Erasing the Past: A New Identity for the Damoclean Pathogen Causing South American Leaf Blight of Rubber

<div><p>Background</p><p>South American leaf blight (SALB) of rubber has been the main constraint to production in its neotropical centre of origin since commercial plantations were first established. The fungal causal agent was identified and described more than a century ago but its precise placement within the Ascomycota still remains uncertain. Indeed, such is the ambiguity surrounding the pathogen that each of the spore morphs would, according to their present classification, be placed in different ascomycete families: the <i>Microcyclus</i> sexual morph in the Planistromellaceae and the two purported asexual morphs - <i>Fusicladium</i> and <i>Aposphaeria</i> – in the Venturiaceae and Lophiostomataceae, respectively. Given the historical importance of the fungus and the ever-menacing threat that it poses to rubber production in the Palaeotropics – and, thus to the rubber industry and to the global economy – its phylogeny, as well as its biology, should be resolved as a matter of urgency.</p><p>Methods and Results</p><p>Here, six genomic regions (LSU rRNA, mtSSU, MCM7, EF-1α, Act and ITS) were used for reconstructing the molecular phylogeny of the SALB fungus based on material collected throughout Brazil. The analyses support the classification of the fungus in the family Mycosphaerellaceae s. str. (Capnodiales, Dothideomycetes) and place it firmly within the clade <i>Pseudocercospora</i> s. str., now accepted as one of the distinct genera within Mycosphaerellaceae. The new combination <i>Pseudocercospora ulei</i> is proposed and the life cycle of the fungus is confirmed, based on both experimental and phylogenetic evidence, with the <i>Aposphaeria</i> morph shown to have a spermatial rather than an infective-dispersal function.</p><p>Conclusions</p><p>Because the phylogeny of the SALB fungus has now been clarified, new insights of its epidemiology and genomics can be gained following comparison with closely-related, better-researched crop pathogens.</p></div

Hypothetical life cycle of <i>Pseudocercospora ulei</i>.

<p><b>A</b>. Asexual morph with conidiophores and conidia (Bar =  35µm) and conidiogenous cells with conidia at different stages of conidial formation. Pictures: Lesions to which the asexual morph is associated (left) and close-up of leaf bearing typical lesions (right). <b>B</b>. Spermogonial morph with stroma, spermogonia (Bar =  30 µm) and spermatia (Bar =  7 µm). Pictures: Lesions to which the spermogonial morph is associated (left), and close-up of the same lesions (right). <b>C</b>. Sexual morph with stroma, pseudothecia, asci and ascospores (Bar = 60µm). Pictures: Lesions to which the sexual morph is associated (left), and close-up of stromata (right). Dotted arrows indicate that both ascospores and conidia can infect young leaves.</p

Bayesian analysis showing the phylogenetic relationships of <i>Microcyclus ulei</i> based on the MCM7 sequence alignment.

<p>Bayesian posterior probabilities are given at the nodes and coded according to the colored scale bar. The black line scale bar shows 0.2 expected changes per site. The tree was rooted with <i>Aspergillus niger</i>.</p

Bayesian analysis showing the phylogenetic relationships of <i>Microcyclus ulei</i> based on the LSU sequence alignment.

<p>Bayesian posterior probabilities are given at the nodes and coded according to the colored scale bar. The black line scale bar shows 0.2 expected changes per site. The tree was rooted with <i>Aspergillus niger</i>.</p

Origin of the <i>Microcyclus ulei</i> isolates used in the phylogenetic study.

1<p>Brazilian states: Acre (AC), Bahia (BA), Espírito Santo (ES), Mato Grosso (MT), Minas Gerais (MG) and Rondônia (RO).</p

Pleomorphic development of the life cycle of <i>Pseudocercospora ulei</i>.

<p>Weighted average based on the score of conidial sporulation intensity and spermogonia and ascostromata density during the leaf development of RO38 rubber clone grown under field conditions. Assessments were made from December 15, 2011 to February 24, 2012 (A); and from September 19 to December 3, 2012 (B).</p

Bayesian analysis showing the phylogenetic relationships of <i>Microcyclus ulei</i> based on the mtSSU sequence alignment.

<p>Bayesian posterior probabilities are given at the nodes and coded according to the colored scale bar. The black line scale bar shows 0.2 expected changes per site. The tree was rooted with <i>Aspergillus niger</i>.</p

Phylogenetic relationships of <i>Microcyclus ulei</i> based on the combined ITS, EF-1α and ACT sequences alignment.

<p>Bayesian posterior probabilities are given at the nodes and coded according to the colored scale bar. The black line scale bar shows 0.08 expected changes per site. The tree was rooted with <i>Passalora eucalypti</i>.</p