Trichinella spiralis—A potential anti-tumor agent

Murine forestomach carcinoma (cell line MFC), ascitic hepatoma (cell line H22) and sarcoma (cell line S180) solid tumor models were used to test the anti-tumor effect of Trichinella spiralis in vivo. Mice previously infected by oral administration of 400 viable T. spiralis larvae per mouse for 7 days were grafted with various solid tumor cell lines. Other groups of tumor-bearing mice were given caudal vein injection of crude extracts of adult and newborn larvae at 17.5, 35.0 or 70.0 mg kg^?1. These treatments to inhibit tumor growth were dose-dependent (p < 0.05). The anti-proliferative activity of crude T. spiralis extract was examined in vitro at 0.035, 0.070 or 0.140 mg ml^?1 using MFC, H22, S180, human chronic myeloid leukemia cell line (K562) and hepatoma cell line (H7402), tumor cell proliferation in vitro was measured by methyl thiazolium stain and was inhibited in dose-dependent manner (p < 0.05). At the same doses, crude T. spiralis extracts induced apoptosis of K562 and H7402 as detected by DNA fragmentation. Cell cycle analysis indicated that crude T. spiralis extracts, at 0.140 mg ml^?1, arrested the cell cycle of K562 and H7402 in G1 or S phase. It is concluded that T. spiralis contains anti-tumor active agent.     

Strain difference in susceptibility to Trichinella spiralis infection between dark agouti and albino oxford rats

The influence of host genetics on the susceptibility to primary Trichinella spiralis infection has been extensively studied in a mouse model, but has not been clarified for rats. Analyses of interstrain and intrastrain genetic variation in response to infectious agents could be beneficial not only for elucidating the genetic basis of host resistance/susceptibility, but for revealing immune response mechanisms as well. The aim of this study was to analyse interstrain differences in worm burdens and cytokine production between Albino Oxford (AO) and Dark Agouti (DA) rats in muscle phase of T. spiralis infection. Clear strain-dependent variation was observed in the number of T. spiralis larvae per gram (lpg) of muscle tissue where values for DA rats (626.7 ± 171 lpg) vastly exceeded those found in AO rats (49.8 ± 25.9 lpg, p < 0.001). Differences between the strains were also noticed in key cytokine levels. In infected AO rats, the cytokine production remained in favor of Th1 type response, while infected DA rats showed a shift towards a Th2 type response. The level of regulatory IL-10 was significantly increased only in T. spiralis infected DA rats. Our results provide evidence that DA rats express higher susceptibility to T. spiralis infection in comparison to AO rats with respect to muscle larvae burden. The infection in DA rats was accompanied by the production of anti-inflammatory cytokines, while the response of AO rats was characterized by a proinflammatory type of immune response.     

Complement membrane attack complex formation and infectivity of Trichinella spiralis and T. nativa in rats

Rats readily become infected with Trichinella spiralis but are more resistant to T. nativa. We infected complement factor C6-deficient (C6?) rats and control (C6+) rats with T. spiralis and T. nativa to compare the effects of membrane attack complex on these parasites in vivo. The 2000 larvae infection dose per rat yielded 652 lpg (larvae per gram) in the C6? group and 608 lpg in the C6+ group with T. spiralis, whereas with T. nativa the corresponding figures were only 1.05 and 1.87 lpg. The difference between the Trichinella species was evident, but the infection intensity was unaffected by the C6 deficiency. When newborn larvae were incubated in C6-deficient and control rat sera for 24 h in vitro, no changes in viability were observed. Immunohistochemistry revealed that the musculature of cross-sectioned adults and certain stichocytes bound human complement factors C3, C8 and C9, but not C1q. Interestingly, the outermost layer of the cuticle and the newborn larvae did not show any binding activity. Similar findings were obtained with immunofluorescence microscopy of intact newborn larvae. These results indicate that both T. spiralis and T. nativa have efficient mechanisms to protect themselves against complement attack. The difference in infectivity for rats between the two species, however, is not due to a differential resistance to complement membrane attack complex.     

Evaluation of the risk of transmission of Trichinella in pork production systems in Argentina

Recently, there has been interest in programs that certify pork production practices that minimize the risk of exposure of pigs to Trichinella spiralis. Certification might be useful for reducing the risk of human trichinellosis from pork in Argentina, but more information is needed on pig production practices and sources of Trichinella infection in Argentinian pigs. In this study, 21 pig farms were assessed for Trichinella infection including some farms using total and partial confinement management, and others with pigs raised exclusively outdoors. A total of 3224 muscle samples were collected from pigs raised on these farms and tested to determine the presence of T. spiralis larvae by artificial digestion. Serum samples from the same 3224 pigs were tested for antibodies to T. spiralis by ELISA. For each farm, a questionnaire was completed summarizing information about management factors and this information was used to assess risk factors for exposure of T. spiralis. Based on the results, pigs raised outdoors were more likely to be infected than pigs raised in total or partial confinement (p ≤ 0.05). Pigs fed waste products containing meat were 12.5 times more likely to be infected than pigs not fed waste containing meat (p < 0.01). The role played by rats in transmission of Trichinella is unclear; however, on farms with evidence of wild animals and access of pigs to wildlife carcasses, the prevalence of Trichinella infection was significantly higher. All pigs raised under good hygienic and sanitary conditions were negative for Trichinella infection by both artificial digestion and ELISA.     

NTPDase and 5′-nucleotidase as inflammatory markers in cattle naturally infected by Eurytrema coelomaticum

The aim of this study was to evaluate seric NTPDase and 5′nucleotidase activities of cattle naturally infected by Eurytrema coelomanticum, as well as to correlate them to histopathological lesions in the pancreas and the degree of parasitism. Blood samples and pancreas of 51 bovines were collected on a slaughterhouse in Southern Brazil: 33 from cattle naturally infected by E. coelomanticum (the Group A), and 18 from uninfected animals (the Group B). Infected animals showed an average of 532 parasites per pancreas. In the pancreatic histology, ducts displayed hyperplasia, stenosis, proliferation of fibrous tissue, and interstitial inflammatory infiltration of lymphocytes. The serum from infected animals showed an increase in NTPDase activity when ATP was used as substrate (P < 0.001). For the ADP substrate, there was no difference between groups regarding NTPDase activity (P = 0.37), as well as 5′-nucleotidase activity (P = 0.27). Correlating NTPDase activity (ATP substrate) with the degree of histopathological lesions (rho = 0.66, P < 0.001) and the parasitic load on the pancreas (rho = 0.65, P < 0.001), a positive correlation was observed. Similar results were found between the degree of histopathological lesions and NTPDase activity (ADP substrate; rho = 0.29, P = 0.03), and 5′nucleotidase activity (rho = 0.35, P = 0.01). Based on the results of NTPDase and 5′nucleotidase enzymes in cattle naturally infected by E. coleomanticum, it is possible to suggest that these enzymes are involved in the modulation of inflammation, and they can act as markers of inflammatory response.     

Humoral immune response of mice infected with low doses of Trichinella spiralis muscle larvae

Serological techniques are frequently used to detect parasite status and to monitor epidemiology and disease prevalence in important reservoir hosts of zoonotic diseases. Small mammals present the most important link in the epidemiological chain in the spread of trichinellosis. In experimental studies, high infective doses are used to provoke strong immune response of laboratory animals. Wild animals, however, could be infected with very low numbers of Trichinella larvae. The aim of this work was to reveal the size of infective doses that can evoke an adequate immune response with detectable level of specific antibodies in mice. Sixty inbred (Balb/c) mice were infected with 50 L1 and 60 outbred (ICR) mice were infected with 5 L1 T. spiralis. The total larval burdens (TLB) in the intestinal and muscle phases, reproductive capacity index (RCI), and the kinetics of development of specific antibodies by iELISA with different conjugates were determined. In the first 10 days post infection (dpi), more adults were found in the intestines of inbred mice. In both mice strains, the first muscle larvae were observed at 20 dpi. The RCI was significantly higher in outbred mice. Sero-conversion of IgM antibodies was detected at 30 dpi. The IgG antibodies appeared at 40 dpi in inbred mice, and at 50 dpi in outbred mice. Using a polyvalent conjugate, the earliest sero-conversion was recorded at 30 dpi. Antibody levels increased until the end of the experiment (80 dpi). Our results support the suitability of ELISA in large epidemiological surveys to detect low-level infection in naturally infected small mammals, and are useful in epidemiological studies of the sylvatic circulation of trichinellosis to determine likely modes of transmission.     

Trichinella species circulating among wild and domestic animals in Romania

Trichinellosis is one of the most important zoonotic diseases in Romania. Even though the disease is a serious public health concern, only a limited number of Trichinella isolates have been identified at the species level; in the past, all larvae were assumed to be Trichinella spiralis. The present study was conducted to identify Trichinella spp. circulating among wild and domestic animals in Romania, using PCR-based methods. Trichinella spp. larvae originating from 54 wild and 23 domestic mammals were examined. No Trichinella spp. larvae were detected in muscle samples of 182 birds. T. spiralis and Trichinella britovi were the only two species identified in the 40 isolates that yielded a positive PCR result. Overall, T. britovi was more prevalent (n = 26; 65%) than T. spiralis (n = 14; 35%). T. spiralis was the predominant species found in domestic animals (n = 9; 75%), while T. britovi was more prevalent in wildlife (n = 24; 86%). No mixed infections were found. The highest prevalence of Trichinella infection was detected in wolves (11/35; 31%), in European wild cats (4/28; 14%), and in red foxes (5/71; 7%). The distribution of Trichinella spp. in Romania does not show a species-specific clustering; both of the two species found were present over the entire range of counties studied.     

Immune and inflammatory responses in pigs infected with Trichuris suis and Oesophagostomum dentatum

The aim of the present study was to investigate parasite induced immune responses in pigs co-infected with Trichuris suis and Oesophagostomum dentatum as compared to mono-species infected pigs. T. suis is known to elicit a strong immune response leading to rapid expulsion, and a strong antagonistic effect on O. dentatum populations has been observed in co-infected pigs. Forty-eight helminth naïve pigs were allocated into 4 groups in a 2-factorial design. Two groups were trickle inoculated with either 10 T. suis eggs/kg/day (Group T) or 20 O. dentatum L3/kg/day (Group O). Group OT was infected with same levels of both T. suis and O. dentatum (Group OT) and Group C remained uninfected. In each group, six pigs were necropsied after 35 days and the remaining pigs after 71 days. Parasite E/S-antigen specific serum antibodies were quantified by an in-direct ELISA. qPCR was used to measure the expression of immune function related genes in the mucosa of proximal colon and the draining lymph node. Highly significant interactions were identified for O. dentatum specific IgG1 (p < 0.0001) and IgG2 (p < 0.0006) antibodies with a remarkable 2-fold higher antibody response in group OT pigs as compared to group O. These findings indicated that T. suis enhanced the antibody response against O. dentatum in Group OT. The gene expression data confirmed a strong Type 2 response to T. suis (e.g. marked increase in IL-13, ARG1 and CCL11) and clearly weaker in amplitude and/or delayed onset response to O. dentatum in the single infected group. Interactions were found between the two nematodes with regard to several cytokines, e.g. the increase in IL-13 observed in Group T was absent in Group OT (p = 0.06, proximal colon mucosa, 35 and 71 p.i.). Some of these immune response-related interactions may support, or even partially explain, the observed interactions between the two worm populations in co-infected pigs.     

Evaluation of three immunoserological techniques in the detection of porcine trichinellosis

Three immunoserological tests (IST) used for the detection of porcine trichinellosis, immunofluorescence (IF), enzyme-inmunoanalysis (EIA), and Western blot (WB), were compared. Three groups of animals were analyzed: Group 1, animals naturally infected with parasite burdens (PB) of <1 muscle larvae (ML)/g (n = 18); Group 2, animals naturally infected with PB of ≥2 ML/g (n = 23); Group 3, animals raised and home-slaughtered on farms in Argentina (n = 59). Animals from Groups 1 and 2 were identified in outbreaks and were analyzed by individual artificial digestion (AD) of ≥30 g of muscle. Animals in Group 3 were subjected to AD of 5 g of muscle.The detection percentages in sera of swine with the lower PB were 100% for IF, 72% for EIA, and 50% for WB. Eighty-three percent of the animals were serologically positive by two or three techniques. In pigs with the higher PB, the detection percentage was similar for IF and EIA (100% vs. 91%, respectively), and was lower for the WB (61%). Ninety-six percent of the animals were serologically positive by two or three techniques. Group 3 animals had similar detection percentages for the three techniques (IF, 30%; EIA, 29%; WB, 42%). Twenty-five percent of the animals were serologically positive by two or three techniques. Two animals were positive by AD with PB of 0.33 and 2.4 ML/g, and were positive for IF and WB, or IF, EIA, and WB. Results indicate that the sensitivity of each technique depends on the PB, and always ranked in sensitivity as IF > EIA > WB. For the lower PB, the decrease in the sensitivity is more pronounced for the EIA. Although the WB has a low sensitivity, the detection of the specific bands for Trichinella spiralis makes it a useful confirmatory tool. Considering that more than 83% of the parasitologically positive animals had 2 or 3 positive serological results using the techniques tested here, for the diagnosis of porcine trichinellosis, pigs positive by two of these serological techniques must be regarded as truly infected pigs.     

Integrating genomics and phylogenetics in understanding the history of Trichinella species

In 2004, funding was received by Washington University's Genome Sequencing Center through NHGRI, to completely sequence several nematode genomes as part of a holistic effort to advance our understanding of the human genome and evolution within the Metazoa. Trichinella spiralis was among this group of worms because of its strategic location at the base of the phylum Nematoda, and the belief that extant species represented an ancient divergent event that occurred as early as the Paleozoic. At the same time, a concerted effort was put forth to solidify the phylogeny of extant species of Trichinella based upon molecular analyses of a multi-gene system to understand the history of the genus and thereby enhance utilization of the forthcoming sequence data. Since the inception of this research, several findings have emerged: (1) the size of T. spiralis genome estimated by flow cytometry (71.3 Mb) is substantially smaller than originally predicted (270 Mb); (2) to date, a subset of the total of 3,534,683 sequences have been assembled into a 59.3 Mb unique sequence; (3) 19% of the assembled sequence is comprised of repetitive elements; and (4) sequence data are predicated upon extant T. spiralis which probably diverged as little as 20 million years ago. Thus, the utility of the T. spiralis genome as representative of an archaic species must be tempered with the knowledge that encapsulated and non-encapsulated clades probably separated during the mid-Miocene as temperate ecosystems changed.     

Cytokine profiles,apoptosis and pathology of experimental Pasteurella multocida serotype A1 infection in mice

Mice were experimentally infected with Pasteurella multocida serotype A1 to study the cytokine profiles, host cell apoptosis and sequential pathology at different hours of post-infection. Infected mice were dull, anorectic and depressed. A transient leukocytopenia followed by progressive leukocytosis was observed in the course of infection. Serum cytokine profiles showed significantly (P < 0.01) higher amount of pro-inflammatory cytokines (TNF-α, IL-1β, IL-6 and mouse KC) in the infected mice when compared to control mice. The circulating lymphocytes were apoptotic on annexin V staining. Apoptotic nuclei were detected in splenocytes, hepatocytes and infiltrating leukocytes of the lungs on TUNEL staining. The lungs were grossly congested and hemorrhagic, and showed infiltration with polymorphonuclear cells at early and mononuclear cells in the late hours of infection. Alveolar epithelia, inter-alveolar septa and capillary endothelium of the lungs showed ultrastructural changes. Liver had degenerative changes in histological and ultrathin sections.     

Iron metabolism in hamsters experimentally infected with Leptospira interrogans serovar Pomona: Influence on disease pathogenesis

The aim of this study was to analyze the classic iron markers associated to the storage process in hamsters experimentally infected by Leptospira interrogans serovar Pomona. Four groups with six hamsters each were used; two were negative controls (C7 and C14) and two were composed by infected animals (T7 and T14). Blood samples were collected on the seventh (C7 and T7) and fourteenth days (C14 and T14) post-inoculation. Iron availability was determined in sera samples through the assessment of iron, ferritin, transferrin, and iron binding capacity, whereas the bone marrow was also evaluated for the presence of iron by Pearl's reaction. Additionally, the total antioxidant capacity (TAC) and total oxidant status (TOS) were assessed, along with hepcidin and IL-6 levels. Based on the results, it was possible to observe the onset of an anemic profile, predominantly hemolytic and regenerative. Also, The other parameters showed an increase in seric iron (P < 0.01) and ferritin (P < 0.01), and a positive Pearl's reaction in T7 and T14, when compared with the control groups. Transferrin levels decreased (P < 0.05) in animals of T14 with saturation index. TAC was increased in both periods (P < 0.01), while TOS was increased only on T14 (P < 0.05). Hepcidin and IL-6 were increased on T7 and T14 (P < 0.01). Therefore, it was observed that the serum profile from infected animals showed a strong hemolytic pattern, with some demonstration of ferric tissue sequestration when the infection tended to become chronic. The results show that iron metabolism is activated in hamsters infected by L. interrogans serovar Pomona.     

A minimally invasive approach to spleen histopathology in dogs: A new method for follow-up studies of spleen changes in the course of Leishmania infantum infection

Severe forms of zoonotic visceral leishmaniosis (ZVL) are associated with disruption of the spleen structure. However, the study of spleen histology requires splenectomy or necropsy. In this work, we present a minimally invasive cell-block technique for studying spleen tissue histology in dogs with ZVL. We examined 13 dogs with and seven dogs without Leishmania infantum infection. The dogs with Leishmania infection had a lower frequency of lymphoid follicles (2/13, Fisher’s test, P < 0.02) and a higher density of plasma cells (score 3, Fisher’s test, P < 0.02) than uninfected dogs (5/7 exhibiting lymphoid follicles and a plasma cell score of 1). The dogs with Leishmania infection also presented with granulomas (8/13) and infected macrophages (5/13). These differences in the histological presentations of spleen tissue from infected and uninfected dogs corresponded to changes observed in conventional histology. Hence, the cell-block technique described here may be used in the follow-up care and study of dogs with ZVL and other diseases in both clinical practice and research.     

Trichinellosis in wolves from Croatia

The aim of the present study was to investigate the prevalence of Trichinella infection in wolves (Canis lupus) in a 17,468 km² area in Croatia. Muscle samples were collected from 67 wolves between 1996 and 2007 and analyzed by artificial digestion. Muscle larvae were detected in 21 wolves (31%) and genotyped by multiplex PCR. Trichinella britovi was the predominant species confirmed in 90% (19 wolves) while Trichinella spiralis was detected in 9% (2 wolves). The presence of the so called “domestic” Trichinella species was a surprise since, to date, only T. britovi had been reported in wild animals in this region. The larval burdens in infected animals ranged from 0.3 to 45.9 larvae per gram. The prevalence of infected animals varied by geographic region; infected animals were found in the region of Gorski Kotar (20%) which has very similar environment to the region of Lika, where almost all wolves were found infected. Interestingly, this is the first report of infected wolves in Dalmatia.     

Micro-environmental conditions modulate protein secretion and infectivity of the Trichinella spiralis L1 larva

After digestion of infected meat the free L1 of Trichinella spp. penetrate the intestinal mucosa where they moult to the mature adult stage. We have used proteomics to identify changes in protein secretion during in vitro culture of free T. spiralis muscle larvae under different environmental conditions, and to correlate these changes with their infectivity in mice. Muscle larvae were cultured in different media (RPMI-1640, C-199 and HBSS) under conditions of anaerobiosis, microaerobiosis and in 5% CO₂ at 37 °C. Following incubation the larval excretory/secretory proteins were analysed by two-dimensional gel electrophoresis and the larvae were used to orally infect naïve CD1 mice. For all culture media tested, infectivity of the L1 was preserved following incubation in anaerobic conditions. In contrast, the infectivity of worms cultured in nutrient-rich media was almost completely abolished in both microaerobiosis and in the presence of 5% CO₂. Some infectivity was retained in poor or reduced culture media. Comparative analysis of larval infectivity and protein secretion showed that loss of infectivity correlated with the appearance of non-tyvelosylated proteins that in turn may be related to the onset of moulting.     

Immunophenotypical characterization in Andalusian horse: Variations with age and gender

Assessment of lymphocyte subsets is an effective method for characterizing disorders such as leukemia, lymphomas, autoimmune and infectious diseases. In order to clinically interpret these parameters, normal reference values should be set, estimating age- and gender-related variations. This research aimed to: (1) characterize lymphocyte subpopulations in Andalusian horse, and (2) evaluate age and gender-related variations of lymphocyte subsets.Jugular blood samples were obtained from 159 animals, 77 males and 82 females, belonging to four age groups—1: 1–2 years (N = 39; 21 males and 18 females), 2: 2–3 years (N = 38; 16 males and 22 females), 3: 3–4 years (N = 41; 19 males and 22 females) and 4: 4–7 years (N = 41; 21 males and 20 females). T lymphocytes subsets were quantified by flow cytometry with monoclonal antibodies specific for CD2, CD4 and CD8 cell markers. B and NK cell counts were estimated by using a mathematical formula. No variations were found in T, B lymphocytes and NK cells between males and females. Animals of group 1 and 2 had a higher number of CD2, T, CD4+, CD8+, B lymphocytes and NK cells than animals of groups 3 and 4.The percentage of CD2 in group 1 was significantly lower than in group 4. The percentage of T and CD4+ lymphocytes in the group 1 were significantly higher than groups 2 and 3, respectively. Whereas the percentage of B cells calculated by flow cytometry was significantly lower in group 2 compared to group 4, the percentage of B cells calculated by a mathematical formula was higher in group 1. NK cells percentage was significantly lower in group 3 and 4 than in younger animals.In conclusion, in Andalusian horse, gender does not influence absolute numbers and percentages of T, B and NK. There is an age-related decline in absolute number of CD2, T, CD4+ and CD8+ lymphocytes, B lymphocytes and NK cells, with increasing percentage of CD2, T, CD4+ and B lymphocytes, and a decrease in NK with no differences in CD4/CD8 ratio. The decline of lymphocyte population numbers with age is a natural process in many animal species, and could be the origin for immune dysfunction observed in geriatric individuals.     

Effects of inoculum size on cell-mediated and humoral immune responses of foals experimentally infected with Rhodococcus equi: A pilot study

The objective of this pilot study was to compare the cytokine profile as well as cell-mediated and antibody responses of foals infected with a low inoculum of virulent Rhodococcus equi resulting in subclinical pneumonia to that of foals infected with a high inoculum resulting in severe clinical pneumonia. The mean (±SD) ratio of post-infection to pre-infection anti-R. equi IgG(T) concentration was significantly (P = 0.002) higher in foals infected with the high inoculum (195 ± 145; range 62–328) compared to foals infected with the low inoculum (3.9 ± 4.5; range 0.5–11). Similarly, mean (±SD) ratio of post-infection to pre-infection IgM concentration was significantly (P = 0.002) higher in foals infected with the high inoculum (12 ± 4.0; range 7.4–14) compared to foals infected with the low inoculum (2.5 ± 1.5; range 1.2–4.7). Proliferative responses to R. equi antigens as well as expression of mRNA for IL-2, IL-4, IL-10, and IFN-γ in BLN were not significantly different between the two groups. There was a tendency (P = 0.073) towards a higher IFN-γ/IL-4 ratio in the low inoculum group. This study demonstrates that the size of inoculum modulates the IgG subisotype response and possibly the cytokine profile of foals.     

Expansion of intracellular IFN-γ positive lymphocytes during Mycoplasma agalactiae infection in sheep

A method to assess the expansion of antigen-specific intracellular IFN-γ positive T cell subsets during the infection will be helpful for a better understanding of mycoplasmal infections physiopathology in the sheep. We analysed the percentage of antigen-specific lymphocytes positive for intracellular IFN-γ during the infection of sheep with Mycoplasma agalactiae by culturing peripheral blood mononuclear cells of infected or uninfected animals with irradiated M. agalactiae. The expansion of antigen-specific IFN-γ positive lymphocytes in infected sheep was initially sustained by CD4⁺ T cells at day 15 after infection, when antigen specific IgG start to be detectable, followed by CD8/IFN-γ double positive cells. γδ T-cells were not expanded at any time point analysed. IFNγ⁺ T cells disappear 60 days after infection, suggesting that antigen specific IFNγ⁺ T cells, mainly detected in the early phase of the disease, could be useful to understand the role of cell-mediated immunity during M. agalactiae infection.     

Pathogenesis of Trypanosoma (brucei) evansi in small East African goats

Trypanosoma evansi is the cause of surra, a camel disease which is the most important single cause of economic losses in camel rearing areas. Sheep and goats herded with camels are the most likely hosts for T evansi. Upon intravenous infections goats developed erratic parasitaemia, lost weight and their packed cell volume dropped significantly (P<0–001). Trypanosomes were demonstrated by direct microscopy in extravascular locations such as synovial, peritoneal and cerebrospinal fluids and also in lymph by subinoculations into mice. The carcases were emaciated and pale. Histologically there was lymphatic tissue hyperplasia, muscular atrophy and nephrotic changes. Two animals had necrotic foci in the liver, kidneys, lymph nodes, spleen and lungs and also bronchopneumonia. Histologically there was depopulation of lymphocytes in lymphatic tissues, destruction of hepatocytes in the liver with infiltration by inflammatory cells in the liver, lymph nodes, spleen and the kidneys.     

Evaluation of serological cross-reactivity between canine visceral leishmaniasis and natural infection by Trypanosoma caninum

In order to evaluate if the presence of Trypanosoma caninum can lead to a confuse diagnosis of canine visceral leishmaniasis (CVL), we investigated the serological status of dogs infected by T. caninum and assessed the serological cross-reactivity with CVL. A set of 117 serum samples from dogs infected by T. caninum, Leishmania chagasi and not infected dogs (n = 39 in each group) was tested using commercial kits – indirect immunofluorescence (IFI-LVC), ELISA (EIE-LVC) and immunochromatographic test (DPP) – and in house tests with T. caninum (IIF-Tc and ELISA-Tc) and L. chagasi antigens (IIF-Lc and ELISA-Lc). IIF-Tc and ELISA-Tc presented sensitivity of 64.1% and 94.9% and specificity of 23.1% and 35.9%, respectively. The sensitivity of the IFI-LVC, EIE-LVC and DPP tests was 100% and the specificity was 70.5%, 68% and 97.5% respectively. The concordance between the tests was considered as satisfactory. The specificities of IFI-LVC, EIE-LVC and DPP were higher when the group Tc was excluded, with significant values for IFI-LVC (χ² = 4.36, P-value = 0.036), thus suggesting that the infection by T. caninum can confuse the diagnosis of CVL.