Virus Incidence in Four Olea Europaea Cultivars Evaluated by Mechanical Inoculation and Immunological Assays

Félix, Maria R.; Leitão, F.; Serrano, José Moreno

doi:10.17660/actahortic.2002.586.154

Cited by 3 publications

(3 citation statements)

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“…Virus surveys carried in Mediterranean countries and others, concerning eight olive-infecting viruses (SLRSV, Strawberry latent ringspot virus ; ArMV, Arabis mosaic virus ; CLRV, Cherry leafroll virus ; OLRSV, Olive latent ringspot virus ; CMV, Cucumber mosaic virus ; OLYaV, Olive leaf yellowing-associated virus ; OLV-1, Olive latent virus 1 ; OLV-2, Olive latent virus 2 ), have shown high levels of infection, such as 75% in Tunisia, 51% in Syria, 33% in Italy, 31% in Lebanon, 25% in Croatia, and 31% of necroviruses infection in Portugal (Al Abdullah et al, 2005; Fadel et al, 2005; Faggioli et al, 2005; Varanda et al, 2010; Luigi et al, 2011; Zellama et al, 2018). However, the high presence of double stranded RNA (dsRNA) in olive orchards suggests even higher levels of infection (Félix et al, 2002; Saponari et al, 2002) that may be identified with new and more sensitive diagnostic techniques.…”

Section: Introductionmentioning

confidence: 99%

Establishment of a Sensitive qPCR Methodology for Detection of the Olive-Infecting Viruses in Portuguese and Tunisian Orchards

et al. 2019

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Sensitive detection of viruses in olive orchards is actually of main importance since these pathogenic agents cannot be treated, their dissemination is quite easy, and they can have eventual negative effects on olive oil quality. The work presented here describes the development and application of a new SYBR ® Green-based real-time quantitative PCR (qPCR) analysis for specific and reliable quantification of highly spread olive tree viruses: Olive latent virus 1 (OLV-1) , Tobacco necrosis virus D (TNV-D), Olive mild mosaic virus (OMMV), and Olive leaf yellowing-associated virus (OLYaV). qPCR methodology revealed high specificity and sensitivity, estimated in the range of 0.8–8 copies of the virus genome, for the studied viruses. For validation of the method, total RNA and double strand RNA (dsRNA) from naturally infected trees were used. In a first trial, dsRNAs from trees of cv. “Galega vulgar” from a Portuguese orchard, were subjected to qPCR and from the 30 samples tested, 26 were TNV-D and/or OMMV-positive and 25 were OLV-1 positive. In a second trial, total RNA from trees of different cultivars from Tunisian orchards, were here tested by qPCR and all viruses were detected. From the 33 samples studied, the most prevalent virus detected in Tunisia orchards was OLV-1 (31 samples diagnosed), followed by OLYaV (20 samples diagnosed), and finally the combination in last TNV-D and/or OMMV (12 samples diagnosed). In both trials, qPCR demonstrated to be effective and sensitive, even when using total RNA as template. qPCR through the use of a SYBR ® Green methodology enabled, for the first time, a reliable, sensitive, and reproducible estimation of virus accumulation in infected olive trees, in which viruses are usually in low titres, that will allow gaining new insights in virus biology essential for disease control and give an important contribution for establishment of sanitary certification of olive propagative material.

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Section: Introductionmentioning

confidence: 99%

Establishment of a Sensitive qPCR Methodology for Detection of the Olive-Infecting Viruses in Portuguese and Tunisian Orchards

et al. 2019

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“…Main disadvantages of this assay are the variable amounts of virus in the olive tissues throughout the year and content in virus inactivator compounds that may lead to erratic and unreliable results. Yet it has been useful in past surveys aiming at sanitary selection of vegetative material in Portugal, where mechanically transmissible virus were found in up to 100% of samples in several sites and allowed the recovery of isolate collections of the three necrovirus species maintained in our laboratory for further studies (Clara et al 1997;Martelli 1999;Félix et al 2002;Félix and Clara 2008).…”

Section: Biological Assaysmentioning

confidence: 99%

Biology and molecular characterization of necroviruses affecting Olea europaea L.: a review

2011

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Three necroviruses, family Tombusviridae, are known to infect Olea europaea L., Olive latent virus 1 (OLV-1), Olive mild mosaic virus (OMMV) and Tobacco necrosis virus D (TNV-D), often asymptomatically. OLV-1 has a wide geographical distribution whereas that of OMMV and TNV-D is not known as their discovery is recent, dating from 2005 and 2009, respectively. OMMV has received special attention because of its origin, likely to have resulted from a recombination event between TNV-D, with which it shares a coat protein (CP) amino acid identity of 86% and OLV-1 sharing an RNA-dependent RNA polymerase amino acid identity of 91%. Due to these properties OMMV and TNV-D are indistinguishable by serology or Reverse Transcription -Polymerase Chain Reaction assays that use primers hybridizing in the CP gene region, rendering ambiguous earlier TNV-D identifications based on such tests. OLV-1 and OMMV are transmitted through the soil to roots of host plants in the absence of any vectors but the transmission rate of OMMV increases to more than double in the presence of Olpidium brassicae zoospores that vector this virus in an in vitro manner. A point mutation leading to an amino acid substitution located in an inner region of the virion is responsible for the loss of biologically assisted transmission, probably by altering the particle conformation rendering the binding site inaccessible to the zoospore receptors. The field occurrences of necrovirus multiple infections are common and increase the chances for recombination events, the epidemiological implications of which are still uncertain. The biological and molecular properties of the necroviruses identified in olive trees are the main focus of this review.

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“…TNV-like viruses, that were able to be assigned to TNV-D or TNV-A, have been detected in herbaceous plants such as tobacco, tulip, bean, cucumber and potato, as well as woody crop plants such as pear and apple, citrus, grape (Kassanis, 1970;Kurstak, 1981) and olive (Félix and Clara, 2002b). OMMV was identified for the first time in olive and genomic sequences of OMMV, published in the Genbank database, were recovered from tulips affected by the Augusta disease, leading to its identification as the causal agent of that disorder (Pham et al, 2009 Mixed infections of these three viruses occur naturally quite frequently (Félix and Clara, 2002a;Varanda et al, 2010). These alphaand betanecroviruses share important similarities.…”

Section: Introductionmentioning

confidence: 99%

A degenerate pair of primers for simultaneous detection of four alpha- and betanecroviruses

Varanda

Cardoso

Oliveira

et al. 2014

Journal of Virological Methods

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a b s t r a c tThe high infection levels due to Olive latent virus 1 (OLV-1), Olive mild mosaic virus (OMMV) (alphanecrovirus) and Tobacco necrosis virus D (TNV-D) (betanecrovirus) in Portuguese olive orchards prompted us to develop a rapid PCR-based assay for the simultaneous detection of these viruses aimed at the sanitary selection and marketing of plant material in compliance with European Union regulations. A pair of degenerate oligonucleotide primers, parRdRp5 and parCoat3 was designed based on conserved regions located in the RNA-dependent RNA polymerase (RdRp) and coat protein (CP) genes of these viruses and one other alphanecrovirus, Tobacco necrosis virus A. Its use in RT-PCR assays generated a product of ca. 2000 bp for the 4 viral species tested. These primers were compared with virus specific primers in multiplex RT-PCR, and identical results were obtained. Its application to dsRNA extracted from 54 olive field growing trees originated the expected ca. 2000 bp amplicon in 17 trees. The virus identity was determined by sequencing the cloned RT-PCR products. No TNV-A was found.The RT-PCR assay using the degenerate primers described in this study were shown to be reliable in detecting any of the above-mentioned alpha-and betanecroviruses, and it is as sensitive as that which uses virus specific primers in multiplex assays. Therefore, this assay is well suited for the rapid screen of virus-free plant material in selection and improvement crop programmes. Additionally, it has the potential to reveal virus diversity and the presence of new viruses, provided the RT-PCR generated amplicon is further sequenced.

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Virus Incidence in Four Olea Europaea Cultivars Evaluated by Mechanical Inoculation and Immunological Assays

Cited by 3 publications

References 0 publications

Establishment of a Sensitive qPCR Methodology for Detection of the Olive-Infecting Viruses in Portuguese and Tunisian Orchards

Establishment of a Sensitive qPCR Methodology for Detection of the Olive-Infecting Viruses in Portuguese and Tunisian Orchards

Biology and molecular characterization of necroviruses affecting Olea europaea L.: a review

A degenerate pair of primers for simultaneous detection of four alpha- and betanecroviruses

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