Multiple diagnostic tests demonstrate an increased risk of canine heartworm disease in northern Queensland, Australia

Background Canine heartworm (Dirofilaria immitis) is a life-threatening infection of dogs with a global distribution. Information on the prevalence of D. immitis and associated risk factors for canine heartworm antigen positivity—and thus disease—in Australia is scarce or outdated. The current reference method for D. immitis diagnosis in dogs is via the detection of heartworm antigen in blood using commercially available microwell-based enzyme-linked immunosorbent assays (ELISAs). Heat treatment of canine plasma prior to testing has been suggested to increase test sensitivity. The aim of the current study was to estimate the prevalence of D. immitis in dogs confined to shelters in Queensland, Australia. The impact of heat treatment on antigen test results was also assessed. Methods Blood samples (n = 166) were collected directly from dogs in seven shelters across Queensland (latitudinal span of approx. 1700 km) into EDTA blood collection tubes. A commercially available ELISA (DiroCHEK®) was used to detect canine heartworm antigen in untreated and heat-treated plasma. Whole blood was concurrently tested for the presence of microfilariae and D. immitis DNA using a modified Knott’s test and real-time PCR, respectively. Risk factors (age, gender, source, location) associated with the odds of positivity for canine heartworm were assessed using binary logistic regression models. Results A total of 16 dogs (9.6%; 95% confidence interval [CI]: 5.9–15.2%) were positive for canine heartworm based on combined test results. Heat treatment did not impact on the positivity of D. immitis antigen within samples (Cohen’s kappa = 0.98), but the optical density was significantly increased in paired plasma samples for D. immitis antigen-positive samples (Wilcoxon matched-pairs signed rank test, two-tailed P < 0.01). Location of the dog in a shelter in northern Queensland was the only risk factor significantly associated with the odds of a dog being more likely to be D. immitis antigen positive (odds ratio: 4.39; 95% CI: 1.26–13.51). All samples positive for the modified Knott’s test were also positive for D. immitis DNA by PCR. Conclusions This study demonstrated the presence of heartworm-positive dogs in shelters in Queensland, with positive animals significantly more likely to occur in northern Queensland than southern Queensland. Sustained testing for the presence of D. immitis microfilariae and antigen remain important diagnostic tools in areas with known and re-emerging canine heartworm activity. Graphical Abstract

Case Report: Molecular Detection of Dirofilaria repens in an Italian Patient after a Stay in Tanzania

A 35-year-old man was admitted to a hospital in the south of Italy because of a periocular nodule and subpalpebral edema. The patient reported having been stayed in Tanzania five months before. Hematologic parameters were within the normality range, the Acanthocheilonema viteae ELISA did not detect significant levels of antifilarial IgG, and no further symptoms were described. The surgical inspection of the nodule led to the isolation of two filarioid parasites, identified as Dirofilaria repens by SEM, and then by molecular assays. Knott’s test did not reveal microfilaremia, whereas loop-mediated isothermal amplification and PCR detected D. repens DNA. The patient was treated with doxycycline, and he was found no more positive at the follow-up.

Prevalence of Dirofilaria immitis in Dogs from Shelters in Vojvodina, Serbia

Background: Dirofilaria immitis is vector borne parasite of carnivores, with zoonotic potential, endemic in many parts of the world, including Europe. The aim of this study was to determine the prevalence of Dirofilaria immitis infection in dogs from shelters, especially compared to their lifestyle. Dogs living in shelters in Serbia may be at high risk of acquiring vector borne pathogens, mainly because most of them live outside in pens and backyards, in contact with vectors. Also, dogs in shelters are not always regularly treated against ectoparasites, thus, representing an easy feeding source for the vectors. The objective of this study was to determine the prevalence of Dirofilaria immitis infection in dogs from 5 shelters in South Bačka and Central Banat districts, in Autonomous Province of Vojvodina, Northern part of Serbia. Also, the objective was to compare the relation of infection with Dirofiaria immitis with age, sex, type of keeping the animals and preventive treatment in dogs.Materials, Methods & Results: Between May 2017 and October 2019, blood samples were collected from 336 randomly selected dogs from 5 shelters in 2 districts, South Bačka and Central Banat districts, in Autonomous Province of Vojvodina, Northern part of Serbia. The epidemiological survey has been conducted with all of the dogs involved in this research. The survey was designed to collect data about sex, age, lifestyles, food type, treatment against mosquitoes with insecticides and filarioid worms with macrocyclic lactones, regular testing for Dirofilaria infections. The presence of circulating microfilariae was examined using a modified Knott’s test. For the presence of circulating adult female Dirofilaria immitis antigen, serum samples were tested by commercially available enzyme-linked immunosorbent assay, which reacts to antigen of female Dirofilaria. In total, 336 dogs were examined for the presence of Dirofilaria immitis antigen. For that dog population which came from 5 shelters, total prevalence was 25.30%. Most of the positive findings were observed in a shelter where dogs lived exclusively outdoors in fenced yards in big groups and they were partly tested for heartworm infections from time to time. These dogs were not treated with macrocyclic lactones, against mosquitoes with insecticides or filarioid worms. The prevalence in this shelter was 56.36%. On the contrary to that, the lowest positive findings were detected in the shelter, where dogs were allowed to move freely between outside and indoors and they were also provided with accommodation indoors. These dogs have been regularly tested for Dirofilaria infections and treated against mosquitoes with insecticides and filarioid worms with macrocyclic lactones. In this shelter the seroprevalence was 7.69%. Microfilariae of Dirofilaria immitis were detected, by modified Knott’s test, in all of the antigen positive dog samples; except in 2 dogs from one shelter. Discussion: This study shows persistence of cardiopulmonary dirofiariosis in shelter dogs under different maintaining conditions. By comparing the data during the last 17 years, it can be stated that there is a constant increase of prevalence for Dirofilaria immitis in dogs in northern part of Serbia over the years. The results gained in this study are important from the veterinary point of view, but also from the Public Health point of view.

Usual or unusual presentations of Dirofilaria repens in two sibling dogs: a case report

This study describes two different manifestations of Dirofilaria repens infection in sibling dogs with microfilaremia. Dog 1, asymptomatic, harbored a gravid female of D. repens on the parietal layer of tunica vaginalis of one testicle and showed a marked circulating eosinophilia (3.3·103/μL). Both testicles were normal in shape and size without any gross lesions. Dog 2 had a pyotraumatic dermatitis. The cases were confirmed by PCR and sequencing. The sequences obtained showed 100% identity with those of D. repens isolated from human scrotum in Croatia. The treatment with moxidectin 2.5% and imidacloprid 10%/kg was effective in eliminating microfilariae after just one application, as demonstrated by negative modified Knott’s tests and PCR analyses of blood samples. This status was maintained during the post-treatment observation period. The classical localization of D. repens in dogs is in subcutaneous tissues, within nodules or free; however, it can also occur with some frequency in testicles, as described in humans. The infection can be associated with circulating eosinophilia or pyotraumatic dermatitis, as reported in this study. Thus, in endemic areas, it is advisable to carefully inspect the removed testicles at neutering since parasite localization can take place without any macroscopic changes. Moreover, in the case of circulating eosinophilia or pyotraumatic dermatitis, investigations should include modified Knott’s test and PCR to ensure that D. repens is not the cause of these alterations. Rapid and sensitive tests for the early detection of infected animals would help to prevent or limit the spread of this zoonosis.

Canine Dirofilariasis: A Case Report and Review of the Literature

Dirofilariasis is a disease of clinical importance in dogs. It is to this end that a case of a seven-year-old mongrel with dirofilariasis was presented to examine the unique features and presentation in the canine patient in question. The dog had clinical signs consistent with the disease. Further diagnostic tests were performed to establish the presence and severity of the disease and make an appropriate treatment plan. The Knott’s test revealed Dirofilaria species in the circulating blood and radiography showed right ventricular hypertrophy with pulmonary arterial enlargement and increased bronchial opacification. The treatment instituted was ivermectin therapy by subcutaneous injection every two weeks for six months, cardiac glycoside and antibiotics for 14 days. The clinical signs resolved after completion of the treatment. A general overview of heartworm infection in dogs was also done to update current knowledge of the disease. Though the risk of significant propagation of Dirofilaria immitis is considered low, with the climate change and international pet travel regulations, this emerging zoonosis remains a threat.

Comparison of Diagnostic Tools for the Detection of Dirofilaria immitis Infection in Dogs

In the last two decades, reports of canine heartworm (HW) infection have increased even in non-endemic areas, with a large variability in prevalence data due to the diagnostic strategy employed. This study evaluated the relative performance of two microtiter plate ELISA methods for the detection of HW antigen in determining the occurrence of Dirofilaria immitis in a dog population previously tested by the modified Knott’s test and SNAP 4Dx Plus test. The prevalence of this infection in the sheltered dog population (n = 363) from a high-risk area for HW infection was 44.4% according to the modified Knott’s test and 58.1% according to a point-of-care antigen ELISA. All serum samples were then evaluated by a microtiter plate ELISA test performed with and without immune complex dissociation (ICD). The prevalence increased from 56.5% to 79.6% following ICD, indicating a high proportion of samples with immune complexing. Comparing these results to that of the modified Knott’s test, the samples negative for microfilariae (mfs) and those positive only for D. repens mfs demonstrated the greatest increase in the proportion of positive results for D. immitis by ELISA following ICD. While the ICD method is not recommended for routine screening, it may be a valuable secondary strategy for identifying HW infections in dogs.

The current situation of canine dirofilariosis in Hungary

Between April and September 2017, blood samples were collected from 344 randomly selected dogs older than 1 year in 180 settlements of 19 counties in Hungary. The dogs lived exclusively outdoors, had never travelled and had neither been examined for Dirofilaria infection nor treated against mosquitoes with insecticides or/and filarioid worms with macrocyclic lactones. Dirofilaria infection was examined with a modified Knott’s test for microfilariae, DiroCHEK®, for the presence of D. immitis antigen, as well as by multiplex and conventional PCR. Altogether, 77 (22.4%) dogs living in 58 settlements of 17 counties were found to be infected with one or both Dirofilaria species based on the PCR techniques. Twenty-eight (8.1%) and 38 (11.1%) dogs were infected with D. immitis and D. repens, respectively. Coinfections were recorded in 11 samples (3.2%) collected in 11 locations of 8 counties. The results confirmed that both dirofilarioses are endemic in dogs and the eastern areas of the country are hyperendemic for heartworm disease. Temperature showed a significant association with the prevalence of D. immitis (OR 2.41, 95% CI 1.24–4.86, p = 0.012) but not with that of D. repens (OR 1.37, 95% CI 0.78–2.47, p = 0.286). The prevalence of neither D. immitis (OR 0.99, 95% CI 0.98–1.00, p = 0.213) nor D. repens (OR 1.01, 95% CI 0.99–1.01, p = 0.094) showed a significant correlation with precipitation. The number of yearly growing degree days (GDD) based on the lifecycle of Dirofilaria in mosquitoes ranged between 3.73 and 7.57 for the Hungarian districts. The GDD showed a significant positive association with the prevalence of D. immitis (OR 2.38, 95% CI 1.43–4.15, p = 0.001) and a non-significant positive relationship with that of D. repens (OR 1.25, 95% CI 0.83–1.95, p = 0.291).

Evaluation of production and quality traits in interspecific hybrids of ornamental pepper

ABSTRACT The cultivation of potted peppers as ornamental plants has increased considerably throughout the planet. In Brazil, this crop is more recent and still lacks cultivars. In order to increase the available variability and obtain new cultivars, the interspecific hybridization is very useful in add a desirable attribute that occurs in one species to another species, resulting in a new cultivar of agronomic interest. The objective of this study was to characterize parents and interspecific hybrids based on 27 quantitative traits and evaluate their genetic diversity by multivariate procedures. Parents of seven pepper genotypes were crossed, and, seven hybrids were generated. The experiment was conducted in a completely randomized design. Data were subjected to analysis of variance, and means were subsequently grouped by Scott-Knott’s method. Tocher’s method was utilized based on Mahalanobis distance, and the relative importance was evaluated by Singh’s method. The effects of treatment were significant by F test at 1 and 5% probability for all studied traits, except for anther length and titratable acidity. According to Scott-Knott’s test, the genotypes were grouped into two to eight classes. By Tocher’s method, the genotypes were separated into four groups. The first three canonical variables explained 92.02% of the total variance. By Singh method, fruit yield per plant was the trait that most contributed to the divergence explaining 21% of the total variance. The studied parents and hybrids diverged for the evaluated traits; however, there was difficulty in obtaining good interspecific hybrids with traits of importance, wherein combinations HS1×L7, L2×L6, and HS1×L2 met these requirements.

First report of heartworm (Dirofilaria immitis) infection in an imported dog in Lithuania

SummaryOver the past decade, increasing numbers of autochthonous cases of heartworm infection have been reported in the countries of Eastern Europe where previously only imported cases were described. In this report we have described the first clinical case of Dirofilaria immitis infection in an imported dog in Lithuania.In 2018, a 5-year-old male Spanish greyhound (Spanish galgo) was imported to Lithuania from southern Spain and referred to a small animal veterinary clinic in Vilnius for wellness screening. Circulating microfilariae and female antigens of D. immitis were detected using the Knott’s test and SNAP 4Dx Plus Test (IDEXX Laboratories, Portland, USA). The diagnosis was confirmed using molecular analysis. Treatment according to the guidelines recommended by the American Heartworm Society was applied. This is the first confirmed report of canine heartworm infection in an imported dog in Lithuania. Heartworm-infected dogs transported to North-Eastern Europe from endemic areas could act as microfilarial reservoirs for the local mosquito population, which could increase the risk of spreading the disease.