First Report of Cophinforma atrovirens Causing Stem Rot and Dieback of Cashew Plants in Brazil

Cashew tree (Anacardium occidentale L.) is a plant of great economic and social importance for northeastern Brazil. In 2015, while conducting a survey in a newly established 90-ha cashew orchard in Barra do Corda, State of Maranhão (05°30′20″S; 45°14′36″W), a shoot and stem rot followed by dieback symptoms were observed in about 30% of plants (CP 76 and BRS 189 clonal cultivars). Isolations from infected tissues revealed a whitish, cottony fungal colony, which later turned gray on potato dextrose agar (PDA). Upon the absence of sporulation under PDA media, mycelial disks from the edge of the colony were transferred to autoclaved fruits of custard apple (Annona squamosa) and incubated at 25°C (Cardoso et al. 2002). Pycnidia were observed after 4 weeks of incubation over the surface of entire fruits. Conidia were hyaline, unicellular, fusiform, and truncated at the base when newly formed, and 33.95 to 53.44 (avg. 47.38) μm in length and 9.13 to 11.47 (avg. 10.59) μm in width (n = 50). No sexual structure was observed. Genomic DNA was extracted from mycelia and the fragments of the genomic regions of the rDNA internal transcribed spacers (ITS), β-tubulin (TUB2), and the translation elongation factor 1-α (EF-1α) were amplified by PCR using primers ITS1/ITS4, βT2a/βT2b, and EF1-688F/EF1-1251R, respectively (Coutinho et al. 2017). The PCR products were sequenced and the sequences obtained were deposited in GenBank under the accession numbers MF521897 (ITS), MF538773 (βT), and MG209371 (EF1). Multiple alignments of the combined data set of the genomic regions and representative sequences obtained from GenBank were submitted to phylogenetic analysis of maximum parsimony (MP) and maximum likelihood (ML) and the tree topology tested by the bootstrap method with 1,000 replicates. For the phylogenetic tree, the isolated fungus (BOT456) was grouped in a clade with isolates CMM 1390, CBS 117444, and CMW 13433 of Cophinforma atrovirens with 100% bootstrap support for MP and ML (Dissanayake et al. 2016; Phillips et al. 2013). Morphological and molecular information presented in this study confirmed this species as C. atrovirens. The pathogenicity was evaluated in cashew tree seedlings (clone BRS 189) by injecting a conidial suspension (6.96 × 10⁵ conidia ml⁻¹) into young shoots with insulin syringe. Five inoculated plants and five noninoculated plants (injected with sterile distilled water as control) were kept in a greenhouse at 28°C under natural sunlight and irrigated daily. The onset of symptoms was observed 3 days after inoculation, evolving to complete rot and decaying death of the shoots at 7 days. A fungus with the same morphological characteristics as the original isolate was successfully reisolated from infected tissues, confirming Koch’s postulates. C. atrovirens is a fungus of the Botyosphaeriaceae family, which has been reported in association with dieback and cankers in woody plants (Dissanayake et al. 2016; Phillips et al. 2013). The presence of this pathogen in a cashew-growing region, which is commonly subject to water stress, poses a threat to cashew industry development. This is the first report of C. atrovirens on cashew plants in Brazil.