Current State of Canine Heartworm in Portugal.

The favourable geo-climatic conditions in Portugal have made it highly conducive to the development of Dirofilaria immitis in dogs, leading to its identification as an endemic region. This nematode is rapidly spreading across Europe, particularly in northeastern countries. The objective of this study was to provide an updated assessment of the prevalence of this disease in Portuguese dogs, analysing the results in relation to epidemiological and geo-environmental factors, and to identify potential risk factors. A total of 1367 dogs from all continental and insular districts were included in the study and tested for D. immitis antigens. The overall prevalence was found to be 5.9%. It was observed that the disease is spreading northward, with previously unaffected districts now reporting cases, and that the prevalence in coastal districts exceeded that of inland ones. Notably, the Aveiro district exhibited a significant increase in D. immitis prevalence, while in certain districts such as Setúbal, Santarém, Madeira, or Faro, a stabilisation or decrease in prevalence was noted. Furthermore, outdoor and older dogs were found to be at a higher risk of infection. No positive cases were detected in the Azores. Most of the infected dogs were located in areas of high and medium risk of infection. This study underscores the importance of implementing pharmacological prophylaxis, vector control strategies, and public awareness programs to control the spread of this zoonotic disease.

Prediction and validation of potential transmission risk of Dirofilaria spp. infection in Serbia and its projection to 2080.

Animal and human dirofilariosis is a vector-borne zoonotic disease, being one of the most important diseases in Europe. In Serbia, there are extensive studies reporting the presence of Dirofilaria immitis and D. repens, mainly in the north of the country, where the human population is concentrated and where there is a presence of culicid mosquitoes that transmit the disease. Ecological niche modeling (ENM) has proven to be a very good tool to predict the appearance of parasitosis in very diverse areas, with distant orography and climatologies at a local, continental, and global level. Taking these factors into account, the objective of this study was to develop an environmental model for Serbia that reflects the suitability of the ecological niche for the risk of infection with Dirofilaria spp. with which the predictive power of existing studies is improved. A wide set of variables related to the transmission of the parasite were used. The potential number of generations of D. immitis and the ecological niche modeling method (ENM) were used to estimate the potential distribution of suitable habitats for Culex pipiens. The highest probability of infection risk was located in the north of the country, and the lowest in the southern regions, where there is more orographic relief and less human activity. The model was corroborated with the location of D. immitis-infected dogs, with 89.28% of the country having a high probability of infection. In addition, it was observed that the percentage of territory with optimal habitat for Culex spp. will increase significantly between now and 2080. This new model can be used as a tool in the control and prevention of heartworm disease in Serbia, due to its high predictive power, and will serve to alert veterinary and health personnel of the presence of the disease in the animal and human population, respectively.

Ecological niche modeling analysis (Cx. pipiens), potential risk and projection of Dirofilaria spp. infection in Greece.

Vector-borne diseases continue to increase worldwide. Dirofilariosis is one of the most common vector-borne zoonotic diseases, mainly caused by Dirofilaria spp. (D. immitis and D. repens) and spread by culicid mosquitoes of different species. Greece is one of the countries in southern Europe where it is traditionally endemic, and its distribution is not homogeneous. The aim of this study was to develop an environmental model for Greece that reflects the suitability of the ecological niche for Dirofilaria spp. infection risk and its projection until 2080. For this purpose, we used the potential distribution of suitable habitats for Culex pipiens calculated using an ecological niche model (ENM) and the potential number of generations of Dirofilaria spp. The ecological niche model of Cx. pipiens in Greece showed good predictive power (AUC=0.897) with the parasite at a resolution of 1 km2. The variables that contributed most to the model were mean annual temperature, rivers and human footprint. The highest risk of infection was found in coastal areas and in riverside areas of the main river basins, as well as in irrigated areas of the mainland and peninsular regions and in the whole territory of island areas, and the lowest risk was found in areas of higher altitude. A positive relationship was found between the risk of dirofilariosis and the location of infected dogs, with 86.65% located in very high and high risk areas. In 2080, the percentage of territory gained by Cx. pipiens will increase by 261.52%. This model provides a high predictive value, predicted presence, and risk of Dirofilaria spp. infection and can serve as a tool for the management and control of this disease.

Assessment Heartworm Disease in the Canary Islands (Spain): Risk of Transmission in a Hyperendemic Area by Ecological Niche Modeling and Its Future Projection.

Heartworm disease is a vector-borne zoonotic disease caused by Dirofilaria immitis. The Canary Islands (Spain), geolocated close to the coast of Western Sahara, is an archipelago considered hyperendemic where the average prevalence in domestic dogs is high, heterogeneous, and non-uniform. In addition, Culex theileri has been reported as a vector of the disease on two of the most populated islands. Our aim was to develop a more accurate transmission risk model for dirofilariosis for the Canary Islands. For this purpose, we used different variables related to parasite transmission; the potential distribution of suitable habitats for Culex spp. was calculated using the ecological niche model (ENM) and the potential number of generations of D. immitis. The resulting model was validated with the geolocation of D. immitis-infected dogs from all islands. In addition, the impact of possible future climatic conditions was estimated. There is a risk of transmission on all islands, being high in coastal areas, moderate in midland areas, and minimal in higher altitude areas. Most of the dogs infected with D. immitis were geolocated in areas with a high risk of transmission. In 2080, the percentage of territory that will have been gained by Culex spp. is small (5.02%), although it will occur toward the midlands from coastal areas. This new model provides a high predictive power for the study of cardiopulmonary dirofilariosis in the Canary Islands, as a hyperendemic area of the disease, and can be used as a tool for its prevention and control.

Current Risk of Dirofilariosis Transmission in the Iberian Peninsula (Spain and Portugal) and the Balearic Islands (Spain) and Its Future Projection under Climate Change Scenarios.

Dirofilariosis is a vector-borne zoonotic disease whose distribution is linked to the presence of culicid mosquitoes. Spain and Portugal are considered endemic countries; however, the distribution of dirofilariosis is not uniform. Our aim was to develop a more accurate risk model of dirofilariosis transmission for the Iberian Peninsula (Spain and Portugal) and the Balearic Islands (Spain). To do this, we used a set of key variables related to parasite transmission: the potential distribution of suitable habitats for Culex pipiens calculated via an ecological niche model (ENM) and the potential number of Dirofilaria spp. generations. The resulting model was validated with the prevalence and geolocation of D. immitis-infected dogs from all provinces and districts. In addition, the impact of possible future climatic conditions was estimated. A quantitative estimate of the risk of infection by Dirofilaria spp. was obtained at a resolution of 1 km2. The entire analyzed territory was susceptible to contact with the parasite. The highest risk of infection was found throughout the eastern coastal strip and the south of the Iberian Peninsula and the Balearic Islands, as well as in the areas surrounding the basins of the main rivers, and the lowest risk was located in the higher-altitude areas. We found a robust and positive relationship between the risk of dirofilariosis and the observed prevalence of infested dogs in the study area (ß ± SE = 3.32 ± 1.43 p < 0.05). In 2080, the percentage of territory gain for Cx. pipiens will increase to 49.98%, which will increase the risk of infection. This new model provides a high predictive value for the current and predicted presence and risk and can serve as a tool for the management and control of dirofilariosis.

Involvement of the excretory/secretory and surface-associated antigens of Dirofilaria immitis adult worms in the angiogenic response in an in-vitro endothelial cell model.

Angiogenesis is a process by which new vessels are formed from pre-existing ones when the physiological conditions of the vascular endothelium are altered. Heartworm disease, caused by Dirofilaria immitis, causes changes in the vascular endothelium of the pulmonary arteries due to obstruction, friction, and hypoxia. The aim of this study was to analyze whether the excretory/secretory and surface-associated antigens of adult worms interact and modulates the angiogenic mechanism, viable cell number and cell migration, as well as the formation of pseudo-capillaries. Cultures of human vascular endothelial cells (HUVECs) stimulated with excretory/secretory antigens (DiES), surface-associated antigens (Cut) from D. immitis adult worms, VEFG-A (Vascular Endothelial Growth Factor A), as well as DiES+VEFG-A and Cut+VEFG-A were used. The production of VEFG-A and other proangiogenic [soluble VEFGR-2 (sVEFGR-2), membrane Endoglin (mEndoglin)] and antiangiogenic [VEFGR-1/soluble Flt (sFlt), soluble Endoglin (sEndoglin)] molecules was assessed using commercial ELISA kits. Cell viability was analyzed by live cell count and cytotoxicity assays by a commercial kit. In addition, viable cell number by MTT-based assay, cell migration by wound-healing assay carrying out scratched wounds, and the capacity of pseudo-capillary formation to analyze cell connections and cell groups in Matrigel cell cultures, were evaluated. In all cases, non­stimulated cultures were used as controls. DiES+VEFG-A and Cut+VEFG-A significantly increased the production of VEFG-A and sVEFGR-2, and only Cut+VEFG-A significantly increased the production of VEFGR-1/sFlt compared to other groups and non-stimulated cultures. Moreover, only DiES+VEFG-A produced a significant increase in viable cell number and significant decrease cell migration, as well as in the organization and number of cell connections. Excretory/secretory and surface-associated antigens of adult D. immitis activated the angiogenic mechanism by mainly stimulating the synthesis of proangiogenic factors, and only excretory/secretory antigens increased viable cell number, activated cell migration and the formation of pseudo-capillaries. These processes could lead to vascular endothelial remodeling of the infected host and favor the long-term survival of the parasite.

Effect of somatic antigens of Dirofilaria repens adult worms on angiogenesis, cell proliferation and migration and pseudo-capillary formation in human endothelial cells.

BACKGROUND: Angiogenesis is defined as the formation of new vessels by sprouting of endothelial cells from pre-existing vessels in response to stimuli, such as hypoxia or inflammation. Subcutaneous dirofilariasis, caused by Dirofilaria repens, is a zoonotic disease characterized by the formation of subcutaneous nodules with the presence of at least one encapsulated worm, showing perivascular vascularization around it. The aim of this study is to analyze whether the somatic antigen of adult D. repens worms interacts with and modulates the angiogenic mechanism, cell proliferation and migration, and formation of pseudo-capillaries. METHODS: The expression of VEGF-A, VEGFR-1/sFlt, VEGFR-2, mEnd and sEnd in cultures of human vascular endothelial cells stimulated with somatic antigen of adult worms of D. repens (DrSA), vascular endothelial growth factor (VEGF) and DrSA + VEGF were evaluated by using ELISA commercial kits. Cellular viability was analyzed by live cell count, cytotoxicity assays by using a commercial kit, cell proliferation by MTT-based assay, cell migration by wound-healing assay carried out by scratching wounds and capacity of formation of pseudo-capillaries analyzing cell connections and cell groups in Matrigel cell cultures. In all cases unstimulated cultures were used as controls. RESULTS: DrSA + VEGF significantly increased the expression of VEGF-A, VEGFR-2 and mEndoglin compared to other groups and unstimulated cultures. Moreover, DrSA + VEGF produced cell proliferation and migration and increased the formation of pseudo-capillaries. CONCLUSIONS: Somatic antigen of adult D. repens worms activated the proangiogenic mechanism, cell proliferation and cell migration as well as formation of pseudo-capillaries in this in vitro human endothelial cell model. These processes could be related to the survival of adult D. repens in subcutaneous nodules in infected hosts.

Comprehensive Map of Canine Angiostrongylosis in Dogs in Spain.

Canine angiostrongylosis is an emerging disease caused by Angiostrongylus vasorum, mainly affecting wild carnivores and dogs. In Spain, there are studies reporting infections in foxes, wolves, and badgers in different regions of the country. However, there are hardly any publications on its prevalence in dogs. The aim of this study was to complete and update the epidemiologic map of A. vasorum in dogs in Spain. A total of 5619 canine blood samples from all autonomous cities and provinces of Spain were collected and tested for the presence of circulating A. vasorum antigens. The overall apparent prevalence of canine A. vasorum infection in Spain was 1.39%. No significant differences were found for sex or age, but significant differences between outdoor and indoor/outdoor dogs were found. A high prevalence was also observed in the northern third of the country, where an oceanic climate prevails, being humid and rainy and where abundant vegetation can be found, thus favoring the proliferation of intermediate hosts. The results suggest that A. vasorum canine infections are heterogeneously present in a large part of the territory, demonstrating its expansion throughout the country, and therefore, awareness and prevention campaigns for this disease should be promoted.

What Has Happened to Heartworm Disease in Europe in the Last 10 Years?

Heartworm disease caused by Dirofilaria immitis is a vector-borne disease that affects canids and felids, both domestic and wild, throughout the world. It is a chronic disease which causes vascular damage in pulmonary arteries, and in advanced stages, the presence of pulmonary hypertension and right-sided congestive heart failure can be evidenced. Moreover, pulmonary thromboembolism is caused by the death of the worms, which can be lethal for the infected animal. Furthermore, it is the causative agent of human pulmonary dirofilariosis, being a zoonotic disease. The aim of this review was to update the current epidemiological situation of heartworm in Europe in dogs, cats, wild animals, and vectors insects, and to analyse the factors that may have contributed to the continuous spread of the disease in the last decade (2012-2021). In Europe, the disease has extended to eastern countries, being currently endemic in countries where previously only isolated or imported cases were reported. Furthermore, its prevalence has continued to increase in southern countries, traditionally endemic. This distribution trends and changes are influenced by several factors which are discussed in this review, such as the climate changes, presence of vectors in new areas, the appearance of new competent vector species in the continent, increased movement of pets that travelled to or originated from endemic countries, the urbanisation of rural areas leading to the formation of so-called "heat islands", or the creation of extensive areas of irrigated crops. The continuous expansion of D. immitis must be monitored, and measures adapted to the situation of each country must be carried out for adequate control.

Seroprevalence of Feline Heartworm in Spain: Completing the Epidemiological Puzzle of a Neglected Disease in the Cat.

Feline heartworm is a vector-borne zoonotic disease caused by Dirofilaria immitis. It is a cosmopolitan disease that is continuously expanding. Spain is considered an endemic country; however, although there are many published studies in dogs, feline heartworm has been poorly studied in this country. Thus, the objective was to analyze the exposure to D. immitis throughout Spain to complete the epidemiological map in the feline species. For this, 6,588 feline serum samples were analyzed for the presence of D. immitis antigens and antibodies against D. immitis and Wolbachia. The results were analyzed according to sex, age, breed, habitat, origin (owned or shelter cats), presence of clinical signs, use of preventive, location and climatology. The results showed a prevalence of 0.5% and a seroprevalence of 9.4%. The highest antibody seroprevalences were reported in the Canary Islands and the Balearic Islands (19.2 and 16%, respectively), as well as in the autonomous communities located on the Mediterranean coast (9.2-11.2%). Seropositive cats were found in both indoor and outdoor cats, and from 6 months of age. Furthermore, only 5.8% of cats received regular prophylactic treatment. The results show that feline dirofilariasis is widely distributed throughout the national territory and corroborate that, where infected dogs are present, there are cats exposed to the parasite. It is necessary to implement efficient awareness and prophylaxis measures to control the incidence and expansion of feline heartworm in Spain.

Expansion of Canine Heartworm in Spain.

The climate of Spain has favourable characteristics for the development of D. immitis in dogs, being an endemic country. Given that vector-borne diseases are spreading rapidly through Europe, due to factors such as climate change, the expansion of vectors and the increased mobility of reservoir animals, the aim was to update the epidemiology of heartworm in dogs and analyse the results based on climate and other epidemiological and geo-environmental factors. To this aim, 9543 blood samples from dogs from all provinces and autonomous cities of Spain were analysed for the detection of antigens of D. immitis, obtaining a prevalence of 6.47%. The northwestern and southern provinces showed the highest prevalences, as well as in the Balearic and Canary Islands. Prevalences were higher in dogs outdoors. Furthermore, most of the positive dogs were found in regions with high humidity and water availability. This study shows, for the first time, positive cases in provinces and islands where no cases had previously been described and demonstrates the continuous expansion and consolidation of heartworm in Spain. Considering its zoonotic character, the implementation of control and awareness programmes for the prevention of D. immitis in pets is necessary.

Serological Survey of Canine Vector-Borne Infections in North-Center Spain.

Various factors are currently causing an increase in vector-borne parasitic diseases at a global scale; among them, some stand out, such as climatic disturbances derived from global change, the increase in movements of reservoir animals, or changes in land made by human activity. In the European continent, there have been an increasing number of epidemiological studies focused on the detection of these diseases, especially in dogs. In Spain, there are few epidemiological studies focused on the evaluation of the biotic and abiotic factors that may influence the distribution, such as climatic zones, orography, or presence of water reservoirs. The aim of this study was to analyze the prevalence and distribution of several canine vector-borne diseases caused by Dirofilaria immitis, Leishmania infantum, Anaplasma platys, and Ehrlichia canis in the autonomous community of Castilla y León, the largest region of the Iberian Peninsula, providing a geospatial approach based on a geographic information system (GIS) analysis. Blood from a total of 1,475 domestic dogs from the nine provinces of Castilla y León were analyzed. Also, a GIS analysis of the sample locations was carried out, taking into account the most important predictor variables. The prevalence in dogs infected by D. immitis was 7.19%, and the seroprevalence by L. infantum was 4.61 and 1.56% for A. platys and E. canis. Most of the infected animals were located in areas with stagnant water, irrigated agriculture, or riverbanks, always close to forest and woodland vegetation. These results indicate that dogs living in Castilla y León should take prophylactic measures to avoid infections.