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Summary Chromosome rearrangements and the way that they impact genetic differentiation and speciation have long raised questions from evolutionary biologists. They are also a major concern for breeders because of their bearing on chromosome recombination. Banana is a major crop that derives from inter(sub)specific hybridizations between various once geographically isolated Musa species and subspecies. We sequenced 155 accessions, including banana cultivars and representatives of Musa diversity, and genotyped‐by‐sequencing 1059 individuals from 11 progenies. We precisely characterized six large reciprocal translocations and showed that they emerged in different (sub)species of Musa acuminata, the main contributor to currently cultivated bananas. Most diploid and triploid cultivars analyzed were structurally heterozygous for 1 to 4 M. acuminata translocations, highlighting their complex origin. We showed that all translocations induced a recombination reduction of variable intensity and extent depending on the translocations, involving only the breakpoint regions, a chromosome arm, or an entire chromosome. The translocated chromosomes were found preferentially transmitted in many cases. We explore and discuss the possible mechanisms involved in this preferential transmission and its impact on translocation colonization., Significance Statement Chromosome rearrangements, the way that they emerged and their potential impact on speciation and breeding raise important questions. We report on the characterization of several large reciprocal translocations in banana, Musa acuminata, identify the genetic group in which they emerged, characterize their impacts on chromosome recombination, and explore possible mechanisms allowing their colonization.

International audience; Most banana cultivars are triploid seedless parthenocarpic clones derived from hybridization between Musa acuminata subspecies and sometimes M. balbisiana. M. acuminata subspecies were suggested to differ by a few large chromosomal rearrangements based on chromosome pairing configurations in intersubspecies hybrids. We searched for large chromosomal rearrangements in a seedy M. acuminata ssp. malaccensis banana accession through mate-pair sequencing, BAC-FISH, targeted PCR and marker (DArTseq) segregation in its progeny. We identified a heterozygous reciprocal translocation involving two distal 3 and 10 Mb segments from chromosomes 01 and 04, respectively, and showed that it generated high segregation distortion, reduced recombination and linkage between chromosomes 01 and 04 in its progeny. The two chromosome structures were found to be mutually exclusive in gametes and the rearranged structure was preferentially transmitted to the progeny. The rearranged chromosome structure was frequently found in triploid cultivars but present only in wild malaccensis ssp. accessions, thus suggesting that this rearrangement occurred in M. acuminata ssp. malaccensis. We propose a mechanism for the spread of this rearrangement in Musa diversity and suggest that this rearrangement could have played a role in the emergence of triploid cultivars.

We report a reference map of Musa acuminata from 180 individuals of BORLI population. This F1 population derived from the cross between M.a. "Borneo" x M.a. "Pisang Lilin" which was chosen for mapping mainly because of the great heterozygosity of both parents (76 % each). For map construction we genotyped 180 SSR markers on 180 individuals, 12 RGA-RFLP probes on 112 individuals and 380 DArT markers on 92 individuals. Using the software Joinmap 4 at LOD 5, Kosambi calculation distance and the regression mapping algorithm, we defined 11 main linkage groups, as expected, on Borneo parent and 10 on Pisang lilin due to one translocation event in the male parent. Despite structural differences between the parents, a consensus map was built. In order to check genotyping results and to analyse structural rearrangements, we developed an original method using Neighbor Joining algorithm implemented in the genetic diversity software Darwin5. It allows visualizing graphically disruptions in alignments arising from these structural rearrangements. (Texte intégral)

As several other plants, the genome of banana and plantain contains integrations of Banana streak virus (BSV) sequences even though integration is not an essential step in the replication cycle of this virus. In banana two types of BSV integrants exist. Ones are non functional sequences present in both common Musa species, Musa acuminata (denoted A) and Musa balbisiana (denoted B) and it is now assumed that the integrants of the other type, containing the complete viral genome and restricted to M. balbisiana genome, become infectious by reconstituting a complete replication-competent viral genome. Thereby, an increasing record of BSV outbreaks was observed fifteen years ago among banana breeding lines and micro propagated inter-specific Musa hybrids, worldwide. Today, three widespread BSV species, Banana streak Obino l'Ewai virus (BSOIV), Banana streak Imové virus (BSImV) and Banana streak Golfinger virus (BSGfV) are known to occur as infectious integrants in the M. balbisiana genome. However, even though such integrations are known to be infectious, their presence is not sufficient to induce infection. We demonstrated that the process of genetic hybridization and abiotic stresses such as micropropagation by in vitro culture contributed in triggering episomal expression from EPRVs. Two mechanisms at least are involved in the BSV expression: the ploidy of the M. balbisiana in Musa genotypes and an additional genetic factor called BEL for BSV expressed locus concerning the triploids (Musa AAB) resulting from inter-species genetic crosses between virus-free diploid M. balbisiana (BB) and tetraploid M acuminata (AAAA) parents. Then, diploids M. balbisiana such as PKW and Pisang Batu harboring pathogenic BSV EPRVs are resistant to any multiplication of BSV while haploid genotypes such as triploids (AAB, French clair) or tetraploids (AAAB, FHIA 21) expressed BSV. Thereby, we characterized the segregation of three BSV species appearance among the AAB F1 progeny as a monogenic allelic system conferring the role of carrier to the M. balbisiana diploid parent. BSOIV and BSImV appeared in almost all infected hybrids (50% of the progeny) depending of BEL regulation while BSGfV are restricted in only half of these hybrids and subordinated by BEL. Three BAC libraries from accession of M.acuminata Cavendish subgroup cv petite naine (AAA), a wild M. acuminata subsp burmannicoides Calcutta 4 (AA) and a wild M. balbisiana PKW (BB) are explored for the pattern of integration of infectious BSV EPRVs by testing a set of different viral probes representing each time the BSOIV, lm and Gf complete genome. BSV positive BAC clones are characterised by RFLP fingerprints approaches. This analysis revealed that the three BSV species represent low-copy loci and their integration is specific to the PKW Musa balbisiana genome. BSGfV EPRVs in PKW is twice and are fully annotated after sequencing. Each BSGfV is composed of back-to-back viral sequences representing more than a whole BSV genome very similar each other. We developed molecular markers (PCR, PCR-RFLP) to distinguish each others and analysed the BSGfV EPRV segregation in the AAB F1 progeny. BSGfV EPRVs are found to be allelic, located at the same locus. In theory, both allelic EPRVs could be involved in the restitution of virions through a set of recombination events. (Texte intégral)

Chromosome pairing studies at meiosis have revealed that translocations are frequent in banana genomes. In Musa acuminata, seven groups of translocation have been identified (Standard, North Malaysia, Malaysia Mountains, North A, North B, Indonesia, and East Africa). Each group comprises accessions having the same chromosome structure. Translocations increase the difficulty of constructing a genetic map, studying the inheritance of agronomical characters and breeding in general. The objectives of our study were to develop a tool to characterize translocations on chromosomes based on fluorescent in situ hybridization of BAC clones (BAC-FISH) and to use this tool to characterize the translocations in the accessions 'Calcutta 4' (2n=2x=22, translocation group North A) and 'Madang' (2n=2x=22, translocation group Standard). To achieve this, a genetic map and a 'Calcutta 4' BAC library were built, the BAC-FISH technology was adapted to bananas and a banana cytogenetic map was initiated. The genetic map was based on a F2 population from a 'Calcutta 4' x 'Madang' cross. It encompasses 120 markers (20 RFLPs, 81 AFLPs and 19 SSR markers) distributed over 14 linkage groups and covering 597 cM. The 'Calcutta 4' BAC library comprises 55 152 clones with an average insert size of 100 Kb. About 1.5% of these clones present chloroplast and mitochondrial DNA inserts. This library covers 9 to 10 times the banana genome. Linkage group Il (LGII) was chosen to search for translocations since some of its characteristic (such as a high number of distorted markers) suggested that this group contained chromosomes with translocations. Four BAC clones corresponding to 3 RFLPs and 1 SSR loci distributed on LG II were localized on the 'Calcutta 4' and 'Madang' chromosomes. The results showed that the markers involved in the LG II were localized on three chromosomes, whose structure was different in 'Calcutta 4' and 'Madang', with two linked translocations observed in 'Calcutta 4'. In 'Madang', the markers mMaCIR161-rMaCIR 560, rMaCIR 1125 and rMaCIR 36 were localized on three chromosomes (A, B and C). In 'Calcutta 4' these markers were localized on only two chromosomes (A and B). The marker rMaCIR 1125 localized on the 'Madang' C chromosome, was localized in the proximal position of the 'Calcutta 4' B chromosome. We also initiated a global cytogenetic map of 'Calcutta 4' that comprises 16 loci (14 BAC clones selected by RFLP and SSR markers and the ribosome probes 45S and 5S). The 14 linkage groups of the 'Calcutta 4' x 'Madang' genetic map are anchored to this cytogenetic map. (Texte intégral)

Recently introduced inter-specific #Musa# hybrids, bred for improved yield and resistance to diseases, have been found to be widely infected with several Banana streak badnavirus (BSV-Obino I'Ewai (BSV-OI), BSV-Imové (BSV-Im) and BSV-GoId Finger (BSV-Gf) strains), the causal agent of banana streak disease (BSD). One hypothesis suggests that BSD occurrence in these inter-specific hybrids results from activation of corresponding BSV endogenous pararetrovirus sequences (EPRVs) integrated into the Musa genome rather than from external sources of infection. The process of genetic hybridisation and abiotic stress such as micropropagation by #in vitro# culture may be responsible in triggering episomal expression of these BSV integrants. In order to test the first hypothesis we carried out a genetic analysis of banana streak disease incidence in a F1 triploid (AAB) population produced by hybridisation between virus- and disease-free #M. balbisiana# Pisang Klutug Wulung PKW (BB diploid) and #M. acuminata# IDN 110T (AAAA tetraploid) parents. Molecular AFLP markers segregating with the presence or absence of episomal viral particles were identified in the #M. balbisiana# genome only. These data indicate that a genetic mechanism is involved in BSV appearance, and suggest that a monogenic allelic system confers the role of carrier to the #M. balbisiana# parent The three different strains were detected by IC-PCR in diseased hybrids. The BSV-OI and BSV-Im strains appeared in almost all diseased hybrids while the BSV-Gf strain appeared only in about half of these hybrids. Genetic analysis resulting from AFLP results showed that BSV-OI EPRVs' regulation depends on a genetic factor: BEL (BSV Expressed Locus) (Lheureux et al. 2003). Further experiments showed that BSV-lm EPRVs' regulation depends on the same genetic factor too. Even if BSV-Gf appears only in half of diseased hybrids, some elements suggest that BSV-Gf strain appearance and the activation of its corresponding EPRV(s) seem to be subordinated to the BEL factor. Three BAC libraries from cv Cavendish (AAA), cv Calcutta 4 (AA) and PKW (BB) were constructed in order to studies genome structure and evolution of #Musa# species. We have explored the pattern of integration for the different BSV EPRV suspected infectious by testing a set of different probes representing each time the BSV-OI, Im and Gf complete viral genome. This analysis revealed that the three EPRVs under analysis represent low-copy loci and their integration is specific to the PKW #Musa balbisiana# genome. These studies provide preliminary results towards characterisation of infectious BSV EPRV and definition of mechanism under laying their potential activation or nonactivation upon stress conditions in banana. (Texte intégral)

Inter-specific #Musa# hybrids have been found to be infected with several Banana streak pararetrovirus strains (BSV-Obino l'Ewai (BSV-Ol), BSV-Imové (BSV-Im) and BSV-Gold Finger (BSV-GF) strains) resulting from activation of BSV endogenous pararetrovirus sequences (EPRVs) integrated into the #Musa# genome rather than from external sources of infection. The process of genetic hybridisation may be responsible in triggering episomal expression of the BSV integrants. In order to test this hypothesis we carried out a genetic analysis of BSD incidence in a F1 triploid (AAB) population produced by hybridisation between virus- and disease-free #M. balbisiana# (BB) and #M. acuminata# (AAAA) parents. Molecular AFLP markers were identified in the #M. balbisiana# genome only. Analysis of the segregation of these markers allowed the construction of a genetic map containing the locus associated with BSV infection called BSV Express locus (BEL). The BSV-Ol and BSV-Im strains appeared in almost all diseased hybrids while the BSV-GF strain appeared only in about half of these hybrids. Genetic analysis resulting from AFLP results showed that BSV-Ol and BSV-Im EPRVs' expression depends on the same genetic factor, the BEL locus. Although the BSV-GF is not genetically linked to the BEL locus, its appearance and the activation of corresponding EPRV(s) seem to be also subordinated to the BEL locus. So, the discovery of several BSV EPRVs linked to BEL locus implicates that BEL differs in nature from BSV EPRVs.

Les bananes comestibles sont issues pour l'essentiel de deux espèces sauvages diploïdes : #Musa acuminata# et #Musa balbisiana#. Il convient de distinguer les productions destinées au marché d'exportation de celles qui s'adressent aux marchés intérieurs. Les programmes d'amélioration génétique ont pour objectif la création de variétés résistantes aux maladies et aux ravageurs. La stérilité des bananiers cultivés oblige à la création de variabilité par voie sexuée (hybrides tétraploïdes et triploïdes). D'autres méthodes sont également utilisées : la mutagenèse, la variation somaclonale et la transformation génétique.

La maladie de la mosaïque en tirets des bananiers BSD frappe aujourd'hui la majorité des zones productrices de bananes. Le virus responsable BSV est un badnavirus. Ces dernières années, la véritable contrainte a résidé dans l'association étroite de l'apparition de la maladie avec l'obtention et la diffusion de nouvelles variétés de bananiers. Des cultivars sains multipliés par CIV ou des géniteurs utilisés lors de croisements génétiques restituent soit des vitroplants soit des hybrides contaminés. Un pourcentage non expliqué de plants BSD voisin de 50% a été observé dans les descendances d'hybridation de géniteurs sains. Ceci a été interprété comme le résultat d'une ségrégation à caractère mendélien de l'expression de la maladie. Au moins 4 souches BSV (Mys, Wu, Gf et Im) existent sous diverses formes : libres comme particule épisomale infectieuse, ou intégrées sous forme de séquences partielles ou totales dans le génome B. Le modèle actuel BSV-Wu suggère une double recombinaison conduisant à l'excision des deux fragments internes et à la circularisation aboutissant à la forme superenroulée transcriptionnellement active. L'analyse du motif activable n'a pas montré d'homologie avec les rétrotransposons pour leurs éléments majeurs. Afin d'identifier et d'analyser les mécanismes moléculaires induisant la multiplication de la souche BSV-Wu au cours de l'amélioration conventionnelle, nous avons tenté de définir un marqueur génétique viral utilisable par les généticiens pour identifier les bananiers sains. Des marqueurs moléculaires liés à la présence ou à l'absence de virus épisomal ont été recherchés par Bulk segregant analysis et par AFLP dans la F1 de deux croisements interspécifiques (IDN110 4A x PKW 2B). 11 marqueurs polymorphes relies au parent B ont été isolés : 7 ségrègent avec le facteur déclenchant et sont corrélés à l'expression du virus épisomal, 4 avec le phénotype 'absence de virions'. Leur analyse a permis d'établir une carte génétique du locus impliqué dans l'expression du virus. Le marqueur de la maladie le plus proche est à OcM du facteur déclenchant. L'analyse de sa séquence révèle une homologie de 86% avec le rétrotransposon 'Monkey' identifié chez #Musa Acuminata#. Ce pathosystème constitue du fait de la présence simultané de séquences intégrées et d'un système de réplication du virus, un candidat idéal pour une hypothèse forte de gene silencing.

Cultivated bananas have first been susPected to originate from four different wild species : Musa acuminata (A), M. balbisiana (B), M. schizocarpa (S) and the Australimusa (T) species. Banana germplasm includes di, tri and tetraploid clones. Cultivated clones have been classified in genomic groups by Simmonds and Shepherd according to their chromosome number and morphological traits; the main groups are AA, AAA, AAB, ABB, AS and AAT. Biotechnology now allows a precise and early characterisation of the genomic nature of the accessions. Flow cytometry allows a rapid determination of ploidy level of a clone and in some extend of its genomic group. Analysis of the nuclear genome using only 8 STMS allows the classification of an accession in a genomic group and subgroup. Recently, genomic in situ hybridisation has permitted to determine the specific origin - A, B, S or T- of the chromosomes of some interspecific clones. Molecular markers have also led to a better understanding of the origin of cultivated bananas, particularly through the analysis of the A genome variability. The maternal and paternal origin of the clones may be inferred respectively from the RFLP analysis of the chloroplast and mitochondria genomes. RFLP analysis of the nuclear genome of cultivated bananas reveals all the wild acuminata subspecies at their origin. For instance, this study has shown that fruit quality is related to the nature of the A genome: cooking types with a M a. bankii / errans origin, and dessert types with a M a. malaccensis / zebrina origin

Banana production is severely threatened by the Black Leaf Streak (BLS) disease caused by Mycosphaerella fijiensis. Numerous chemical applications are needed to control the fungus's impact on leaves which reduces yields and causes premature ripening of fruits. Genetic improvement for resistance to BLS disease appears as the most durable, cost and environmentally efficient control. Screening for resistance has led to the identification of two forms of resistance reported as High Resistance (HR, hypersensitive response) and Partial Resistance (PR). In connection with the breeding program, a genome mapping approach for genetic and QTL analyses was developed to characterise the HR form. An F2 segregating population of 153 plants was obtained from a cross between two wild seeded banana accessions, Musa acuminata burmannicoides type Calcutta 4 and M. a. banksii type Madang. A linkage map was constructed with 110 AFLP markers in association with 39 codominant RFLP and SSR markers to anchor the linkage groups to the banana genetic core map. Results showed significant allelic distorted segregations in the F2 progeny for 58 markers which highlight the use of molecular studies for inheritance analyses. A first mapping concluded to join the markers in 11 linkage groups. Correlations made with field observations, ranked into six classes of infection severity, led to the identification of one RFLP marker strongly associated to the resistance and a second QTL mapped onto a different linkage group with lower significance level

The genomes of four different wild Musa species are involved in cultivated bananas : the A, B, S and T genomes related to M. acuminata, M. balbisiana, M. schizocarpa and the Australimusa species, respectively. Most cultivated clones are triploid or diploid. They have been classified into genomic groups according to chromosome numbers and morphological traits. The main groups are AA, AAA, AAB, ABB, AS or AT. Flow cytometry reveals that the sizes of A, B, S and T genomes are significantly different. M. balbisiana has the smallest genome (1.03 pg/2C, 2n=2x=22), followed by M. acuminata (1.11 pg/2C, 2n=2x=22) and M. schizocarpa (1.18 pg/2C, 2n=2x =22). The highest value was obtained for the Australimusa species M. textilis (1.27 pg/2C, 2n=2x=20). Genome size can thus be used to determine the species involved in interspecific diploid clones. Genomic in situ hybridization has been developed to characterize the specific origin of the chromosomes in interspecific clones. This technique enables us to differentiate the chromosomes from the genomes A, B, S and T and is being used to determine the exact genome structure of cultivated clones and synthetic hybrids. (Texte intégral)

A completed map, based on two selfed progenies from two banana cultivars (M53 and SFB5) is presented (roughly 1200 cM). More than three hundred markers are linked in 11 linkage groups representing the genorne (2n=22) of Musa acuminata. Roughly one third of the markers are codominant restriction fragment polymorphisms (RFLPs; one hundred) or microsatellites (thirty). Two thirds of the markers are dominant amplified fragment length polymorphisms (AFLPs; 10% could be considered to be co-dominant). The mean linkage distance is 3 cM, but marker density still should be increased on a couple of linkage groups. Particularities for a mapping job in banana are discussed. Due to Musa acuminata sub-species specific translocations, up to 36% of all the markers tested show important segregation distortions. The need for a cooperative mapping initiative based on a proposed "frame-map" harbouring evenly spaced co-dominant "anchor" markers is proposed. CIRAD has published 45 sequence tagged microsatellite sites (STMS) in the EMBL database which are accessible at : "http : //www.ebi.ac.uk/' using the keywords LAGODA and MICROSATELLITE (EMBL accessions X87258 to X87265, X90740 to X90750 and Z85950 to Z85977).

Les bananes comestibles sont issues pour l'essentiel de deux espèces sauvages diploïdes : #musa acuminata# et #Musa balbisiana#. Il convient de distinguer les productions destinées au marché d'exportation de celles qui s'adressent aux marchés intérieurs. Les programmes d'amélioration génétique ont pour objectif la création de variétés résistantes aux maladies et aux ravageurs. La stérilité des bananiers cultivés oblige à la création de variabilité par voie sexuée (hybrides tétraploïdes et triploïdes). D'autres méthodes sont également utilisées : la mutagenèse, la variation somaclonale et la transformation génétique

Bananas belong to the #Musaceae# family and are among the tallest monocotyledons. Main dessert banana cultivars are triploid, highly sterile, parthenocarpic and atonally propagated. They are susceptible to several diseases that seriously threaten plantations. In order to better understand banana genetics, more knowledge must be gained about their genomes. This would help to develop new improvement strategies. Major aims of the molecular studies, in progress at CIRAD at present, are the construction of genetic maps, a better understanding of chromosome structural differentiation in #Musa#, the analysis of quantitative characters as well as a comprehensive study of molecular genetic diversity. A first genetic linkage map based on RFLP markers has already been published by our lab (Fauré et al., 1993, TAG 87 : 517-526). In order to saturate this map for QTL analysis, we checked the banana genome for VNTR markers. We report here preliminary results on dinucleotide microsatellite sequences in the banana genome. Microsatellite gene tagging and targeting are discussed.

Bananas belong to the #Musaceae# family and are among the tallest monocotyledons. Main dessert banana cultivars are triploid, highly sterile, parthenocarpic and atonally propagated. They are susceptible to several diseases that seriously threaten plantations. In order to better understand banana genetics, more knowledge must be gained about their genomes. This would help to develop new improvement strategies. Major aims of the molecular studies, in progress at CIRAD at present, are the construction of genetic maps, a better understanding of chromosome structural differentiation in #Musa#, the analysis of quantitative characters as well as a comprehensive study of molecular genetic diversity. A first genetic linkage map based on RFLP markers has already been published by our lab (Fauré et al., 1993, TAG 87 : 517-526). In order to saturate this map for QTL analysis, we checked the banana genome for VNTR markers. We report here preliminary results on dinucleotide microsatellite sequences in the banana genome. Microsatellite gene tagging and targeting are discussed. (Texte intégral)

Les analyses RFLP ont pour but d'établir la carte génétique du génome diploïde de Musa acuminata, de permettre une meilleure compréhension de la différenciation structurale des chromosomes chez Musa, et de développer une base de données concernant la variabilité génétique. Une banque nucléaire EcoRI contenait 67% de clones correspondant à des séquences uniques. Sur dix enzymes de restriction testées, EcoRI, EcoRV, DraI et HindIII ont révélé les plus hauts niveaux de polymorphisme et ont été sélectionnées pour la cartographie. Avec ces quatre enzymes, on a révélé jusqu'à 91% de polymorphisme sur huit clones de Musa en utilisant des sondes correspondant à des séquences uniques. Les parents du croisement ayant généré la population F2 cartographiée ont été choisis pour avoir un niveau maximal de polymorphisme. Cinq combinaisons sonde/enzyme utilisant des sondes correspondant à des séquences répétées ont suffi à distinguer les clones de bananiers entre deux sous-groupes représentatifs de la diversité génétique des Musa. La banque actuelle de 97 sondes polymorphes correspondant à des séquences uniques est portée à 250. Ce travail a fourni le point de départ pour la construction de la première carte génétique du bananier, et un premier aperçu du potentiel des marqueurs RFLP pour l'éclaircissement des relations génétiques entre taxa. (d'après résumé d'auteur)