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Molecular Pathogens
MP2011, Vol.2, No.3
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Research Article Open Access
Comparison Between Serological And Molecular Detection of Citrus Canker
Pathogen (
Xanthomonas axonopodis
pv.
citri
)
Pongpan Songwattana
1
, Mariena Ketudat-Cairns
1,2
1. School of Biotechnology, Institute of Agricultural Technology, 111 University Avenue, Nakhon Ratchasima, 30000, Thailand
2. Embryo and Stem cell Research Center, Suranaree University of Technology (SUT), 111 University Avenue, Nakhon Ratchasima, 30000, Thailand
Corresponding author; ketudat@sut.ac.th;
Authors
Molecular Pathogens 2011, Vol 2 No 3 DOI: 10.5376/mp.2011.02.0003
Received: 25 May, 2011
Accepted: 21 Jun., 2011
Published: 01 Jul., 2011
This is an open access article published under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and
reproduction in any medium, provided the original work is properly cited.
Preferred citation for this article as:
Songwattana and Ketudat-Cairns, 2011, Comparison between serological and molecular detection of citrus canker pathogen (
Xanthomonas axonopodis
pv.
citri
),
Molecular Pathogens, Vol.2 No.3 (doi: 10.5376/mp.2011.02.0003)
Abstract
The specificity and sensitivity of serological and molecular tools for the detection of citrus canker pathogen
(
Xanthomonas axonopodis
pv.
citri
;
Xac
) were investigated and compared. Virulence
Xac
BP210 was used as antigen for antisera
production. The sensitivity of 1:2,000 diluted antisera were at 10
6
CFU/mL for live cell and 10
5
CFU/mL for dead cells. The
cross-reaction of the antisera was observed only with
X. campestris
pv.
vesicatoria
but not other
Xanthomonas
nor other unrelated
bacteria tested. Molecular tool was performed using specific primer to the
rpf
gene on
Xac
. The PCR amplification indicated that, all
Xac
isolates amplified a product of 581 bp which was not seen in other
Xanthomonas
sp.
and unrelated bacteria tested. The
sensitivity of these specific primers was down to at least 1 cell which was effective to detect the pathogen in both infected
symptomatic and asymptomatic lime tissues. The serological tool was able to detect the pathogen only on infected leaves of day 4
post inoculation when the symptoms were already detected by eye. The serological tool can be used to detect and quantify the present
of
Xac
to study the disease development on symptomatic tissues.
Keywords
Serological tool; Citrus canker; Antiserum; Pathogen detection
Background
Citrus canker is one of the serious diseases in citrus
and some citrus relatives. This disease has been
caused by
Xanthomonas axonopodis
pv.
citri
(
Xac
)
bacteria or
Xac
pathotype A (
Xac
-A) which is the most
wide spread pathogen in Asia and other citrus growing
areas (Verniere et al., 1998). Most commercial citrus
including Mexican lime, grapefruits and commercial
lime in Thailand (Pan lime) are susceptible host for
Xac
. This pathogen can infect leaves, twigs and fruits
through stomata and wounds (Graham et. al., 2004).
Moreover, this disease directly reduce fruit quality and
quantity of yield by defoliation, blemished fruit,
premature fruit drop, die-back of twigs and general
debilitation of the tree (Das, 2003). Normally, the
pathogenic bacteria can survive from one crop season
to the next as latent infection in propagation organs or
epiphytic populations on plant surfaces. The
equipments for harvesting are also the cause of disease
spread (Graham et al., 2000). Therefore, the method
for their early detection is necessary. Currently, many
methods have been developed and are available in
laboratories. Polymerase chain reaction (PCR) method
is a high sensitive detection method (Cubero et al.,
2001) but, it requires laboratory equipments and
specialized training. In contrast, immunodiagnostic
test kit is an easy method to use at the site where
disease is suspected. It does not require neither special
equipment nor training to perform the detection. The
immunodiagnostic test is based on the ability of an
antibody to recognize and bind to a specific antigen, a
substance associated with the plant pathogen. This
method is known as enzyme-linked immunosorbent
assay (ELISA). In this study the specificity and
sensitivity of serological and molecular tools for the
detection of citrus canker pathogen (
X. axonopodis
pv.
citri
) were investigated and compared.
1 Results and discussions
1.1 Bacteria isolation and pathogenicity test
Xanthomonas axonopodis
pv.
citri
(
Xac
) was isolated