Real Time PCR Based Quantification of Banana Bunchy Top Virus (BBTV) Titre in Banana cv. Grand Naine (Musa acuminata)

Main Article Content

N. Tanuja
A. Ramanathan
S. Varanavasiappan
E. Kokila Devi
L. Arul
D. Sudhakar
S. Vanitha
K. Soorianathasundaram
K. K. Kumar

Abstract

Banana Bunchy Top Disease (BBTD) is one of the most severe viral diseases affecting major banana growing belts in India. Banana Bunchy Top Virus (BBTV) is transmitted by a black banana aphid (Pentalonia nigronervosa L Coquerel) in a persistent manner. BBTV virions are limited to the phloem tissue of banana resulting in low titre in banana. A reliable method to quantify the BBTV in the banana will be useful for monitoring the insect vector mediated BBTV transmission in banana, an essential requirement for characterizing the transgenic banana transformed for BBTV resistance. A protocol for real time PCR based absolute quantification of BBTV is reported in the present study. The partial BBTV coat protein gene (459 bp) was isolated, cloned into a plasmid vector and used to construct a standard curve using an SYBR green-based assay with known copies of BBTV coat protein gene. Using the standard curve, BBTV viral load was estimated in BBTV infected symptomatic and asymptomatic leaf samples of banana cultivar Grand Naine through SYBR green-based quantitative Polymerase Chain Reaction (qPCR). The study demonstrated that a higher viral tire was associated with BBTV disease symptoms appearance, whereas the low titre resulted in asymptomatic plants.

Keywords:
Grand Naine, BBTV, coat protein, PCR, real time quantification.

Article Details

How to Cite
Tanuja, N., Ramanathan, A., Varanavasiappan, S., Kokila Devi, E., Arul, L., Sudhakar, D., Vanitha, S., Soorianathasundaram, K., & K. Kumar, K. (2019). Real Time PCR Based Quantification of Banana Bunchy Top Virus (BBTV) Titre in Banana cv. Grand Naine (Musa acuminata). Advances in Research, 20(4), 1-7. https://doi.org/10.9734/air/2019/v20i430163
Section
Original Research Article

References

Magee CJ. Investigation on the bunchy top disease of bananas. Bulletin of the Council for Scientific and Industrial Research in Australia. 1927;30.

Hu JS, Wang M, Sether D, Xie W, Leonhardt KW. Use of polymerase chain reaction (PCR) to study transmission of banana bunchy top virus by the banana aphid (Pentalonia nigronervosa). Annals of Applied Biology. 1996;128(1):55-64.

King AMQ, Adams MJ, Lefkowitz EJ. Virus taxonomy: Classification and nomenclature of viruses: Ninth report of the international committee on taxonomy of viruses. Springer, Wein; 2011.

Karan M, Harding RM, Dale JL. Evidence for two groups of banana bunchy top virus isolates. Journal of General Virology. 1994;75(12):3541-3546.

Watanabe S, Greenwell A, Bressan A. Localization, concentration, and transmission efficiency of Banana bunchy top virus in four asexual lineages of Pentalonia aphids. Viruses. 2013;5(2):758-776.

Furuya N, Somowiyarjo S, Natsuaki KT. Virus detection from local banana cultivars and the first molecular characterization of Banana bunchy top virus in Indonesia. J Agri Sci Tokyo. 2004;49(3):75–81.

Mansoor S, Qazi J, Amin I, Khatri A, Khan IA, Raza S, et al. A PCR-based method, with internal control, for the detection of Banana bunchy top virus in banana. Molecular Biotechnology. 2005;30(2):167-169.

Chen Y, Hu X. High-throughput detection of banana bunchy top virus in banana plants and aphids using real-time TaqMan® PCR. Journal of Virological Methods. 2013;193(1):177-183.

Fu HC, Hu JM, Hung TH, Su HJ, Yeh HH. Unusual events involved in Banana bunchy top virus strain evolution. Phytopathology. 2009;99(7):812-822.

Jebakumar RM, Balasubramanian V, Selvarajan R. Virus titre determines the efficiency of Pentalonia nigronervosa (Aphididae: Hemiptera) to transmit banana bunchy top virus. Virus Disease. 2018; 29(4):499-505.

Doyle JJ, Doyle JL. Isolation of plant DNA from fresh tissue. Focus. 1990;12:13.

Xue B, Guo J, Que Y, Fu Z, Wu L, Xu L. Selection of suitable endogenous reference genes for relative copy number detection in sugarcane. International Journal of Molecular Sciences. 2014;15(5): 8846-8862.

Bernard Ouma. Development of Banana bunchy top virus resistance in bananas: RNAi approach. Ph. D. thesis. Queensland University of Technology; 2016.

Selvarajan R, Balasubramanian V, Anuradha C. Population Structure and Diversity of Banana Bunchy Top Virus and Banana Bract Mosaic Virus. In Plant Viruses. 2018;149-170. CRC Press.

Stainton D, Martin DP, Muhire BM, Lolohea S, Halafihi MI, Lepoint P, et al. The global distribution of Banana bunchy top virus reveals little evidence for frequent recent, human-mediated long-distance dispersal events. Virus Evolution. 2015; 1(1).

Yu NT, Xie HM, Zhang YL, Wang JH, Xiong Z, Liu ZX. Independent modulation of individual genomic component transcription and a cis- acting element related to high transcriptional activity in a multipartite DNA virus. BMC Genomics. 2019;20:573.