Molecular studies of the poplar rust Melampsora Medusae-Populina, an interspecific hybrid between M.Larici-Populina and M.Medusae

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Inglés. In Europe, poplar rusts are caused by three Melampsora species, M. larici-populina, M. allii-populina and M. medusae, which are distinguished by morphological traits and host range.Recently, Spiers and Hopcroft (1994, Mycol. Res. 98: 889-903) described a new Melampsoraspecies in New-Zealand, named M. medusae-populina. According to several morphologicalfeatures, especially the urediniospore ornamentation, these authors hypothesized that M.medusae-populina was an interspecific hybrid between M. larici-populina and M. medusae. In1997, we received poplar leaves from South Africa, bearing uredinia of the newlydescribed species, according to our scanning electron microscope observations. In order toconfirm the hybrid status of this new species, we began a molecular study of theinterspecific variability of several Melampsora species. The Internal Transcribed Spacer(ITS) of the ribosomal DNA of several isolates of M. larici-populina, M. medusae, M.medusae-populina and M. allii-populina was amplified by PCR. In the case of M. medusae-populina, the formation of heteroduplex molecules was detected, suggesting the presenceof at least two partially homologous ITS sequences within each isolate. In order to separatethese molecules, amplification products were cloned in a bacterial vector. Heteroduplexanalysis of the cloned ITS regions proved that each isolate of M. medusae-populinacontained two ITS sequences, one homologous to that of M. larici-populina, and the otherhomologous to that of M. medusae. These results were confirmed by sequencing the clonedITS regions. Comparison of both types of sequences revealed 12% sequence divergence,including a 16-bp region within ITS2 which was only present in M. larici-populina. Thisregion, presenting a palindromic 12-bp core region, could be partly responsible for theformation of heteroduplexes. Furthermore, genomic DNA fingerprinting with severalRAPD primers showed that the banding pattern of M. medusae-populina consisted of anaddition of M. larici-populina-specific and M. medusae-specific fragments, thus confirmingthat M. medusae-populina was an interspecific hybrid between M. larici-populina and M.medusae. Besides, preliminary pathogenicity tests suggested that the hybrid combines thehost ranges of both parental species, and presents a high aggressiveness on several poplarclones. Therefore, M. medusae-populina should be considered as a serious threat for poplarcultivation in Europe in case of introduction or apparition through hybridization betweenM. larici-populina and M. medusae, which may occur on larch, the common alternate host.