Human seroprevalence data indicate other factors than climatic conditions influencing dirofilariosis transmission in the Russian Federation.

The species of the genus Dirofilaria are filarial parasites causing zoonotic infections in humans with an increasing incidence in temperate and tropical areas of the world. Due to its classification as a vector-borne disease, the most important factors influencing dirofilariosis transmission are those related to climate, such as temperature and humidity. However, other factors linked with human behaviour can influence the distribution of the parasite. Although the Russian Federation could be considered as a non-suitable area for Dirofilaria spp. transmission due to its climatic conditions, one third of the human cases of dirofilariosis have been declared in this country. Here, seroepidemiological data on human dirofilariosis for five different regions distributed throughout the Russian Federation (Rostov, Moscow, Ekaterinburg, Yakutia and Khabarovsk) were obtained. A total of 940 serum samples from totally random donors living in these areas were analysed by enzyme-linked immunosorbent assay for the detection of anti-Dirofilaria immitis immunoglobulin G antibodies. Similar seroprevalence data ranging from 3.41% in Yakutia to 6.95% in Khabarovsk, with no significant correlation with climatic data of yearly average temperature and rainfall from these regions were found. These results suggest that other factors probably related to human behaviour, and not only climatic conditions, might be facilitating the spread of human dirofilariosis in these areas.

Ticks (Acari: Ixodidae) infesting dogs in Nigeria: epidemiological and public health implications.

Ticks are haematophagous arthropods that exert direct and indirect effects on their hosts. Their global importance as reservoirs and vectors of diseases of veterinary and public health importance is well recognized. However, the level of understanding of their role in disease epidemiology varies from one country to the other based on available data. Information on ticks infesting dogs across Nigeria and the public health significance is scarce. Therefore, this study aimed to provide information on ixodid ticks infesting dogs in Nigeria. Ticks were collected from 608 owned dogs presented to veterinary clinics and hospitals in 10 out of 36 states of Nigeria over a 14-month period and identified using taxonomic descriptions and morphological keys. In all, 1196 ticks belonging to three genera were identified. Rhipicephalus (including the subgenus Boophilus) ticks were collected from dogs from all the states surveyed and accounted for 95.2% of the ticks collected, followed by Haemaphysalis (3.7%) and Amblyomma species (1.2%). The brown dog tick, Rhipicephalus sanguineus sensu lato was the only tick identified in all the climatic zones of Nigeria. There is a statistically significant association between tick infection rate and rainy season, female animals, local and cross breed against exotic animals, total lack of control practice by dog owners, frequency of the control and with traditional methods of tick control but not the age of the dogs. The epidemiological and public health implications of these findings were discussed.

Numerous Fasciola plasminogen-binding proteins may underlie blood-brain barrier leakage and explain neurological disorder complexity and heterogeneity in the acute and chronic phases of human fascioliasis.

Human fascioliasis is a worldwide, pathogenic food-borne trematodiasis. Impressive clinical pictures comprising puzzling polymorphisms, manifestation multifocality, disease evolution changes, sequelae and mortality, have been reported in patients presenting with neurological, meningeal, neuropsychic and ocular disorders caused at distance by flukes infecting the liver. Proteomic and mass spectrometry analyses of the Fasciola hepatica excretome/secretome identified numerous, several new, plasminogen-binding proteins enhancing plasmin generation. This may underlie blood-brain barrier leakage whether by many simultaneously migrating, small-sized juvenile flukes in the acute phase, or by breakage of encapsulating formations triggered by single worm tracks in the chronic phase. Blood-brain barrier leakages may subsequently occur due to a fibrinolytic system-dependent mechanism involving plasmin-dependent generation of the proinflammatory peptide bradykinin and activation of bradykinin B2 receptors, after different plasminogen-binding protein agglomeration waves. Interactions between diverse parasitic situations and non-imbalancing fibrinolysis system alterations are for the first time proposed that explain the complexity, heterogeneity and timely variations of neurological disorders. Additionally, inflammation and dilation of blood vessels may be due to contact system-dependent generation bradykinin. This baseline allows for search of indicators to detect neurological risk in fascioliasis patients and experimental work on antifibrinolytic treatments or B2 receptor antagonists for preventing blood-brain barrier leakage.

Pathological, immunological and parasitological study of sheep vaccinated with the recombinant protein 14-3-3z and experimentally infected with Fasciola hepatica.

In this study, the immunogenicity and protective capacity of a new recombinant vaccine candidate, the rFh14-3-3z protein was analysed in sheep experimentally challenged with Fasciola hepatica, in terms of fluke burden, faecal egg counts, hepatic damage and humoral immune response. Three groups of 8 animals each were used for study, group 1 was immunised with the rFh14-3-3z in Montanide adjuvant, whereas group 2 and 3 remained as adjuvant control and infection control groups, respectively. The parasitological analysis showed that no significant reduction in fluke burden, fluke size and faecal egg counts was detected. The extent of hepatic damage was very similar between groups. Nonetheless, animals immunised with the rFh14-3-3z protein induced the development of specific IgG1 and IgG2, being the IgG1 the predominant antibody; which confirms the immunogenicity of this protein in sheep. This is the first report of the 14-3-3z proteins as vaccine against the infection with F. hepatica.

Current status of canine dirofilariosis in an endemic area of western Spain.

Since dirofilariosis caused by Dirofilaria immitis is a vector-borne disease, its distribution depends on environmental conditions as well as demographic factors and the management of pets by humans. In the province of Salamanca (west-central Spain) the disease has been known for many years, appearing in an area with extensive irrigated crops along the Tormes river. Because recent demographic changes have occurred in this area, the present study has been carried out with the aim of monitoring the distribution and prevalence of the disease in the canine population of this area. For that purpose, 191 dogs were analysed through antigen and microfilaria tests and geo-referenced in a map. The overall prevalence was 5.8%, although the disease was only present in dogs from municipalities with irrigated crops in which the prevalence was 16.7%. These results indicate that D. immitis continues to be present in the province of Salamanca, and that it is associated with the presence of irrigation, but with a clear decrease in the prevalence. Causes of the decrease in prevalence, as well as the potential zoonotic risk, are discussed.

Evaluation of cardiopulmonary biomarkers during classic adulticide treatment versus the American Heartworm Society recommended treatment protocol in dogs infected by Dirofilaria immitis.

Adulticide treatment of dogs with canine heartworm disease causes the death of the adult Dirofilaria immitis lodged in the vascular system of the host. During the death of the worms, pulmonary thromboembolisms (PTE), pulmonary inflammation, congestive heart failure, or renal disease are possible consequences. The aim of this study was to evaluate cardiopulmonary biomarkers and renal parameters during adulticide treatment of canine heartworm to compare the classic two-injection treatment protocol versus the American Heartworm Society (AHS) recommended protocol. Fourteen heartworm-infected dogs with high parasite burdens were divided in three groups and allocated to adulticide protocols as follows: Group A (n=5) was treated with the classic two-injection protocol; group B (n=5) was treated using the AHS recommended protocol, and group C (n=4) was treated as for group B but received diminishing anti-inflammatory doses of prednisone. To assess cardiorespiratory status, cardiac troponin I (cTnI), myoglobin, and D-dimer were measured. Renal function was evaluated by measuring urea, creatinine, and urine protein:creatinine (UP:C). Serum and urine samples were collected day 0 (day of diagnosis), 7 and 14 days after the first dose of adulticide, and 1 month after the last adulticide injection. Dogs that received classic treatment presented pathologic concentrations of D-dimer more frequently and showed higher average D-dimer levels, which may indicate the presence of more severe PTE. Group C showed the highest levels of D-dimer during treatment, which may be due to an exacerbation of PTE caused by the administration of prednisone. CTnI and myoglobin values remained above reference values in all groups during the study but reached the lowest values 1 month after the last injection. Levels of urea and creatinine were within normal ranges in all groups, and 28.5% of the dogs were proteinuric on day 0, progressing to better UP:C values at the end of the treatment, except in group A. The results of this study justify the treatment of canine heartworm disease using the AHS recommended adulticide guidelines and recommends re-evaluation of the role of glucocorticosteroids in the prevention and treatment of PTE.

Proteomic analysis of the somatic and surface compartments from Dirofilaria immitis adult worms.

Dirofilaria immitis (hearthworm) is a filarial roundworm transmitted by mosquitoes to different vertebrate hosts (dogs, cats and humans, among others), causing dirofilariosis. The adult worms reside in the pulmonary arteries affecting vessels and tissues and resulting in different pathological manifestations. Worms migrate to the heart and surrounding major vessels in heavy infections. Dirofilariosis can result in serious damage to affected hosts. In the last few years, a re-emergence of the disease driven by the climate change has been pointed out. Very recently, the knowledge at molecular level of this parasite has been extended by the published studies on its genome and transcriptome. Nevertheless, studies on the expression of defined protein sets in different parasite compartments and the corresponding role of those proteins in the host-parasite relationship have been relatively scarce to date. These include the description of the adult worm secretome, and some of the proteins eliciting humoural immune responses and those related with plasminogen binding in secreted and surface extracts of the parasite. Here, we investigate by proteomics the somatic and surface compartments of the D. immitis adult worm, adding new information on protein expression and localization that would facilitate a deeper understanding of the host-parasite relationships in dirofilariosis.

Utility of cardiac biomarkers during adulticide treatment of heartworm disease (Dirofilaria immitis) in dogs.

Heartworm disease (Dirofilaria immitis) is a parasitic disease of dogs and other carnivores, characterized by the presence of adult worms in the pulmonary arteries and right ventricle, leading to pulmonary hypertension which may progress to congestive heart failure. Cardiac biomarkers are biological parameters that can be objectively measured as indicators of pathological processes, or to assess the response to therapeutic interventions. To evaluate the myocardial damage during the adulticide treatment in 15 heartworm-infected dogs with ivermectin, doxycycline and melarsomine, measurements of cardiac troponin I (cTnI), myoglobin, MB isoenzyme of creatine kinase (CK-MB) and aspartate aminotransferase (AST) were carried out on days 0, 60 and afterwards weekly on days 67, 75, 82, 91, 106, 113 and 120. Dogs were divided by low parasite burden (n=9) and high parasite burden (n=6). On day 0, dogs with high worm burden showed increased cTnI concentrations (3.62 ± 4.78 ng/ml) while dogs with low worm burden had concentrations similar to those of healthy dogs (0.78 ± 0.22 ng/ml), CK-MB concentrations were increased only in dogs with high parasite burden as well (54.4 ± 54.2U/l) and 26.6% (4/15) of the dogs showed pathological concentrations of myoglobin. On day 91, most dogs showed pathological concentrations of myoglobin, CK-MB and AST, probably due to the myositis associated to the intramuscular injection of melarsomine. The rest of the measurements made in the study, the biomarkers concentrations were within normal values, except for cTnI in dogs with high parasite burden, which remained above reference concentrations for healthy dogs during all the study. The evaluation of cardiac biomarkers seems to be a helpful test in the assessment of the myocardium in dogs with heartworm disease during the adulticide treatment.

Variation of D-dimer values as assessment of pulmonary thromboembolism during adulticide treatment of heartworm disease in dogs.

Dirofilaria immitis adult worms lodged in the pulmonary arteries are the causative agent of heartworm disease, characterized by endarteritis and obstruction of the blood flow. Dying worms form embolic fragments which trigger thrombosis. Thus, pulmonary thromboembolism is an inevitable consequence of successful adulticide therapy. Clinical signs of pulmonary thromboembolisms are highly variable and non-specific, and its diagnosis is often difficult to obtain. The D-dimer assay is considered the marker of choice for dogs with a suspicion of pulmonary thromboembolism. The aim of the present study was to evaluate the variation of D-dimer and platelets in 15 heartworm-infected dogs during the adulticide treatment with ivermectin, doxycycline and melarsomine. Nine dogs were considered to have a low parasite burden and 6 had a high burden. Measurements were carried out on days 0, 60, and afterwards weekly on days 67, 75, 82, 91, 98, 106, 113, 120, and finally on day 271 after treatment. 40% of dogs showed pathological concentrations of D-dimer and 40% showed slight or mild thrombocytopenia on day 0. The levels of D-dimer decreased by day 60, probably due to the joint action of the ivermectin, doxycycline and exercise restriction. All dogs with high parasite burden showed elevated values of D-dimer on several occasions during the treatment, reaching the highest values the first and second week after the injections of melarsomine. Only 33.3% of the dogs with low parasite burden showed elevated D-dimer levels at some point during the treatment, and all of them showed undetectable levels from the third week after the first injection of melarsomine. On day 271, all dogs showed undetectable levels of D-dimer. There was no correlation between thrombocytopenia and levels of D-dimer during the treatment. The evaluation of serum D-dimer appears to be a supportive test in the assessment and monitoring of pulmonary thromboembolism in dogs with heartworm disease during the adulticide treatment.

D-dimer deposits in lungs and kidneys suggest its use as a marker in the clinical workup of dogs with heartworm (Dirofilaria immitis) disease.

It has been reported that dogs with heartworm disease (Dirofilaria immitis) show increased plasma levels of D-dimer, a fibrin degradation product present in the blood after a blood clot is degraded by fibrinolysis. In the present study the authors show that, in dogs with both experimental and natural infections with D. immitis, D-dimer deposits in lungs and kidneys are associated with pulmonary thromboembolism and microfilariemic status, as well as there was a clear association between increased plasma values of D-dimer and positive staining in immunohistochemistry. Results suggest that the monitoring of D-dimer levels in infected dogs could be useful in evaluating the presence of pulmonary thromboembolism in the lungs and that microfilariae may induce microthrombosis in kidneys, thus contributing to renal pathology.

Heartworm Disease (Dirofilaria immitis) and Their Vectors in Europe - New Distribution Trends.

Cardiopulmonary dirofilariasis is a cosmopolitan disease caused by Dirofilaria immitis, which affects mainly canids and felids. Moreover, it causes zoonotic infections, producing pulmonary dirofilariasis in humans. Heartworm disease is a vector-borne transmitted disease, thus transmission depends on the presence of competent mosquito species, which is directly related to favorable climate conditions for its development and survival. Cardiopulmonary dirofilariasis is mainly located in countries with temperate and tropical climates. Europe is one of the continents where animal dirofilariasis has been studied more extensively. In this article we review the current prevalence of canine and feline cardiopulmonary dirofilariasis in the European continent, the transmission vectors, the current changes in the distribution and the possible causes, though the analysis of the epidemiological studies carried out until 2001 and between 2002 and 2011. The highest prevalences have been observed in the southern European countries, which are considered historically endemic/hyperendemic countries. Studies carried out in the last 10 years suggest an expansion of cardiopulmonary dirofilariasis in dogs toward central and northern Europe. Several factors can exert an influence on the spreading of the disease, such as movement of infected animals, the introduction of new species of mosquitoes able to act as vectors, the climate change caused by the global warming, and development of human activity in new areas. Veterinary controls to prevent the spreading of this disease, programs of control of vectors, and adequate protocols of prevention of dirofilariasis in the susceptible species should be carried out.

Adult Dirofilaria immitis excretory/secretory antigens upregulate the production of prostaglandin E2 and downregulate monocyte transmigration in an "in vitro" model of vascular endothelial cell cultures.

Canine and feline cardiopulmonary dirofilariosis caused by Dirofilaria immitis is a chronic and potentially fatal disease. Adult worms live in the pulmonary arteries of infected immunocompetent hosts for years. The aim of the present study is the identification of the influence of the metabolic products (excretory/secretory antigens, DiE/S) of D. immitis on the vascular endothelial cells, because the vascular endothelium interplays in a direct manner with the parasite and their products. For this purpose, HAAE-1 vascular endothelial cells were treated with DiE/S, using non-treated cultures as negative controls. Significant increases in the COX-2, 5-LO expression and PGE(2) level were detected in the treated cells compared with the control cells. Moreover, DiE/S decreases monocyte transmigrations across vascular endothelial cell monolayers. Treatment with DiE/S does not have a cytotoxic effect and do not alter apoptosis, necrosis or cell cycle of vascular endothelial cells. These results suggest that the DiE/S stimulates the production of mediators and mechanisms that favor the survival of the parasite, in vascular endothelial cells, contributing to restrict vascular and lung damages in the infected host, without altering the basic physiologic processes of endothelial cells.

Zoonotic Dirofilaria immitis infections in a province of Northern Spain.

Dirofilaria immitis is the causal agent of canine and feline cardiopulmonary dirofilariasis. Moreover, the existence of canine dirofilariasis implies a risk for human populations living in an endemic area in which, the parasite can cause pulmonary dirofilariasis. The Spanish epidemiological situation is not well understood, lacking data from many central and Northern provinces. In our study, epidemiological data on canine and human dirofilariasis for La Rioja (Northern Spain) have been obtained for the first time. The overall prevalence of D. immitis in dogs was 12% (9% of patent and 3% of occult infections), being 11.6% the seroprevalence of human residents in this province. The geographic distribution of both canine and human D. immitis infections in La Rioja is restricted to humid and irrigated areas near the Ebro Valley River, being absent in the rest of the province where hills and mountains predominate.