Utilization of infectious clones to visualize Cassava brown streak virus replication in planta and gain insights into symptom development

Katie R Tomlinson,Susan E Seal,Gary D Foster,Andy M Bailey

doi:10.1007/s11262-019-01697-5

Katie R Tomlinson, Susan E Seal + Show 2 more

Open Access

https://doi.org/10.1007/s11262-019-01697-5

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Abstract

Cassava brown streak disease (CBSD) is a leading cause of cassava yield losses across eastern and central Africa and is having a severe impact on food security across the region. Despite its importance, relatively little is known about the mechanisms behind CBSD viral infections. We have recently reported the construction of Cassava brown streak virus (CBSV) and Ugandan cassava brown streak virus (UCBSV) infectious clones (IC), which can be used to gain insights into the functions of viral proteins and sequences associated with symptom development. In this study, we perform the first reporter gene tagging of a CBSV IC, with the insertion of green fluorescent protein (GFP) sequence at two different genome positions. Nicotiana benthamiana infections with the CBSV_GFP ICs revealed active CBSV replication in inoculated leaves at 2–5 days post inoculation (dpi) and systemic leaves at 10–14 dpi. We also constructed the chimera CBSV_UCP IC, consisting of the CBSV genome with a UCBSV coat protein (CP) sequence replacement. N. benthamiana infections with CBSV_UCP revealed that the CBSV CP may be associated with high levels of viral accumulation and necrosis development during early infection. These initial manipulations pave the way for U/CBSV ICs to be used to understand U/CBSV biology that will inform vital CBSD control strategies.

Highlights

Cassava brown streak disease (CBSD) is currently having a severe impact on cassava production in East and Central Africa [1]
The localization of Cassava brown streak virus (CBSV) in epidermal and mesophyll cells of N. benthamiana is consistent with untagged CBSV localization to epidermal and mesophyll cells and phloem, detected using immune-histochemical staining of CBSD infected cassava [28]
Green fluorescence during CBSV_GFP1/2 infections of N. benthamiana appeared to peak at 14 dpi, which may correspond to a peak in CBSV replication, as was identified during untagged CBSV infections (Fig. 3)

Summary

Introduction

Cassava brown streak disease (CBSD) is currently having a severe impact on cassava production in East and Central Africa [1]. Across Africa, cassava is the second most important crop in terms of per-capita calories consumed [2]. Cassava produces carbohydrate rich storage roots that are consumed by the grower or sold at markets to generate income [3]. Unlike other staple food crops, cassava can be harvested throughout the year, grows on marginal soil and Edited by Karel Petrzik. Tolerates unpredictable rainfall [3] and so is predicted to provide opportunities for climate change adaptation in Africa [4]. CBSD symptoms include storage root necrosis, radial root constrictions, foliar chlorosis, brown streaks on stems and stunting [5]

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