Biology and Interaction of the Natural Occurrence of Distinct Monopartite Begomoviruses Associated With Satellites in Capsicum annum From India.

Megha Mishra,Pradeep Sharma,R K Gaur,Avinash Marwal,Rakesh Kumar Verma

doi:10.3389/fmicb.2020.512957

Abstract

Chili (Capsicum annuum L.) is an important vegetable and spice crop of tropical and sub-tropical regions. Chili plants showing upward leaf curling, leaf crinkling, and leaf yellowing symptoms, collected from Sikar district of Rajasthan, India, were found to be associated with begomovirus and satellite molecules. The presence of virus was confirmed by PCR using virus-specific primer. The full-length genomic DNA-A of three begomovirus (MM-1, CS-1 and RV-1) and two satellites (MM-2 and MM-3) were cloned which was identified from single symptomatic chili plant. The genome organization of isolated three viruses is similar to those of other Old World monopartite begomoviruses. The comparison of the sequences and closest phylogenetic relationships for the begomoviruses, betasatellite and alphasatellite DNAs revealed that MM-1 was designated as DNA-A of Chili leaf curl virus (ChiLCV), CS-1 is considered to be a new distinct species of Tomato leaf curl Gujrat virus (ToLCGV) whereas RV-1 as a new strain of Cotton leaf curl Multan virus (CLCuMuV). The DNA-A component of ChiLCV showed 8.6%, ToLCGV of 16.6% and CLCuMuV of 7.7% average evolutionary divergence, concomitantly, the betasatellite and alphasatellite molecule had 9.9% and 5.9% overall sequence divergence, respectively. Interestingly, most of the begomoviruses were found to be intra-species recombinants. The dN/dS ratio and Tajima D value of all viral DNA-A component and their associated betasatellite showed their selective control on evolutionary relationships. The nucleotide substitution rates were determined for the DNA-A genomes of ChiLCV (7.22 × 10–4 substitutions site–1 year–1), CLCuMuV (1.49 × 10–4 substitutions site–1 year–1), ToLCGV (7.47 × 10–4 substitutions site–1 year–1), the genome of associated ChiLCB (4.20 × 10–4 substitutions site–1 year–1) and CLCuMuA (1.49 × 10–4 substitutions site–1 year–1). Agro-inoculation studies indicate that the presence of DNA betasatellite induce severe symptoms in N. benthamiana and chili, suggesting prerequisite association for typical disease development.

Highlights

Geminiviruses are critical plant DNA viruses infecting a variety of crops, including both monocotyledonous and dicotyledonous plants (Kumar, 2019)
To identify the causal begomovirus infection in symptomatic chili plants, using polymerase chain reaction (PCR) based amplification of ∼560 bp fragment with universal primers confirm the presence of begomoviral (Wyatt and Brown, 1996) and associated betasatellite (Briddon et al, 2002; Bull et al, 2003) components, but amplification was not optimal with the presence of non-specific amplicons
The monopartite begomoviral genomes were cloned from the rolling circle amplification technology (RCA)-amplified products obtained using total DNA that was extracted from chili plants collected from Sikar, Rajasthan

Summary

Introduction

Geminiviruses are critical plant DNA viruses infecting a variety of crops, including both monocotyledonous and dicotyledonous plants (Kumar, 2019). The DNA-A component encodes for AV1 and AV2 ORFs are present on the virion sense strand and encodes coat protein (CP) and movement protein (MP) respectively, complementary sense strand has AC1, AC2, AC3, and AC4 ORFs encoding replication associated protein (Rep), transcriptional activator protein (TrAP), replication enhancer protein (REn) and AC4 protein, respectively (Zerbini et al, 2017). DNA satellites associated with monopartite begomoviruses have been described as alphasatellite, betasatellite, and deltasatellitets (Fiallo-Olivé et al, 2016) forming disease complexes and emerged as a serious threat to agriculture globally (Rodríguez-Negrete et al, 2019). Alphasatellite belongs to the Geminialphasatellitinae subfamily, a member of the family Alphasatellitidae, comprising a single gene encoding alpha-rep protein in the virion-sense, and a hairpin structure at their origin of replication (Briddon et al, 2018). Earlier finding, suggested that the alphasstellite play an important role in the epidemiology of begomovirus/betasatellite complex (Xie et al, 2010)

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