Diagnóstico e descoberta de badnavírus associados a cará-moela (Dioscorea bulbifera L.) no Brasil via sequenciamento Illumina

Abstract

Dioscorea bulbifera L., commonly known as air yam, is an edible crop belonging to the botanical family Dioscoreaceae and it has increasingly attracted attention due to its socioeconomical relevance for smallholder farmers in Brazil. Although its natural resistance to diseases, air yam is affected by plant pararetroviruses into the genus Badnavirus (family Caulimoviridae). Badnaviruses have double-stranded, semicircular, DNA genomes of 7.4 to 9.0 kb in size, encapsidated into non-enveloped, bacilliform particles and are mainly transmitted by mealybugs (Pseudococcidae). To molecularly characterize the species diversity and complete genomes of badnaviruses associated with D. bulbifera in Brazil, plant samples (n=60) were collected from different growing regions. Partial sequences of the reverse transcriptase (RT) and ribonuclease H (RNase H) domains (~530 bp) were amplified by PCR and Sanger sequenced from 26 positive samples, while full-length genome sequences were recovered from three representative samples. The new partial RT-RNase H sequences were identified as belonging to the badnaviruses Dioscorea bacilliform AL virus, Dioscorea bacilliform SN virus, and Dioscorea bacilliform TR virus, and the endogenous pararetrovirus Dioscorea rotundata endogenous virus eDBV12. Further, a putative species novel tentatively named Dioscorea bacilliform BL virus was partially characterized, sharing highest nucleotide identity with eDBV12 sequences, at 73.4-79.9%. The Bayesian phylogenetic tree based on partial RT-RNase H sequences showed that the new sequences were clustered in three different clades, with the new species being more closely related to eDBV12. Finally, three Illumina-based badnaviral genomes were assembled from 2,240 to 30,668 reads and a coverage depth of 74 to 963 times. The new genomes ranged from 7,208 to 7,420 bp in size and showed typical badnaviral genomic organization with three main open reading frames (ORFs 1-3). The complete RT-RNase H sequences (~1,230 bp) retrieved from the new viral genomes were more closely related to Dioscorea bacilliform AL virus (n=1) and Dioscorea bacilliform SN virus (n=2), at 85.1-86.6 and 82.2-83.9% nucleotide identity, respectively. These results reinforce the high badnaviral species diversity usually observed associated with Dioscorea spp. and constitute the first report of Dioscorea bacilliform TR virus (DBTRV) in D. bulbifera. Also, these are the first complete genome sequences of yam-infecting badnaviruses described in Brazil.