Evaluation of a non-radioactive dot-blot nucleic acid hybridization as a detection tool for citrus leaf blotch virus in Cuba

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Keywords:
budwood, certification, Citrivirus, CLBV, diagnosis, Flexiviridae, Fortunella

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Lester Hernández-Rodríguez
Instituto de Investigaciones en Fruticultura Tropical
Gabriel García-García
Instituto de Investigaciones en Fruticultura Tropical
Juana María Pérez-Castro
Instituto de Investigaciones en Fruticultura Tropical
Lochy Batista-Le Riverend
Instituto de Investigaciones en Fruticultura Tropical
Victoria Zamora-Rodríguez
Instituto de Investigaciones en Fruticultura Tropical
Inés Peña-Barzaga
Instituto de Investigaciones en Fruticultura Tropical
Pedro Luis Ramos-González
Instituto de Investigaciones en Fruticultura Tropical

Abstract

A non-radioactive dot-blot nucleic acid hybridization method was evaluated for detecting citrus leaf blotch virus (CLBV, Flexiviridae: Citrivirus) in total RNA extracts from kumquat trees. DNA fragments partially encompassing the open reading frames encoding the polymerase (RdRp) and the movement (MP coat (CP) proteins of CLBV were used to generate digoxigenin (DIG-11-dUTP) probes by PCR. Probes were individually generated using cloned DNA fragments of the Cuban CLBV isolate as templates. In the assays, several working conditions, including the use of different membranes, hybridization and washing buffers were tested. The method detected up to 40 pg of a plasmid containing the complete CLBV genome, as well as viral genomic sequences in RNA extracts of naturally infected trees of 'Nagami' kumquat (Fortunella margarita) and 'Round' kumquat (F. japonica). Results support considering the method developed an appropriated tool to be used after its validation by the programs of sanitation, quarantine, and certification of citrus propagation materials in Cuba.

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How to Cite
Hernández-Rodríguez, L., García-García, G., Pérez-Castro, J. M., Batista-Le Riverend, L., Zamora-Rodríguez, V., Peña-Barzaga, I., & Ramos-González, P. L. (2020). Evaluation of a non-radioactive dot-blot nucleic acid hybridization as a detection tool for citrus leaf blotch virus in Cuba. Revista De Protección Vegetal, 34(3). Retrieved from https://revistas.censa.edu.cu/index.php/RPV/article/view/1057
Issue
Vol. 34 No. 3 (2019): september-december
Section
ORIGINAL ARTICLES

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References

Navarro L, Pina JA, Ballester-Olmos JF, Moreno P, Cambra M. A new graft transmissible disease found in Nagami kumquat. In: Proc. 9th Conf. of IOC; 1984; Riverside: C.A. Timmer LW and. Dodds JA (eds.); p. 234-240.

Galipienso L, Navarro L, Ballester-Olmos JF, Pina JA, Moreno P, Guerri J. Host range and symptomatology of a graft transmissible pathogen causing bud union crease of citrus on trifoliate rootstocks. Plant Pathol. 2000; 49:308-314.

Guardo M, Sorrentino G, Marletta T, Caruso A. First Report of Citrus leaf blotch virus on Kumquat in Italy. Am. Phytopathol. Soc. 2007; 91(8):1054.

Harper J, Chooi KM, Pearson MN. First Report of Citrus leaf blotch virus in New Zealand. Am. Phytopathol. Soc. 2008; 92(10):1470.

Hajeri S, Ramadugu C, Keremane M, Vidalakis G, Lee R. Nucleotide sequence and genome organization of Dweet mottle virus and its relationship to members of the family Betaflexiviridae. Arch. Virol. 2010; 155:1523-1527.

Hernández-Rodríguez L, Pérez-Castro JM, García-García G, Ramos-González PL, Zamora-Rodríguez V, Ferriol-Marchena X, Peña-Bárzaga I, Batista-Le Riverend L. Citrus leaf blotch virus in Cuba: first report and partial molecular characterization. Trop. Plant Pathol. 2016; 41:147-154.

Cao MJ, Yu Y Q, Tian X, Yang FY, Li RH, Zhou CY. First report of Citrus leaf blotch virus in lemon in China. Plant Dis. 2017; 101:1561.

Ping Li, Min LI, Song Zhang, Jun Wang, Fang-yun Yang, Meng-ji Cao, Zhong-an LI. Complete genome sequences of four isolates of Citrus leaf blotch virus from citrus in China. Journal of Integrative Agriculture. 2018; 17(3):712-715.

Chavan RR, Blouin AG, Cohen D, Pearson MN. Characterization of the complete genome of a novel citrivirus infecting Actinidia chinensis. Arch. Virol. 2013; 158(8):1679-1686.

Wang J, Zhu D, Tan Y, Zong X, Wei H, Liu Q. First Report of Citrus leaf blotch virus in Sweet Cherry. Plant Dis. 2016; 100(5):1027.

Zhu CX, Wang GP, Zheng YZ, Yang ZK, Wang LP, Xu WX, et al. RT-PCR detection and sequence analysis of coat protein gene of Citrus leaf blotch virus infecting kiwifruit trees. Acta Phytopathologica Sinica. 2016; 46:11-16.

Gress JC, Smith S, Tzanetakis IE. First report of Citrus leaf blotch virus in peony in the USA. Plant Dis. 2017; 101:637.

Guerri J, Pina JA, Vives MC, Navarro L, Moreno P. Seed transmission of Citrus leaf botch virus: Implications in quarantine and certification programs. Plant Dis. 2004; 88:906-907.

Vives MC, Martín S, Ambrós S, Renovell A, Navarro L, Pina JA, Moreno P, Guerri J. Development of a full-genome cDNA clone of Citrus leaf blotch virus and infection of citrus plants. Mol. Plant Pathol. 2008; 9:787-797.

Guardo M, Potere O, Castellano MA, Savino V, Caruso A. A new herbaceous host of citrus leaf blotch virus. J. Plant Pathol. 2009; 91:485-488.

Agüero J, Vives MC, Velázquez K, Ruiz-Ruiz S, Juárez J, Navarro L, et al. Citrus leaf blotch virus invades meristematic regions in Nicotiana benthamiana and citrus. Mol. Plant Pathol. 2013; 14:610-6.

Adams MJ, Candresse T, Hammond J, Kreuze JF, Martelli GP, Namba S, et al. Family Betaflexiviridae. In King AMQ, Adams MJ, Carstens EB, and Lefkowitz EJ, eds.: Virus Taxonomy: IXth Report of the International Committee on Taxonomy of Viruses. Elsevier Academic Press, London. 2012; p. 920-941.

Galipienso L, Vives MC, Moreno P, Milne RG, Navarro L, Guerri J. Partial characterization of citrus leaf blotch virus, a new virus from Nagami kumquat. Arch. Virol. 2001; 146:357-368.

Vives MC, Galipienso L, Navarro L, Moreno P, Guerri J. The nucleotide sequence and genomic organization of Citrus leaf blotch virus: a candidate type species for a new virus genus. Virology. 2001; 287: 225-233.

Vives MC, Galipienso L, Navarro L, Moreno P, Guerri J. Characterization of two kinds of subgenomic RNAs produced by citrus leaf blotch virus. Virology. 2002; 295:328-336.

Renovell A, Gago S, Ruiz-Ruiz S, Velázquez K, Navarro L, Moreno P, et al. Mapping the subgenomic RNA promoter of the Citrus leaf blotch virus coat protein gene by Agrobacterium-mediated inoculation. Virology. 2010; 406:360-369.

Galipienso L, Vives MC, Navarro L, Moreno P, Guerri J. Detection of Citrus leaf blotch virus using digoxigenin-labeled cDNA probes and RT-PCR. Eur. J. Plant Pathol. 2004; 110:175-181.

Vives MC, Moreno P, Navarro L, Guerri J. Citrus leaf blotch virus. In: Rao GP, Myrta A, Ling K (eds) Characterization, Diagnosis and Management of Plant Viruses. Studium Press, Houston. 2008; p. 55-67

Ruiz-Ruiz S, Ambrós S, Vives MC, Navarro L, Moreno P, Guerri J. Detection and quantitation of Citrus leaf blotch virus by TaqMan real-time RT-PCR. J. Virol. Methods. 2009; 160:57-62.

López MM, Llop P, Olmos A, Marco-Noales E, Cambra M, Bertolini E. Are molecular tools solving the challenges posed by detection of plant pathogenic bacteria and viruses? Curr. Issues Mol. Biol. 2009; 11(1):13-46.

Ramos-González PR, Hernández-Rodríguez L, Banguela-Castillo A. Plataformas genéricas para el diagnóstico a gran escala en fitopatología [Generic platforms for high-throughput diagnosis in phytopathology]. CitriFrut. 2011; 28(1):25-7.

Roistacher CN. Graft-transmissible Diseases of Citrus. Handbook for detection and diagnosis. Food & Agriculture Org. FAO, Rome. 1991; 286 p.

Vives MC, Rubio L, Galipienso L, Navarro L, Moreno P, Guerri J. Low genetic variation between isolates of Citrus leaf blotch virus from different host species and different geographical origins. J. Gen. Virol. 2002; 83:2587-2591.

Lion T, Haas OA. Nonradioactive labeling of probe with digoxigenin by polymerase chain reaction. Anal. Biochem. 1990; 188(2):335-337.

Sambrook J, Fritsch EF and Maniatis T. Molecular hybridization. In: Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory. 1989: p. 931-937.

Holtke HJ, Sagner G, Kessler C, Schmitz G. Sensitive chemiluminescent detection of digoxigenin-labeled nucleic acids: a fast and simple prototocol and its applications. Biotechniques. 1992; 12:104-113.

Yadav N, Khurana SMP. Plant Virus Detection and Diagnosis: Progress and Challenges. In: Frontier Discoveries and Innovations in Interdisciplinary Microbiology. Springer India, New Delhi. 2016; p. 97-132.

OIE. OIE Standard for Management and Technical Requirements for Laboratories Conducting Tests for Infectious Diseases. OIE Quality Standard and Guidelines for Veterinary Laboratories: Infectious Diseases. OIE, Paris, France. 2008; p. 1-31.

Llanes-Alvarez Y, Hernández-Rodríguez L, Peña-Bárzaga I. La validación de métodos de diagnóstico como herramienta en los programas de vigilancia y manejo de fitopatógenos. CitriFrut. 2017; 34(1):46-54.

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