INVESTIGADORES
FARBER Marisa Diana
congresos y reuniones científicas
Título:
Molecular diagnostic tools for simultaneous detection of Babesia bovis, Babesia bigemina and Anaplasma marginale.
Autor/es:
DE LA FOURNIÈRE S; PAOLETTA M; GUILLEMI E; LOPEZ ARIAS L; WILKOWSKY, S; FARBER MD
Lugar:
Ciudad del Cabo
Reunión:
Congreso; 12th Biennial Conference of the Society for Tropical Veterinary Medicine (STVM) and the VIII International Conference on Ticks and Tick-borne Pathogens (TTP-8); 2014
Institución organizadora:
STVM
Resumen:
Bovine Babesiosis and Anaplasmosis are
tick-borne diseases that impose important constraints for livestock industry in
tropical and subtropical areas. In endemic regions, Babesia bovis, B. bigemina
and Anaplasma marginale are often
found together within a single bovine host. Given the importance of
simultaneous detection for epidemiological evaluation and the detection and
quantification during outbreaks, we developed a multiplex PCR (mPCR) and a
quantitative Real Time PCR (qPCR) assays respectively.
The mPCR was rationally designed targeting
specific multi-copy genes for each microorganism (VESA, Rap-1 and XXX); the
products generated were easily detected in a 2% agarose gel due to their
different sizes. The assay sensitivity curve tested on artificial mixtures of reference
DNA samples was equivalent to the single PCR for each pathogenNevertheless a
reduced sensitivity was observed when the mPCR was tested on field samples.
In addition, a qPCR involving the use of SYBR
Green/ROX was developed to easily quantify field samples and compared with
previous protocols for the three hemoparasites. Particularly, for B.a bovis and B. bigemina, we demonstrated
that the already reported qPCR based on mitochondrial cytochrome B (cytb) was not enough reproducible, as cytb copy number varies between samples
and also in the same sample within a period of time. Thus, we set up a new qPCR
targeting single copy genes with equivalent sensitivity to the described cytb qPCR.
The ongoing development of these methodologies
would assist to the rapid and specific detection of a large number of coinfected
field samples from endemic areas where these parasites are prevalent