Isolation, genome analysis and tissue localization of Ceratobasidium theobromae, a new encounter pathogen of cassava in Southeast Asia.

In Southeast Asia (SEA) fastidious fungi of the Ceratobasidium genus are associated with proliferation of sprouts and vascular necrosis in cacao and cassava, crops that were introduced from the tropical Americas to this region. Here, we report the isolation and in vitro culture of a Ceratobasidium sp. isolated from cassava with symptoms of witches' broom disease (CWBD), a devastating disease of this crop in SEA. The genome characterization using a hybrid assembly strategy identifies the fungus as an isolate of the species C. theobromae, the causal agent of vascular streak dieback of cacao in SEA. Both fungi have a genome size > 31 Mb (G+C content 49%), share > 98% nucleotide identity of the Internal Transcribed Spacer (ITS) and > 94% in genes used for species-level identification. Using RNAscope® we traced the pathogen and confirmed its irregular distribution in the xylem and epidermis along the cassava stem, which explains the obtention of healthy planting material from symptom-free parts of a diseased plant. These results are essential for understanding the epidemiology of CWBD, as a basis for disease management including measures to prevent further spread and minimize the risk of introducing C. theobromae via long-distance movement of cassava materials to Africa and the Americas.

Ceratobasidium sp. is associated with cassava witches' broom disease, a re-emerging threat to cassava cultivation in Southeast Asia.

Cassava witches' broom disease (CWBD) is a devastating disease of cassava in Southeast Asia (SEA), of unknown etiology. Affected plants show reduced internodal length, proliferation of leaves and weakening of stems. This results in poor germination of infected stem cuttings (i.e., planting material) and significant reductions in fresh root yields and starch content, causing economic losses for farmers and processors. Using a metagenomic approach, we identified a fungus belonging to the Ceratobasidium genus, sharing more than 98.3-99.7% nucleotide identity at the Internal Transcribed Spacer (ITS), with Ceratobasidium theobromae a pathogen causing similar symptoms in cacao. Microscopy analysis confirmed the identity of the fungus and specific designed PCR tests readily showed (1) Ceratobasidium sp. of cassava is strongly associated with CWBD symptoms, (2) the fungus is present in diseased samples collected since the first recorded CWBD outbreaks in SEA and (3) the fungus is transmissible by grafting. No phytoplasma sequences were detected in diseased plants. Current disease management efforts include adjustment of quarantine protocols and guarantee the production and distribution of Ceratobasidium-free planting material. Implications of related Ceratobasidium fungi, infecting cassava, and cacao in SEA and in other potential risk areas are discussed.

Mitochondrial Genetic Diversity of Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) Associated with Cassava in Lao PDR.

Cassava Mosaic Disease (CMD) caused by Sri Lankan cassava mosaic virus (SLCMV), has rapidly spread in Southeast Asia (SEA) since 2016. Recently it has been documented in Lao PDR. Previous reports have identified whitefly species of B. tabaci as potential vectors of CMD in SEA, but their occurrence and distribution in cassava fields is not well known. We conducted a countrywide survey in Lao PDR for adult whiteflies in cassava fields, and determined the abundance and genetic diversity of the B. tabaci species complex using mitochondrial cytochrome oxidase I (mtCOI) sequencing. In order to expedite the process, PCR amplifications were performed directly on whitefly adults without DNA extraction, and mtCOI sequences obtained using nanopore portable-sequencing technology. Low whitefly abundances and two cryptic species of the B. tabaci complex, Asia II 1 and Asia II 6, were identified. This is the first work on abundance and genetic identification of whiteflies associated with cassava in Lao PDR. This study indicates currently only a secondary role for Asia II in spreading CMD or as a pest. Routine monitoring and transmission studies on Asia II 6 should be carried out to establish its potential role as a vector of SLCMV in this region.