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Domenico Otranto - One of the best experts on this subject based on the ideXlab platform.
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Dirofilariosis
Arthropod Borne Diseases, 2017Co-Authors: Filipe Dantas-torres, Emanuele Brianti, Domenico OtrantoAbstract:Dirofilarioses are diseases caused by spirurid nematodes of the genus Dirofilaria , which are transmitted by several mosquito species. The most important species causing dirofilariosis in dogs are Dirofilaria immitis and Dirofilaria repens , which may also affect other mammals, including cats and humans. Dirofilaria immitis causes heartworm disease in dogs and may eventually lead to the formation of pulmonary nodules in humans, which may be often confounded with malignant tumors. Dirofilaria repens in dogs is usually asymptomatic, but in humans the parasite may localize in different tissues and organs (e.g., the eyes, lungs, and testicles) and cause disease. The diagnosis of cardiopulmonary dirofilariosis is usually based on clinical signs and laboratorial tests, including the modified Knott test for the detection of circulating microfilariae and rapid serological tests, targeting antigens of or antibodies to the parasite. Molecular tools have also been recently developed but are still mostly restricted to research centers and universities. This chapter reviews general aspects related to the etiology, epidemiology, clinical signs, diagnosis, treatment, and control of dirofilariosis.
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the mitochondrial genomes of the zoonotic canine filarial parasites Dirofilaria nochtiella repens and candidatus Dirofilaria nochtiella honkongensis provide evidence for presence of cryptic species
PLOS Neglected Tropical Diseases, 2016Co-Authors: Esra Yilmaz, Domenico Otranto, Moritz Fritzenwanker, Nikola Pantchev, Mathias Lendner, Sirichit Wongkamchai, Inge Kroidl, Martin Dennebaum, Thanh Hoa Le, Tran Anh LeAbstract:Background Cutaneous dirofilariosis is a canine mosquito-borne zoonosis that can cause larva migrans disease in humans. Dirofilaria repens is considered an emerging pathogen occurring with high prevalence in Mediterranean areas and many parts of tropical Asia. In Hong Kong, a second species, Candidatus Dirofilaria hongkongensis, has been reported. The present study aimed to compare mitochondrial genomes from these parasites and to obtain population genetic information. Methods and Findings Complete mitochondrial genomes were obtained by PCR and Sanger sequencing or ILLU-MINA sequencing for four worms. Cytochrome oxidase subunit 1 sequences identified three as D. repens (all from Europe) and one as C. D. hongkongensis (from India). Mitochondrial genomes have the same organization as in other spirurid nematodes but a higher preference for thymine in the coding strand. Phylogenetic analysis was in contradiction to current taxonomy of the Onchocercidae but in agreement with a recent multi-locus p hylogenetic analysis using both mitochondrial and nuclear markers. D. repens and C. D. hongkongensis sequences clustered together and were the common sister group to Dirofilaria immitis. Analysis of a 2.5 kb mitochondrial genome fragment from macrofilaria or canine blood samples from Europe (42), Thailand (2), India (1) and Vietnam (1) revealed only small genetic differences in the D. repens samples including all European and the Vietnam sample. The Indian C. D. hongkongensis and the two Thai samples formed separate clusters and differences were comparatively large. Conclusion Genetic differences between Dirofilaria spp. causing cutaneous disease can be considerable whereas D. repens itself was genetically quite homogenous. C. D. hongkongensis was identified for the first time from the Indian subcontinent. The full mitochondrial genome sequence strengthens the hypothesis that it represents an independent species and the Thai samples might represent another cryptic species, Candidatus Dirofilaria sp. 'Thailand II', or a quite divergent population of C. D. hongkongensis.
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current surveys on the prevalence and distribution of Dirofilaria spp and acanthocheilonema reconditum infections in dogs in romania
Parasitology Research, 2015Co-Authors: Angela Monica Ionică, Nikola Pantchev, Ioana Adriana Matei, Viorica Mircean, Mirabela Oana Dumitrache, Gianluca Damico, Adriana Győrke, Giada Annoscia, Kateřina Albrechtova, Domenico OtrantoAbstract:During the last decades, Dirofilaria spp. infection in European dogs has rapidly spread from historically endemic areas towards eastern and northeastern countries, but little or no information is available from these geographical regions. The present study provides a picture of filarial infections in dogs from Romania and compares two tests for the diagnosis of Dirofilaria immitis. From July 2010 to March 2011, blood samples were collected from 390 dogs from nine counties of Romania and serological SNAP tests were performed for the detection of D. immitis antigen. The remaining blood clots were subsequently used for DNA extraction followed by multiplex PCR for assessing filarioid species diversity (i.e. D. immitis, Dirofilaria repens and Acanthocheilonema reconditum). Based on molecular detection, an overall prevalence of 6.92 % (n = 27; 95 % confidence interval (CI) 4.70–10.03 %) for D. repens, 6.15 % (n = 24; 95 % CI 4.07–9.14 %) for D. immitis and 2.05 % (n = 8; 95 % CI 0.96–4.16 %) for A. reconditum was recorded, with significant variations according to sampling areas. Coinfections of D. immitis and D. repens were recorded in 23.91 % (n = 11) positive dogs. A slightly higher prevalence for D. immitis was detected at the SNAP test (n = 28, 7.17 %; 95 % CI 4.91–10.33 %), but this difference was not statistically significant (p = 0.66). However, only 53.57 % (n = 15) of antigen-positive dogs were confirmed by PCR, while other dogs (n = 9) PCR positive for D. immitis were negative at the serology. The present study shows that Dirofilaria species are endemic in the southern and southeastern areas of Romania, This article also provides, for the first time, an epidemiological picture of the distribution of A. reconditum in Romania.
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dirofilariosis in the americas a more virulent Dirofilaria immitis
Parasites & Vectors, 2013Co-Authors: Filipe Dantastorres, Domenico OtrantoAbstract: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 Para 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.
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a duplex real time polymerase chain reaction assay for the detection of and differentiation between Dirofilaria immitis and Dirofilaria repens in dogs and mosquitoes
Veterinary Parasitology, 2012Co-Authors: Maria Stefania Latrofa, Marco Genchi, Filipe Dantastorres, Giada Annoscia, Donato Traversa, Domenico OtrantoAbstract:Abstract This study describes a duplex real-time polymerase chain reaction (PCR) assay for the detection and differentiation between Dirofilaria immitis and Dirofilaria repens in dog blood and mosquitoes. Regions of a cytochrome oxidase 1 (cox1) mitochondrial DNA fragment and the second internal transcribed spacer (ITS-2) of nuclear ribosomal DNA were amplified from microfilariae and adult worm samples, using a sensitive SsoFast™ EvaGreen® based real-time PCR method coupled with melting-curve analysis. The limit of the real-time PCR in detecting microfilaria and adult worm DNA was also tested both in dog blood and in artificially infected microfilarial. Two peaks at different melting temperatures (Tm) for D. immitis (mean ± SD = 75.7 ± 0.3 °C) and D. repens (mean ± SD = 70 ± 0.7 °C), respectively, were obtained for microfilarial and adult positive controls of both species when examined separately and together. The real-time PCR protocol was also efficient in detecting microfilarial and adult DNA of both species when tested in samples spiked with DNA from Aedes albopictus, in Aedes aegypti experimentally infected by D. repens and in Culex pipiens naturally infected by D. repens and D. immitis. The high sensitivity of real-time PCR confirmed its reliability in detecting small amounts of genomic DNA either in dog blood or mosquitoes (2.5 pg/μl and 3 × 10−1 pg/μl for D. immitis and D. repens, respectively). This assay is proposed as a tool for the epidemiological surveillance of the two most important Dirofilaria species in areas where they are endemic and sympatric.
Michele Mortarino - One of the best experts on this subject based on the ideXlab platform.
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Mapping and modeling Dirofilaria infections in Europe
Parasites & Vectors, 2014Co-Authors: Laura Rinaldi, Michele Mortarino, Vincenzo Musella, G. Marzatico, Genchi Claudio, Giuseppe CringoliAbstract:Climate change and increasing temperatures are a global phenomenon that can influence the dynamics of a number of hematophagous arthropods, vectors of pathogens with importance in human and veterinary medicine. In fact, climatic changes, together with an increase in the movement of dogs across Europe, have caused an increase in the geographical range of Dirofilaria immitis and D. repens infections. A Geographic Information System based on thermal regimen was constructed to identify areas potentially suitable for Dirofilaria transmission in Europe. These models are based on evidence that: i) there is a threshold of 14 °C below which Dirofilaria development will not proceed in mosquitoes; ii) there is a requirement of 130 growing degree-days for larvae to reach infectivity, and; iii) there is a maximum life expectancy of 30 days for a mosquito vector. The output of these models predicted that the summer temperatures (with peaks in July and August) are sufficient to facilitate extrinsic incubation of Dirofilaria even at high latitudes. Recently, an additional model was constructed to verify the influence of temperature in the course of three decades (1980-1989, 1990-1999 and 2000-2012) on the risk of infection by Dirofilaria in Italy. The results showed an expected increasing trend of temperatures, an increase of the Dirofilaria generation numbers into the mosquitoes and a significant extension of the infection risk from 5-6 months (1980-1989) to 6.5 months (1990-1999), up to more than 7 months (2000-20012). These findings show that geospatial tools are very useful for mapping, monitoring, forecasting and surveillance of both heartworm and subcutaneous Dirofilariasis.
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rapid differentiation of Dirofilaria immitis and Dirofilaria repens in canine peripheral blood by real time pcr coupled to high resolution melting analysis
Veterinary Parasitology, 2014Co-Authors: Francesca Albonico, C Genchi, Marco Genchi, G Gioia, Monica Loiacono, Michele MortarinoAbstract:Abstract Dirofilaria immitis and D. repens are the principal causative agents of canine filariosis and, although the number of dogs subjected to specific prevention is increasing, the prevalence of these parasites remains high in many areas of the world. The discrimination between the two Dirofilaria species using the classical diagnostic methods can be difficult and may lead to misdiagnosis especially on samples from areas where both Dirofilaria are present. Over the last years, several molecular methods with higher sensitivity and specificity compared to classical microscopy and ELISA assays were designed. Nevertheless, a need for simple, rapid, and cost-effective molecular protocols to accurately discriminate between D. immitis and D. repens still remains. High resolution melting analysis coupled to real-time PCR (real-time PCR–HRMA) is a widely used technique to target sequence polymorphisms of the same gene in different species without the need to perform DNA sequencing or to use species-specific probes. In this work, a fast and cost-effective real-time PCR–HRMA protocol to detect and differentiate simultaneously and unequivocally D. immitis and D. repens microfilarial DNA extracted from peripheral dog blood samples is described. The present method is simpler to use than most other DNA-based methods and provides comparable discrimination between the two sibling species.
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changing climate and changing vector borne disease distribution the example of Dirofilaria in europe
Veterinary Parasitology, 2011Co-Authors: C Genchi, Michele Mortarino, Laura Rinaldi, Giuseppe Cringoli, Giorgio Traldi, Marco GenchiAbstract:Abstract Climatic changes, together with an increase in the movement of dogs across Europe, have caused an increase in the geographical range of Dirofilaria infections. The present paper is focuses on northeastern European countries, where survey data have shown an increase of Dirofilaria repens infections both in animals and humans. A growing degree day-based forecast model has been developed to predict the occurrence. The model is based on evidence that there is a threshold of 14 °C below which Dirofilaria development will not proceed in mosquitoes, there is a requirement of 130 growing degree-days (GDDs) for larvae to reach infectivity, and there is a maximum life expectancy of 30 days for a mosquito vector. The output of this model predicted that the summer temperatures (with peaks in August) are sufficient to facilitate extrinsic incubation of Dirofilaria even at latitudes of 56°N and longitudes of 39°E. Despite the fact that both Dirofilaria immitis and D. repens have the same temperature requirement for extrinsic incubation in mosquitoes, empirical data has shown that D. repens is the main cause of Dirofilarial infections in both humans and animals. Clinical signs are absent in most canine infections with D. repens. Furthermore, diagnosis is problematic and in-clinic serological tests, such as those for D. immitis, do not exist. Therefore, most infections go undiagnosed, allowing the infection to spread undetected.
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highly sensitive multiplex pcr for simultaneous detection and discrimination of Dirofilaria immitis and Dirofilaria repens in canine peripheral blood
Veterinary Parasitology, 2010Co-Authors: G Gioia, C Genchi, Marco Genchi, L Lecova, E Ferri, Michele MortarinoAbstract:Dirofilaria immitis and Dirofilaria repens are the most common species of filarial nematodes described in the dogs with increasing spread into new geographical areas. The diagnosis of canine dirofilariosis is usually based upon the microscopical detection and identification of circulating microfilariae together with ELISA detection of serum circulating heartworm antigens or antibodies. The identification of the parasite species using the traditional approaches sometimes can be difficult and can lead to misdiagnosis especially on samples from areas where both Dirofilaria are present. In this paper we report a new molecular method based on single-step multiplex PCR to detect and differentiate simultaneously and unequivocally D. immitis and D. repens on DNA extracted from canine peripheral blood. The amplification was performed using a set of primers designed on a portion of the small subunit ribosomal RNA gene of the mitochondrion (12S rDNA). The single-step multiplex PCR here described ensured high (4 mf/ml) sensitivity and specificity with reduced cost and time saving. The multiplex PCR assay represents an additional tool for epidemiological studies and routine disease assessment in areas co-endemic for the two Dirofilaria species.
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climate and Dirofilaria infection in europe
Veterinary Parasitology, 2009Co-Authors: C Genchi, Michele Mortarino, Laura Rinaldi, Marco Genchi, Giuseppe CringoliAbstract:Climatic changes, together with an increase in the movement of cats and dogs across Europe, have caused an increase in the geographical range of several vector borne parasites like Dirofilaria, and in the risk of infection for animals and humans. The present paper reviews the effects of climate and other global drivers on Dirofilaria immitis and Dirofilaria repens infections in Europe and the possible implications on the transmission and control of these mosquito-borne nematodes. In the last several years, growing degree day (GDD)-based forecast models, which use wide or local scale temperature data, have been developed to predict the occurrence and seasonality of Dirofilaria in different parts of the world. All these models are based on the fact that: there is a threshold of 14 °C below which Dirofilaria development will not proceed; and there is a requirement of 130 GDD for larvae to reach infectivity and a maximum life expectancy of 30 days for a vector mosquito. The output of these models predicts that the summer temperatures (with peaks in July) are sufficient to facilitate extrinsic incubation of Dirofilaria even at high latitudes. The global warming projected by the Intergovernmental Panel on Climate Change suggests that warm summers suitable for Dirofilaria transmission in Europe will be the rule in the future decades and if the actual trend of temperature increase continues, filarial infection should spread into previously infection-free areas. These factors not only favour incubation of Dirofilaria, but also impact on mosquito species. Recent findings have also demonstrated that Aedes albopictus is now considered to be an important, competent vector of Dirofilaria infections. This mosquito species could spread from southern to northern European countries in the near future, changing the epidemiological patterns of dirofilariosis both in humans and animals.
Marco Genchi - One of the best experts on this subject based on the ideXlab platform.
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rapid differentiation of Dirofilaria immitis and Dirofilaria repens in canine peripheral blood by real time pcr coupled to high resolution melting analysis
Veterinary Parasitology, 2014Co-Authors: Francesca Albonico, C Genchi, Marco Genchi, G Gioia, Monica Loiacono, Michele MortarinoAbstract:Abstract Dirofilaria immitis and D. repens are the principal causative agents of canine filariosis and, although the number of dogs subjected to specific prevention is increasing, the prevalence of these parasites remains high in many areas of the world. The discrimination between the two Dirofilaria species using the classical diagnostic methods can be difficult and may lead to misdiagnosis especially on samples from areas where both Dirofilaria are present. Over the last years, several molecular methods with higher sensitivity and specificity compared to classical microscopy and ELISA assays were designed. Nevertheless, a need for simple, rapid, and cost-effective molecular protocols to accurately discriminate between D. immitis and D. repens still remains. High resolution melting analysis coupled to real-time PCR (real-time PCR–HRMA) is a widely used technique to target sequence polymorphisms of the same gene in different species without the need to perform DNA sequencing or to use species-specific probes. In this work, a fast and cost-effective real-time PCR–HRMA protocol to detect and differentiate simultaneously and unequivocally D. immitis and D. repens microfilarial DNA extracted from peripheral dog blood samples is described. The present method is simpler to use than most other DNA-based methods and provides comparable discrimination between the two sibling species.
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a duplex real time polymerase chain reaction assay for the detection of and differentiation between Dirofilaria immitis and Dirofilaria repens in dogs and mosquitoes
Veterinary Parasitology, 2012Co-Authors: Maria Stefania Latrofa, Marco Genchi, Filipe Dantastorres, Giada Annoscia, Donato Traversa, Domenico OtrantoAbstract:Abstract This study describes a duplex real-time polymerase chain reaction (PCR) assay for the detection and differentiation between Dirofilaria immitis and Dirofilaria repens in dog blood and mosquitoes. Regions of a cytochrome oxidase 1 (cox1) mitochondrial DNA fragment and the second internal transcribed spacer (ITS-2) of nuclear ribosomal DNA were amplified from microfilariae and adult worm samples, using a sensitive SsoFast™ EvaGreen® based real-time PCR method coupled with melting-curve analysis. The limit of the real-time PCR in detecting microfilaria and adult worm DNA was also tested both in dog blood and in artificially infected microfilarial. Two peaks at different melting temperatures (Tm) for D. immitis (mean ± SD = 75.7 ± 0.3 °C) and D. repens (mean ± SD = 70 ± 0.7 °C), respectively, were obtained for microfilarial and adult positive controls of both species when examined separately and together. The real-time PCR protocol was also efficient in detecting microfilarial and adult DNA of both species when tested in samples spiked with DNA from Aedes albopictus, in Aedes aegypti experimentally infected by D. repens and in Culex pipiens naturally infected by D. repens and D. immitis. The high sensitivity of real-time PCR confirmed its reliability in detecting small amounts of genomic DNA either in dog blood or mosquitoes (2.5 pg/μl and 3 × 10−1 pg/μl for D. immitis and D. repens, respectively). This assay is proposed as a tool for the epidemiological surveillance of the two most important Dirofilaria species in areas where they are endemic and sympatric.
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changing climate and changing vector borne disease distribution the example of Dirofilaria in europe
Veterinary Parasitology, 2011Co-Authors: C Genchi, Michele Mortarino, Laura Rinaldi, Giuseppe Cringoli, Giorgio Traldi, Marco GenchiAbstract:Abstract Climatic changes, together with an increase in the movement of dogs across Europe, have caused an increase in the geographical range of Dirofilaria infections. The present paper is focuses on northeastern European countries, where survey data have shown an increase of Dirofilaria repens infections both in animals and humans. A growing degree day-based forecast model has been developed to predict the occurrence. The model is based on evidence that there is a threshold of 14 °C below which Dirofilaria development will not proceed in mosquitoes, there is a requirement of 130 growing degree-days (GDDs) for larvae to reach infectivity, and there is a maximum life expectancy of 30 days for a mosquito vector. The output of this model predicted that the summer temperatures (with peaks in August) are sufficient to facilitate extrinsic incubation of Dirofilaria even at latitudes of 56°N and longitudes of 39°E. Despite the fact that both Dirofilaria immitis and D. repens have the same temperature requirement for extrinsic incubation in mosquitoes, empirical data has shown that D. repens is the main cause of Dirofilarial infections in both humans and animals. Clinical signs are absent in most canine infections with D. repens. Furthermore, diagnosis is problematic and in-clinic serological tests, such as those for D. immitis, do not exist. Therefore, most infections go undiagnosed, allowing the infection to spread undetected.
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highly sensitive multiplex pcr for simultaneous detection and discrimination of Dirofilaria immitis and Dirofilaria repens in canine peripheral blood
Veterinary Parasitology, 2010Co-Authors: G Gioia, C Genchi, Marco Genchi, L Lecova, E Ferri, Michele MortarinoAbstract:Dirofilaria immitis and Dirofilaria repens are the most common species of filarial nematodes described in the dogs with increasing spread into new geographical areas. The diagnosis of canine dirofilariosis is usually based upon the microscopical detection and identification of circulating microfilariae together with ELISA detection of serum circulating heartworm antigens or antibodies. The identification of the parasite species using the traditional approaches sometimes can be difficult and can lead to misdiagnosis especially on samples from areas where both Dirofilaria are present. In this paper we report a new molecular method based on single-step multiplex PCR to detect and differentiate simultaneously and unequivocally D. immitis and D. repens on DNA extracted from canine peripheral blood. The amplification was performed using a set of primers designed on a portion of the small subunit ribosomal RNA gene of the mitochondrion (12S rDNA). The single-step multiplex PCR here described ensured high (4 mf/ml) sensitivity and specificity with reduced cost and time saving. The multiplex PCR assay represents an additional tool for epidemiological studies and routine disease assessment in areas co-endemic for the two Dirofilaria species.
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climate and Dirofilaria infection in europe
Veterinary Parasitology, 2009Co-Authors: C Genchi, Michele Mortarino, Laura Rinaldi, Marco Genchi, Giuseppe CringoliAbstract:Climatic changes, together with an increase in the movement of cats and dogs across Europe, have caused an increase in the geographical range of several vector borne parasites like Dirofilaria, and in the risk of infection for animals and humans. The present paper reviews the effects of climate and other global drivers on Dirofilaria immitis and Dirofilaria repens infections in Europe and the possible implications on the transmission and control of these mosquito-borne nematodes. In the last several years, growing degree day (GDD)-based forecast models, which use wide or local scale temperature data, have been developed to predict the occurrence and seasonality of Dirofilaria in different parts of the world. All these models are based on the fact that: there is a threshold of 14 °C below which Dirofilaria development will not proceed; and there is a requirement of 130 GDD for larvae to reach infectivity and a maximum life expectancy of 30 days for a vector mosquito. The output of these models predicts that the summer temperatures (with peaks in July) are sufficient to facilitate extrinsic incubation of Dirofilaria even at high latitudes. The global warming projected by the Intergovernmental Panel on Climate Change suggests that warm summers suitable for Dirofilaria transmission in Europe will be the rule in the future decades and if the actual trend of temperature increase continues, filarial infection should spread into previously infection-free areas. These factors not only favour incubation of Dirofilaria, but also impact on mosquito species. Recent findings have also demonstrated that Aedes albopictus is now considered to be an important, competent vector of Dirofilaria infections. This mosquito species could spread from southern to northern European countries in the near future, changing the epidemiological patterns of dirofilariosis both in humans and animals.
G Cancrini - One of the best experts on this subject based on the ideXlab platform.
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canine Dirofilaria infections in two uninvestigated areas of serbia epidemiological and genetic aspects
Vector-borne and Zoonotic Diseases, 2012Co-Authors: Aleksandar Tasic, Simona Gabrielli, Suzana Tasicotasevic, Natasa Miladinovictasic, Aleksandra Ignjatovic, Jovana đorđevic, Sanda Dimitrijevic, G CancriniAbstract:In 2009 canine filarial infections were investigated in two northern areas of Serbia (Pancevo and Veliko Gradiste), applying morphometry, biochemical staining, and immunological kit to detect Dirofilaria immitis antigens, and two home-made ELISAs to detect antibodies to D. repens and D. immitis somatic/metabolic polyproteins. Moreover, molecular tools were applied to analyze the phylogenetic relationships of the isolates. The microfilariae detected in 21/122 dogs (17.2%) were identified as D. repens (n=21) and D. immitis (n=2). D. immitis antigens were found in another 13 animals with occult infection. All of the 15 heartworm-positive dogs also had antibodies to this parasite, which were detected in another 13 subjects, indicating an overall D. immitis seroprevalence rate of 22.9%. Serology for D. repens revealed evidence of antibodies in 42.6% of the dogs, but was negative for 4 microfilaremic dogs. As for the two different areas, the prevalence of microfilariae and/or D. immitis antigens, mainly due to D. repens microfilaremic animals, was not significantly higher in Veliko Gradiste (33.3%) than in Pancevo (22%). However, serology showed a different epidemiological picture. Heartworm infection occurred more often in both areas, and antibodies to Dirofilarial nematodes were detected in 72.9% of dogs living in Pancevo, a rate higher than in those living in Veliko Gradiste (57.1%). No risk factors for infection were found, confirming the uselessness of prophylactic drugs against D. repens, and suggesting the presence in these areas of sunrise- or sunset-biting mosquitoes as important vectors. The results indicate the need for both appropriate entomological studies and further research on the intra-species variability shown by D. repens.
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aedes albopictus and culex pipiens implicated as natural vectors of Dirofilaria repens in central italy
Journal of Medical Entomology, 2007Co-Authors: G Cancrini, Simona Gabrielli, L Toma, M Di Paolo, Paola Scaramozzino, Roberto RomiAbstract:To identify the natural vectors of Dirofilaria repens Railliet et Henry, entomological samplings were carried out in four sites within the Lazio region, foci of canine subcutaneous Dirofilariasis. Collections were made in 2002–2003 by means of dog-baited and miniature Centers for Disease Control and Prevention traps as well as on humans. Microscopy identified 1,576 attracted mosquito females as belonging to six species, but molecular diagnostics detected filarial DNA only in Culex pipiens L. and Aedes albopictus (Skuse, 1894). Dirofilaria immitis Leidy DNA, D. repens DNA, or both were found in the head and thorax of both mosquitoes. The simultaneous presence of vectors showing diurnal and nocturnal activity patterns is of concern for animal and human health. The finding of D. immitis DNA in mosquitoes in areas where only D. repens was been recovered in dogs also demonstrates that this filarial parasite circulates among carnivores (wild or domesticated pets).
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Ribosomal DNA second internal transcribed spacer sequence studies of Culicid vectors from an endemic area of Dirofilaria immitis in Spain
Parasitology Research, 2006Co-Authors: M. D. Bargues, G Cancrini, R Morchon, J. M. Latorre, S. Mas-coma, F SimonAbstract:Studies were performed in an endemic area of Dirofilaria immitis in Spain to genetically characterize the potential mosquito vector species present by means of DNA sequencing and elucidate which of these species may be involved in the transmission. The rDNA ITS-2 sequences of two Culex pipiens haplotypes, H1 and H2, Aedes ( Aedimorphus ) vexans , Fredwardsius vittatus , Ochlerotatus ( Ochlerotatus ) caspius , Anopheles ( Anopheles ) atroparvus , and Anopheles ( Anopheles ) plumbeus were obtained. F. vittatus and An. plumbeus were detected for the first time. Results on abundance, presence, and activity suggest that the month of August is the period of higher transmission risk, with C. pipiens , Ae. vexans , and An. atroparvus present simultaneously. Population studies indicate that C. pipiens may be considered the most important potential vector, while Ae. vexans , An. atroparvus , and O. caspius being involved in transmission only sporadically. The absence of larval Dirofilarial infection agrees with the very low prevalences known in endemic areas.
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first finding of Dirofilaria repens in a natural population of aedes albopictus
Medical and Veterinary Entomology, 2003Co-Authors: G Cancrini, Roberto Romi, Simona Gabrielli, L Toma, M Di Paolo, Paola ScaramozzinoAbstract:. The invasive mosquito Aedes albopictus (Skuse) (Diptera: Culicidae) has become widespread in Italy during the past decade. Also Italy has foci of canine filariasis caused by Dirofilaria (Spirurida: Onchocercidae), due to subcutaneous D. repens Railliet & Henry as well as the dog heartworm D. immitis (Leidy) transmitted by various vector mosquitoes (Diptera: Culicidae). In 2002, at Fiumicino, west of Rome (Lazio Region), 17% of dogs were found to have D. repens microfilariae in peripheral blood. To evaluate the role of Ae. albopictus as a vector of Dirofilaria in this area, female mosquitoes were collected daily, June–October 2002, landing on dog or human bait in a rural house at Focene. Mosquitoes were maintained at 27°C and 70% RH for 6 days, to allow development or purging of filaria larvae, then identified and frozen for subsequent molecular assay with filaria-specific ribosomal S2-S16 primers. To distinguish specimens harbouring infective L3 Dirofilaria larvae, DNA was extracted separately from the mosquito abdomen and head-thorax. Dirofilaria species were identified by sequencing, confirmed by polymerase chain reaction of positive specimens using primers specific for D. immitis and D. repens. Dirofilaria DNA was detected in 3/154 (2%) of Ae. albopictus females examined: D. repens DNA in head-thorax and abdomen of one collected 27th July; D. immitis in the abdomen of one collected 24th September; DNA of both D. immitis and D. repens in the head-thorax of one collected 11th October 2002. Thus Ae. albopictus is a potential vector of both Dirofilarias in Italy, representing risks for veterinary and human health.
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polymerase chain reaction identification of Dirofilaria repens and Dirofilaria immitis
Parasitology, 1996Co-Authors: Guido Favia, G Cancrini, A Lanfrancotti, A Della Torre, M ColuzziAbstract:On the basis of known DNA sequences of Dirofilaria repens and D. immitis we designed specific primers for the amplification by Polymerase Chain Reaction (PCR) of the DNA from the two species. The PCR-based identification was found to be unambiguous and allowed specific diagnosis of microfilariae in blood samples, of developing larvae in the mosquito vector and of immature adults in bioptic material, overcoming the serious constraints of the morphological separation of these filarial parasites at the pre-adult stages. The technique was found to be very sensitive and applicable to samples stored either dry or in various preservation media, with the exception of formalin. The reliable identification of D. repens and D. immitis from bioptic material is expected to greatly enhance the chances of detecting human infections and to further clarify the role of the two parasites as pathogens of man. The possibility of routine identification of developing larvae in the vector will substantially improve the perspectives for epidemiological investigations, particularly in Southern European regions, such as Italy, where the two nematode species are largely sympatric.
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the occurrence of the filarial nematode Dirofilaria repens in canine hosts from maio island cape verde
Journal of Helminthology, 2017Co-Authors: Ricardo Marcos, C Genchi, Cristina Pereira, Joao P M C Maia, Maria Joao Santos, C Luzzago, Stefania Lauzi, Augusto M R Faustino, Pablo PuentepayoAbstract:The prevalence of canine Dirofilaria infection in Maio Island (Cape Verde) was analysed by serology, morphological and molecular identification of the parasite species. Blood and sera were collected from 150 dogs and 80 cats aged over 6 months from various localities of the island. DNA was extracted from blood and samples were screened by polymerase chain reaction (PCR) using microfilaria-specific primers. No Dirofilaria immitis was found in dogs while D. repens microfilariae were found in 5.3% of dogs and 6% were positive by PCR. The species identity was confirmed by sequencing of PCR products, which showed almost 100% homology with D. repens European sequences published in GenBank. No difference in Dirofilaria infection was observed between males and females or in dogs with different weights. However, older dogs and those from the western part of Maio Island were more frequently infected. No Dirofilaria was found in cats. This study represents the first evidence of D. repens in Cape Verde (West Africa) and highlights the need for implementing control measures and for a better surveillance of dirofilariosis in Africa.
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canine and human Dirofilaria infections in the balkan peninsula
Veterinary Parasitology, 2015Co-Authors: Suzana Tasicotasevic, Simona Gabrielli, Marija Trenkic Božinovic, C GenchiAbstract:Abstract Dirofilaria immitis and Dirofilaria repens infections are mosquito-borne diseases, mainly of dogs. Both parasites are zoonotic, and they sometimes cause serious infections in humans. The aim of this short review was to examine the situation in the Balkan Peninsula, from where it is not always easy to obtain suitable data, often reported in journals and other publications difficult to be retrieved and with poor or no visibility. The review included data from international and regional literature, doctoral theses, and conference proceedings.
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rapid differentiation of Dirofilaria immitis and Dirofilaria repens in canine peripheral blood by real time pcr coupled to high resolution melting analysis
Veterinary Parasitology, 2014Co-Authors: Francesca Albonico, C Genchi, Marco Genchi, G Gioia, Monica Loiacono, Michele MortarinoAbstract:Abstract Dirofilaria immitis and D. repens are the principal causative agents of canine filariosis and, although the number of dogs subjected to specific prevention is increasing, the prevalence of these parasites remains high in many areas of the world. The discrimination between the two Dirofilaria species using the classical diagnostic methods can be difficult and may lead to misdiagnosis especially on samples from areas where both Dirofilaria are present. Over the last years, several molecular methods with higher sensitivity and specificity compared to classical microscopy and ELISA assays were designed. Nevertheless, a need for simple, rapid, and cost-effective molecular protocols to accurately discriminate between D. immitis and D. repens still remains. High resolution melting analysis coupled to real-time PCR (real-time PCR–HRMA) is a widely used technique to target sequence polymorphisms of the same gene in different species without the need to perform DNA sequencing or to use species-specific probes. In this work, a fast and cost-effective real-time PCR–HRMA protocol to detect and differentiate simultaneously and unequivocally D. immitis and D. repens microfilarial DNA extracted from peripheral dog blood samples is described. The present method is simpler to use than most other DNA-based methods and provides comparable discrimination between the two sibling species.
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changing climate and changing vector borne disease distribution the example of Dirofilaria in europe
Veterinary Parasitology, 2011Co-Authors: C Genchi, Michele Mortarino, Laura Rinaldi, Giuseppe Cringoli, Giorgio Traldi, Marco GenchiAbstract:Abstract Climatic changes, together with an increase in the movement of dogs across Europe, have caused an increase in the geographical range of Dirofilaria infections. The present paper is focuses on northeastern European countries, where survey data have shown an increase of Dirofilaria repens infections both in animals and humans. A growing degree day-based forecast model has been developed to predict the occurrence. The model is based on evidence that there is a threshold of 14 °C below which Dirofilaria development will not proceed in mosquitoes, there is a requirement of 130 growing degree-days (GDDs) for larvae to reach infectivity, and there is a maximum life expectancy of 30 days for a mosquito vector. The output of this model predicted that the summer temperatures (with peaks in August) are sufficient to facilitate extrinsic incubation of Dirofilaria even at latitudes of 56°N and longitudes of 39°E. Despite the fact that both Dirofilaria immitis and D. repens have the same temperature requirement for extrinsic incubation in mosquitoes, empirical data has shown that D. repens is the main cause of Dirofilarial infections in both humans and animals. Clinical signs are absent in most canine infections with D. repens. Furthermore, diagnosis is problematic and in-clinic serological tests, such as those for D. immitis, do not exist. Therefore, most infections go undiagnosed, allowing the infection to spread undetected.
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highly sensitive multiplex pcr for simultaneous detection and discrimination of Dirofilaria immitis and Dirofilaria repens in canine peripheral blood
Veterinary Parasitology, 2010Co-Authors: G Gioia, C Genchi, Marco Genchi, L Lecova, E Ferri, Michele MortarinoAbstract:Dirofilaria immitis and Dirofilaria repens are the most common species of filarial nematodes described in the dogs with increasing spread into new geographical areas. The diagnosis of canine dirofilariosis is usually based upon the microscopical detection and identification of circulating microfilariae together with ELISA detection of serum circulating heartworm antigens or antibodies. The identification of the parasite species using the traditional approaches sometimes can be difficult and can lead to misdiagnosis especially on samples from areas where both Dirofilaria are present. In this paper we report a new molecular method based on single-step multiplex PCR to detect and differentiate simultaneously and unequivocally D. immitis and D. repens on DNA extracted from canine peripheral blood. The amplification was performed using a set of primers designed on a portion of the small subunit ribosomal RNA gene of the mitochondrion (12S rDNA). The single-step multiplex PCR here described ensured high (4 mf/ml) sensitivity and specificity with reduced cost and time saving. The multiplex PCR assay represents an additional tool for epidemiological studies and routine disease assessment in areas co-endemic for the two Dirofilaria species.