Introduction: A serological survey carried out between 2005 and 2008 by the “Istituto
Zooprofilattico Sperimentale” of Sardinia; showed that CAEV infection is spread throughout
the regional territory with a prevalence of positive farms around 80% and a seropositivity
of more than 70% of tested animals. The CAE is a serious threat to
goat-sheep heritage and is likely to compromise an economy that is a fundamental
source of income for breeders, especially in those areas considered "agronomically"
difficult. It can be inferred therefore that the disease will become more important in
the coming years than it has been in the past. Currently, there are no vaccines for the
control of the infection, thus the only prophylaxis that can be implemented is hygienic-
sanitary. Prevention measures are based on the removal of healthy infected heads.
In this context, laboratory testing for CAEV-positive samples plays a crucial role in
the entire diagnostic procedure. The aim of the work was to evaluate a molecular system
based on real-time PCR amplification on relatively gag gene sequences in the
SRLV, as already indicated by several authors [1,2].
Methods: A total of 50 blood samples from farms in southern Sardinia with a clinical
diagnosis for CAEV infections have been anlyzed and proviral DNA was obtained from
buffy-coat by GeneProof Pathogen Kit (Brno-Czech Republic), 2 μl of DNA extract
was used in a real-time PCR reaction by using the kit SYBR Premix (TaKara-Clontech
®) with a LightCycler II Roche® apparatus following the manufacture istructions
[3]. The PCR target region was identified by multiple sequence alignment program
Geneious Biomatters Ltd., on gag gene sequences available in the DNA data Bank
(GenBank). Melting curve analysis showed two different peaks after real time PCR
reaction. Positive samples showed a Tm peak ranged from 82 °C to 88 °C, while the
negative sample was identified for a unique peak positioned from 73 °C to 77 °C, Figure
1. These samples resulted 90% CAEV positives and the analysis of the melting curve revealed several multiple-infection (different CAEV variants in the same subject),
in this case the most detected sequences are referenced to genotypes B3 and E
(subtype Volterra, Fonni, Roccaverano, Seui) GenBank accession n. JF502417,
JF502416, EU293537, GQ381130
Conclusions: The presented method/approach could be suitable for a rapid and complete
laboratory diagnosis of CAEV infection