A newly emerging alphasatellite affects banana bunchy top virus replication, transcription, siRNA production and transmission by aphids

Banana bunchy top virus (BBTV) is a six-component ssDNA virus (genus Babuvirus, family Nanoviridae) transmitted by aphids, infecting monocots (mainly species in the family Musaceae) and likely originating from South-East Asia where it is frequently associated with self-replicating alphasatellites. Illumina sequencing analysis of banana aphids and leaf samples from Africa revealed an alphasatellite that should be classified in a new genus, phylogenetically related to alphasatellites of nanoviruses infecting dicots. Alphasatellite DNA was encapsidated by BBTV coat protein and accumulated at high levels in plants and aphids, thereby reducing helper virus loads, altering relative abundance (formula) of viral genome components and interfering with virus transmission by aphids. BBTV and alphasatellite clones infected dicot Nicotiana benthamiana, followed by recovery and symptomless persistence of alphasatellite, and BBTV replication protein (Rep), but not alphasatellite Rep, induced leaf chlorosis. Transcriptome sequencing revealed 21, 22 and 24 nucleotide small interfering (si)RNAs covering both strands of the entire viral genome, monodirectional Pol II transcription units of viral mRNAs and pervasive transcription of each component and alphasatellite in both directions, likely generating double-stranded precursors of viral siRNAs. Consistent with the latter hypothesis, viral DNA formulas with and without alphasatellite resembled viral siRNA formulas but not mRNA formulas. Alphasatellite decreased transcription efficiency of DNA-N encoding a putative aphid transmission factor and increased relative siRNA production rates from Rep- and movement protein-encoding components. Alphasatellite itself spawned the most abundant siRNAs and had the lowest mRNA transcription rate. Collectively, following African invasion, BBTV got associated with an alphasatellite likely originating from a dicot plant and interfering with BBTV replication and transmission. Molecular analysis of virus-infected banana plants revealed new features of viral DNA transcription and siRNA biogenesis, both affected by alphasatellite. Costs and benefits of alphasatellite association with helper viruses are discussed. Author summarySelf-replicating alphasatellites are frequently associated plant ssDNA viruses. Their origin and costs versus benefits for helper virus replication, antiviral defense evasion and transmission by insect vectors are poorly understood. Here we describe identification in Africa and in depth molecular and biological characterization of a newly emerging alphasatellite of BBTV, a multicomponent ssDNA babuvirus causing one of the most economically-important diseases of monocotyledonous bananas and plantains. Phylogenetically, this alphasatellite represents a novel genus and is more related to alphasatellites of nanoviruses infecting dicot hosts than to other BBTV alphasatellites previously identified only in Asia. Consistent with its hypothetical dicot origin, cloned alphasatellite and BBTV can establish systemic infection in a model dicot plant, followed by recovery and symptomless alphasatellite persistence. In banana plants, alphasatellite competes for the host replication and transcription machinery and accumulates at high levels, thereby reducing loads of the helper virus, modifying relative abundance of its components and interfering with its acquisition and transmission by aphids. On the other hand, plant antiviral defenses silence alphasatellite gene expression at both transcriptional and posttranscriptional levels, generating highly-abundant 21, 22 and 24 nucleotide small interfering RNAs, suggesting that alphasatellite may serve as a decoy protecting its helper virus from gene silencing.