Autor(es):
Roriz, Mariana 
; Santos, Carla ; Vasconcelos, Marta W.
Data: 2011
Identificador Persistente: http://hdl.handle.net/10400.14/7640
Origem: Veritati - Repositório Institucional da Universidade Católica Portuguesa
Assunto(s): Bacteria; Bursaphelenchus xylophilus; Nematode; Pine wilt disease; Pinus pinaster; Symptoms
Descrição
For a long time it was thought that Bursaphelenchus xylophilus was the only agent of the pine wilt disease.
Recently, it was discovered that there are bacteria associated with the nematodes that contribute to the
pathogenesis of this disease, mainly through the release of toxins that promote the death of the pines.
Among the species most commonly found, are bacteria belonging to the Bacillus, Pantoea, Pseudomonas
and Xanthomonas genera.
The main objective of this work was to study the effect of inoculation of maritime pine (Pinus pinaster)
with four different nematode isolates, in the bacterial population of nematodes and trees, at different
stages of disease progression. The monitoring of progression of disease symptoms was also recorded.
Also, the identification of bacteria isolated from the xylem of trees and the surface of nematodes was performed
by classical identification methods, by the API20E identification system and by sequencing of bacterial
DNA.
The results showed that for the symptoms progression, the most striking difference was observed for
the pines inoculated with the avirulent isolate, C14-5, which led to a slower and less severe aggravation
of symptoms than in pines inoculated with the virulent isolates. In general, it was found that bacterial
population, inside the tree, increased with disease progression. A superior bacterial quantity was isolated
from pines inoculated with the nematode isolates HF and 20, and, comparatively, few bacteria were
isolated from pines inoculated with the avirulent isolate. The identification system API20E was insufficient
in the identification of bacterial species; Enterobacter cloacae species was identified in 79% of the
isolated bacterial colonies and seven of these colonies could not be identified by this method. Molecular
identification methods, through bacterial DNA sequencing, allowed a more reliable identification: eleven
different bacterial species within the Bacillus, Citrobacter, Enterobacter, Escherichia, Klebsiella, Paenibacillus,
Bacillus, Citrobacter, Enterobacter, Escherichia, Klebsiella, Paenibacillus, Pantoea and Terribacillus genera
were identified. General bacterial diversity increased with the progression of the disease. Bacillus spp.
were predominant at the earlier stage of disease progression and Klebsiella oxytoca at the later stages. Furthermore,
bacterial species isolated from the surface of nematodes were similar to those isolated from the
xylem of pines.
In the present work new bacterial species were identified which have never been reported before in
this type of study and may be associated with their geographical origin (Portugal). P. pinaster, the pine
species used in this study, was different from those commonly grown in Japan and China. Furthermore,
it was the first time that bacteria were isolated and identified from an avirulent pine wood nematode
isolate.
