Antibody and lymphoblastogenic responses of dogs experimentally infected with Trypanosoma cruzi isolates from North American mammals

The humoral and cellular immune responses of dogs infected with either a non-pathogenic Trypanosoma cruzi isolated from a North American dog (Tc-D) or a pathogenic T. cruzi isolate from an opossum (Tc-O) were studied over a 240 day period. Antibody to T. cruzi epimastigote antigens prepared from Tc-O or Tc-D isolates were first detected by ELISA by Day 26 post infection (PI), peaked by day 175 PI and remained elevated throughout the experimental period in both Tc-O and Tc-D infected dogs. Differences in antibody levels between infected groups were not detected. Western blot analyses were performed using Tc-O and Tc-D epimastigote antigens probed with pooled sera and sera from individual Tc-O and Tc-D infected dogs prior to infection (Day 0), and during the acute (Day 16-35 PI), indeterminate (Day 50-135 PI) and chronic (Day 235 PI) stages of infection. Generally, the patterns, number of protein bands, and temporal appearance of the protein bands identified by pooled sera and sera from individual dogs within each antigen preparation were similar. However, similarities and differences were present in antibody responses between sera from Tc-O and Tc-D infected dogs. Blastogenic responses of peripheral blood mononuclear cells (PBMC) from Tc-O and Tc-D infected dogs to mitogens (concanavalin A, phytohemagglutinin and pokeweed) were not significantly different from controls at any time during the experimental period. The PBMC from both groups of dogs were unresponsive to epimastigote antigens during the acute stage of infection. Statistically significant differences (P less than 0.05) in PBMC responsiveness from controls were observed on Days 70 and 175 PI. Responses decreased to pre-infection levels by Day 240 PI. These studies demonstrate that although two North American T. cruzi isolates have markedly different virulence for dogs, some aspects of their cellular and humoral immune responses are similar while other responses, such as antibody recognition of specific T. cruzi antigens, vary.     

Clinical, clinicopathologic, and parasitologic observations of trypanosomiasis in dogs infected with North American Trypanosoma cruzi isolates

Nineteen purebred Beagles of various ages (4, 5, 13, and 47 weeks) were inoculated with North American Trypanosoma cruzi isolates obtained from an opossum (Tc-O), an armadillo (Tc-A), or a dog (Tc-D). Dogs were grouped on the basis of clinical outcome of infection. During the acute stage of disease, dogs of group 1 (n = 7 inoculated with Tc-O or Tc-A) died or were euthanatized because of the severity of disease. Dogs of group 2 (n = 5 inoculated with Tc-O or Tc-A) developed acute disease, but survived to develop chronic disease. Dogs of group 3 (n = 7 Tc-D-inoculated dogs) developed neither acute nor chronic disease. Dogs of group 4 (n = 4--2 dogs 13 weeks old and 2 dogs 47 weeks old) served as noninoculated controls. Clinical signs associated with severe acute myocarditis developed in dogs of groups 1 and 2 between postinoculation day (PID) 15 and 28. Generalized lymphadenopathy and lymphocytosis were observed in all dogs of groups 1, 2, and 3 between PID 14 and 17. Serum alanine transaminase and aspartate transaminase activities and urea nitrogen concentration were high, and glucose concentration was low prior to death of dogs in group 1. Serum activities of isoenzymes of creatine kinase were significantly (P less than 0.05) high in only 1 dog (group 1), whereas serum lactate dehydrogenase isoenzyme activities were not significantly high in any dog.(ABSTRACT TRUNCATED AT 250 WORDS)     

Chronic dilatative myocarditis caused by Trypanosoma cruzi in two dogs

Trypanosoma cruzi was believed responsible for causing chronic dilatative myocarditis in 2 female hunting dogs. Clinical signs included ascites, respiratory distress, thoracic effusion, cyanosis, and weak pulse with ventricular arrhythmias. Electrocardiography indicated first-degree heart block, chamber enlargement, and ventricular-based arrhythmias unresponsive to treatment. M-mode echocardiography of 1 dog confirmed bilateral cardiac enlargement and septal and left ventricular free wall thinning. Multifocal infiltrates of plasma cells, lymphocytes, and histiocytes, cardiocyte degeneration, and multifocal fibrosis were the predominant histologic lesions. Trypanosoma cruzi pseudocysts were infrequently found.     

Haematological and histopathological findings after ovariectomy in Trypanosoma cruzi infected mice

The aim of this study was to investigate the influences of ovariectomy on histopathological and hematological parameters during the course of Trypanosoma cruzi infection. Hematological and immunological homeostasis is influenced by gonadal steroid hormones. Ovariectomy exerts profound influences on parasitic diseases including T. cruzi infection through modulation of the host's immune response. Three groups of female Mus musculus were infected with 4000 blood trypomastigotes of the Y strain of T. cruzi. One group was subjected to ovariectomy, another to simulated surgery before the infection, and a third group of unoperated animals were used as controls. Marked differences were detected in the responses of blood and tissue parasites. On day 9, post-infection parasitism was significantly higher in ovariectomized animals (P<0.05). These results were confirmed by histopathological studies, in which ovariectomized animals displayed hearts with higher number of amastigote burdens, increased inflammatory infiltrate, enhanced tissue fibers disorganization and decreased lytic antibody percentage, when compared to their counterparts. On day 9 the hematological changes were more apparent, with a decrease in erythrocytes, platelets and leucocytes for ovariectomized infected animals. Simulated surgery, as a stressful agent, did not cause any imbalance in parasitism or in the hemogram profile. The results confirm the importance of the female steroids in resistance against T. cruzi infection.     

Antileishmanial antibody profile in dogs naturally infected with Leishmania chagasi

Visceral leishmaniasis (VL) presents vigorous Th2 immune response, which is mainly characterized in human by augmented expression of Il-4, polyclonal B cell activation, intense hypergammaglobulinemia and production of antileishmanial IgE antibodies. However, few aspects of this type of immune response have been demonstrated in studies of canine visceral leishmaniasis (CVL). This work investigated by ELISA and western immunoblotting the production of antileishmanial IgE antibodies (IgE Ab) in symptomatic and asymptomatic dogs naturally infected by Leishmania chagasi, and also compared this IgE immune response with those of IgG, IgG1 and IgG2 antibodies. Three groups of dogs were evaluated: 12 VL dogs with positive Leishmania biopsies (GI), 44 dogs with a positive leishmanial indirect fluorescent antibody test (IFAT), 30 of them presenting clinical signs of VL and 14 asymptomatic (GII) and 21 healthy dogs living in kennels located in leishmaniasis endemic areas (GIII), which were seronegative in the IFAT. Eighteen dogs from an area free of CVL were used as controls (GIV). Antileishmanial IgE antibodies were detected in 4 of 12 VL dogs from group I (33%) and 14 of 30 symptomatic dogs from group II (47%). While all asymptomatic dogs from group II (100%) were seronegative for antileishmanial IgE Ab, 7 of 21 healthy animals from group III (33%) had these immunoglobulins. A strong correlation was verified between antileishmanial IgG and IgG2 antibody titers in all symptomatic dogs, but only 15 of these 42 animals (36%) produced simultaneously IgE, IgG, IgG1 and IgG2 antibodies to Leishmania. IgE antibodies recognized leishmanial antigens of 12, 36, 61, 81 and 118 KDD, while a more complex pattern of immunoblotting was verified mainly for IgG and IgG2 antibodies from symptomatic animals. IgG1 and IgG2 antibodies shared the recognition of L. chagasi polypeptides of 118, 81, 61, 36, 18, 14 and 12 KDD, being more intense the immune reactions between IgG1 Ab and the leishmanial polypeptides of 61 and 36 KDD, and also between IgG2 antibodies and the antigens of 26, 21, 18, 14 and 12 KDD. Our results suggest that the polyclonal production of antileishmanial antibodies that includes IgE Ab could characterize a Th2 immune response in CVL and can help the laboratory diagnosis of this disease.     

Idiotype expression of IgG1 and IgG2 in dogs naturally infected with Leishmania infantum

Here we analyzed, by Western blot analysis, the idiotype expression of IgG1 and IgG2 in 109 canine sera corresponding to 50 dogs from endemic areas of leishmaniosis in order to detect markers related to Leishmania infantum infection and clinical condition (asymptomatic or symptomatic). Twenty-four dogs from an area free of leishmaniosis were used as controls. IgG1 and IgG2 responses in symptomatic and asymptomatic L. infantum infections differed mainly in subclass production (ELISA values), with higher IgG2 production occurring particularly in symptomatic dogs. Nevertheless, we observed little difference in the idiotype expression of these IgG subclasses, which, in general, recognized the same antigenic fractions. While early L. infantum infection was characterized by recognition of polypeptide fractions of low molecular weight, mainly fractions of 14, 16 and 18 kDa by IgG1 and 14 and 16 kDa by IgG2, symptomatology was associated with recognition by both IgG subclasses of a 24 kDa fraction and other antigens belonging to the AG24 family.     

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.     

Erythrocyte response to Trypanosoma brucei in experimentally infected dogs.

Trypanosoma brucei infection produced an acute and fatal disease in Nigerian mongrel dogs due to a rapidly developing anaemia. Infected dogs responded with increased reticulocytosis, which was not sustained with chronicity. In comparison the response to artificially-induced haemolytic anaemia was progressive, marked and sustained. The anaemia of T. brucei infection of dogs was either normocytic normochromic in acute infection or microcytic normochromic in chronic infection. Artificially-induced haemolytic anaemia was either macrocytic normochromic or normocytic normochromic. The erythropoietic potential of plasma in vivo in mice increased in T. brucei-infected dogs except at the terminal parasitaemia. The anaemia in Trypanosoma brucei-infected dogs is therefore initially responsive but becomes poorly involved with chronicity.     

Parasitaemia and clinical manifestations in Trypanosoma brucei infected dogs.

Four dogs were infected with Trypanosoma brucei (Mkar strain) while another four were used as uninfected controls. Two of the dogs showed acute disease and died in the first wave of parasitaemia on days 7 and 8 post infection (PI) while the other two died from the sub-acute disease on days 24 and 28 PI corresponding to the second wave of parasitaemia. In the first wave of parasitaemia there was a sharp decrease in the packed cell volume, red blood cell, haemoglobin, total leucocytes, eosinophil, neutrophil and lymphocyte values, but during the period of low parasitaemia there was a slight recovery of the values of total leucocytes and lymphocytes although these and the other values showed a continuous decrease during the second wave of parasitaemia. In contrast, there was a consistent monocytosis in both acute and sub-acute diseases. The general picture was that of loss of condition, anaemia, leucopenia, monocytosis, ocular impairment, elevated temperature, pulse and respiratory rates, the difference between the acute and sub-acute diseases being in the degree of intensity. The degree of anaemia noted and the circulatory disturbances associated with the infection could have caused the death of all the infected dogs.     

Electrocardiographic and echocardiographic features of trypanosomiasis in dogs inoculated with North American Trypanosoma cruzi isolates.

Purebred Beagles were inoculated with Trypanosoma cruzi isolates from a North American opossum or armadillo (Tc-W), and dog (Tc-D). Although Tc-D established infection in dogs, the dogs did not develop cardiac abnormalities. Dogs inoculated with Tc-W developed acute myocarditis associated with increases in P-R interval, atrioventricular block, depression of R wave amplitude and shifts in mean electrical axis. Echocardiograms were normal during this stage. Three Tc-W-inoculated dogs died during the acute stage. Following the acute stage, 5 of 8 Tc-W-inoculated dogs entered an indeterminate stage in which ECG changes were minor and echocardiograms were normal. Progression to the chronic stage in 5 of the 8 Tc-W-inoculated dogs was indicated by development of ventricular-based arrhythmias, mainly ventricular premature contractions, between postinoculation days 60 and 170. In some dogs, ventricular premature contractions were multifocal. Electrocardiographic abnormalities progressively degenerated to various forms of ventricular tachycardia. Worsening ECG coincided with loss of left ventricular function as measured by echocardiography. Mean percent ejection fraction and percentage of fractional shortening decreased to 63% and 52% of control values, respectively. The left ventricular free wall (LVFW) thickness decreased and % septal: % LVFW thickening ratio increased, indicating a relative preservation of septal wall motion and LVFW hypokinesis.     

Relapses in dogs experimentally infected with Trypanosoma brucei and treated with diminazene aceturate or isometamidium chloride

Twenty dogs of mixed local East African breeds were used. Five of the dogs were uninfected controls and 15 were infected with T. brucei (ILRAD 273). Five of the infected dogs were untreated controls, five were treated with a high curative dose of diminazene aceturate, (7 mg kg-1 body weight (wt.), and five were given a subcurative dose of isometamidium chloride (1 mg kg-1 body wt.). The drugs, given at 8 days post infection (d.p.i..), led to apparent recovery. The antibody titres, however, remained high in both groups and at 42-49 d.p.i. there was at least one relapse in each treatment group. Parasite populations from relapsed animals were more resistant to the drugs than the original infecting populations. The implications of these findings are discussed.     

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.     

Diminazene aceturate residues in the tissues of healthy, Trypanosoma congolense and Trypanosoma brucei brucei infected dogs

The tissue distribution and residue profile of diminazene aceturate was investigated in healthy dogs and in dogs infected with Trypanosoma congolense and Trypanosoma brucei brucei. The drug was administered at 3.5 mg/kg i.m. and tissue samples were taken post mortem from the animals at 48, 72, 120, 168 and 240 h after injection. The drug was distributed to various organs and tissues of the body with the highest concentrations occurring in liver and kidney. Higher drug levels were obtained in the tissues of healthy dogs compared with trypanosome infected animals except in the brain. The levels of residues in the healthy animals were significantly different (P less than 0.05) from those of the infected dogs. The drug residues were still detectable in the tissues of the animals 10 days after drug administration.     

Cerebral state monitoring in Beagle dogs sedated with medetomidine

OBJECTIVE: To provide experience of monitoring the level of hypnosis with the Cerebral State Monitor (CSM), a device extracting a single numerical variable between 0 and 100 from the electroencephalogram in dogs sedated with medetomidine during dental scale removal. STUDY DESIGN: Prospective observational study. Animals Nine female Beagle dogs weighing 13.3 +/- 1.3 kg. METHODS: Cerebral state index (CSI) and burst suppression ratio (BSR) were recorded from sub-dermal needle electrodes in dogs sedated after subcutaneous injection of 114 +/- 11 microg kg(-1) medetomidine. Ten minutes after injection CSI monitoring began, and after 5 minutes, dental scale removal with an ultrasonic probe was started. After approximately 30 minutes, the effects of medetomidine were antagonized with atipamezole. RESULTS: The CSI had a median value of 43 (range 40-56) in undisturbed sedated dogs. During dental scale removal, CSI increased to a median value of 99 (range 92-100). The BSR in undisturbed sedated dogs ranged from 2 to 15, but fell to zero during dental scale removal. CONCLUSIONS: Stimulation during dental scale removal might be expected to reduce the level of sedation and hypnosis in dogs to which medetomidine had been administered. The concurrent increase in CSI and decrease in BSR suggested that a higher CSI was associated with arousal from sedation and a reduction in the depth of hypnosis. More studies are needed to validate CSI in order to better understand the functioning of this monitor. CLINICAL RELEVANCE: The CSM shows promise for monitoring the degree of sedation and hypnosis during anaesthesia, and after adequate validation, could contribute to the refinement of anaesthetic techniques in animals.     

Immunoglobulin isotype distribution of antinuclear antibodies in dogs with leishmaniasis

A modified indirect immunofluorescence method, using rat liver as substrate, was developed to determine the immunoglobulin isotypes forming antinuclear antibodies in sera from 12 antinuclear antibody-positive dogs out of 121 dogs with natural Leishmania infection. Immunoglobulin M was found to be the most frequent component of antinuclear antibodies (91·7 per cent), followed by IgG (41·7 per cent) and IgA (33·2 per cent). When these immunoglobulin isotypes were titrated, IgG antinuclear antibodies showed higher titres (1:200) than IgM and IgA antinuclear antibodies (1:50 and 1:20 respectively). Most of the antinuclear antibody-positive dogs simultaneously had two immunoglobulin isotypes, whereas none had all three immunoglobulin isotypes at the same time. Spearman rank correlation analysis showed no significant correlation between antinuclear antibody titres and circulating immune complexes or immunoglobulin levels. The low incidence of antinuclear antibodies and the absence of a clear relationship between isotype titres and clinical signs suggest a minor pathogenic role of antinuclear antibodies in canine leishmaniasis.     

A follow-up of Beagle dogs intradermally infected with Leishmania chagasi in the presence or absence of sand fly saliva

In this study, we compare the development of infection and/or disease in Beagle dogs intradermally infected with Leishmania chagasi, in the presence or absence of Lutzomyia longipalpis saliva, with those of intravenously infected animals.Spleen samples of all the animals inoculated with parasites had positive polymerase chain reaction tests for Leishmania DNA. Positive spleen cultures for Leishmania were detected earlier (P < or = 0.018) and were more frequent (five out of the five animals) in intravenously infected animals than in the intradermally infected animals, in presence (two out of the six animals) or absence (three out of the five animals) of salivary gland lysate of L. longipalpis. Significant increase in serum antibodies against Leishmania was observed only in the intravenously infected group (P = 0.004). In addition, dogs with infection confirmed by isolation of amastigotes or detection of parasite DNA were, nevertheless, negative for anti-Leishmania antibodies up to 5 months or more after infection. Only animals of the intravenously infected group developed progressive decreases in hematocrit (Pearson r = -0.8076, P = -0.0026) and hemoglobin (Pearson r = -0.8403, P = 0.0012) during the infection period. No significant difference in the course of infection was observed between groups of intradermally infected animals. The data presented herein confirms that the intradermal inoculation of dogs with Leishmania produces an asymptomatic form of infection. It also fails to show an advantage in using L. longipalpis saliva as an infection-enhancing agent in experimental canine leishmaniasis.     

Hematologic values in calves infected with Sarcocystis cruzi

Calves were inoculated with 2 X 10(5) Sarcocystis cruzi sporocysts. Red cell mass decreased dramatically between Days 21 and 35 post-infection and plasma volume increased concurrently, so that blood volume did not change significantly. Mild reticulocytosis and increased pyrimidine 5' nucleotidase activity in erythrocytes occurred between Days 35 and 42. Antiglobulin tests with anti-bovine IgG, IgM and C3 were negative, with the exception of a positive test for C3 in 1 of 6 infected calves.     

Serum resistance is correlated with encapsulation of avian strains of Pasteurella multocida

Encapsulated avian strains of Pasteurella multocida possessing an A-type capsule were shown to be resistant to the bactericidal action of turkey serum, whereas unencapsulated variants as well as other unencapsulated strains were not. Removal of the capsule from serum-resistant strain P1059-1 resulted in this strain becoming susceptible to the bactericidal effects of turkey serum. Since complement was consumed when encapsulated or unencapsulated strain P1059-1 was incubated in turkey serum, we conclude that the capsule acts to shield the outer membrane rather than prohibiting the generation of an effective membrane attack complex.