Innovative tools for the diagnosis of Echinococcus granulosus
in definitive hosts
Maria Paola Maurelli
Received: 25 February 2018 /Accepted: 25 May 2018
Springer-Verlag GmbH Germany, part of Springer Nature 2018
The aim of this study was to develop and validate an innovative protocol for the diagnosis of Echinococcus granulosus and other
Taeniidae in dogs. For this purpose, three experiments were performed, using faecal samples from naturally infected dogs. Firstly,
the FLOTAC technique was calibrated with five flotation solutions: saturated sodium chloride (specific gravity, s.g. = 1.20), zinc
sulphate (s.g. = 1.35), zinc chloride (s.g. = 1.45), Breza (s.g. = 1.30) and modified Breza (s.g. = 1.40). Then, FLOTAC was
compared with four techniques of flotation in centrifuge using: zinc sulphate (s.g. = 1.20), Breza (s.g. = 1.30), modified Breza
(s.g. = 1.40), and zinc chloride (s.g. = 1.45). Finally, four different protocols of DNA extraction were compared for Taeniidae
identification: QIAamp Tissue Kit and QIAamp Stool from eggs; QIAamp Stool and Wizard Magnetic Purification System for
Food from faeces. FLOTAC with zinc sulphate was the most efficient method to detect Taeniidae eggs, showing highest mean of
eggs per gram (EPG) of faeces. The QIAamp Stool, using eggs concentrated by FLOTAC, was the best method for DNA
extraction. The combination of these protocols provided the highest number of positive samples with PCR, i.e., 47/50
(94.0%). The three negative samples showed a low faecal egg count (2 EPG) below the detection limit (4 EPG) of the protocol.
From sequencing of the 47 positive samples: 6 samples were identified as E. granulosus sensu stricto (s.s.), 28 as Taenia
hydatigena and6asT. pisiformis. A co-infection between different genera of Taeniidae was found in 7 samples.
Keywords Echinococcus granulosus
Echinococcosis is a zoonosis, caused by the small cestodes of
the genus Echinococcus. Echinococcus granulosus is the spe-
cie most widespread and of major public health importance
especially in the Mediterranean area (reviewed in Deplazes et
al. 2017). Its lifecycle involves carnivorous (mainly dogs) as
definitive hosts and a broad spectrum of mammalians, includ-
ing humans, as intermediate hosts (McManus et al. 2003).
Dogs become infected feeding the offal of intermediate hosts
(especially sheep) with hydatid cysts and expel out the eggs
with faeces. When the intermediate hosts eat the eggs, the
oncospheres inside penetrate the mucosae of the intestine
and through the blood and lymphatic system reach the target
organs (e.g., liver and lungs) where the larval stage
(metacestode) develops and causes cystic echinococcosis
(CE) (Craig et al. 2007).
In Europe, dogs and sheep play a key role in the transmis-
sion of CE. Therefore, monitoring and surveillance of echino-
coccosis in dogs is useful for epidemiological studies and
control programs. It can also indicate the potential risk to
humans of being infected with hydatid disease (Huang et al.
2007; Craig et al. 2017). However, ante-mortem diagnosis of
infection with E. granulosus in definitive hosts is difficult,
because the eggs of all genera of Taeniidae cannot be differ-
entiated morphologically by light microscopy (Carmena and
Cardona 2013) and the characteristic small segments of E.
granulosus may be absent from the faeces or can be easily
overlooked (Eckert et al. 2001). Moreover, even with a high
parasite burden, echinococcosis in definitive hosts is asymp-
tomatic (Carmena and Cardona 2013).
Conventionally, the evaluation of echinococcosis preva-
lence in dogs was based on purging them with arecoline
* Maria Paola Maurelli
Department of Veterinary Medicine and Animal Production,
University of Naples Federico II, CREMOPAR Campania Region,