First report of pepper leaf curl Bangladesh virus (PepLCBV) associated with cotton leaf curl Multan betasatellite on kid
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First report of pepper leaf curl Bangladesh virus (PepLCBV) associated with cotton leaf curl Multan betasatellite on kidney bean (Phaseolus vulgaris) in Pakistan Saleem Ur Rahman 1,2 & Muhammad Zubair 1,3 & Nasim Ahmed 1,2 & Muhammad Zuhaib Khan 1 & Ghulam Raza 1 & Imran Amin 1 & Shahid Mansoor 1 Received: 6 May 2019 / Accepted: 17 January 2020 # Società Italiana di Patologia Vegetale (S.I.Pa.V.) 2020
Keywords Begomovirus . Pepper leaf curl Bangladesh virus . Cotton leaf curl Multan betasatellite . Kidney bean
Kidney bean is an important legume crop grown throughout the world and its production is badly affected by begomoviruses (Collins et al. 2009). To better understand the diversity of begomoviruses causing infection in kidney beans in Pakistan, a field survey was conducted during year 2016 and typical symptoms of begomovirus infection (downward leaf curling, yellow mosaic and stunting) were observed. A total of 11 symptomatic samples were collected from the field. Total nucleic acids were extracted using cetyl trimethyl ammonium bromide method (Doyle and Doyle 1987) and circular molecules were enriched with rolling circle amplification (RCA). The RCA product was used as template in PCR with chilli leaf curl virus specific primer (SAZ-CHLCV1-F: CATATGCG TCGTTGGCAGA; SAZ-CHLCV1-R: CATATGCC GAGGCAATCAATTCCAGG) for chilli leaf curl virus and β01/β02 (Briddon et al. 2002) for betasatellite. The products of ~2.8 kb and ~1.4 kb were amplified by using these primers. These PCR fragments were cloned in T/A cloning vector (pTZ57R/T; Thermofisher Scientific). Two of the clones, SR12 and SR-33 for begomovirus and associated satellite were sequenced by Applied Biosystems 3730XL DNA sequencer
(USA). The sequences were submitted to NCBI and are available under Accession Nos. MG564658 and MG913377 for clones SR-12 and SR-33, respectively. The Sequence Demarcation Tool (SDT) analysis of SR-12 showed maximum pairwise sequence identity of 93.2% with pepper leaf curl Bangladesh virus (PepLCBV). Following the ICTV nomenclature, it is a novel strain of PepLCBV identified from kidney beans. The clone SR-33 showed 99.85% identity with cotton leaf curl Multan betasatellite (CLCuMuB; FN432359). All symptomatic samples were PCR positive for the presence of CLCuMuB. A strong hybridization signal was observed for all symptomatic samples by Southern blot analysis using a probe based on the Rep gene of PepLCBV; these results were confirmed by sequencing the amplicons obtained with PepLCBVspecific primers from the same samples. PepLCBV (AM691745) has been previously reported from chilli plants (Capsicum annuum) in Pakistan. There are no reports of identification of PepLCBV and CLCMuB from kidney beans. To the best of our knowledge, this is the first report of PepLCBV associated with CLCuMuB identified from kidney beans in Pakistan.
* Saleem Ur Rahman [email protected]
Imran Amin [email protected] Shahid Mansoor [email protected]
Muhammad Zubair [email protected] Nasim Ahmed [email protected]
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