Viral metagenomics of aphids present in bean and maize plots on mixed-use farms in Kenya reveals the presence of three dicistroviruses including a novel Big Sioux River virus-like dicistrovirus

Francis O Wamonje,Luke A Braidwood,Jagger J W Harvey,John P Carr,Appolinaire Djikeng,Joyce N Njuguna,J Musembi Mutuku,George N Michuki

doi:10.1186/s12985-017-0854-x

Abstract

BackgroundAphids are major vectors of plant viruses. Common bean (Phaseolus vulgaris L.) and maize (Zea mays L.) are important crops that are vulnerable to aphid herbivory and aphid-transmitted viruses. In East and Central Africa, common bean is frequently intercropped by smallholder farmers to provide fixed nitrogen for cultivation of starch crops such as maize. We used a PCR-based technique to identify aphids prevalent in smallholder bean farms and next generation sequencing shotgun metagenomics to examine the diversity of viruses present in aphids and in maize leaf samples. Samples were collected from farms in Kenya in a range of agro-ecological zones.ResultsCytochrome oxidase 1 (CO1) gene sequencing showed that Aphis fabae was the sole aphid species present in bean plots in the farms visited. Sequencing of total RNA from aphids using the Illumina platform detected three dicistroviruses. Maize leaf RNA was also analysed. Identification of Aphid lethal paralysis virus (ALPV), Rhopalosiphum padi virus (RhPV), and a novel Big Sioux River virus (BSRV)-like dicistrovirus in aphid and maize samples was confirmed using reverse transcription-polymerase chain reactions and sequencing of amplified DNA products. Phylogenetic, nucleotide and protein sequence analyses of eight ALPV genomes revealed evidence of intra-species recombination, with the data suggesting there may be two ALPV lineages. Analysis of BSRV-like virus genomic RNA sequences revealed features that are consistent with other dicistroviruses and that it is phylogenetically closely related to dicistroviruses of the genus Cripavirus.ConclusionsThe discovery of ALPV and RhPV in aphids and maize further demonstrates the broad occurrence of these dicistroviruses. Dicistroviruses are remarkable in that they use plants as reservoirs that facilitate infection of their insect replicative hosts, such as aphids. This is the first report of these viruses being isolated from either organism. The BSRV-like sequences represent a potentially novel dicistrovirus infecting A. fabae.

Highlights

Aphids are major vectors of plant viruses
Following PCR of aphid DNA using primers specific for Cytochrome oxidase 1 (CO1), a basic local alignment search tool (BLAST) [34] search of the generated CO1 gene sequences showed that the samples were all A. fabae
In this article, we describe DNA-based species identification of winged aphids collected from farms in Kenya and the detection, using viral metagenomics, of dicistrovirus sequences in these aphids and in maize leaf samples

Summary

Introduction

Aphids are major vectors of plant viruses. Common bean (Phaseolus vulgaris L.) and maize (Zea mays L.) are important crops that are vulnerable to aphid herbivory and aphid-transmitted viruses. We used a PCR-based technique to identify aphids prevalent in smallholder bean farms and generation sequencing shotgun metagenomics to examine the diversity of viruses present in aphids and in maize leaf samples. Aphids are themselves infected by viruses, which may cause disease in the insects [1, 3]. Both RhPV and ALPV can be transmitted between insects (horizontal transmission) and transovarially (vertical transmission) [3] These aphid-infecting viruses are transmitted horizontally through plants, which serve as infection reservoirs but are not considered to be hosts, since they do not support virus replication [3]. We carried out viral metagenomic studies on aphid and plant samples collected on smallholder farms in from different agroecological zones in Kenya, focusing on farms where common bean and/or maize were grown

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