Species diversity of dermal microfilariae of the genus Cercopithifilaria infesting dogs in the Mediterranean region

Abstract

Following the recent description of microfilariae of a Cercopithifilaria sp. in a dog from Sicily, Italy, (herein after referred to as Cercopithifilaria sp. I), numerous skin samples were collected from dogs in the Mediterranean region. In addition to Cercopithifilaria sp. I (185·7 ± 7·2 μm long), microfilariae of 2 other species were identified, namely Cercopithifilaria grassii (651·7 ± 23·6 μm long) and a yet undescribed microfilaria, Cercopithifilaria sp. II (264·4 ± 20·2 μm long, with evident lateral alae). The morphological differentiation among the 3 species of dermal microfilariae was confirmed by differences in cytochrome c oxidase subunit 1 and ribosomal 12S sequences examined (mean level of interspecific pairwise distance of 11·4%, and 17·7%, respectively). Phylogenetic analyses were concordant in clustering these with other sequences of Cercopithifilaria spp. to the exclusion of Dirofilaria spp., Onchocerca spp. and Acanthocheilonema spp. Dermal microfilariae collected (n = 132) were morphologically identified as Cercopithifilaria sp. I (n = 108, 81·8%), Cercopithifilaria sp. II (n = 17, 12·9%), whereas only 7 (5·3%) were identified as C. grassii. Mixed infestations were detected in all sites examined. The great diversity of these neglected filarioids in dogs is of biological interest, considering the complex interactions occurring among hosts, ticks and Cercopithifilaria spp. in different environments.

Diversity of Cercopithifilaria species in dogs from Portugal

“…II sensu Otranto et al , 2013 have been morphologically and molecularly differentiated in canine populations in Europe [7,9-11]. Microfilariae of these three species can be distinguished from each other by their length and general morphological features and also by the inter-specific genetic distance of the mitochondrial cytochrome c oxidase subunit I ( cox 1) and 12S rRNA genes [6,10]. …”

“…Skin snips of around 0.01 cm 3 (0.2 × 0.2 × 0.2 cm) were collected from the forehead region using a disposable scalpel and soaked in saline solution (pH 7.4) for 12 h in an incubator at 30°C. Dermal microfilariae were searched for in 20 μl sediment placed on a glass slide, using a light microscope (magnifications of ×40 and ×100), and identified according to their morphological features, as previously described [6,10]. Microfilariae were photographed with a digital camera mounted on the microscope (DP20, Olympus, Japan) and measurements (in micrometers) taken with the Live Cell Imaging software (Olympus).…”

“…Following morphological analysis of microfilariae, sediments scoring positive were stored in 70% ethanol and species identification was subsequently assessed by PCR amplification and DNA sequencing [6,10]. Microfilariae were isolated and genomic DNA extracted from individual specimens using a commercial kit (DNeasy Blood & Tissue Kit, Qiagen, Germany) in accordance with the manufacturer’s instructions.…”

“…Infections with C. bainae and Cercopithifilaria sp. II were detected in dogs from the Italian regions of Basilicata [10] and Sardinia [15], whereas C. bainae and C. grassii were detected in Sicily [10]. Sympatric C. bainae , C. grassi and Cercopithifilaria sp.…”

Diagnostic challenges and the unwritten stories of dog and cat parasites

“…Nowadays, scientists are aware that dogs may be infected by at least three species of Cercopithifilaria, namely C. grassii, Cercopithifilaria sp. II sensu Otranto et al (2012a) (a formal taxonomic description of adults for this species is lacking) and C. bainae (Otranto et al, 2013b). In particular, C. bainae is prevalent in canine and tick populations from the Mediterranean (i.e., Spain, Greece and southern Italy) (Otranto et al, 2013b), as well as from Australia, Brazil, Malaysia and South Africa , and overlaps the distribution of its main arthropod vector, the brown dog tick Rhipicephalus sanguineus sensu lato Ramos et al, 2013Ramos et al, , 2014.…”