Molecular characterization of polish blueberry red ringspot virus isolate

In this study, we determined the complete sequence of the genomic DNA of a Polish isolate of Blueberry red ringspot virus (BRRSV24) and compared it with a Czech (Darrow 5), and the US isolates of the virus and those of other Caulimoviridae family. The genomic DNA of BRRSV24 consists of 8,265 nucleotides and encodes eight open reading frames (ORFs). The sequence homologies of the eight ORFs of BRRSV24 were from 95 to 98% in respect of Darrow 5 and from 91 to 98% in respect of the US isolates at the amino acid level. This high level of amino acid sequence identity within the coding regions among the Czech, the US and Polish BRRSV isolates is suggestive of their common origin

Zróżnicowanie genetyczne genu kodującego białko płaszcza izolatów Prunus necrotic ringspot virus

We have obtained and described the nucleotide sequences corresponding to ilarvirus Prunus necrotic ringspot virus (PNRSV] coat protein (CP) gene in eleven Polish isolates, from different Prunus and Rosa species and four isolates provided in plant material from Australia, Hungary and Italy. Virus identification was possible using specific primers allowed amplifying the 700 bp amplicon of coat protein gene. The product was obtained used IC-RT-PCR. The results indicated no association between the host species or the geographic origin and the PNRSV CP sequence specificity. The CP gene nucleotide sequence of studied isolates allowed for clustering them into the previously reported PV32-I, PV96-II and PE5-III phylogroups.W pracy uzyskano i opisano sekwencje genu kodującego białko płaszcza 11 izolatów wirusa nekrotycznej pierścieniowej plamistości wiśni (Prunus necrotic ringspot virus, PNRSV) z różnych gatunków roślin z rodzaju Prunus i Rosa pochodzących z Polski oraz czterech izolatów wirusa uzyskanych w materiale roślinnym z Australii, Węgier i Włoch. Wykorzystując specyficzne startery, uzyskano produkt 700 par zasad obejmujący gen kodujący białko płaszcza wirusa. Do amplifikacji zastosowano technikę IC-RT-PCR. Wykazano brak korelacji pomiędzy gatunkiem, pochodzeniem izolatu, a sekwencją nukleotydów białka jego płaszcza. Sekwencje genu kodującego CP badanych izolatów przyporządkowano do trzech głównych filogenetycznych grup wzorcowych: PV32-I, PV96-II i PE5-III

Population genetics analysis of Garlic virus A, Garlic virus B, Garlic virus C and Garlic virus X

Garlic virus A (GarV-A), Garlic virus B (GarV-B), Garlic virus C (GarV-C) and Garlic virus X (GarV-X) are members of the genus Allexivirus in the family Alphaflexiviridae. In this study, we collected 10, 30, 10 and 14 isolates of GarV-A, GarV-B, GarV-C and GarV-X, respectively, from different parts of Poland. All sequences of coat protein (CP) and nucleic-acid binding protein (NABP) regions of Allexivirus isolates available in GenBank were also included in this study. The nucleotide and amino acid sequences identities within each population differed substantially depending on the region of the genome and virus species. The results of selection pressure analysis showed that populations of each Allexivirus underwent negative selection, but the extent of the negative selection varied. It was also concluded that the GarV-A and GarV-C populations underwent a decrease in population size or balancing selection, while the GarV-B and GarV-X populations underwent an increase in population size. It was concluded that both populations of GarV-X evolved independently in each respective area, in contrast to populations of GarV-A, GarV-B and GarV-C

First Report of <i>Blueberry scorch virus</i> in Elderberry in Poland

Blueberry scorch virus (BlScV) is a member of the genus Carlavirus and one of the most widespread pathogens of highbush blueberry (Vaccinium corymbosum L.). The virus was first reported in the United States and has been reported in several countries in Europe, including Italy, Germany, the Netherlands, and Poland. Symptoms of scorch disease in highbush blueberry include necrosis of flower blossoms and leaves, shoot blight, and chlorosis. Sometimes BlScV infection is symptomless or limited to single blossoms and shoots, but all highbush blueberry cultivars are susceptible to virus infection. Cranberry (V. macrocarpon L.) and wild black huckleberry (V. membranaceum L.) are known as natural and symptomless hosts of BlScV (1). In June 2012, during the research concerning the occurrence of BlScV in plants outside Vaccinium sp., 15 leaf samples from five elderberry bushes (Sambucus nigra L., family Adoxaceae) were randomly collected from the Lodzkie region in Central Poland and three were positive in double antibody sandwich (DAS)-ELISA using specific antiserum (Agdia Inc., Elkhart, IN). To confirm the presence of the virus, total nucleic acid was extracted from ELISA-positive elderberry samples according to established protocol (T. Malinowski. Proc. 4th Int. EFPP Symposium, 445, 1996) and used in one step reverse transcription PCR. Primers were developed against the published NJ-2, BC-1, and BC-2 sequences of BlScV (GenBank Accession Nos. NC_003499, AY941198, and AY941199, respectively). The forward primer, RDP_1 (5′-ATGGCACTCACATACAGAAGTCC-3′), and the reverse primer, RDP_2 (5′-TGCCTCTTCAATGCACGATGTTC-3′), were used to amplify a 420-bp fragment of the RNA-dependent RNA polymerase gene of the virus. Amplicons of expected size were obtained from three DAS-ELISA-positive samples, while no products were observed for the negative control (DAS-ELISA-negative elderberry tissues). Sequence of one selected PCR product revealed 100, 88, and 87% nucleotide sequence identity and 100, 96, and 96% amino acid sequence identity with BC-2, NJ-2, and BC-1, respectively. BlScV-infected elderberry bushes were asymptomatic. As BlScV is transmitted by aphids in a non-persistent manner, infected elderberry bushes near highbush blueberry plantings may play an important role in virus spread. The potential for BlScV infection of plants outside family Ericaceae should be investigated. To the best of our knowledge, this is the first report of BlScV infecting elderberry. Reference: (1) R. R. Martin et al. Viruses 4:2831, 2012. </jats:p

Wykrywanie wirusa oparzeliny borówki wysokiej w różnych okresach roku

Viral diseases are a worldwide problem of blueberry which a major limiting factor for production. A survey for Blueberry scorch virus (BlScV) by DAS-ELISA in various organs of highbush blueberry conducted from May 2010 to April 2011, showed the occurrence of these virus in cvs Bluecrop and Herbert, which showing virus-like symptoms. Samples of plant materials (bud flower, flower, leaf, bark) were collected individually from each highbush blueberry plant of every cultivar. It was established that the detection of virus of each the investigated bushes cvs Bluecrop and Herbert depended on the tested plant materials as well as the period in which the tests were performed. The effectiveness of the virus detection varied for the investigated cultivars. The presence of the BlScV was confirmed in leaves samples with specific primer pair which amplifies a 430 bp fragment of the 5’-proximal ORF I [RNA-dependent RNA polymerase (RdRp)].Celem przeprowadzonych badań było wykrywanie i identyfikacja wirusa oparzeliny borówki wysokiej (Blueberry scorch virus, BlScV) w różnych organach pobieranych z krzewów borówki wysokiej odmian Bluecrop i Herbert rosnących na plantacji produkcyjnej zlokalizowanej w centralnej Polsce. Badania byáy prowadzone w okresie od maja 2010 do kwietnia 2011 r. przy użyciu testu serologicznego DAS-ELISA. Próby materiału roślinnego (pąki kwiatowe, kwiaty, liście, kora) pobierano indywidualnie z krzewów kaĪdej z badanych odmian. Ustalono, że wykrywanie wirusów w krzewach odmian Bluecrop i Herbert zależało od testowanego organu oraz terminu, w którym przeprowadzono test. Obecność BlScV w krzewach badanych odmian potwierdzono przy pomocy techniki RT-PCR z wykorzystaniem starterów amplifikujących fragment 5’ genu kodującego polimerazą RNA zależną od RNA

Ocena stanu porażenia przez allexiwirusy roślin czosnku w Polsce

Garlic (Allium sativum L.) plants may be infected in the field by viruses of the genera Potyvirus, Carlavirus and Allexivirus. These viruses are transmitted by vegetative propagation and by vectors. Detection and identification of allexiviruses was carried out in 2011–2012. It was based on ELISA and RT-PCR assays. Samples of plant material were collected from 26 garlic production fields located in different regions of Poland. Garlic virus D, Garlic virus B and Garlic virus X were the most abundant viruses in all examined regions and were identified in 79%, 64% and 59% of all garlic samples, respectively. Garlic virus A and Garlic virus C were identified in all studied regions with low frequency. Garlic virus E was detected with 100% frequency in east-central Poland. None of the tested garlic samples were infected with Shallot virus X. Allexiviruses were always present in garlic plants in mixed infections.Czosnek (Allium sativum L.) może być porażany przez wiele gatunków wirusów należących do rodzajów: Potyvirus, Carlavirus i Allexivirus, które przenoszone są z materiałem rozmnożeniowym, a w okresie wegetacji za pośrednictwem wektorów. Badania przeprowadzono w latach 2011–2012. Ich celem było wykrywanie allexiwirusów porażających rośliny czosnku w Polsce oparte na teście serologicznym ELISA oraz technice RT-PCR. Próby materiału roślinnego pobierano losowo z 26 pól produkcyjnych czosnku. Wirus D czosnku, wirus B czosnku oraz wirus X czosnku były wykrywane we wszystkich badanych rejonach w, odpowiednio, 79, 64 i 59% badanych prób. Wirus A czosnku oraz wirus C czosnku występowały w mniejszej liczbie prób. Wirus E czosnku został wykryty we wszystkich próbach pobranych z pól zlokalizowanych w centralno-wschodniej Polsce. W żadnej z badanych prób nie stwierdzono obecności wirusa X szalotki. Allexiwirusy w roślinach czosnku zawsze występowały w mieszanych infekcjach

COMPARISON OF MOLECULAR ASSAYS FOR THE DETECTION OF ALLEXIVIRUSES IN GARLIC PLANTS

Allexiviruses and several Carla- and Potyvirus species are the most widespread viral pathogens of Allium crops world-wide. Simultaneous infection by these viruses causes significant reduction of bulb weight. Production of species-specific antisera against allexiviruses is complicated. Production of virus-free propagation material requires the use of highly specific, sensitive and rapid virus detection methods. The aim of the present study therefore was to examine primer sequences using RT-PCR and IC-RT-PCR method and testing the dot-blot hybridization for detection of allexiviruses. Electron microscope was also used for selective garlic plants infected with Allexiviruses