Immitis Infection Research Articles

Immunoblotting analysis of somatic components of Dirofilaria immitis.

SDS-PAGE analysis of Dirofilaria immitis extracts demonstrated the complexicity of protein components of microfilariae, similar to that of adult male and female worms. Immunoblotting analysis using sera from microfilaremic and amicrofilaremic dogs with D. immitis infection suggest that antigenic components in the low molecular weight region may be related to the anti-microfilarial mechanism of the host.

Comparison of three diagnostic tests for Dirofilaria immitis in a low-incidence area.

was obtained, tests were repeated by a different technologist. Microfilaria identification using the alkaline phosphatase technique was done to differentiate between D. immitis and Dipetalonema reconditum on all samples in which microfilariae were observed. Clinical follow-up was obtained for 11 of the 12 dogs that had any positive test result by reviewing the medical record or discussing the case with the clinician. The results of the laboratory tests are summarized in Table 1. Samples from 1,347/1,359 dogs (99.12% of samples submitted) were negative for microfilariae and adult D. immitis antigen using both commercial kits. Microfilariae, subsequently identified as D. immitis, were observed in blood samples from 7 dogs. Four of these dogs also had adult D. immitis antigen detected by both kits. Two microfilaremic dogs were negative for D. immitis adult antigen with both kits. Blood from 1 dog that was positive for microfilariae was serologically positive for adult antigen with only 1 of the commercial kits. Five dogs were serologically positive for adult D. immitis antigen but negative for microfilariae. The 5 dogs that had positive tests for adult D. immitis antigen but negative filter tests were considered otherwise healthy. They had no evidence of circulatory or respiratory compromise. Adulticide treatment was recommended for each of these dogs, but owners chose to administer ivermectin or milbemycin oxime monthly to their pets. Owners were advised to observe their dogs carefully and to restrict their pets’ exercise for at least 2 days following the administration of the monthly heartworm preventative. Adverse reactions were not reported in any of these dogs. Six of the 7 dogs that tested positive for D. immitis microfilariae underwent treatment for the elimination of adult D. immitis and microfilariae, regardless of the results of the adult antigen test. Additional information could not be obtained for 1 dog that was positive on all 3 tests. Treatment in 1 dog that had positive results with all 3 tests was initiated as an emergency procedure when the dog developed acute respiratory distress. Two dogs that were microfilaremic were adult antigen negative using both kits, and 1 microfilaremic dog was antigen negative with 1 kit but antigen positive with the other. Such results are likely to occur if the adult worm burden is low. In these cases, the sensitivity of the tests for detection of adult antigen is decreased. 2,3,6,8,9 Based on physical examination and radiographs, the clinicians strongly suspected that 2 of these 3 dogs had few adult D. immitis in their right ventricles, and the dogs were treated with adulticides before beginning preventative therapy. The presence of adult D. immitis infection in the right ventricle in the third dog was questionable, yet the owner preferred to have the dog treated with adulticide before preventative therapy was begun. Although the number of infected dogs detected in this study was small, infection in 5 of 12 animals would have gone undetected if only a filter test, accompanied by acid phos

Range of Heartworm (Dirofilaria immitis) Infection in Montana Dogs

The serological presence of heartworm in dogs of Montana was studied in a 3-yr survey. Serum samples were provided by veterinary practitioners throughout Montana or were submitted to the state diagnostic laboratory. Sera from 3,490 dogs were tested using an enzyme-linked immunosorbent assay (ELISA) for circulating adult heartworm antigen. Twenty-four serum samples were positive for heartworm antigen. Two were from dogs that had never been outside the state. Nineteen additional positive dogs, 7 of which presumably had never been out of the state, were reported through correspondence with veterinary practitioners. Because suitable vectors for Dirofilaria immitis exist in Montana, there is potential for propagation of heartworm.

Variation of mosquito (Diptera: Culicidae) relative abundance and Dirofilaria immitis (Nematoda: Filarioidea) vector potential in coastal North Carolina.

At an enzootic focus of Dirofilaria immitis in coastal North Carolina, mosquito populations were sampled June-September 1985 and on several occasions during August-October 1986 and June-August 1987, to identify local vectors and to determine relative abundance and D. immitis infection rates. Predominant species collected were Anopheles bradleyi King (66.6%), Culex salinarius Coquillett (15.9%), Aedes taeniorhynchus (Wiedemann) (8.2%), and Aedes sollicitans (Walker) (4.9%). Population abundance varied within and among seasons. D. immitis infection was found in An. bradleyi (1.2%), Ae. taeniorhynchus (0.9%), and Ae. sollicitans (0.9%). Of infected An. bradleyi, 88% were collected June-2 July, whereas the highest number of infected Ae. taeniorhynchus and Ae. sollicitans were found during mid-July 1985. Ae. sollicitans and Ae. taeniorhynchus were the only infected species found during 1986 and 1987. However, based on overall relative abundance and infective rate, An. bradleyi appeared to be the primary vector of D. immitis in the study area.

Prophylactic effect of milbemycin oxime against Dirofilaria immitis infection in dogs: optimum dose and administration time.

To examine the prophylactic efficacy of milbemycin oxime against Dirofilaria immitis infection, 42 filaria-free beagles were inoculated with 100 infective larvae of the parasite. Milbemycin oxime was administered orally at doses of 0.05 to 1.0 mg/kg body weight one month after inoculation, and at a single dose of 0.25 mg/kg body weight on days 1 to 90 after inoculation. All dogs were euthanized and autopsied to evaluate the prophylactic effect of the drug against D. immitis infection 6 months after inoculation. It was confirmed that a dose of 0.25 mg/kg of milbemycin oxime completely-protected the infection 15 to 60 days after infection.

Immunoglobulin E recognition of Dirofilaria immitis antigens is more specific than immunoglobulin G

The chronological development of the serum IgE and IgG response to microfilaria, third and fourth stage larvae, and male and female adult Dirofilaria immitis was examined by enzyme-linked immunosorbent assay (ELISA) and enzyme-linked immunoelectrotransfer blot (EITB). Dirofilaria immitis-specific IgE and IgG levels peaked 16–18 weeks post-infection after increasing in response to the fourth larval molt. Specific IgG levels plateaued after patency, while IgE continued to decline. The use of ammonium sulfate cut sera showed there was no quenching or blocking of IgE binding by IgG in the ELISA and EITB methods used in this study. IgE-specific EITB showed 30–49 bands for the five respective extracts that were identified by M r or relative mobility. Eighty-five to 100 bands were visualized by IgG-specific EITB for the same five extracts. The isotype-specific ELISA and EITB were shown to be closely related by significant correlations ( P < 0.0001) between S/N ratios and the number of bands found on blots. The isotype-specific EITB bands non-specifically recognized were greater in size than 21 kDa for IgG and 45 kDa for IgE. Recognition of bands changed over time with some bands being recognized only by prepatent sera. Ten antigen bands of seven M r were consistently and specifically recognized by IgE in the five-stage extracts by sera from prepatent and patent infections; only one such M r at 13.9 kDa, was described for IgG. A potentially diagnostic 31.9 kDa antigen band was identified on the IgE-specific EITB of D. immitis female extract and was shown to be recognized by IgE in sera from all infected dogs at all time points examined from 2 weeks until 1 year post-inoculation. Overall, IgE reactivity was more specific for D. immitis infections than IgG reactivity.

Dirofilaria immitis: A Rare, Increasing Cause of Pulmonary Nodules

Dirofilariasis is an unusual but increasing cause of solitary pulmonary nodules. In this study, we reviewed the entire experience with dirofilariasis at our institution. Five such patients were identified. In all patients, the Dirofilaria immitis infection manifested as a solitary pulmonary nodule, and all patients underwent thoracotomy for diagnosis. None required systemic treatment. D. immitis is found in dog, cat, wolf, coyote, and fox populations throughout the United States, but the highest concentrations have been noted in the eastern, southeastern, and southern coastal states. The distribution of human cases of D. immitis infection has a similar pattern. Pulmonary dirofilariasis should be included in the differential diagnosis of peripheral noncalcified pulmonary nodules, especially in endemic areas.

Natural infections of Dirofilaria immitis in Aedes (Stegomyia) polynesiensis and Aedes (Finlaya) samoanus and their implication in human health in Samoa

Dirofilaria immitis infections were observed in Aedes polynesiensis and Ae. samoanus in Samoa, together with Wuchereria bancrofti infections, in a study on sub-periodic bancroftian filariasis during 1978–1980. In the 4 indicator villages, the infection rate in Ae. polynesiensis was 0·46% and the infective rate 0·09% (15 223 mosquitoes were dissected). The infection rate in Ae. samoanus was 0·20% and the infective rate 0·08% (10 089 dissected). In 45 selected villages throughout the country, Ae. polynesiensis infection and infective rates were 0·92% and 0·29% (7575 dissected) and the rates for Ae. samoanus were 0·21% and 0·07% (9093 dissected). Infection with D. immitis was comparable in degree and distribution to that with W. bancrofti. There was a steady and consistent exchange of parasites between the human and canine populations, creating conditions favourable for human dirofilariasis in Samoa. Clinicians are warned against this probable human infection.

Serological Survey for Heartworm (Dirofilaria immitis) in Dogs in Washington

The serologic prevalence of Dirofilaria immitis infection in 601 dogs in Washington was investigated in 1989-1990. Blood samples for serum were obtained from dogs approximately 2 yr of age or older in humane society shelters (n = 392) or veterinary clinics (n = 209). Serum samples were tested for heartworm infection using an enzyme-linked immunosorbent assay antigen test. Three (0.5%) dogs were positive for D. immitis infection, all of which were born and lived several years in states other than Washington. Heartworms have not been detected during the last 9 yr in the 1,203 dogs examined at the Washington Animal Disease Diagnostic Laboratory, Pullman, Washington. Consequently, the likelihood of indigenous heartworm infections in dogs in Washington remains low at this time.

Primary diagnosis of disseminated fungal disease by fine-needle aspiration of soft-tissue lesions

Two cases of disseminated fungal infections diagnosed by fine-needle aspiration of soft-tissue masses are presented. Both patients were initially treated on an outpatient basis for a presumed musculoskeletal disorder before the diagnostic biopsy was obtained. The first patient apparently contracted Coccidioides immitis infection while residing in Arizona, but the diagnosis was rendered on an aspirate of a buttock mass performed in Boston. The second patient, a non-insulin-dependent diabetic, was diagnosed with a cryptococcal abscess on an aspirate of a shoulder mass performed in the emergency room. The diagnosis of coccidioidomycosis was confirmed by culture and that of Cryptococcus by serology. Fungal infection was clinically unsuspected in both patients and the cytologic diagnosis facilitated rapid implementation of therapy.

The Maternal Immune Response in Coccidioidomycosis: Is Pregnancy a Risk Factor for Serious Infection?

Seven subjects with prior coccidioidal disease and three with active Coccidioides immitis infection during their first trimester were studied during pregnancy and postpartum to determine their general and antigen-specific cell-mediated immune status. All ten were white and carried their pregnancies to term without incident. Decreases in total lymphocytes and T-helper and T-suppressor subsets were noted during the third trimester, presumably secondary to an increase in plasma volume. Lymphocyte responses to the mitogens phytohemagglutinin, concanavalin A, and pokeweed were mildly decreased late in pregnancy, with significant intrasubject and intersubject variation. Responses to tetanus antigen were consistently and significantly lower as pregnancy progressed, rising above first trimester levels by 12 weeks postpartum. A similar pattern of response was noted with spherulin antigen for the seven subjects with previously demonstrated coccidioidal immunity. The three subjects with active coccidioidomycosis either failed to mount a significant spherulin immune response or demonstrated an early response that fell as pregnancy progressed. This antigen-specific immune suppression continued for up to 16 months postpartum despite the fact that there was no clinical evidence of coccidioidal activity beyond the first trimester. Thus, while all three completed pregnancy without complication, the data suggest that significantly increased maternal risk may be present when active coccidioidomycosis and pregnancy occur together. This risk may be greatest among darker-skinned individuals who become infected during the latter half of pregnancy.

Review of human and animal cases of coccidioidomycosis diagnosed in Canada

The first Canadian case of coccidioidomycosis in a human was reported in 1952 and 11 more cases since then. This study provides details of other cases of coccidioidomycosis that have been diagnosed in Canada. Based on clinical details, isolation of Coccidioides immitis, detection of a specific antibody (F band) for coccidioidomycosis by macro- or microimmunodiffusion tests, concurrently used with the complement fixation procedure, and histopathological findings, 116 more cases of this disease were verified. The great majority (94%) of these cases were diagnosed in the western Canadian provinces of British Columbia, Alberta, Saskatchewan and Manitoba, and the others in Quebec, Ontario and Nova Scotia (5, 1, and 1 cases, respectively). Available information indicates that the C. immitis infections were contracted during visits to endemic areas in the United States (Arizona, California and New Mexico), Mexico, and Bolivia. Pulmonary infections were the most common type of coccidioidomycosis (93%) followed by the disseminated or meningeal types C. immitis infections occurred in individuals with or without predisposing factor(s) and were more common in males than in females. The exoantigen procedure was very useful and reliable in the accurate and rapid identification of suspected C. immitis isolates. Two cases of coccidioidomycosis were reported in animals in Ontario, Canada.

Seronegative disseminated coccidioidomycosis in patients with HIV infection

Serologic testing for complement-fixing antibodies to Coccidioides immitis is commonly employed to assist in the diagnosis and management of this infection, but its usefulness in an HIV-coinfected population is unknown. In this study we reviewed all the mycologically or histologically proven cases of disseminated C. immitis infection after 1982. Disseminated C. immitis and proven HIV infection were present in eight patients. We performed serum complement-fixing antibody titers on all eight patients, six of whom gave positive tests, while two patients (25%) gave repeatedly negative results despite widely disseminated disease. We conclude that histopathology and culture remain the most reliable methods for the diagnosis of disseminated coccidioidomycosis in the HIV-infected host.

Entomological Aspects of Human Dirofilariasis and Dirofilaria immitis Infections

Entomological Aspects of Human Dirofilariasis and Dirofilaria immitis Infections

Experimental Dirofilaria immitis-Associated Glomerulonephritis Induced in Part by in Situ Formation of Immune Complexes in the Glomerular Capillary Wall

Eight dogs were immunized with an aqueous-soluble extract of adult Dirofilaria immitis. Subsequent to at least 7-fold increases in antibody titer, the left renal artery of each dog was infused with 6 mg of D. immitis antigen. Fourteen days after infusion, the left kidney was compared to the right kidney and preinfusion biopsies. All dogs developed glomerular lesions in the left kidney characterized by 1 or more of the following: mesangial cell proliferation, neutrophil infiltration, increased periodic acid-Schiff-positive staining of the mesangium and glomerular basement membrane (GBM), fibrin deposition, and thickening of the GBM. Left kidney glomerular immunofluorescence was positive in 7 of the 8 dogs using polyclonal antisera for canine IgG and C3 in a linear or fine granular pattern. Ultrastructural lesions were present in the left kidney of all dogs and consisted of irregular GBM thickening, intramembranous and mesangial electron-dense deposits, and mesangial and endothelial cell proliferation. Antibodies directed against D. immitis antigen were demonstrated in all kidney eluates from the left kidney. The right kidneys of 3 of the dogs developed lesions; however, in comparison to the left kidney, the lesions in the right kidneys were inconsistent, mild, and focal. The histologic findings in the left kidney were similar to those observed in dogs with naturally occurring D. immitis infections. In sham-immunized control dogs, renal arterial infusion of D. immitis antigen did not cause consistent immune complex glomerulonephritis; however, antigen adherence to glomerular capillary walls was observed by immunofluorescent microscopy.(ABSTRACT TRUNCATED AT 250 WORDS)

Dirofilaria immitis: Alteration of endothelium-dependent relaxation in the in vivo canine femoral artery

Pathogenic mechanisms in filarial diseases are complex and poorly understood. While examining endothelium-dependent vasodilatory responses in the in vivo canine femoral artery, we noticed that dogs with Dirofilaria immitis infection had altered vascular responsiveness. The results reported here extend our original observations on vascular reactivity in dogs with D. immitis infection ( L. Kaiser, J. F. Williams, E. A. Meade, and H. V. Sparks, 1987, American Journal of Physiology, 253, H1325–H1329). In noninfected dogs, acetylcholine binds to the luminal endothelial cell muscarinic receptor. This results in release of a nonprostaglandin endothelium-derived relaxing factor. The relaxing factor causes an increase in vascular smooth muscle guanylate cyclase and relaxation. However, in dogs with D. immitis infection the mechanism of relaxation to acetylcholine is different. At least two endothelium derived relaxing factors are involved: the major factor is a prostaglandin; the second factor works through vascular smooth muscle cGMP. These data suggest that adult D. immitis release pharmacologically active factors that can alter distal endothelial cell function. The notion that filarial products may alter the physiological function of endothelial cells should be considered in the pursuit of improved understanding of pathogenic mechanisms of filariasis.

Influence of immunosuppressants against Dirofilaria immitis infection in dogs.

Two immunosuppressive drugs, azathiopurine (AZP) and prednisolone (PDS), were examined on dogs experimentally infected with Dirofilaria immitis in order to estimate the involvement of immunological assail in rejecting the parasite by the host. AZP was orally administered to 3 dogs daily at a dosage of 1-10 mg/kg for a period from 3 days before infection until the end of the experiment. The dose was then varied and transiently ceased according to the severity of the side effects. PDS was subcutaneously administered daily to 2 dogs. They were administered 10 mg/kg of PDS from 3 days before infection to day 15 and 5.0-8.5 mg/kg from day 60 after infection to day 70. The serum D. immitis-specific antibody level assessed by an indirect hemagglutination test was steadily decreased in the AZP-medicated dogs. However, in the PDS-medicated dogs, the antibody titer was decreased until day 32 and, thereafter, was recovered. When all dogs were sacrificed between days 145-148, an average recovery rate of worms in both the AZP- and PDS-medicated dogs was 52.5% and 49.6%, respectively, while the controls showed 42.5%. The statistical analysis revealed no significant difference among the groups, indicating that the administration of AZP and PDS was not effective in protecting the larvae from the host's immune attack.

Influence of immune treatments against Dirofilaria immitis infection in dogs.

Mongrel dogs were inoculated with two kinds of antigenic substances. The first was a phosphate buffered saline extract of whole Dirofilaria immitis mixed with aluminum hydroxide gel (group 1), and the second was an orally administered live Metastrongylus apri infective larvae (L3) (group 2). Both groups were then infected with D. immitis L3. Indirect hemagglutination (IHA) tests showed that the antibody was produced by these inoculations before the infection was introduced, even in dogs inoculated with M. apri. This suggests a cross-reactivity between D. immitis and M. apri. The initial passive cutaneous anaphylaxis (PCA) antibody production was markedly delayed by about 70 days in group 2 compared with the production in the infected control dogs (group 3). The appearance of microfilaremia was also delayed by about one month in group 2 compared with that in the above control group. All dogs were sacrificed after the termination of the observation and worms recovered from the right ventricle and pulmonary arteries were counted and measured. The results indicated that immunization resulting from the homologous worm-somatic antigen might accelerate the growth of the infected larvae, whereas immunization resulting from the heterologous worm antigen, but cross-reactive to D. immitis, might disadvantageously affect the growth.

Epizootiology of endoparasitic infections in pet dogs and cats presented to a veterinary teaching hospital.

Prevalences of, and risk factors associated with, protozoan and helminth parasite infections were determined for pet dogs and cats presented to the teaching hospital of the University of Pennsylvania School of Veterinary Medicine. Of 2294 canine fecal specimens, 34.8% were found to contain one or more kinds of parasite, consisting of: hookworms (14.4%); Trichuris vulpis (12.3%); Giardia (7.2%); ascarids (5.5%); coccidia (2.7%); cestodes (1.6%). The following risk factors were identified with infection in dogs: age less than 2 years; urban locality; male sex. Gonadectomy was associated with decreased parasite prevalences in both male and female dogs. Significant seasonal variations in prevalences of hookworm, ascarid and Giardia infections were found in dogs. Of 452 feline fecal specimens tested, 24.6% were positive for parasites, including: ascarids (16.4%); Giardia (3.5%); coccidia (2.9%); cestodes (2.4%); hookworm (1.1%). Host age of less than 2 years was a significant risk factor associated with parasitism in cats. Of 1571 dogs tested for Dirofilaria immitis microfilaremia, 3.7% were positive. Non-urban locality was a significant risk factor for D. immitis infection in dogs.

Enzyme immunoassay of Dirofilaria immitis specific IgE in infected dogs

An ELISA procedure was developed for monitoring the specific IgE response in dogs to Dirofilaria immitis infection. The results of this assay correlated well with, and appeared to be more sensitive than, the passive cutaneous anaphylaxis test. The IgE ELISA values of the positive reference serum and the passive cutaneous anaphylaxis test results showed that a serum to negative absorbance ratio of 1.45 was statistically significally significant for discrinimination and was used to evaluate the specific IgE response in the sera from 90 clinically diagnosed heartworm cases. This ELISA procedure was more sensitive, as it detected 78% of the 90 cases as compared to a detection rate of only 43–47% by IgG ELISA or IFA. Sera obtained from 28 experimentally infected dogs at 4-week intervals for 20 weeks post-infection, were assayed for D. immitis-specific IgE by ELISA. A group of the infected dogs was also treated with diethylcarbamazine during the course of infection. All the experimentally infected dogs developed a specific IgE response, with treated dogs generally responding earlier.