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1.  Native strains of Beauveria bassiana for the control of Rhipicephalus sanguineus sensu lato 
Parasites & Vectors  2015;8:80.
Background
Rhipicephalus sanguineus sensu lato ticks are widespread worldwide due to their adaptability to survive under different environmental conditions. They may act as vectors of a wide range of pathogens to humans and animals and their control is based on the use of chemical products on dogs and in the environment. Alternative control strategies, such as the use of entomopathogenic fungi as bio-control agents have also been investigated. The ability of native strains of Beauveria bassiana sensu lato in causing mortality in different tick species (e.g., Amblyomma cajennense and Rhipicephalus microplus) has been demonstrated. However, limited studies have assessed the use of B. bassiana for the control of R. sanguineus s.l. and none of them have employed native strains of this fungus. Here we investigated the pathogenicity of a native strain of B. bassiana (CD1123) against all developmental stages of R. sanguineus s.l..
Methods
Batches of eggs, larvae, nymphs and adult ticks were immersed in a suspension of 107 conidia/ml of B. bassiana s.l., isolated from a R. sanguineus s.l. engorged female. All treatment and control groups were observed for 20 days, and the biological parameters (i.e., mortality, hatching, moulting percentage, pre-oviposition period, oviposition period and rate, eggs production efficiency, reproductive efficiency and fitness indexes) were assessed.
Results
The effect of the B. bassiana strain tested herein on eggs, larvae, nymphs and adults showed a significantly higher mortality than those of the control groups (p < 0.05) at 5 days post-infection. No infected eggs hatched and no infected larvae moulted. Only 15% of infected nymphs moulted into adults. All biological parameters of treated groups differed significantly (p < 0.001) from those of control groups.
Conclusions
This study demonstrates that a suspension containing 107 conidia/ml of a native B. bassiana strain is highly virulent towards all life-cycle developmental stages of R. sanguineus s.l. and may be of potential interest as a biological control agent against these ticks.
doi:10.1186/s13071-015-0693-9
PMCID: PMC4324834  PMID: 25651851
Beauveria bassiana; Biological control; Rhipicephalus sanguineus s.l.; Entomopathogenic fungus; Vector borne-diseases
2.  Angiostrongylus chabaudi Biocca, 1957: a new parasite for domestic cats? 
Parasites & Vectors  2014;7(1):588.
Background
Natural infection with a species of Angiostrongylus has been reported only once in wildcats from central Italy by Biocca in 1957. The causative species of this infection was identified as Angiostrongylus chabaudi. Following this report, this parasite had never been found in either wild or domestic cats.
Findings
The lungs and the pulmonary arteries of an adult female cat (Felis silvestris catus), road-killed in Sardinia, Italy, were macroscopically examined and dissected under a light microscope for the presence of parasites. A slender nematode was detected and its morphometrical features were consistent with those of A. chabaudi. Morphological data were supplemented by sequencing of the partial cytochrome oxidase c subunit 1 (cox1) gene, as well as the internal transcribed spacer 2 (ITS2) of the rDNA. Nucleotide sequences displayed 99% homology with the ITS2 sequence [GenBank KM216825.1] of a specimen of Angiostrongylus sp. recovered recently from the pulmonary artery of a wildcat in Germany and 91% with cox1 sequence [GenBank GU138118.1] of Angiostrongylus vasorum.
Conclusion
The results of the present study indicate, for the first time, that A. chabaudi may also infect domestic cats, and thus should be considered in the diagnosis of metastrongyloid species infecting their cardio-pulmonary system.
doi:10.1186/s13071-014-0588-1
PMCID: PMC4308007  PMID: 25515026
Angiostrongylus chabaudi; Metastrongyloidea; Angiostrongylidae; Cat; Cardio-pulmonary nematodes
3.  Evaluation of blood and bone marrow in selected canine vector-borne diseases 
Parasites & Vectors  2014;7(1):534.
Background
Bone marrow (BM) is a major hematopoietic organ that can harbour a variety of vector-borne pathogens; however, knowledge of BM pathological changes in dogs infected with vector-borne pathogens is limited. Thus, the aim of the present study was to assess the pathological changes in canine BM associated with natural infections by four vector-borne pathogens, as well as to determine the relationships between such changes and abnormalities of the peripheral blood.
Methods
Cytological disorders and pathological changes of the BM of 83 dogs naturally-infected with one or more of four vector-borne pathogens (i.e., Anaplasma platys, Leishmania infantum, Babesia vogeli and Hepatozoon canis) were evaluated and compared with the corresponding hematological findings.
Results
Dysgranulopoiesis and dysmegakaryocytopoiesis were the most frequently observed BM abnormalities in infected dogs. Erythroid suppression, and lymphocytic, monocytic and macrophage hyperplasia were also observed. Interestingly, associations between suppression and hyperplasia of specific cell lines in the marrow and corresponding changes in numbers of circulating peripheral blood cells were not observed.
Conclusions
Infections with one or more of the vector-borne pathogens examined in this study should be considered as differential diagnoses for secondary dysmyelopoiesis.
doi:10.1186/s13071-014-0534-2
PMCID: PMC4261574  PMID: 25441458
Bone marrow; Cytology; Vector-borne pathogens; Dysplasia; Secondary dysmyelopoiesis
4.  Anaplasma platys in Bone Marrow Megakaryocytes of Young Dogs 
Journal of Clinical Microbiology  2014;52(6):2231-2234.
Anaplasma platys is an obligate intracellular rickettsial pathogen that infects platelets of dogs, forming basophilic intracellular morulae. In the present report, cellular inclusions were documented in bone marrow thrombocyte precursors of two young naturally infected dogs, indicating that A. platys can infect megakaryocytes and promegakaryocytes.
doi:10.1128/JCM.00395-14
PMCID: PMC4042781  PMID: 24622106
5.  Drosophilidae feeding on animals and the inherent mystery of their parasitism 
Parasites & Vectors  2014;7(1):516.
Insect evolution, from a free to a parasitic lifestyle, took eons under the pressure of a plethora of ecological and environmental drivers in different habitats, resulting in varying degrees of interactions with their hosts. Most Drosophilidae are known to be adapted to feeding on substrates rich in bacteria, yeasts and other microfungi. Some of them, mainly those in the Steganinae subfamily, display a singular behaviour, feeding on animal tissues or secretions. This behaviour may represent an evolving tendency towards parasitism. Indeed, while the predatory attitude is typical for the larval stages of a great proportion of flies within this subfamily, adult males of the genera Amiota, Apsiphortica and Phortica display a clearly zoophilic attitude, feeding on the lachrymal secretions of living mammals (also referred as to lachryphagy). Ultimately, some of these lachryphagous species act as vectors and intermediate hosts for the spirurid nematode Thelazia callipaeda, which parasitizes the eyes of domestic and wild carnivores and also humans. Here we review the scientific information available and provide an opinion on the roots of their evolution towards the parasitic behaviour. The distribution of T. callipaeda and its host affiliation is also discussed and future trends in the study of the ecology of Steganinae are outlined.
Electronic supplementary material
The online version of this article (doi:10.1186/s13071-014-0516-4) contains supplementary material, which is available to authorized users.
doi:10.1186/s13071-014-0516-4
PMCID: PMC4243723  PMID: 25404259
Drosophilidae; Steganinae; zoophagy; parasitism; lachryphagy; Phortica variegata; Thelazia callipaeda
6.  Morphological keys for the identification of Italian phlebotomine sand flies (Diptera: Psychodidae: Phlebotominae) 
Parasites & Vectors  2014;7(1):479.
Background
Phlebotomine sand flies are small blood-feeding insects of great medical and veterinary significance. Their identification relies basically on the microscopic examination of key morphological characters. Therefore, identification keys are fundamental to any researcher dealing with these insects. The Italian fauna of phlebotomine sand flies consists of eight species (Phlebotomus perniciosus, Phlebotomus perfiliewi, Phlebotomus ariasi, Phlebotomus neglectus, Phlebotomus papatasi, Phlebotomus mascittii, Phlebotomus sergenti and Sergentomyia minuta), whose morphological delineation may be troublesome for non-taxonomists.
Methods
A total of 8,757 pictures were taken from the 419 selected phlebotomine sand fly specimens collected on different occasions. Twenty-eight characters for the males and 23 for the females were examined, resulting in a database containing over 10,000 entries. Representative phlebotomine sand fly specimens for each species available were selected and relevant characters were drawn with the aid of a camera lucida.
Results
After detailed morphological study of representative specimens, comprehensive identification keys based on key characters (e.g., pharynx and spermathecae of females and male terminalia) were elaborated.
Conclusions
The identification keys provided herein allow the identification of genera and species of phlebotomine sand flies of Italy and they will facilitate future studies on these medically important insects.
doi:10.1186/s13071-014-0479-5
PMCID: PMC4203899  PMID: 25323537
Sand flies; Taxonomy; Vectors; Leishmaniasis
7.  The spread of zoonotic Thelazia callipaeda in the Balkan area 
Parasites & Vectors  2014;7(1):352.
Background
Thelazia callipaeda (Spirurida, Thelaziidae), also known as “oriental eyeworm”, is a small nematode parasite that lives in the conjunctival sac of domestic and wild carnivores, rabbits and even humans, causing mild (e.g., conjunctivitis, epiphora, and ocular discharge) to severe (e.g., keratitis, and corneal ulcers) ocular disease. This study reports, for the first time, the occurrence of T. callipaeda infection in the Balkan regions (i.e., Bosnia and Herzegovina and Croatia), it provides genetic evidence on the origin of the infection in that area and discusses potential expansion pathways in the near future.
Methods
This survey was conducted in two Western Balkan countries, Bosnia and Herzegovina and Croatia. At necropsy, from January 2011 to April 2014, a total of 184 carcasses of red foxes were examined throughout the study area and worms were collected from the conjunctival sac. In the same period, worms were also collected during clinical examination from the conjunctival sac of four dogs and a cat from Bosnia and Herzegovina and two dogs from Croatia. All nematodes collected were morphologically identified and molecularly characterized by sequencing of partial cox1 gene.
Results
T. callipaeda was observed in 51 (27.71%) foxes and the highest prevalence (50.0%) was in the region of East Bosnia. Beside the 4 cases of hyperemia (7.84%), most of the infected animals had no signs of ocular infection (n = 47, 92.15%). A total of 417 adult nematodes collected (364 from foxes, 51 from dogs, 2 from cat) were morphologically and molecularly identified as T. callipaeda haplotype 1.
Conclusion
This is the first report of autochthonous cases of T. callipaeda infection in red foxes, dogs and cat in Bosnia and Herzegovina and Croatia and data presented here suggest that reports of thelaziosis in other Balkan areas are, as yet, not diagnosed most likely due to the lack of awareness of practitioners. In addition, data regarding the spread of the infection in Europe over the last ten years suggests that an increasing pattern in the distribution of this disease in domestic and wild animals should be expected in the future.
doi:10.1186/1756-3305-7-352
PMCID: PMC4124150  PMID: 25078286
Thelazia callipaeda; Red fox; Dog; Cat; Balkan; Bosnia and Herzegovina; Croatia; Balkan regions
8.  ABC transporters are involved in defense against permethrin insecticide in the malaria vector Anopheles stephensi 
Parasites & Vectors  2014;7(1):349.
Background
Proteins from the ABC family (ATP-binding cassette) represent the largest known group of efflux pumps, responsible for transporting specific molecules across lipid membranes in both prokaryotic and eukaryotic organisms. In arthropods they have been shown to play a role in insecticide defense/resistance. The presence of ABC transporters and their possible association with insecticide transport have not yet been investigated in the mosquito Anopheles stephensi, the major vector of human malaria in the Middle East and South Asian regions. Here we investigated the presence and role of ABCs in transport of permethrin insecticide in a susceptible strain of this mosquito species.
Methods
To identify ABC transporter genes we obtained a transcriptome from untreated larvae of An. stephensi and then compared it with the annotated transcriptome of Anopheles gambiae. To analyse the association between ABC transporters and permethrin we conducted bioassays with permethrin alone and in combination with an ABC inhibitor, and then we investigated expression profiles of the identified genes in larvae exposed to permethrin.
Results
Bioassays showed an increased mortality of mosquitoes when permethrin was used in combination with the ABC-transporter inhibitor. Genes for ABC transporters were detected in the transcriptome, and five were selected (AnstABCB2, AnstABCB3, AnstABCB4, AnstABCmember6 and AnstABCG4). An increased expression in one of them (AnstABCG4) was observed in larvae exposed to the LD50 dose of permethrin. Contrary to what was found in other insect species, no up-regulation was observed in the AnstABCB genes.
Conclusions
Our results show for the first time the involvement of ABC transporters in larval defense against permethrin in An. stephensi and, more in general, confirm the role of ABC transporters in insecticide defense. The differences observed with previous studies highlight the need of further research as, despite the growing number of studies on ABC transporters in insects, the heterogeneity of the results available at present does not allow us to infer general trends in ABC transporter-insecticide interactions.
Electronic supplementary material
The online version of this article (doi:10.1186/1756-3305-7-349) contains supplementary material, which is available to authorized users.
doi:10.1186/1756-3305-7-349
PMCID: PMC4124152  PMID: 25073980
Mosquitoes; Bioassays; Insecticide resistance; Culicidae; Vector control; ABC transporters
9.  Efficacy of a slow-release imidacloprid (10%)/flumethrin (4.5%) collar for the prevention of canine leishmaniosis 
Parasites & Vectors  2014;7:327.
Background
The efficacy of a slow-release insecticidal and repellent collar containing 10% imidacloprid and 4.5% flumethrin (Seresto, Bayer Animal Health) in preventing Leishmania infantum infection was evaluated in a large population of dogs living in a hyper-endemic area of Sicily (Italy).
Methods
A total of 219 dogs, negative for L. infantum were enrolled in a multicentre, controlled study. Dogs were divided into two homogeneous groups, defined as G1 (n = 102) and G2 (n = 117). Before the start of the sand fly season, dogs in G1 were treated with the collar while animals in G2 were left untreated, serving as negative controls. Dogs were serially sampled on day D90, D180, D210 and D300 in order to assess Leishmania infection by IFAT, PCR on skin (D210-D300) and bone marrow (D300) and cytology on bone marrow aspirate (D300).
Results
Three dogs (2.9%) in G1 and 41 (40.2%) in G2 became positive for L. infantum in at least one of the diagnostic tests employed in the study. The number of seropositive dogs in G2 increased in the course of the study from 15 (D90) to 41 (D300), with some of them also positive in other diagnostic tests. Eight (19.6%) of the seropositive dogs in G2 showed an increase in antibody titers ranging from 1:160 to 1:1,280. At the last follow-up, some of dogs in G2 displayed overt clinical signs suggestive of leishmaniosis. The mean incidence density rate at the final follow-up was 4.0% for G1 and 60.7% for G2, leading to a mean efficacy of the collar in protecting dogs at both sites of 93.4%.
Conclusions
The slow-release collar tested in this study was shown to be safe and highly effective in preventing L. infantum infection in a large population of dogs. Protection conferred by a single collar (up to eight months) spanned an entire sand fly season in a hyper-endemic area of southern Italy. The regular use of collars, at least during the sand fly season, may represent a reliable and sustainable strategy for the prevention of leishmaniosis in dogs living in or travelling to an endemic area.
doi:10.1186/1756-3305-7-327
PMCID: PMC4105111  PMID: 25023573
Leishmania infantum; Dog; Canine leishmaniosis; Prevention; Control
10.  Ticks infesting humans in Italy and associated pathogens 
Parasites & Vectors  2014;7:328.
Background
Ticks may transmit a large variety of pathogens, which cause illnesses in animals and humans, commonly referred to as to tick-borne diseases (TBDs). The incidence of human TBDs in Italy is underestimated because of poor surveillance and the scant amount of studies available.
Methods
Samples (n = 561) were collected from humans in four main geographical areas of Italy (i.e., northwestern, northeastern, southern Italy, and Sicily), which represent a variety of environments. After being morphologically identified, ticks were molecularly tested with selected protocols for the presence of pathogens of the genera Rickettsia, Babesia, Theileria, Candidatus Neoehrlichia mikurensis, Borrelia and Anaplasma.
Results
Ticks belonged to 16 species of the genera Argas, Dermacentor, Haemaphysalis, Hyalomma, Ixodes and Rhipicephalus, with Ixodes ricinus (59.5%) being the species most frequently retrieved, followed by Rhipicephalus sanguineus sensu lato (21.4%). Nymphs were the life stage most frequently retrieved (41%), followed by adult females (34.6%). The overall positivity to any pathogen detected was 18%. Detected microorganisms were Rickettsia spp. (17.0%), Anaplasma phagocytophilum (0.8%), Borrelia afzelii (0.5%), Borrelia valaisiana (0.3%), C. N. mikurensis (0.5%) and Babesia venatorum (0.6%).
Conclusions
Results indicate that people living in the Italian peninsula are at risk of being bitten by different tick species, which may transmit a plethora of TBD causing pathogens and that co-infections may also occur.
doi:10.1186/1756-3305-7-328
PMCID: PMC4223688  PMID: 25023709
Ticks; Pathogens; Humans; Tick-borne diseases; Italy; Distribution
11.  Parasites of domestic owned cats in Europe: co-infestations and risk factors 
Parasites & Vectors  2014;7:291.
Background
Domestic cats can be infested by a large range of parasite species. Parasitic infestations may cause very different clinical signs. Endoparasites and ectoparasites are rarely explored in the same study and therefore multiparasitism is poorly documented. The present survey aimed to improve knowledge of the prevalence and risk factors associated with ecto- and endoparasite infestations in owned cats in Europe.
Methods
From March 2012 to May 2013, 1519 owned cats were included in a multicenter study conducted in 9 veterinary faculties throughout Europe (Austria, Belgium, France, Hungary, Italy, Romania and Spain). For each cat, ectoparasites were checked by combing of the coat surface associated with otoscopic evaluation and microscopy on cerumen samples. Endoparasites were identified by standard coproscopical examinations performed on fresh faecal samples. Risk factors and their influence on parasitism were evaluated by univariate analysis followed by a multivariate statistical analysis (including center of examination, age, outdoor access, multipet status, and frequency of treatments as main criteria) with logistic regression models.
Results
Overall, 50.7% of cats resulted positive for at least one internal or one external parasite species. Ectoparasites were found in 29.6% of cats (CI95 27.3-32.0%). Otodectes cynotis was the most frequently identified species (17.4%), followed by fleas (15.5%). Endoparasites were identified in 35.1% of the cats (CI95 32.7-35.7%), including gastro-intestinal helminths in 25.7% (CI95 23.5-28.0), respiratory nematodes in 5.5% (CI95 4.2-7.0%) and protozoans in 13.5% (CI95 11.8-15.3%). Toxocara cati was the most commonly diagnosed endoparasite (19.7%, CI95 17.8-21.8%). Co-infestation with endoparasites and ectoparasites was found in 14.0% of the cats, and 11.9% harbored both ectoparasites and gastro-intestinal helminths.
Age, outdoor access, living with other pets, and anthelmintic or insecticide treatments were significantly associated with the prevalence of various parasites.
Conclusions
This survey demonstrates that parasitism is not a rare event in European owned cat populations. The prevalence of multi-parasitism is significantly greater than expected by chance and hence there is tendency for some individual cats to be more prone to infestation by both endo- and ectoparasites due to common risk factors.
doi:10.1186/1756-3305-7-291
PMCID: PMC4082530  PMID: 24965063
Cats; Ectoparasites; Endoparasites; Toxocara cati; Ctenocephalides; Otodectes; Europe; Risk factors; Treatments
12.  Diversity of Cercopithifilaria species in dogs from Portugal 
Parasites & Vectors  2014;7:261.
Background
Filarioids belonging to the genus Cercopithifilaria (Spirurida: Onchocercidae) have been described in dogs in association with Rhipicephalus sanguineus group ticks, which act as their biological vectors. This study represents the first investigation on Cercopithifilaria spp. in dogs from Portugal.
Findings
Dogs (n = 102) from the Algarve region (south of Portugal) were sampled by skin snip collection and tissues were left to soak overnight in saline solution. Sediments were observed under a light microscope and the detected microfilariae identified according to their morphology. Twenty-four dogs (23.5%) were found infected with at least one species of Cercopithifilaria, namely C. bainae (9.8%), C. grassii (3.9%) and Cercopithifilaria sp. II sensu Otranto et al., 2013 (13.7%). Results were confirmed by molecular amplification of partial cytochrome c oxidase subunit I and 12S rRNA genes and sequence analysis. Co-infections with more than one Cercopithifilaria species were detected in 3.9% of the animals.
Conclusions
This is the first report of Cercopithifilaria spp. in dogs from Portugal. The estimated level of infection with C. bainae, C. grassii and Cercopithifilaria sp. II suggests that these filarioids are prevalent in the canine population of southern Portugal.
doi:10.1186/1756-3305-7-261
PMCID: PMC4051959  PMID: 24898125
Cercopithifilaria bainae; Cercopithifilaria grassii; Cercopithifilaria sp. II; Dogs; Portugal
13.  Incidence of Cercopithifilaria bainae in Dogs and Probability of Co-Infection with Other Tick-Borne Pathogens 
PLoS ONE  2014;9(2):e88198.
Background
Cercopithifilaria bainae is a filarioid parasite that infects dogs, being transmitted by Rhipicephalus sanguineus group ticks in many countries of the Mediterranean basin. This study assessed the incidence density rate (IDR) of infection by C. bainae in dogs and the probability of co-infection with other tick-borne pathogens (i.e., Anaplasma platys, Babesia vogeli and Hepatozoon canis), in an area of high endemicity in southern Italy.
Methodology/Principal Findings
From March 2011 to October 2012, a field study involving 58 young dogs naturally exposed to tick infestation was conducted. Skin and blood samples obtained from each dog six times during an 18-month period were tested for C. bainae by parasite detection within skin snip sediments, with subsequent confirmation through PCR and DNA sequencing. Dogs examined monthly for ticks and A. platys, B. vogeli and H. canis were microscopically and/or molecularly diagnosed and after the first and the second summer seasons, the IDR for positive animal-month at risk was 3.8% and 1.7% in November 2011 and October 2012, respectively. All 58 C. bainae-infected dogs were simultaneously infected with at least one other tick-borne pathogen. After the first summer season (assessment in November 2011), a C. bainae-infected dog had a 33% probability of being infected with H. canis or A. platys, whereas after the second tick season (assessment in October 2012) the probability of co-infection was 78%, 22% and 11% for H. canis, A. platys and B. vogeli, respectively.
Conclusions
Our data indicate that tick-infested dogs are at risk of acquiring infection by C. bainae. In addition, the detection of C. bainae microfilariae indicates a prior tick exposure and, should stimulate testing for other tick-borne disease causing pathogens.
doi:10.1371/journal.pone.0088198
PMCID: PMC3912174  PMID: 24498437
14.  The enigma of the dog mummy from Ancient Egypt and the origin of ‘Rhipicephalus sanguineus’ 
Background
Ticks belonging to the Rhipicephalus sanguineus group are amongst the most important vectors of pathogenic microorganisms to dogs and humans. However, the taxonomy of this species group is still the subject of debate, especially because there is no type specimen or reliable morphological description for Rhipicephalus sanguineus sensu stricto. Recently, a comprehensive morphological and genetic study on representative tick specimens from Europe, Africa, Americas, and Oceania, revealed the existence of at least four morphologically and genetically distinct species under the name ‘R. sanguineus’ infesting dogs from different countries.
Methods
Herein, we examined morphologically tick specimens retrieved on a dog mummy from Ancient Egypt (ca. 1st century – 4th century A.D.). The dog mummy and associated ticks were found during an archaeological expedition conducted in El Deir.
Results
Scanning electron micrographs allowed us to assess their identity as belonging to the R. sanguineus group. In addition on the basis of the scutal punctation pattern, spiracular plates, width of dorsal tail of spiracular plates relative to the adjacent festoon, female genital aperture, male adanal plates and accessory shields, these ticks were tentatively identified as Rhipicephalus sp. II (=temperate species).
Conclusions
It can be concluded that R. sanguineus group ticks have infested dogs living in the Mediterranean region since ancient times. This finding represents the oldest record of ticks on any animal species and adds a new piece in the complex puzzle regarding tick parasitism on dogs and humans and their role as vectors of pathogens.
doi:10.1186/1756-3305-7-2
PMCID: PMC3896854  PMID: 24438558
Tick; Rhipicephalus sanguineus; Dog mummy; Archeoparasitology; Origins
15.  Dogs, cats, parasites, and humans in Brazil: opening the black box 
Parasites & Vectors  2014;7:22.
Dogs and cats in Brazil serve as primary hosts for a considerable number of parasites, which may affect their health and wellbeing. These may include endoparasites (e.g., protozoa, cestodes, trematodes, and nematodes) and ectoparasites (i.e., fleas, lice, mites, and ticks). While some dog and cat parasites are highly host-specific (e.g., Aelurostrongylus abstrusus and Felicola subrostratus for cats, and Angiostrongylus vasorum and Trichodectes canis for dogs), others may easily switch to other hosts, including humans. In fact, several dog and cat parasites (e.g., Toxoplasma gondii, Dipylidium caninum, Ancylostoma caninum, Strongyloides stercoralis, and Toxocara canis) are important not only from a veterinary perspective but also from a medical standpoint. In addition, some of them (e.g., Lynxacarus radovskyi on cats and Rangelia vitalii in dogs) are little known to most veterinary practitioners working in Brazil. This article is a compendium on dog and cat parasites in Brazil and a call for a One Health approach towards a better management of some of these parasites, which may potentially affect humans. Practical aspects related to the diagnosis, treatment, and control of parasitic diseases of dogs and cats in Brazil are discussed.
doi:10.1186/1756-3305-7-22
PMCID: PMC3914713  PMID: 24423244
Dogs; Cats; Humans; Zoonosis; Control; South America
16.  Clinical Bovine Piroplasmosis Caused by Babesia occultans in Italy 
Journal of Clinical Microbiology  2013;51(7):2432-2434.
A clinical outbreak of bovine piroplasmosis was reported in Italy. The etiological agent was characterized as Babesia occultans, a parasite regarded as apathogenic and never detected before in continental Europe. This report paves the way for further studies to assess the occurrence of this tick-transmitted protozoan in other European regions.
doi:10.1128/JCM.00713-13
PMCID: PMC3697720  PMID: 23658262
17.  Cutaneous Distribution and Circadian Rhythm of Onchocerca lupi Microfilariae in Dogs 
Background
Among the arthropod-borne nematodes infesting dogs, Onchocerca lupi (Spirurida: Onchocercidae) is of increasing zoonotic concern, with new human cases of infection diagnosed in Turkey, Tunisia, Iran and the USA. Knowledge of the biology of this nematode is meagre. This study aimed at assessing the distribution and periodicity of O. lupi microfilariae from different body regions in naturally infested dogs.
Methodology/Principal Findings
Skin samples were collected from six dogs infested with O. lupi but without apparent clinical signs. Two skin samples were collected from 18 anatomical regions of dog 1 at necropsy. In addition, single skin biopsies were performed from the forehead, inter-scapular and lumbar regions of dogs 2–6, in the morning, afternoon, and at night. Two aliquots of the sediment of each sample were microscopically observed, microfilariae counted and morphologically and molecularly identified. Most of the 1,667 microfilariae retrieved from dog 1 were in the right ear (59.6%), nose (26.5%), left ear (6.7%), forehead (3.0%), and inter-scapular (2.9%) regions. In dogs 2–6, the overall mean number of microfilariae was larger on the head (n = 122.8), followed by the inter-scapular (n = 119.0) and lumbar (n = 12.8) regions. The overall mean number of microfilariae was larger in the afternoon (153.4), followed by night (75.4) and morning (25.8).
Conclusions
Onchocerca lupi microfilariae were more common in the head (i.e., ears and nose) than in the remaining part of the dog's body, indicating they tend to aggregate in specific body regions, which are the best sites to collect skin samples for diagnostic purposes. The periodicity pattern of microfilariae of O. lupi and their concentration in specific body regions is most likely a result of the co-evolution with their as-yet-unknown vector. The detection of skin microfilariae in asymptomatic animals, suggests the potential role of these animals as carriers and reservoirs of O. lupi.
Author Summary
Onchocerca lupi is a little known arthropod-borne helminth infesting dogs of increasing interest to the scientific community due to the recent demonstration of its zoonotic potential. Nonetheless, knowledge of the biology of this nematode is exiguous. In this study the distribution and periodicity of O. lupi microfilariae was investigated from different body regions in naturally infested dogs. Data indicate that O. lupi microfilariae were more common in the head (i.e., ears and nose) followed by the inter-scapular region than in the remaining part of the dog's body suggesting that these parasites aggregate in these anatomical sites. These regions might be the best sites to collect skin samples for diagnostic purposes. Finally, the periodicity pattern of microfilariae of O. lupi and their concentration in specific body regions is most likely a result of the co-evolution with their as-yet-unknown vector.
doi:10.1371/journal.pntd.0002585
PMCID: PMC3861181  PMID: 24349594
18.  Zoonotic Onchocerca lupi Infection in Dogs, Greece and Portugal, 2011–2012 
Emerging Infectious Diseases  2013;19(12):2000-2003.
Onchocerca lupi infection is reported primarily in symptomatic dogs. We aimed to determine the infection in dogs from areas of Greece and Portugal with reported cases. Of 107 dogs, 9 (8%) were skin snip–positive for the parasite. DNA sequences of parasites in specimens from distinct dog populations differed genetically from thoses in GenBank.
doi:10.3201/eid1912.130264
PMCID: PMC3840859  PMID: 24274145
Onchocerca lupi; dogs; zoonoses; infection; Portugal; Greece; parasites
19.  Redescription of Onchocerca lupi (Spirurida: Onchocercidae) with histopathological observations 
Parasites & Vectors  2013;6:309.
Background
Onchocerca lupi is a dog parasite of increasing zoonotic concern, with new human cases diagnosed in Turkey, Tunisia, Iran, and the United States. Information about the morphology of this nematode is scant and a detailed re-description of this species is overdue. In addition, histopathological data of potential usefulness for the identification of O. lupi infections are provided.
Methods
Male and female nematodes, collected from the connective tissue of a dog, were examined using light microscopy and scanning electron microscopy (SEM), and an histological evaluation was performed on biopsy samples from periocular tissues.
Results
The morphological identification was confirmed by molecular amplification and partial sequencing of cytochrome oxidase subunit 1 gene. This study provides the first comprehensive morphological and morphometric description of O. lupi from a dog based on light microscopy, SEM, molecular characterization, and histological observations.
Conclusions
Data herein presented contribute to a better understanding of this little known parasitic zoonosis, whose impact on human and animal health is still underestimated. The presence of granulomatous reactions only around the female adult suggests that the release of microfilariae from the uterus might be the cause of the inflammatory reaction observed.
doi:10.1186/1756-3305-6-309
PMCID: PMC3818983  PMID: 24499611
Onchocerca lupi; Vector-borne disease; Scanning electron microscopy; Light microscopy; Histology; Zoonosis
20.  Cercopithifilaria rugosicauda (Spirurida, Onchocercidae) in a roe deer and ticks from southern Italy☆ 
Graphical abstract
Highlights
•First report of Cercopithifilaria rugosicauda in a roe deer and ticks from Italy.•This study provides new morphological data on this little known nematode.•The genetic data provide for the first time information on mitochondrial genes of C. rugosicauda.
Cercopithifilaria rugosicauda (Spirurida, Onchocercidae) is a subcutaneous filarial nematode of the European roe deer (Capreolus capreolus) transmitted by Ixodes ricinus (Acari, Ixodidae). At the necropsy of a roe deer from the Parco Regionale di Gallipoli Cognato (Basilicata region, southern Italy), two female nematodes of C. rugosicauda were found. Following the necropsy, seven skin snips were sampled from different body regions and 96 I. ricinus ticks were collected. In addition, 240 ticks were collected by dragging in the enclosure where the roe deer lived. Samples were examined for the presence of C. rugosicauda larvae and assayed by PCR targeting cytochrome c oxidase subunit 1 (cox1, ∼300 bp) and 12S rDNA (∼330 bp) gene fragments. Female nematodes, microfilariae from skin samples and eight third stage larvae (L3) from ticks were morphologically and molecularly identified as C. rugosicauda. Phylogenetic analyses clustered this species with other sequences of Cercopithifilaria spp. This study represents the first report of C. rugosicauda in a roe deer and ticks from Italy and provides new morphological and molecular data on this little known nematode.
doi:10.1016/j.ijppaw.2013.09.009
PMCID: PMC3862540  PMID: 24533349
Cercopithifilaria rugosicauda; Ixodes ricinus; Roe deer; Capreolus capreolus
21.  Dirofilariosis in the Americas: a more virulent Dirofilaria immitis? 
Parasites & Vectors  2013;6:288.
Dirofilarioses are widespread diseases caused by filarioid nematodes (superfamily Filarioidea) of the genus Dirofilaria, which are transmitted by a plethora of mosquito species. The principal agent of canine dirofilariosis in the Americas is Dirofilaria immitis, which may also occasionally infest humans, resulting in pulmonary nodules that may be confounded with malignant lung tumours. Because human cases of dirofilariosis by D. immitis are relatively frequent in the Americas and rare in Europe and other eastern countries, where Dirofilaria repens is the main causative agent, the existence of a more virulent strain of D. immitis in the Americas has been speculated. Recently, a case of human ocular infestation by Dirofilaria sp. was diagnosed in Pará State, northern Brazil, where canine heartworm dirofilariosis is endemic. The nematode was shown to be morphologically and phylogenetically related to D. immitis but it was genetically distinct from reference sequences, including those of D. immitis infesting dogs in the same geographical area. This finding raised questions regarding the aetiology of human dirofilariosis in the Americas, since information on the genetic makeup of filarioids infesting dogs and humans is meagre. Further studies would be needed to better characterize filarioids infesting dogs, wild animals, and humans in the Americas and to assess the existence of a more virulent D. immitis strain in this continent. Finally, the competence of different culicid species/strains from Europe and the Americas as vectors of Dirofilaria species should be investigated. Such studies would help us to understand possible variations in transmission patterns and even to predict possible scenarios that may emerge in the future, with the introduction of non-endemic Dirofilaria species/strains in free areas through importation of infested animals, vectors, or both.
doi:10.1186/1756-3305-6-288
PMCID: PMC3851770  PMID: 24274042
Dirofilaria immitis; Americas; Genetics; Pathogenicity; Virulence; Zoonosis
22.  Effects of global changes on the climatic niche of the tick Ixodes ricinus inferred by species distribution modelling 
Parasites & Vectors  2013;6:271.
Background
Global climate change can seriously impact on the epidemiological dynamics of vector-borne diseases. In this study we investigated how future climatic changes could affect the climatic niche of Ixodes ricinus (Acari, Ixodida), among the most important vectors of pathogens of medical and veterinary concern in Europe.
Methods
Species Distribution Modelling (SDM) was used to reconstruct the climatic niche of I. ricinus, and to project it into the future conditions for 2050 and 2080, under two scenarios: a continuous human demographic growth and a severe increase of gas emissions (scenario A2), and a scenario that proposes lower human demographic growth than A2, and a more sustainable gas emissions (scenario B2). Models were reconstructed using the algorithm of “maximum entropy”, as implemented in the software Maxent 3.3.3e; 4,544 occurrence points and 15 bioclimatic variables were used.
Results
In both scenarios an increase of climatic niche of about two times greater than the current area was predicted as well as a higher climatic suitability under the scenario B2 than A2. Such an increase occurred both in a latitudinal and longitudinal way, including northern Eurasian regions (e.g. Sweden and Russia), that were previously unsuitable for the species.
Conclusions
Our models are congruent with the predictions of range expansion already observed in I. ricinus at a regional scale and provide a qualitative and quantitative assessment of the future climatically suitable areas for I. ricinus at a continental scale. Although the use of SDM at a higher resolution should be integrated by a more refined analysis of further abiotic and biotic data, the results presented here suggest that under future climatic scenarios most of the current distribution area of I. ricinus could remain suitable and significantly increase at a continental geographic scale. Therefore disease outbreaks of pathogens transmitted by this tick species could emerge in previous non-endemic geographic areas. Further studies will implement and refine present data toward a better understanding of the risk represented by I. ricinus to human health.
doi:10.1186/1756-3305-6-271
PMCID: PMC3848450  PMID: 24330500
Climatic changes; Future climatic niche; Species distribution modeling; Tick; Vector-borne disease; Lyme disease; Babesiosis; Anaplasmosis; Erlichiosis and tick-borne encephalitis
23.  Efficacy of an imidacloprid/flumethrin collar against fleas, ticks and tick-borne pathogens in dogs 
Parasites & Vectors  2013;6:245.
Background
Tick-borne diseases comprise a group of maladies that are of substantial medical and veterinary significance. A range of tick-borne pathogens, including diverse species of bacteria and protozoa, can infect both dogs and humans. Hence, the control of tick infestations is pivotal to decrease or prevent tick-borne pathogen transmission. Therefore, different commercial products with insecticidal, repellent or both properties have been developed for use on dogs. Recently, a collar containing a combination of imidacloprid 10% and flumethrin 4.5% has proven effective to prevent tick and flea infestations in dogs under field conditions and the infection by some vector-borne pathogens they transmit under laboratory-controlled conditions.
Methods
From March 2011 to April 2012, a field study was conducted in a private shelter in southern Italy to assess the efficacy of the imidacloprid/flumethrin collar against tick and flea infestations and to determine if this strategy would decrease tick-borne pathogen transmission in young dogs. A total of 122 animals were enrolled in the study and randomly assigned to group A (n = 64; collared) or group B (n = 58; untreated controls). Dogs were examined monthly for ticks and fleas and systematically tested for selected tick-borne pathogens.
Results
Compared to controls, the collar provided overall efficacies of 99.7% and 100% against tick and flea infestation, respectively. The overall efficacy for the prevention of tick-borne pathogens (i.e., Anaplasma platys and Babesia vogeli) was 91.6%.
Conclusions
This study demonstrates that the imidacloprid/flumethrin collar is efficacious against flea and tick infestation as well as tick-borne pathogen transmission to dogs under field conditions.
doi:10.1186/1756-3305-6-245
PMCID: PMC3766024  PMID: 23972013
Canine vector-borne diseases; Anaplasma platys; Babesia vogeli; Hepatozoon canis; Prevention; Dog; Imidacloprid; Flumethrin; Tick
24.  Morphological and genetic diversity of Rhipicephalus sanguineus sensu lato from the New and Old Worlds 
Parasites & Vectors  2013;6:213.
Background
The taxonomic status of the brown dog tick (Rhipicephalus sanguineus sensu stricto), which has long been regarded as the most widespread tick worldwide and a vector of many pathogens to dogs and humans, is currently under dispute.
Methods
We conducted a comprehensive morphological and genetic study of 278 representative specimens, which belonged to different species (i.e., Rhipicephalus bursa, R. guilhoni, R. microplus, R. muhsamae, R. pusillus, R. sanguineus sensu lato, and R. turanicus) collected from Europe, Asia, Americas, and Oceania. After detailed morphological examination, ticks were molecularly processed for the analysis of partial mitochondrial (16S rDNA, 12S rDNA, and cox1) gene sequences.
Results
In addition to R. sanguineus s.l. and R. turanicus, three different operational taxonomic units (namely, R. sp. I, R. sp. II, and R. sp. III) were found on dogs. These operational taxonomical units were morphologically and genetically different from R. sanguineus s.l. and R. turanicus. Ticks identified as R. sanguineus s.l., which corresponds to the so-called “tropical species” (=northern lineage), were found in all continents and genetically it represents a sister group of R. guilhoni. R. turanicus was found on a wide range of hosts in Italy and also on dogs in Greece.
Conclusions
The tropical species and the temperate species (=southern lineage) are paraphyletic groups. The occurrence of R. turanicus in the Mediterranean region is confirmed. A consensual re-description of R. sanguineus s.s. and R. turanicus will be necessary to solve the taxonomic problems within the so-called R. sanguineus group.
doi:10.1186/1756-3305-6-213
PMCID: PMC3735430  PMID: 23880226
Rhipicephalus sanguineus; Taxonomy; Morphology; Phylogeny
25.  Redescription of Cercopithifilaria bainae Almeida & Vicente, 1984 (Spirurida, Onchocercidae) from a dog in Sardinia, Italy 
Parasites & Vectors  2013;6:132.
Background
Three species of the genus Cercopithifilaria have been morphologically and molecularly characterized in dog populations in southern Europe: Cercopithifilaria grassii (Noè, 1907), Cercopithifilaria sp. sensu Otranto et al., 2011 (reported as Cercopithifilaria sp. I), and Cercopithifilaria sp. II sensu Otranto et al., 2012. The adults of Cercopithifilaria sp. I have remained unknown until the present study.
Methods
The material originated from a dog from Sardinia (Italy) diagnosed with dermal microfilariae of Cercopithifilaria sp. I. The holotype and three paratypes of Cercopithifilaria bainae Almeida & Vicente, 1984, described from dogs in Brazil, were studied as comparative material. A cox1 (~689 bp) and 12S (~330 bp) gene fragments were amplified and phylogenetic analysis carried out.
Results
The highest numbers of adult nematodes (82%) were collected in the sediment of the subcutaneous tissues of the trunk (n = 37) and forelimbs (n = 36). The morphology of the adult nematodes and microfilariae collected from the dog in Sardinia corresponded to those of C. bainae. All cox1 and 12S gene sequences showed a high homology (99-100%) with sequences from microfilariae of Cercopithifilaria sp. I.
Conclusions
The morphological and molecular identity of the microfilariae of C. bainae overlap those described previously as Cercopithifilaria sp. sensu Otranto et al., 2011 (=Cercopithifilaria sp. I). Therefore, the present study reports the occurrence of C. bainae in Europe, for the first time after its description and the single record in Brazil. C. bainae appears to be highly diffused in dog populations in southern Europe. The phylogenetic analyses based on cox1 and 12S do not reveal the three species of Cercopithifilaria parasitizing dogs as a monophyletic group, which suggests that they have derived independently by host switching.
doi:10.1186/1756-3305-6-132
PMCID: PMC3655055  PMID: 23642161
Canine filarioids; Cercopithifilaria sp. I; Cercopithifilaria sp. II; Cercopithifilaria bainae; Cercopithifilaria grassii; Taxonomy; Europe

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