Simultaneous detection and identification of four pome fruit viruses by one-tube pentaplex RT-PCR

Hassan, M.; Myrta, A.; Polák, J.

doi:10.1016/j.jviromet.2005.11.002

Cited by 51 publications

(29 citation statements)

References 18 publications

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“…Starkrimson was detected by ELISA and confirmed by RT-PCR (data not shown). Hassan et al (2006) found that evaluated trees were simultaneously infected by one or more of three other viruses, ACLSV, ASGV, and Apple stem pitting virus (ASPV), none of which were expected to affect ELISA results in the current study.…”

Section: Resultsmentioning

confidence: 73%

“…The EPPO diagnostic procedure is based on using woody indicator plants, which takes a minimum of two years. ApMV can also be detected using molecular methods (Hassan et al 2006;Lenz et al 2008) and ELISA (Torrance, Dolby 1984;Turk 1996;Matic et al 2008), which can serve as a quick screening approach. The virus concentration in an infected apple tree varies through the year similarly to other apple tree viruses such as Apple chlorotic leaf spot virus (ACLSV) and Apple stem grooving virus (ASGV), and is higher in the first half of the year (Fuchs 1982;Matic et al 2008).…”

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confidence: 99%

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Relative concentration of Apple mosaic virus coat protein in different parts of apple tree

Svobodá¹,

Polák²

2010

Hort. Sci. (Prague)

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Svoboda J., Polak J., 2010. Relative concentration of Apple mosaic virus coat protein in different parts of apple tree. Hort. Sci. (Prague), 37: 22-26.The optimal conditions for detecting Apple mosaic virus were assessed by determining relative concentrations of viral coat protein in different tissues (leaves, flower petals, dormant buds, and phloem) in five selected symptomless Apple mosaic virus-infected apple trees of two cultivars at different terms during the vegetation period. Relative concentrations of Apple mosaic virus were calculated as the reciprocal value of the Apple mosaic virus coat protein titer determined by ELISA. The highest relative virus concentration and therefore the highest reliability of virus detection was obtained with young leaves in April before flowering. The reliability of the detection was proved by repeating tests of 80 apple trees of four cultivars in the two subsequent years. The presence of Apple mosaic virus was tested in young apple leaves before flowering taken from 472 apple trees from selected orchards of the Czech Republic. The association of the outcome with climate is also discussed.

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Section: Resultsmentioning

confidence: 73%

mentioning

confidence: 99%

Relative concentration of Apple mosaic virus coat protein in different parts of apple tree

Svobodá¹,

Polák²

2010

Hort. Sci. (Prague)

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“…Для синтеза первой нити кДНК использовали набор RevertAid H Minus First Strand cDNA Synthesis Kit (Thermo Scientific, EU) согласно рекомендованному протоколу. Идентификацию вирусов ACLSV, ASGV и ASPV выпол-няли по методу, описанному в работе (Hassan et al, 2006). В качестве внутреннего контроля в реакции использовали амплификацию гена nad5 (Menzel et al, 2002).…”

Section: материалы и методыunclassified

Distribution and genetic diversity of three apple viruses in Belarus

Kuzmitskaya¹,

Urbanovich²

2016

Vestn. VOGiS

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Apple chlorotic leaf spot virus (ACLSV), apple stem pitting virus (ASPV), and apple stem grooving virus (ASGV) are common in all apple cultivating countries including Belarus. The aim of this investigation was to study the genetic diversity of the apple-tree viruses in Belarus. Virus-infected apple trees were identified by RT-PCR in modern horticultural plantations and were not found among old trees aged more than 50 years. The genome fragments of the ACLSV, ASPV, and ASGV viruses detected were cloned and sequenced. The analysis of their nucleotide sequences gives evidence of high molecular variability generated mostly by single nucleotide substitutions and rarely by single nucleotide deletions and insertions. Recombination processes could also make some contribution to the existing genetic diversity of the virus populations studied. Estimates of selective pressure on the genome fragments of ACLSV, ASPV and ASGV obtained in this study as well as homologous sequences from the GenBank database indicate negative selection, except for one codon in the sequence of the ACLSV coat protein, which is under positive selection. The study of phylogenetic relationships between viruses isolated in different countries did not reveal any clear-cut relationship between their geographical origin and the degree of similarity of virus genome fragments.Keywords: RT-PCR; ACLSV; ASGV; ASPV; genetic diversity; selection pressure.Вирус хлоротической пятнистости листьев яблони (ACLSV), вирус ямчатости древесины яблони (ASPV) и вирус бороздчатости дре-весины яблони (ASGV) распространены во всех странах, где воз-делывается яблоня. В данной работе исследовалось генетическое разнообразие этих патогенов на территории Беларуси. Заражен-ные различными вирусными инфекциями деревья были обна-ружены с помощью метода ОТ-ПЦР в современных садовых насаж дениях. Среди старых деревьев, возрастом более 50 лет, заражен ных не выявлено. Фрагменты геномов вирусов ACLSV, ASPV и ASGV были клонированы и секвенированы. Анализ их нуклеотидных последовательностей свидетельствует о высокой молекулярной вариабельности, сформированной за счет одно-нуклеотидных замен, реже -однонуклеотидных делеций и инсер-ций. Некоторый вклад в существующее генетическое разнообра-зие популяции вирусов также могли вносить и рекомбинацион-ные процессы. Оценка селекционного давления на фрагменты геномов ACLSV, ASPV и ASGV, полученные в настоящем исследова-нии, а также гомологичные последовательности из базы данных GenBank свидетельствует о том, что они находятся под воздей-ствием отрицательной селекции, за исключением одного кодона в последовательности белка оболочки ACLSV, находящегося под воздействием положительной селекции. При исследовании филогенетических взаимоотношений изолятов вирусов, выделен-ных в разных странах, четкой зависимости между географическим происхождением и степенью идентичности фрагментов геномов вирусов выявлено не было. Ключевые

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“…According to Saade et al, 2000, nonisotopic molecular hybridization and multiplex RT-PCR methodologies were developed that could detect all these viruses simultaneously. Meanwhile, the presence of a number of mixed viral infections at apple, already reported in the Mediterranean (Hassan et al, 2005;Di Terlizzi et al, 2003, etc) raises the need for further investigations on the issue. This study aimed to detect viruses at in vitro propagated plant material based on RT-PCR methodology, provide considerations on the efficacy of in vitro propagation of apple from seeds and buds as a tool for phytosanitation, and offer an indication on possible transmission of the infections from seeds.…”

Section: Introductionmentioning

confidence: 98%

“…Considering the knowledge on actual status of viral disease at apple in Albania, and in accordance to EPPO, 1999, these plants have to be tested. Nowadays the basic methods for the detection are ELISA and RT-PCR (Posnette, et al, 1963;Hassan et al, 2005;Menzel et al, 2002;Kinard et al, 1996;Watpade et al, 2013, etc). According to Saade et al, 2000, nonisotopic molecular hybridization and multiplex RT-PCR methodologies were developed that could detect all these viruses simultaneously.…”

Section: Introductionmentioning

confidence: 99%

Seed and Bud Propagated Plantlets Can Be Used to Double Check the Presence of Mixed Viral Infections at Apple

Grazhdani¹,

Bacu²

2015

AgricultForest

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SUMMARYTwo apple cultivars, Golden delicious and Starking grown in Devoll, Albania, were analyzed to verify the rate of infection by ApMV (Apple Mosaic Virus), ASGV (Apple Stem Growing Virus), ACLSV (Apple Chlorotic Stem Virus) and ASPV (Apple Stem Pitting Virus) using in vitro propagated plantlets, and results taken from different categories of explants and among collections were compared based in the fact that template concentrations were equal. Samples, in total 450 trees, were buds taken from branches collected in early spring from symptomless trees, and seeds taken from fruits at three collection orchards during consecutive years 2013, 2014, 2015. Total RNA was extracted from plantlets grown in vitro from buds and seeds, which were used as template for One-step RT-PCR detection of the viruses. Results showed that viruses are present in both cultivars from the three collection orchards. Concentrations of amplicons varied among cultivars, explants categories and from collection to collection, which suggested different infection rates. The presence of mixed latent viral infection was discussed in terms of a possible mechanism of crossprotection among viral strains already amplified, and concentration of amplicons considered proportionate to the infection rate, was seen as a tool to monitor the situation in collections even in the absence of field symptoms.

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Simultaneous detection and identification of four pome fruit viruses by one-tube pentaplex RT-PCR

Cited by 51 publications

References 18 publications

Relative concentration of Apple mosaic virus coat protein in different parts of apple tree

Relative concentration of Apple mosaic virus coat protein in different parts of apple tree

Distribution and genetic diversity of three apple viruses in Belarus

Seed and Bud Propagated Plantlets Can Be Used to Double Check the Presence of Mixed Viral Infections at Apple

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