Comparison of two immunochromatographic assays and the indirect immunofluorscence antibody test for diagnosis of Trypanosoma cruzi infection in dogs in south central Louisiana

Two rapid tests evaluated in dogs considered to be of high risk of infection with the Chagas parasite Trypanosoma cruzi using two immunochromatographic assays: Trypanosoma Detect™ for canine, InBios, Seattle, WA and CHAGAS STAT-PAK™ assay, Chembio Diagnostic Systems, Medford, NY, in south central Louisiana. For this purpose a serological survey was carried out in a total of 122 dogs and a serum bank was created. These 122 animals were first tested by IFAT that was used as the standard test. From the serum bank 50 samples were tested using the two rapid Chagas assays and results compared to the standard test IFAT. The serological survey using IFAT showed a prevalence of T. cruzi infection in 22.1% of the tested dogs. In the immunochromatographic assays, 13 and 11 animals were positive on rapid assay: Trypanosoma Detect™ for canine, InBios and CHAGAS STAT-PAK™, Chembio Diagnostic Systems, respectively compared to 11 positive by IFAT. These two immunochromatographic tests have shown high susceptibility and specificity compared to our standard method IFAT. The rapid, easy and accurate screening assays used in conjunction with confirmatory tests, would be an excellent tool for veterinarians to diagnose T. cruzi infection. Early detection of T. cruzi infection may prevent complications through an effective treatment. Greater awareness by veterinarians of the risk, clinical findings, history along with diagnostic methods will contribute greatly to an understanding of the true prevalence of Chagas disease in dogs in Louisiana.     

Frequency of IFNγ-producing T cells correlates with seroreactivity and activated T cells during canine Trypanosoma cruzi infection

Vaccines to prevent Trypanosoma cruzi infection in humans or animals are not available, and in many settings, dogs are an important source of domestic infection for the insect vector. Identification of infected canines is crucial for evaluating peridomestic transmission dynamics and parasite control strategies. As immune control of T. cruzi infection is dependent on humoral and cell-mediated immune responses, we aimed to define a serodiagnostic assay and T cell phenotypic markers for identifying infected dogs and studying the canine T. cruzi-specific immune response. Plasma samples and peripheral blood mononuclear cells (PBMCs) were obtained from forty-two dogs living in a T. cruzi-endemic region. Twenty dogs were known to be seropositive and nine seronegative by conventional serologic tests two years prior to our study. To determine canine seroreactivity, we tested sera or plasma samples in a multiplex bead array against eleven recombinant T. cruzi proteins. Ninety-four percent (17/18) of dogs positive by multiplex serology were initially positive by conventional serology. The frequency of IFNγ-producing cells in PBMCs responding to T. cruzi correlated to serological status, identifying 95% of multiplex seropositive dogs. Intracellular staining identified CD4⁺ and CD8⁺ T cell populations as the sources of T. cruzi lysate-induced IFNγ. Low expression of CCR7 and CD62L on CD4⁺ and CD8⁺ T cells suggested a predominance of effector/effector memory T cells in seropositive canines. These results are the first, to our knowledge, to correlate T. cruzi-specific antibody responses with T cell responses in naturally infected dogs and validate these methods for identifying dogs exposed to T. cruzi.     

Sylvatic Transmission of Trypanosoma cruzi Among Domestic and Wildlife Reservoirs in Texas,USA: A Review of the Historical Literature

Chagas disease (Trypanosoma cruzi infection) is one of the most important neglected tropical diseases affecting the Americas. The transmission dynamic of this parasite is a complicated process that involves three genera of Triatominae subfamily and over 100 known mammalian reservoirs composed of domestic, peridomestic and wildlife species. Understanding the complex relationship between vector species and mammalian hosts is important for preventing transmission to humans. We performed a historical literature review to assess the disease burden in the Texas wildlife and domestic animal population. Reports of sylvatic transmission in Texas date back to the 1940s. We found that up to 23 species can serve as reservoirs for T. cruzi in the state with wood rats, raccoons, and wild and domestic canine species most frequently reported as positive for the parasite. We finish with a discussion of the current research gaps, implications for high‐risk populations and future directions for research.     

First Report of Human Trypanosoma cruzi Infection Attributed to TcBat Genotype

Chagas disease is an endemic disease of the American continent caused by Trypanosoma cruzi and divided into six discrete typing units (TcI – TcVI). Nearly 10 million people harbour the infection representing a serious issue in public health. Epidemiological surveillance allowed us to detect a bat‐related T. cruzi genotype (henceforth named TcBat) in a 5‐year‐old female living in a forest area in northwestern Colombia. Molecular tools determined a mixed infection of T. cruzi I and TcBat genotypes. This represents the first report of TcBat infection in humans; the epidemiological consequences of this finding are discussed herein.     

Vaccination with Trypanosoma rangeli induces resistance of guinea pigs to virulent Trypanosoma cruzi

Chagas’ disease, endemic in Latin America, is spread in natural environments through animal reservoirs, including marsupials, mice and guinea pigs. Farms breeding guinea pigs for food are located in some Latin-American countries with consequent risk of digestive infection. The aim of this work was to study the effect of vaccination with Trypanosoma rangeli in guinea pigs challenged with Trypanosoma cruzi. Animals were vaccinated with fixated epimastigotes of T. rangeli, emulsified with saponin. Controls received only PBS. Before being challenged with T. cruzi, parasitemia, survival rates and histological studies were performed. The vaccinated guinea pigs revealed significantly lower parasitemia than controls (p < 0.0001–0.01) and a discrete lymphomonocytic infiltrate in cardiac and skeletal muscles was present. In the chronic phase, the histological view was normal. In contrast, control group revealed amastigote nests and typical histopathological alterations compatible with chagasic myocarditis, endocarditis and pericarditis. These results, together with previous works in our laboratory, show that T. rangeli induces immunoprotection in three species of animals: mice, guinea pigs and dogs. The development of vaccines for use in animals, like domestic dogs and guinea pigs in captivity, opens up new opportunities for preventive tools, and could reduce the risk of infection with T. cruzi in the community.     

The impact of concomitant infections by Trypanosoma cruzi and intestinal helminths on the health of wild golden and golden-headed lion tamarins

A 4-year longitudinal epidemiological study was carried out to evaluate the effect of infection by Trypanosoma cruzi and three intestinal helminth species on the health of golden and golden-headed lion tamarins. We evaluated health using analysis of blood counts, serum proteins, electrophoretograms, electrocardiograms and a health ranking based on physiological parameters. Among the helminths, Trichostrongylidae was demonstrated as the most pathogenic, followed by Prosthenorchis sp.; concomitant infection by Spiruridae may exacerbate the negative effects of the other two helminths. T. cruzi infection was not highly detrimental to the health of the study animals and was correlated with increased resilience to helminths. Tamarins younger than 1-year of age or older than 4-years had lower health condition. Golden-headed lion tamarins were in lower health condition because of higher parasitic prevalence. Our data suggest that when parasite community pathogenicity and prevalence are high, natural selection will allow survival only of lion tamarins in the best health condition.     

Observations on Trypanosoma theileri Infection in Cattle

Naturally occurring Trypanosoma theileri infection was studied in two cattle herds. Herd A was a dairy herd of approximately 250. Herd B was an isolated herd of 32 and contained both dairy and beef breeds. Blood samples were collected from all animals in Herd A during July and August on two successive years. Samples were collected from Herd B at monthly intervals. Total leukocyte and differential counts packed cell volume determinations, and trypanosome cultures were made on each sample.

Infection was detected in all age groups between seven months and fifteen years but it was rare in calves. Infected animals were not consistently positive for trypanosomes on consecutive blood cultures and there was considerable variation between infected individuals. Positive cultures were usually obtained from some animals while others were positive intermittently. No correlation was found between trypanosome isolations and the season of the year.

A correlation was found between trypanosome isolation and lymphocytosis. Of the 920 blood samples examined, approximately one in every five trypanosome positive samples had lymphocyte levels in the Bendixen positive range. Approximately one in every twenty trypanosome negative samples had lymphocyte numbers in the Bendixen positive range. Evidence indicated that trypanosome isolation from animals with lymphocytosis was not caused by increased numbers of infected buffy coat cells in the inoculum cultured.

Eight calves were inoculated intravenously with trypanosome-infected blood. Lymphocyte numbers increased an average of 3549 per cumm above pre-inoculation levels in seven and remained essentially unchanged in one. Prior to inoculation with infective blood, two of the calves were intravenously inoculated with trypanosome-infected blood that had been frozen and thawed to kill the trypanosomes contained in it. Neither developed lymphocytosis following this inoculation.

No clinical disease problems which could be attributed to trypanosome infection were found.

     

Canine Trypanosoma evansi infection introduced into Germany

A 9‐year‐old male Jack Russell Terrier with a history of travel to Thailand was presented with chronic lethargy, weight loss, unilateral anterior uveitis, pancytopenia, hyperglobulinemia, and proteinuria. Numerous trypomastigotes were found on a blood smear, and using molecular methods the parasite was identified as Trypanosoma evansi. After initial response to treatment, the dog experienced a relapse with central neurologic signs 88 days after initial presentation and died. Antibodies to T evansi were detected in both serum and cerebrospinal fluid (CSF) using a card agglutination test (CATT/T evansi), and PCR analysis of CSF for T evansi was positive. Findings at necropsy included marked non‐purulent meningoencephalitis. Chronic infection with T evansi in a dog that returned to Germany following international travel highlights the risk associated with introduction of foreign animal diseases to Europe and the possibility of these infections becoming endemic. Detection of chronic infection and curative therapy of trypanosomiasis are challenging, and infection is usually fatal in the dog.     

Growth hormones therapy in immune response against Trypanosoma cruzi

Growth hormone (GH) is an important hypophyseal hormone that is primarily involved in body growth and metabolism. In mammals, control of Trypanosoma cruzi parasitism during the acute phase of infection is considered to be critically dependent on direct macrophage activation by cytokines. To explore the possibility that GH might be effective in the treatment of Chagas’ disease, we investigated its effects on the course of T. cruzi infection in rats, focusing our analyses on its influences on parasitemia, NO, TNF-α and IFN-γ concentration and on histopathological alterations and parasite burden in heart tissue. T. cruzi-infected male Wistar rats were intraperitoneally treated with 5 ng/10 g body weight/day of GH. Animals treated with GH showed a significant reduction in the number of blood trypomastigotes during the acute phase of infection compared with untreated animals (P < 0.05). For all experimental days (7, 14 and 21 post infection) of the acute phase, infected and GH treated animals reached higher concentrations of TNF-α, IFN-γ and nitric oxide as compared to untreated and infected counterparts (P < 0.05) Histopathological observations of heart tissue revealed that GH administration also resulted in fewer and smaller amastigote burdens, and less inflammatory infiltrate and tissue disorganization, indicating a reduced parasitism of this tissue. These results show that GH can be considered as an immunomodulator substance for controlling parasite replication and combined with the current drug used may represent in the future a new therapeutic tool to reduce the harmful effects of Chagas’ disease.     

Trypanosoma cruzi infection in a bat-eared fox (Otocyon megalotis) with severe focally extensive suppurative myocarditis

Chagas disease is a parasitic illness caused by Trypanosoma cruzi. This is an intracellular parasite that can cause cardiomyopathy, megaesophagus and megacolon in the chronic phase of the disease. One of the characteristics is sudden death due to heart failure. In this study, a 15-month-old, intact female bat-eared fox (Otocyon megalotis) kept in a private collection presented for sudden death with no preceding signs of illness. Annual health exam was unremarkable one month prior. On gross necropsy, yellow-white, irregular lesions were found in the ventricles near the apex of the heart. Histopathology found severe focally extensive suppurative myocarditis and PCR was positive for Trypanosoma cruzi within the heart tissue. To date, this is the only reported case of a bat-eared fox infected with Trypanosoma cruzi.     

Reactivation of a Trypanosoma cruzi infection in a rhesus monkey (Macaca mulatta) experimentally infected with SIV

Trypanosoma cruzi-like flagellates were incidentally noted in blood smears of a routinely monitored rhesus monkey experimentally infected with the simian immunodeficiency virus (SIV). Immunodeficiency in the course of the SIV infection reactivated a chronic infection of Chagas' disease that had been unnoticed when the macaque was imported to Europe. The animal developed no specific clinical symptoms of American trypanosomiasis, but histologically a chagasic myocarditis was detected. Analysis of the small subunit rRNA gene of the trypanosome identified the protozoan as T. cruzi.     

Parasitemia in a neonatal bison calf

A 3-day old female bison calf (Bison bison) was presented in lateral recumbency to the Université de Montréal Veterinary Teaching Hospital. The animal was severely depressed and dehydrated (10%) and died a few hours after admission. Prior to death, blood samples were obtained for CBC, clinical chemistry, and serology tests. Abnormal CBC findings included thrombocytopenia, lymphocytosis, mild monocytosis, and a toxic left shift. Abnormal serum clinical chemistry findings included marked azotemia, hyperphosphatemia, hypernatremia, hyperalbuminemia, hypoglobulinemia, and low gamma-glutamyltransferase activity. Serologic test results for bovine leukosis virus and bovine viral diarrhea virus were negative. Blood smear examination revealed numerous elongated organisms that were tapered at both ends and characterized by an undulating membrane and a long flagellum. The organisms ranged in length from 35 to 40 micro meter, excluding the flagellum, and were identified as Trypanosoma theileri. Postmortem examination revealed that the animal suffered from concurrent mycotic abomasitis and colisepticemia.     

Staphylococcus sciuri peritonitis in a patient on peritoneal dialysis

Staphylococcus sciuri is an occasional cause of human infection that has been described in the flora of numerous animal species and in environmental specimens. Here, we report a rare case of S. sciuri peritonitis in a dialysis patient who had exposure to peridomestic animals.     

Influence sexual dimorphism on the persistence of blood parasites in infected Calomys callosus

Gender has long been known to be a contributory factor in the incidence and progression of disorders associated with immune system disregulation. The aims of this experiment were to verify the influences of sexual dimorphism on the persistence of blood parasites out of the acute phase of infection. Male and female Calomys callosus were separated and infected with two strains of Trypanosoma cruzi, and let age until 120 days. Xenogiagnostic, culture of organs and blood, histopathology and lytic antibody percentages were evaluated on late chronic phase. Xenodiagnosis, hemoculture and lytic antibody percentages were positive from 45 until 120 days. For both strains in adrenal and heart, amastigote burdens were present until 45 days, scarcely found on 60 days and absent on 120 days. Steroid hormones, although having a protective role, does not enable animals to get completely rid of the infection. Even without showing apparent signs of pathological unbalance, parasites persists, hidden throughout the host’s body.     

Spectrum of disease in macaque monkeys chronically infected with SIV/SMM

Twelve rhesus and one pig-tailed macaque have been monitored for 28-41 months following experimental infection with 10(4) TCID of SIV/SMM. Twelve of the 13 animals became virus positive and seroconverted within 3 to 6 weeks of exposure; the remaining animal seroconverted at 6 months, but has remained virus negative. Six of the 13 animals (46%) died between 14 and 28 months post-infection, following prolonged clinical disease characterized by chronic diarrhea and weight loss, peripheral lymphadenopathy and hemogram abnormalities. Histologic findings ranged from prominent follicular hyperplasia to severe lymphoid depletion, with lymphoid tissues often showing an infiltrate of syncytial giant cells. One animal had intestinal cryptosporidiosis and two had brain lesions comparable to those seen in AIDS encephalopathy in humans. Three of the remaining seven animals have an ARC-like disease and are showing gradual deterioration of their clinical condition. These animals, as well as animals that died, had progressive decreases in CD4+ cells and CD4+/CD8+ cell ratios. These observations further document the marked clinical, pathologic and immunologic similarities between human AIDS and the SIV-infected macaque model.     

Isolation of Trypanosoma theileri from the blood of two cows, one leukotic, one exhibiting lymphocytosis

Trypanosoma theileri was isolated by the blood culture method from a leukotic cow in extremis. The parasite could be maintained along with leukocytes with weekly changes of culture medium up to 6 months. In connection with a subsequent transmission experiment a cow, not inoculated with trypanosomes but kept in the same shelter as the inoculated one, commenced to be positive for T. theileri in its blood cultures. This cow had a long history of lymphocytosis but showed no signs of leukosis when slaughtered. The significance of the findings is discussed from the point of view of false positive diagnosis of bovine leukosis and the possible role of trypanosome in the etiology of leukosis.Keyword: Trypanosoma, Trypanosoma theileri, lymphocytosis, leukosis     

Bovine leukosis. IV. Trypanosomiasis, lymphocytosis and DNA synthesis.

The possible influence of natural trypanosome infection on lymphocytosis and DNA synthesizing lymphocyte counts in peripheral blood was determined on 220 cows from two leukosis herds and 25 cows from leukosis free control herd. Trypanosome incidences were determined during summers of 1969 and 1970 by inoculating whole blood onto blood agar slants and incubating at room temperature. Incidence of trypanosomiasis in cattle was found to be variable, possibly due to factors affecting the primary isolation of Trypanosoma theileri. A small trypanosome resembling Trypanosoma uniforme was found occasionally as a concomitant infection with T. theileri. Trypanosomiasis occurred with equal frequency in the animals of the leukosis and the control herds. No correlations were noted between trypanosomiasis, lymphocytosis and DNA synthesizing lymphocytes in peripheral circulation.     

Parasitological prevalence of bovine trypanosomosis in Kindo Koisha district,Wollaita zone,south Ethiopia

A cross sectional survey to determine the distribution and prevalence of trypanosomosis was conducted in Kindo Koisha district, in the Wollaita zone in southern Ethiopia. A total of 1 008 adult cattle was examined at eight different localities. Dark field examination of the buffy coat, as well as stained thin blood film examination and packed cell volume (PCV) evaluation were the diagnostic techniques used. The overall prevalence of bovine trypanosomosis was 15 %. Among the positive animals, 108 (71.1%), 43 (28.4%) and 1 (0.6%) were due to Trypanosoma vivax, Trypanosoma congolense and mixed infection (T. vivax and T. congolense), respectively. The infection rate of T. vivax and T. congolense varied significantly (P < 0.01). The mean PCV of the positive and negative animals ranged between 18.3-32.1% and 26.8-33.4%, respectively. The mean PCV of negative animals (28 %) was significantly higher than the mean PCV of positive animals (22.3%) (P < 0.001). There was an inverse association of PCV with the prevalence of trypanosomosis (P > 0.05). The herd average PCV values of each site decreased with increasing proportion of the positive herds of that particular site. Of the diagnostic tests employed, the microhaematocrit buffy coat technique is relatively sensitive and it has an added advantage of indicating the general condition of the animal by haematocrit measurement. In view of the risk of trypanosomosis, a control intervention through the strategic application of appropriate trypanocidal drugs is recommended. A tsetse fly control scheme to reduce host-tsetse fly contact is equally as important as chemotherapy and chemoprophylaxis against trypanosomosis.     

Relapse infection after chemotherapy in dogs experimentally infected with Trypanosoma brucei brucei

Studies on relapsing Trypanosoma brucei brucei infections in dogs after Berenil treatment revealed that the first relapse occurred 13 to 64 days after chemotherapy and 36 to 79 days after inoculation. A second relapse infection was observed in two dogs 43 and 60 days after a second Berenil treatment. During the aparasitaemic period following chemotherapy in four dogs, successful transmission (as evidenced by subsequent parasitaemia) following the intraperitoneal inoculation of homogenate of brain from two of the dogs into recipient rats was obtained. Transmission with blood collected just before the animals were sacrificed was, however, negative. Hornogenates of other organs (liver, spleen, eyes, testes, kidneys, heart and lymph node) were also non-infective. One dog inoculated with relapsed trypanosomes and treated with Berenil soon after showing parasitaemia was completely cured of the infection. It was considered that the brain is the source of relapse in T b brucei infection after Berenil therapy and that the relapse was not due to drug resistance.     

Relationship between splenic sequestration and thrombocytopenia in Trypanosoma evansi infection in rats

Trypanosoma evansi infections in domestic animals are characterized by anemia and thrombocytopenia. The cause of the platelets decrease is unknown, but researchers suggest that thrombocytopenia may result from damage of the bone marrow, reduced survival of platelets, auto-immune thrombocytopenia, disseminated intravascular coagulation and splenic sequestration. Some of these causes have already been tested by our research group and found to be unrelated. Therefore, this study has the objective of testing the hypothesis that splenic sequestration might be responsible for thrombocytopenia in T. evansi-infected rats. A total of 28 rats assigned to four groups were used in the experiment. Group A rats were splenectomized and infected with T. evansi, group B rats were infected with T. evansi, group C rats were splenectomized, but not infected and group D rats were normal controls. Five days post-infection all rats were anesthetized and blood was collected in order to measure the number of circulating platelets, fibrinogen levels, prothrombin time (PT) and activated partial thromboplastin time (aPTT). The spleens of groups B and D were weighed at necropsy. The infected animals (groups A and B) showed a significant reduction in platelets and increased PT and aPTT when compared to negative control groups (groups C and D). Animals from group A showed increased levels of fibrinogen. The mean weight of spleen differed between group B (2.62 g) and group D (0.55 g). It was concluded that there is no relationship between thrombocytopenia and splenic sequestration in infection by T. evansi.