Canine antibodies against salivary recombinant proteins of Phlebotomus perniciosus:
a longitudinal study in an endemic focus of canine leishmaniasis
in south Italy
Tatiana Kostalova1, Tereza Lestinova1, Petra Sumova1, Michaela Vlkova1, Iva Rohousova1, Eduardo
Berriatua2, Gaetano Oliva3, Eleonora Fiorentino4, Aldo Scalone4, Marina Gramiccia4, Luigi Gradoni4,
Petr Volf1
1Department
of Parasitology, Faculty of Science, Charles University in Prague, Czech Republic 2Animal Health Department, Universidad de Murcia, Spain
3Department of Veterinary Medicine and Animal Production, University Federico II, Naples, Italy 4Unit of Vector-borne Diseases and International Health,
Istituto Superiore di Sanità, Rome, Italy
Introduction
Results
Canine leishmaniasis (CanL) is a widespread zoonosis
caused by the protozoan parasite Leishmania infantum.
CanL is endemic in more than 70 countries including Italy,
where the main vector is Phlebotomus perniciosus.
During blood feeding sand flies deposit into the host skin
immunogenic salivary proteins which elicit a specific
antibody response. These anti-saliva antibodies enable to
estimate the host exposure to sand flies and also the risk
for Leishmania infections. However, the use of whole
salivary antigens has several limitations and therefore
recombinant salivary proteins have been produced and
tested as a tool to replace whole salivary gland lysates. In
this work, we selected two most antigenic P. perniciosus
salivary proteins, 43 kDa yellow protein (rSP03B) and 35.5
kDa apyrase (rSP01) and used them in a longitudinal field
study on dogs naturally exposed to
P. perniciosus.
Methods
 Beagle dogs naturally exposed to P. perniciosus over
two years in a site endemic for CanL (Naples area, Italy)
 canine sera were tested:
1) by ELISA for the IgG antibodies against
salivary gland lysate, rSP03B, rSP03B+rSP01
2) for L. infantum positivity by serology (IFAT),
culture, PCR
Study design
test for
anti-salivary
proteins
antibodies
L. infantum
positivity
year
season without
sand fly activity
pre-immune sand fly season
July
August, September,
October
2nd
-
July, August,
September
1st
2nd
3rd
July
1st
December
January, March
July
March
March
Dynamics of L. infantum infection status in dogs
 at the beginning of the study all
dogs were L. infantum negative
to all tests
 by the end of the study, the
percentage of subpatently and
actively infected dogs sharply
increase
Dynamics of IgG antibody response against P. perniciosus salivary
proteins in dogs
 antibodies against salivary
gland lysate correlates better
with antibodies recognizing
rSP03B (r=0.77) than antibodies
against salivary gland lysate
with antibodies recognizing
rSP03B+rSP01 (r=0.65)
* represents significant change in the median compared to previous sampling
 kinetics of anti-saliva and antirSP03B
IgG
antibodies
developed with similar pattern
and were clearly seasonal
Multivariable relationship between IgG against P. perniciosus saliva
and L. infantum infection
 multilevel models showed strong association between the level of
antibodies against salivary gland lysate and positivity of dogs for active
CanL infection (p<0.005)
 association between active infection and antibodies against rSP03B
and combination of rSP03B+rSP01 did not reach statistical significance
Conclusions
 fluctuation of antibody response against salivary proteins in naturally
exposed dogs was related to the period of activity of vector
Naples area
 our results suggest that P. perniciosus rSP03B protein is a valid
alternative to salivary gland lysate and could be used in large-scale
serological studies
 this novel method could be a practical and economically-sound tool to
detect host exposure to sand fly
Leishmaniasis in a Beagle dog
(Foglia Manzillo et al., 2013)
Distribution of communes with cases of
autochthonous CanL, Italy, 2005-2012
(modified according Gramiccia et al., 2013)
This study was partially funded by Charles University (GAUK – 1642314/2014), by the EU grant FP7-261504 EDENext and
by the EurNegVec COST Action TD1303.
Correspondence: Tatiana Kostalova, Laboratory of Vector Biology, Department of Parasitology, Faculty of Science, Charles
University in Prague, Czech Republic, e-mail: [email protected]