Progress in Plant Protection

The use of quantitative PCR with high resolution melting (qPCR-HRM) analysis to distinguish the Polish mild and yellowing pathotypes of Pepino mosaic virus (PepMV)
Zastosowanie techniki ilościowego PCR z analizą krzywych topnienia (qPCR-HRM) do rozróżniania polskich łagodnych i żółtaczkowych patotypów wirusa mozaiki pepino (Pepino mosaic virus, PepMV)

Julia Minicka, e-mail: J.Minicka@iorpib.poznan.pl

Instytut Ochrony Roślin – Państwowy Instytut Badawczy, Zakład Wirusologii i Bakteriologii, Władysława Węgorka 20, 60-318 Poznań, Polska

Beata Komorowska, e-mail: Beata.Komorowska@inhort.pl

Instytut Ogrodnictwa, Zakład Fitopatologii, Konstytucji 3 Maja 1/3, 96-100 Skierniewice, Polska

Beata Hasiów-Jaroszewska, e-mail: B.Hasiow@iorpib.poznan.pl

Instytut Ochrony Roślin – Państwowy Instytut Badawczy, Zakład Wirusologii i Bakteriologii, Władysława Węgorka 20, 60-318 Poznań, Polska
Streszczenie

Pepino mosaic virus (PepMV) is a highly infectious virus causing significant losses in tomato greenhouse production worldwide. One of the characteristic symptoms is yellowing discolorations of leaf blades, which is often accompanied by damage of the fruits. It has been found that, two independent point mutations in the gene encoding the coat protein (CP) are responsible for the formation of yellowing symptoms: G463AA/A463AA (155 aa) and GA497U/GG497U (166 aa), respectively. In our study, qPCR-HRM was used to distinguish mild and yellowing pathotypes. Appropriate primers, amplifying a fragment of the CP-encoding gene in which both yellowing mutations are located were designed. The analysis was carried out using 18 isolates originated from greenhouses in different region of Poland. Our experiments confirmed the effectiveness of the designed primers to differentiate the analyzed pathotypes. This technique is a valuable tool for detecting the diversity of PepMV.

 

Wirus mozaiki pepino (PepMV) jest groźnym patogenem powodującym znaczne straty w uprawie szklarniowej pomidora w wielu regionach świata. Jednym z charakterystycznych objawów obecności wirusa w roślinie są żółtaczkowe przebarwienia blaszek, którym często towarzyszy uszkodzenie owoców. Za ich powstawanie odpowiedzialne są dwie niezależne mutacje punktowe w genie kodującym białko płaszcza (CP): G463AA/A463AA (155 aa) i GA497U/GG497U (166 aa). W przeprowadzonych badaniach wykorzystano qPCR-HRM do rozróżniania łagodnych i żółtaczkowych patotypów. Zaprojektowano startery, amplifikujące fragment genu kodującego CP, w którym znajdują się obie mutacje żółtaczkowe. Przeanalizowano 18 izolatów, pochodzących z obiektów szklarniowych na terenie całego kraju. Wykonane doświadczenia potwierdziły skuteczność zaprojektowanych starterów do różnicowania analizowanych patotypów. Technika ta może być z powodzeniem stosowana do rozróżniania łagodnych i żółtaczkowych patotypów PepMV na terenie naszego kraju.

Słowa kluczowe
Pepino mosaic virus; CH2 strain; yellowing pathotype; Solanum lycopersicum; szczep CH2; patotyp żółtaczkowy
Referencje

Bester R., Jooste A.E., Maree H.J., Burger J.T. 2012. Real-time RT-PCR high-resolution melting curve analysis and multiplet RT-PCR to detect and differentiate grapevine leafroll-associated virus 3 variant groups I, II, III and VI. Virology Journal 9: 219. DOI: 10.1186/1743-422X-9-219

 

Blystad D.R., van der Vlugt R., Alfaro-Fernandez A., Cordoba M.D., Bese G., Hristova D., Pospieszny H., Mehle N., Ravnikar M., Tomassoli L., Varveri C., Nielsen S.L. 2015. Host range and symptomatology of Pepino mosaic virus strains occurring in Europe. European Journal of Plant Pathology 143 (1): 43–56. DOI: 10.1007/s10658-015-0664-1

 

Córdoba-Sellés M.C., García-Rández A., Alfaro-Fernández A., Jordá-Gutiérrez C. 2007. Seed transmission of Pepino mosaic virus and efficacy of tomato seed disinfection treatments. Plant Disease 91 (10): 1250–1254. DOI: 10.1094/PDIS-91-10-1250

 

Filipiak A., Tomalak M. 2014. Wykorzystanie techniki PCR-HRM do wykrywania kwarantannowego gatunku nicienia Bursaphelenchus xylophilus. [The use of PCR-HRM technique for detection of the quarantine nematode Bursaphelenchus xylophilus]. Progress in Plant Protection 54 (4): 400–402. DOI: 10.14199/ppp-2014-067

 

French C.J., Bouthillier M., Bernardy M., Ferguson G., Sabourin M., Johnson R.C., Masters C., Godkin S., Mumford R. 2001. First report of Pepino mosaic virus in Canada and the United States. Plant Disease 85 (10): 1121. DOI: 10.1094/PDIS.2001.85.10.1121B

 

Gómez P., Sempere R.N., Elena S.F., Aranda M.A. 2009. Mixed infections of Pepino mosaic virus strains modulate the evolutionary dynamics of this emergent virus. Journal of Virology 83 (23): 12378–12387. DOI: 10.1128/JVI.01486-09

 

Hanssen I.M., Mumford R., Blystad D.R., Cortez I., Hasiów-Jaroszewska B., Hristova D., Pagán I., Pereira A.M., Peters J., Pospieszny H., Ravnikar M., Stijger I., Tomassoli L., Varveri C., van der Vlugt R., Nielsen S.L. 2010. Seed transmission of Pepino mosaic virus in tomato. European Journal of Plant Pathology 126 (2): 145–152. DOI: 10.1007/s10658-009-9528-x

 

Hanssen I.M., Paeleman A., Vandewoestijne E., Van Bergen L., Bragard C., Lievens B., Vanacher A.C.R.C., Thomma B.P.H.J. 2009. Pepino mosaic virus isolates and differential symptomatology in tomato. Plant Pathology 58 (3): 450–460. DOI: 10.1111/j.1365-3059.2008.02018.x

 

Hanssen I.M., Paeleman A., Wittemans L., Goen K., Lievens B., Bragard C., Vanachter A.C.R.C., Thomma B.P.H.J. 2008. Genetic characterization of Pepino mosaic virus isolates from Belgian greenhouse tomatoes reveals genetic recombination. European Journal Plant Pathology 121 (2): 131–146. DOI: 10.1007/s10658-007-9255-0

 

Hanssen I.M., Thomma B.P.H.J. 2010. Pepino mosaic virus: a successful pathogen that rapidly evolved from emerging to endemic in tomato crops. Molecular Plant Pathology 11 (2): 179–189. DOI: 10.1111/j.1364-3703.2009.00600.x

 

Hasiów B., Borodynko N., Pospieszny H. 2008. Complete genomic RNA sequence of the Polish Pepino mosaic virus isolate belonging to the US2 strain. Virus Genes 36 (1): 1–8. DOI: 10.1007/s11262-007-0171-3

 

Hasiów-Jaroszewska B., Jackowiak P., Borodynko N., Figlerowicz M., Pospieszny H. 2010. Quasispecies nature of Pepino mosaic virus and its evolutionary dynamics. Virus Genes 41 (2): 260–267. DOI: 10.1007/s11262-010-0497-0

 

Hasiów-Jaroszewska B., Komorowska B. 2013. A new method for detection and discrimination of Pepino mosaic virus isolates using high resolution melting analysis of the triple gene block 3. Journal of Virological Methods 193 (1): 1–5. DOI: 10.1016/j.jviromet.2013.04.022

 

Hasiów-Jaroszewska B., Paeleman A., Ortega-Parra N., Borodynko N., Minicka J., Czerwoniec A., Thomma B.P.H.J., Hanssen I.M. 2013. Ratio of mutated versus wild-type coat protein sequences in Pepino mosaic virus determines the nature and severity of yellowing symptoms on tomato plants. Molecular Plant Pathology 14 (9): 923–933. DOI: 10.1111/mpp.12059

 

Hasiów-Jaroszewska B., Pospieszny H., Borodynko N. 2009. New necrotic isolates of Pepino mosaic virus representing the CH2 genotype. Journal of Phytopathology 157 (7–8): 494–496. DOI: 10.1111/j.1439-0434.2008.01496.x

 

Ling K.S. 2007. Molecular characterization of two Pepino mosaic virus variants from imported tomato seed reveals high levels of sequence identity between Chilean and US isolates. Virus Genes 34 (1): 1–8. DOI: 10.1007/s11262-006-0003-x

 

Ling K., Li R., Bledsoe M. 2013. Pepino mosaic virus genotype shift in North America and development of a loop-mediated isothermal amplification for rapid genotype identification. Virology Journal 10: 117. DOI: 10.1186/1743-422X-10-117

 

Maroon-Lango C.J., Guaragna M.A., Jordan R.L., Hammond J., Bandla M., Marquardt S.K. 2005. Two unique US isolates of Pepino mosaic virus from a limited source of pooled tomato tissue are distinct from a third (European-like) US isolate. Archives of Virology 150 (6): 1187–1201. DOI: 10.1007/s00705-005-0495-z

 

Mehle N., Gutierrez-Aguirre I., Prezelj N., Delić D., Vidic U., Ravnikar M. 2014. Survival and transmission of Potato virus Y, Pepino mosaic virus, and Potato spindle tuber viroid in water. Applied and Environmental Microbiology 80 (4): 1455–1462. DOI: 10.1128/AEM.03349-13

 

Minicka J., Hasiów-Jaroszewska B., Borodynko-Filas N., Pospieszny H., Hanssen I.M. 2016. Rapid evolutionary dynamics of the Pepino mosaic virus – status and future perspectives. Journal of Plant Protection Research 56 (4): 337–345. DOI: 10.1515/jppr-2016-0054

 

Moreno-Pérez M.G., Pagán I., Aragón-Caballero L., Cáceres F., Fraile A., García-Arenal F. 2014. Ecological and genetic determinants of Pepino mosaic virus emergence. Journal of Virology 88 (6): 3359–3368. DOI: 10.1128/JVI.02980-13

 

Noël P., Hance T., Bragard A.C. 2014. Transmission of the Pepino mosaic virus by whitefly. European Journal of Plant Pathology 138 (1): 23–27. DOI: 10.1007/s10658-013-0313-5

 

Pospieszny H., Borodynko N., Palczewska M. 2003. First record of Pepino mosaic virus in Poland. Journal of Plant Disease Protection 100: 97.

 

Roggero P., Masenga V., Lenzi R., Coghe F., Ena S., Winter S. 2001. First report of Pepino mosaic virus in tomato in Italy. Plant Pathology 50 (6): 798. DOI: 10.1046/j.1365-3059.2001.00621.x

 

Schwarz D., Beuch U., Bandte M., Fakhro A., Büttner C., Obermeier C. 2010. Spread and interaction of Pepino mosaic virus (PepMV) and Pythium aphanidermatum in a closed nutrient solution recirculation system: effects on tomato growth and yield. Plant Pathology 59 (3): 443–452. DOI: 10.1111/j.1365-3059.2009.02229.x

 

Shipp J.L., Buitenhuis R., Stobbs L., Wang K., Kim W.S., Ferguson G. 2008. Vectoring of Pepino mosaic virus by bumblebees in tomato greenhouses. Annals of Applied Biology 153 (2): 149–155. DOI: 10.1111/j.1744-7348.2008.00245.x

 

Spence N.J., Basham J., Mumford R.A., Hayman G., Edmondson R., Jones D.R. 2006. Effect of Pepino mosaic virus on the yield and quality of glasshouse-grown tomatoes in the UK. Plant Pathology 55 (5): 595–606. DOI: 10.1111/j.1365-3059.2006.01406.x

 

Tiberini A., Davino S., Davino M., Tomassoli L. 2011. Complete sequence, genotyping and comparative analysis of Pepino mosaic virus isolates from Italy. Journal of Plant Pathology 93 (2): 437–442. DOI: 10.4454/jpp.v93i2.1199

 

van der Vlugt R.A., Stijger C.M., Verhoeven J.T.J., Lesemann D.E. 2000. First report of Pepino mosaic virus on tomato. Plant Disease 84 (1): 103–108. DOI: 10.1094/PDIS.2000.84.1.103C

 

Wittwer C.T., Reed G.H., Gundry C.N., Vandersteen J.G., Pryor R.J. 2003. High-resolution genotyping by amplicon melting analysis using LCGreen. Clinical Chemistry 49 (6): 853–860. DOI: 10.1373/49.6.853

Progress in Plant Protection (2020) : 0-0
Data pierwszej publikacji on-line: 2020-02-07 11:13:55
http://dx.doi.org/10.14199/ppp-2020-001
Pełny tekst (.PDF) BibTeX Mendeley Powrót do listy