Name: Walkíria Andrade Amorim
Type: PhD thesis
Publication date: 27/09/2016
Advisor:

Namesort descending Role
Jose Aires Ventura (M/D) Advisor *

Examining board:

Namesort descending Role
Diolina Moura Silva Internal Examiner *
Francisco Murilo Zerbini Junior External Examiner *
Jose Aires Ventura (M/D) Advisor *
Poliane Alfenas Zerbini External Examiner *
Ricardo Pinto Schuenck Internal Examiner *

Summary: Pineapple mealybug wilt associated virus species (PMWaV) in association with
Dysmicoccus brevipes cause mealybug wilt of pineapple. Virus dissemination
occurs primarily through infected and asymptomatic propagative material. In
this work, we validated a molecular methodology by RT-qPCR, to detect of the
PMWaV-1, PMWaV-2 and PMWaV-3 virus in asymptomatic and symptomatic
plants. Leaves and tissue culture plantlets from pineapple cultivars Smooth
Cayenne, Pérola and Vitória were analysed by conventional RT-PCR and
reverse transcription quantitative real-time (qRT-PCR). Mealybugs D. brevipes
obtained from symptomatic wilt plants were analyzed. A specific genes for each
virus of PMWaV was found in response to infection and was not expressed in
healthy plant. In silico assessment four housekeeping were selected for
accurate gene expression analysis in pineapple during biotic stress by RT-
qPCR. PMWaV-3 virus was prevalent in symptomatic and asymptomatic
plants, and associated with PMWaV-2 virus in the symptomatic wilt plants.
PMWaV-1 virus did not exhibited direct relation with the disease. The nutritional
status of plants influenced the onset of symtoms and presence of PMWaV-2
virus. The PMWaV-1, PMWaV-2 and PMWaV-3 virus were detected in
mealybugs colleted from diseased plants. Our results demonstrated
transmission of the PMWaV-2 and PMWaV-3 by mealybugs. However, D.
brevipes was not a good vector of PMWaV-1. This research provided a method
for indexing plants for production or certification healthy propagative material.
This prevent the spread of virus by propagation material to new crops areas and
an important tool for epidemiological study of the mealybug wilt of pineapple.
These is the first validation of an accurately method to detect three PMWaVs
virus in asymptomatic and symptomatic plants.
Keywords: Ananas comosus var. comosus, propagative material, qRT-PCR,
housekeeping, Dysmicoccus. Brevipes.

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