnamese banana accessions, from the AA, AAA, AAB, ABB, AB genome groups and some wild accessions, were evaluated for resistance and/or tolerance to Meloidogyne spp. and Pratylenchus coffeae under greenhouse conditions. No source of resistance to Meloidogyne spp. was found. All the tested genotypes were found to be at least as susceptible to Meloidogyne spp. as the susceptible reference cv. Grand Nain (AAA) and the final nematode population in the roots was always much higher than the initial inoculum. The intensity of galling was less in the cvv. Man (AAB), Tay (ABB), Ngu Thoc (AA) and Yangambi Km 5 (AAA). Possible sources of resistance/tolerance to P. coffeae were found in cw. Yangambi Km 5 (AAA), Tieu Xanh (AAA), Tieu Mien Nam (AA), Gros Michel (AAA), Com Chua (AAB), Com Lua (AA), Man (AAB), Ngu Thoc (AA) and Grand Nain (AAA).

CRYOPRESERVATION OF PLANT GERMPLASM

CRYOPRESERVATION OF PLANT GERMPLASM

Agronomic performance of IITA's improved Musa germplasm at CRBP, Njombé, Cameroon

Black Leaf Streak Disease (BLSD) constitutes one of the major biological threats to plantain and banana production in sub-Saharan Africa, where over 100 million people rely on these crops for their food energy. In order to overcome this constraint, IITA (International Institute of Tropical Agriculture) developed resistant germplasm that has been evaluated in various production areas in Africa. Six of these hybrids (TMPx 7002-1, TMPx 1621-1, TMPx 4479-1, TMPx 7152-2, TMBx 1378 and TMBx 5295-1) recently made available through the INIBAP Transit Center (ITC), were evaluated in the climatic conditions of CRBP-Njombé (Cameroon) for their agronomic performance and their response to the disease. The trial consisted of a randomised complete block design with 2 replicates of 10 plants per hybrid. Observations were carried out for two production cycles. The results showed that the plantain hybrids TMPx 1621-1 and TMPx 7002-1 performed well in Njombé, both having an average bunch weight of 17.7 kg. In terms of yield potential, TMPx 7002-1 outyielded TMPx 1621-1 because of its shorter growth cycle. The very precocious cooking banana hybrid TMBx 5295-1 (9.4 months) is highly productive with over 50 t.ha-1 yr-1. Reaction to black sigatoka of the hybrids tested varied from "susceptible" (S) to "highly resistant" (HR). These results emphasise the need for multilocational testing of newly bred germplasm before their release in production areas. RÉSUMÉ La Maladie des Raies Noires (MRN) est l'une des contraintes majeures à la production des bananes et des plantains qui constituent une importante source alimentaire pour plus de 100 millions de personnes en Afrique au sud du Sahara. De nombreux hybrides résistants à cette maladie ont été créés par le programme d'amélioration génétique des bananiers de l'IITA et évalués dans plusieurs localités de la région. Six de ces hybrides (TMPx 7002-1, TMPx 1621-1, TMPx 4479-1, TMPx 7152-2, TMBx 1378 et TMBx 5295-1) récemment mis à disposition par l'INIBAP après indexation virale ont été évalués dans les conditions climatiques de Njombé (Cameroun) pour leurs performances agronomiques et leur réaction vis à vis de la maladie. Les hybrides ont été plantés selon le dispositif expérimental des blocs de Fisher constitués de 2 répétitions de 10 bananiers par traitement. Les observations ont été réalisées au cours des 2 premiers cycles de production. Les caractéristiques agronomiques montrent que deux hybrides de plantain, TMPx 1621-1 et TMPx 7002-1, ayant des régimes de poids moyen de 17,7 kg, se comportent mieux que les autres à Njombé. Cependant, l'hybride TMPx 7002-1 est plus productif que le TMPx 1621-1 à cause de son cycle plus court. L'hybride de banane à cuire TMBx 5295-1 qui est très précoce (9,4 mois) est très productif avec un rendement potentiel atteignant 50 kg/ha/an. Le comportement des hybrides testés vis à vis de la MRN va du phénotype S (sensible) au phénotype TR (très résistant). Ces résultats impliquent la nécessité des évaluations multilocales de matériel végétal nouvellement créé avant leur diffusion pour la production. (African Crop Science Journal 2001 9(3): 481-486)

TAXONOMIC VALUE OF CALCIUM OXALATE CRYSTALS IN MUSA GERMPLASM

Epidermal and histological characterization of Musa species and cultivars were carried out to investigate the descriptor value of calcium oxalate between groups and subgroups of cultivars. Epidermal peels of the bracts of selected plantain and banana cultivars stained with 0.01% safranin solution and permanent mounts of microtometric sections of fruits, bracts and peduncles stained with hydrogen peroxide and silver nitrate were microscopically examined for their crystal content and features. Bract epidermal features discriminate between the ‘AAB’ plantains and ‘AAA’ bananas. The major form of occurrence of calcium oxalate in the vegetative and reproductive tissues is bundle raphide. The bract adaxial epidermis of bananas show the occurrence of raphide idioblasts which contain a bundle raphide each. The idioblasts and bundles are more-or-less absent in the plantain bracts. Calcium oxalate occurs in the fruit peel as bundle raphides and in the pulp as intra-amylar crystals. Histological characterization of Musa germplasm helps to elucidate the limits of diversity in the germplasm.

Fruit quality evaluation of plantains, plantain hybrids, and cooking bananas

Abstract Triploid plantains (Musa AAB) are among the most important horticultural crops of West Africa. However, all landraces are susceptible to black Sigatoka, a fungal disease caused by Mycosphaerella fijiensis, Morelet. To ameliorate the spread of this epidemic, exotic triploid cooking bananas (ABB), with resistance to black Sigatoka, were introduced into West Africa and new disease resistant polyploid Musa (AAAB) germplasm developed through conventional cross-breeding of French plantains (AAB). To ensure that released materials would compete with landraces on a fruit quality basis, an evaluation methodology was developed to compare exotic and hybrid Musa genotypes with West African triploid landraces. Tests showed that plantain hybrids and cooking bananas had heavier bunches than plantain landraces. The plantain hybrids generally produced more but smaller fruit compared with False Horn plantain (AAB). The fruit ripening time of plantain hybrids and the cooking banana cultivar Pelipita was longer than that of plantain landraces, and selected plantain hybrids had fruit with a dry matter content similar to that of the female parent. The fruit of ripe plantain landraces had a firmer texture than plantain hybrids and the landraces Agbagba and Bobby Tannap had lower levels of soluble solids at specific stages of ripening compared with the other Musa germplasm. These results showed that simple physical tests can be used to select for higher quality cooking bananas and plantain hybrids and that specific plantain hybrids, within the test group, showed a higher potential for market acceptance.

THE BIOTECHNOLOGY CASE HISTORY FOR MUSA

Banana and plantain (Musa spp. L.) is a major tropical food crop and world commodity. Being a polyploid and vegetatively-propagated crop, its genetic improvement by conventional hybridisation is complex and difficult, yet not impossible. Simple tissue culture techniques, such as shoot-tip and embryo culture, are well developed and have greatly enhanced Musa germplasm handling and breeding. Plant regeneration by somatic embryogenesis is used to recover transgenic plants from single cells transformed by Agrobacterium or biolistic techniques for fungal and viral disease resistance. The potential of somaclonal variation as a source of novel and useful variability is limited, but it may be used as a secondary tool in Musa improvement. Molecular genetic techniques are being applied for improving the efficiency of Musa breeding. RFLPs appear to have limited application due to the low frequency of detected polymorphisms. The RAPD technique holds great promise for genome analysis. It detects high levels of polymorphism, yet still suffers from a number of practical drawbacks. Microsatellites are used to study genetic behaviour and structure in Musa, for fingerprinting hybrids, and to develop marker-assisted breeding systems. Candidate markers for such complex and important characters as parthenocarpy, earliness and regulated suckering are being tested. PCR primers have also been developed for virus diagnostics, already providing a specific and sensitive method to detect banana streak virus among in vitro plants. Reliable diagnostic protocols are increasingly needed for safe international movement of germplasm. Musa improvement requires a holistic approach to produce better cultivars. Hence, it is advocated that multidisciplinary public Musa breeding programmes should be central to any improvement effort. Such programmes will benefit from three biotechnologies that are clearly valuable in Musa breeding - tissue culture, diagnostics and molecular markers. Realistic, well defined targets for genetic engineering are occasionally warranted for inclusion within innovative breeding programmes.

AMMI and Stability Analyses of Bunch Mass in Multilocational Testing of Musa Germplasm in Sub-Saharan Africa

There is a genuine need within a plantain and banana (Musa spp.) breeding program to assess thoroughly the experimental materials through a sequence of trials. This will result in the selection of promising clones as potential new cultivars in the targeted agroecozone. Stability analyses and the additive main effects and multiplicative interaction (AMMI) model provide together a means for the identification of clones with 1) homeostatic responses to environmental changes, 2) a genotypic response to environmental changes, and 3) adaptation to specific niches. Fourteen polyploid clones (10 tetraploid hybrids and 4 triploid cultivars) were evaluated in a broad range of environments in sub-Saharan Africa to determine the value of stability and AMMI analyses in Musa trials. The interpretation of the results, especially those concerning the genotype × environment interaction, was facilitated by the combination of stability and AMMI analyses. Tetraploid hybrids combining heavy and stable bunch mass were identified. The results also suggested that a clone should be assessed in the ratoon cycle because plantain and banana are perennial crops. Likewise, high yielding clones with specific adaptation should be selected in environments showing the respective environmental or biotic stress.

Genetic diversity, hybrid performance, and combining ability for yield in Musa germplasm

Genome size variation occurs within and across generations in Musa spp., which reduces the predictive accuracy of parental performance on progeny value for yield and other traits with complex inheritance. Parental selection through progeny testing of prospective parents is required to achieve further genetic gains. This was carried out in this study, using a factorial mating design involving five 4x females and five 2x males. Genetic differences among offspring families were essentially due to differences in additive effects of the parents. Thus, little recombinative heterosis can be expected upon 4x-2x cross-breeding, and breeding strategies should target the development of 4x and 2x cultivars by accumulation of favorable alleles through recurrent selection within each ploidy pool. Offspring yield was positively correlated with parental GCAs but not with mid-parent values. Hybrid performance was also associated but not significantly correlated with genetic similarity indices based on both pedigree and molecular data. This study further suggests that current genetic models may not be adequate for populations with intergeneration genome size polymorphism.

Multivariate pattern of quantitative trait variation in triploid banana and plantain cultivars

Plantains and bananas ( Musa spp. L.) are inter- or intraspecific triploid hybrids derived from crosses between M. acuminata Colla. (A genome) and M. balbisiana Colla. (B genome). Cultivars have been assigned to different taxonomic groups (AA, BB, AAA, AAB, ABB, etc.) based on morphological qualitative descriptors. Principal component analysis of 15 quantitative traits was carried out to establish a more objective taxonomic relationship of cultivar groups and subgroups in the Musa germplasm. Fruit traits, number of neutral flowers, total number of leaves, plant girth at 50 cm, and days to flowering and harvest were the major discriminating traits in the germplasm. Principal components analyses grouped Musa germplasm into AAB plantains, AAA dessert bananas and ABB cooking bananas. The AAB starchy bananas appear to be separated into two subgroups with one being close to the AAB plantains and the other being close to the ABB cooking bananas. The dwarf French AAB plantain cultivar ‘Njock Kon’ appears to be a mutant of a giant AAB plantain cultivar.

Host response to black sigatoka in Musa germplasm of different ages under natural inoculation conditions

The development of an early evaluation method for host response of different Musa clones to black sigatoka was investigated under natural inoculation conditions in southeastern Nigeria. Two months old tissue-culture-derived plants of three plantain hybrids and parents of the hybrids were exposed to the natural inoculum of Mycosphaerella fijiensis in a field planted with a susceptible plantain cultivar. The response to M. fijiensis was evaluated on these young plants and on the same clones of mature, field-established plants. Young plants expressed similar resistance responses as mature plants and allowed an earlier evaluation of host-plant response to M. fijiensis. The male diploid banana parent, ‘Calcutta 4’, was highly resistant to black sigatoka. The three hybrids displayed partial resistance before flowering while the female parent, a triploid plantain, was susceptible. Disease responses of both mature and young plants ranked in the same order and were highly correlated ( P = 0.01). Young-plant response to black sigatoka in Musa germplasm was shown to be a reliable indicator of mature-plant response and a quick means of disease reaction assessment. This could accelerate progress made by breeding programs when producing new black sigatoka-resistant germplasm.

Improved polypoid Musa germplasm developed through ploidy manipulation

Improved polypoid Musa germplasm developed through ploidy manipulation

Morphological variation in Musa germplasm

Musa cultivars (bananas and plantains) are important crops in the humid tropics of Africa, America and Asia. This paper reports the extent of morphological variation of the Musa germplasm maintained in the gene-bank of the International Institute of Tropical Agriculture in southeastern Nigeria. Qualitative and quantitative descriptors were used to evaluate AA, BB, AB, AAA, AAB, AAAA, AAAB and AABB bananas, AAA and ABB cooking bananas, AAA beer bananas and AAB plantains and a few wild species. Univariate and principal component (PCA) analyses were performed to identify the most important descriptors to characterize and classify Musa germplasm collections. The most important qualitative morphological descriptors were persistence of male bud and hermaphrodite flowers, pigmentation in pseudostem, foliage, petiole and male flower, pseudostem blotching and waxiness, and leaf orientation. Furthermore, the most important quantitative morphological descriptors included pseudostem girth, height of tallest sucker, number of fruits and fruit sizes. These quantitative descriptors have a high heritability (>0.8), high repeatibility (>2.0) and low coefficient of variation (9–15%) with the exception of the height of the tallest sucker. The implications of the germplasm grouping revealed by PCA are briefly discussed in relation to the evolution of the triploid cultivars. This paper also proposes a new scientific nomenclature for the triploid Musa cultivars.

Molecular breeding in the genus Musa: a strong case for STMS marker technology

Musa species are among the tallest monocotyledons and include major food-producing species. The principal cultivars, derived from two major species Musa acuminata (‘A’ genome) and Musa balbisiana (‘B’ genome), are polyploid hybrids (mainly AAA, AAB and ABB triploids), medium to highly sterile, parthenocarpic and clonally propagated. Bananas and plantains are crops to which molecular breeding is expected to have a positive impact. In order to better understand banana genetics, more knowledge has to be accumulated about the complex genome structure of hybrids and cultivars. Therefore, the aim of our work is to develop molecular markers that are codominant, reliable, universal, highly polymorphic and that are applicable to collaborative Musa germplasm genotyping and mapping. Two size-selected genomic libraries have been screened for the presence of simple sequence repeats (SSR). Our data demonstrate that SSR are readily applicable to the study of Musa genetics. Our comprehensive analyses of a significant number of banana sequence tagged microsatellite sites (STMS) will add to our knowledge on the structure and phylogeny of genomes of the Musa species, and suggest that microsatellites be used as anchor markers for a banana genetic core map. Additional markers, such as e.g. CAPS have also been tested in order to increase the detection of polymorphisms exceeding that revealed by STMS technology. The utility of PCR-derived markers for collaborative genetic analyses of the banana genome, and the transferability of 'streamlined’ laboratory techniques and data analysis to Developing Countries are discussed.