Trypanosoma vivax in and outside cattle blood: Parasitological, molecular, and serological detection, reservoir tissues, histopathological lesions, and vertical transmission evaluation.

This study reports assessment of the sensitivity of diagnostic techniques to detect T. vivax in experimentally infected cattle. Additionally, it describes T. vivax extravascular parasitism during the acute and chronic phases of trypanosomosis and congenital transmission. The T. vivax diagnosis was compared using blood samples collected from the jugular, coccygeal and ear tip veins. For this study, 13 males and two females were infected with ≈ 1 × 106 viable T. vivax trypomastigotes (D0). One animal was kept as a negative control during the entire study. The 13 infected males were euthanized between 14 and 749 days post-infection (DPI). After confirming the cyclicity of both females (9 months of age), they were naturally mated with a bull. One female was euthanized at 840 DPI, and the other at 924 DPI. The two calves, one from each female, were euthanized at six months of age (924 DPI), and the negative control at 924 DPI. During this period, T. vivax in blood was assessed using direct methods (Woo test, cPCR, microscopic examination of fresh wet blood films and parasite quantification - Brener method), and serological methods (IFAT, ELISA, and IA). Tissue samples were collected from the liver, spleen, brain, cerebellum, heart, testicles, epididymis, kidneys, eyeballs, pre-scapular lymph nodes, ear tips, mammary glands, uterus, and ovaries. The protozoan DNA was examined using LAMP. There was no difference in the detection of T. vivax using the Woo test and Brener method among the jugular, coccygeal, and ear tip veins. The sensitivity of the detection methods varied depending on the disease phase. Direct methods (Woo test, Brener method, and cPCR) demonstrated higher sensitivity during the acute phase, while serological methods (IFAT, ELISA, and IA) were more sensitive during the chronic phase. Anti-T. vivax antibodies were detected up to 924 DPI. Tissue evaluation using LAMP demonstrated the presence of T. vivax DNA and associated histopathological changes up to 840 or 924 DPI. Only in mammary glands and ovaries was no DNA detected. The most frequently observed histopathological alteration was lymphohistioplasmocytic inflammatory infiltrate. No transplacental transmission of T. vivax was observed.

Evaluation of mechanical transmission of Trypanosoma vivax by Stomoxys calcitrans in a region without a cyclic vector.

This work investigated the mechanical transmission of Trypanosoma vivax by Stomoxys calcitrans to cattle in a region without a cyclic vector. The study involved two experiments, one with calves experimentally infected with T. vivax, in the acute phase of trypanosomosis (Experiment 1) and the other in the chronic phase (Experiment 2). In both experiments, two transmission methods were used with flies that had not fed for 24 h or had never fed: (i) Method 1: flies released freely in cattle pens (≈3,300 flies/pen for 10 days); and (ii) Method 2: flies placed in a feeding chamber (12 flies/animal). To develop Method 1 in the two experiments (acute and chronic phases), T. vivax-positive animals were kept with T. vivax-negative animals. Periodically, the Brener method, Woo method, blood smears, cPCR, ELISA, IFAT, and Imunoteste® were performed to detect T. vivax in the animals. We also recorded the animals' head tossing and hoof stomping and the number of flies near the pens' inner walls. Subsequently, biological testing was performed using lambs. For Method 2 in both experiments, flies inside the feeding chamber first fed on T. vivax-positive animals and later on negative animals. In both experiments and methods, we examined the flies for the presence of T. vivax through blood smears and cPCR of the proboscis and abdomen. In Experiment 2 (chronic phase), a test was conducted to determine how long trypomastigotes forms could survive on the blood of animals with different levels of parasitemia. None of the animals (calves and lambs) became infected with T. vivax or showed antibodies against it. During the evaluation period, the animals in the presence of the flies exhibited more hoof stomping and head tossing compared to those without flies (control). Additionally, there was an increase in the number of flies in the pens during the experiment. Only in Experiment 1 (acute phase) were T. vivax trypomastigotes and DNA found in the abdomen of the flies but not in the proboscis. In Experiment 2 (chronic phase), higher concentrations of trypomastigotes per milliliter of blood were associated with a shorter the lifespan of this stage of the parasite. In conclusion, under the variable conditions of the experiments (hosts, number of flies, and level of parasitemia), S. calcitrans was unable to mechanically transmit T. vivax to cattle.

"Lysis and Concentration Technique" improves the parasitological diagnosis of Trypanosoma vivax.

The "Lysis and Concentration Technique" (LCTe) involves lysing red blood cells and concentrating parasites to increase the chances of visualization in low parasitemia and in scenarios of evaluators with less knowledge. The lysis of red blood cells reduced the time of diagnosis by 21 s, showing that the effect produced by the treatment is comparable to the effect of the experience in parasitological examination. In addition, the concentration of parasites was 39.18% higher for slides with high parasitemia and 131.03% for slides with low parasitemia in relation to the standard slide. LCTe proved to be inexpensive, with a total cost of approximately US$0.07 per slide made, which allows it to be easily implemented in most laboratories.

Presence of Trypanosoma vivax DNA in cattle semen and reproductive tissues and related changes in sperm parameters.

The present work investigated the presence of Trypanosoma vivax in semen and reproductive tissues of experimentally infected cattle and evaluated changes in seminal parameters. Two groups of cattle were established: T01 - experimentally infected with T. vivax (n = 8) and T02 - not experimentally infected with T. vivax (n = 8). After infection, blood (every seven days until 182 days post-infection - DPI), semen (7, 14, 35, 56, 70, 120 and 182 DPI) and reproductive tissue (after euthanasia, 182 DPI) were collected to search for T. vivax using different techniques, including PCR, Woo and Brener. Seminal parameters, including turbulence, motility, concentration, and vigor, were also analyzed. Packed cell volume (PCV) of the animals was determined weekly and weight gain was calculated. The PCR revealed T. vivax DNA in 7/56 semen samples of post-infection T01 cattle. Trypanosoma vivax DNA was detected in the semen of 5/8 animals at 7, 14, 56, 70 and 120 DPI, in the testis of four, and in the epididymis and fat located around the testis of two others. Trypomastigote forms of T. vivax were not found in any semen sample. Sperm of T01 cattle had lower turbulence (p ≤ 0.05) at 7, 14, 35, 56, 120 and 182 DPI, lower vigor (p ≤ 0.05) at 120 DPI and more sperm abnormalities (p ≤ 0.05) than T02. Digital dermatitis was observed among T01 cattle. Animals of T01 had lower PCV values than did those of T02 for most of the evaluations performed and T02 animals gained more weight during the experiment. The results highlight the presence of T. vivax DNA in semen of infected cattle and the importance of this disease for male breeding cattle. Further research is needed to determine whether T. vivax can be sexually transmitted in cattle.

How many cattle can be infected by Trypanosoma vivax by reusing the same needle and syringe, and what is the viability time of this protozoan in injectable veterinary products?

It was investigated how many cattle become infected with Trypanosoma vivax by subcutaneous (SC), intramuscular (IM) and intravenous (IV) routes, using the same syringe and needle from an animal with acute T. vivax infection. Besides, the T. vivax viability in 109 injectable veterinary drugs (antibiotics, antiparasitics, reproductive hormones, vitamin complex and derivatives, vaccines, anaesthetics, anti-inflammatory/antipyretics, antitoxics). In the field assay, four groups were performed: T01, T02 and T03 animals that received saline solution with the same syringe and needle contaminated with T. vivax via SC, IM and IV routes, respectively, and T04 control animals that received only saline solution with the same syringe and needle IV. In the laboratory, drugs had their pH measured and T. vivax viability verified. The number of cattle infected with T. vivax via SC (3/20) was lower (P ≤ 0.05) compared to via IM (9/20), which was lower (P ≤ 0.05) compared to IV (15/20). The solution pH did not influence T. vivax viability. In 44% (48/109) of the products, T. vivax remained viable regardless of time, stooding out that in 100% of oxytocins the protozoan was verified, at some evaluation times. The mean of T. vivax quantified in foot-and-mouth and brucellosis vaccines and in doramectin-based products were higher (P ≤ 0.05) than found in blood + saline solution.

A Preliminary Study on the Relationship between Parasitaemia and Cytokine Expression of Peripheral Blood Cells in Trypanosoma vivax-Experimentally Infected Cattle.

Trypanosoma vivax outbreaks have been reported with increasing frequency worldwide, causing significant economic losses in livestock. Though several studies have suggested that cytokine responses may influence infection caused by Trypanosoma sp., their exact role remains unclear and may vary according to the animal species and parasite strain. The present study aimed to evaluate cytokine expression of peripheral blood cells from three Girolando dairy cows experimentally infected with T. vivax. For this purpose, blood samples were collected prior to the inoculation on the day of inoculation (D0), the day after inoculation (D1), and then every seven days up to 119 days after infection (DAI). Each animal presented a unique pattern of cytokine expression. While a tendency of a Th1 cytokine response was observed during the patent phase (presence of circulating parasites), an increase of Th2 cytokine expression was found at the beginning of the sub-patent phase (low parasitaemia or aparasitaemic periods). In animals that presented a better control of parasitaemia, IL-6 and IFNγ increased during most of the trial period. On the other hand, the cow that presented reduction of IL-1ß, IL-2, and TNFα during the entire period did not control parasitaemia properly. A balance between the Th1 and Th2 profile is beneficial for parasite control and animal health. The results found in the present study are a first step towards elucidating the dynamics of cattle's inflammatory response against T. vivax, requiring future studies focusing on the role of key cytokines on the controlling of parasitaemia in different stages of bovine trypanosomosis.

In vitro and in vivo effectiveness of disinfectants against Trypanosoma vivax.

Trypanosoma vivax causes bovine trypanosomosis in cattle and resulting in economic losses to farmers. In Brazil, shared contaminated materials is the main transmission pathway. To evaluate the effectiveness of different disinfectants for T. vivax, in vitro and in vivo analyses were performed. At the laboratory, 21 disinfectants were tested. The disinfectants were placed in microtubes containing blood with approximately 1.0 × 106 trypomastigotes of T. vivax. The viability and motile of trypomastigotes after 30 s, one, 10, 15 and 30 min was evaluated by the thick drop method and the efficacy calculated. Disinfectants that showed 100% effectiveness were used in in vivo tests. Thirty calves negative for T. vivax were divided into six groups and were inoculated with disinfectant solutions (46% alcohol, 70% alcohol, or 0.5% iodine) + 1 × 106 trypomastigotes of the protozoa. Blood from each animal was collected at seven, 14 and 21 days after inoculation to verify the viability and presence of the protozoan by Woo, Brener, PCR, and LAMP methods. In the in vitro step, 13 of the 21 disinfected solutions exhibited 100% effectiveness against T. vivax at all evaluation times. In contrast, 70% alcohol and 0.5% iodine solutions exhibited 100% effectiveness in the in vivo tests and can be used to disinfect needles and syringes. The use of disinfectants is a rapid and efficient procedure to disinfect materials utilized in the field and concomitantly could help to reduce the dissemination of T. vivax in the cattle herd in cases of iatrogenic transmission.

Delayed reduction of Anaplasma marginale parasitemia and packed cell volume normalization despite prolonged enrofloxacin treatment of cattle co-infected with Trypanosoma vivax.

Although co-infections of Trypanosoma vivax, Anaplasma spp., and Babesia spp. have been reported, knowledge gaps remain that need to be addressed. The present study evaluated the efficacy of enrofloxacin (7.5 mg/kg) against A. marginale in naturally infected cattle and cattle experimentally co-infected with T. vivax by observation of the variation in A. marginale parasitemia and packed cell volume (PCV) for 39 days. Bovines were distributed into two groups, each with six calves: T01 = animals immunosuppressed with dexamethasone and with latent anaplasmosis; T02 = animals immunosuppressed with dexamethasone, with latent anaplasmosis and experimentally co-infected with T. vivax on day 0 (D0). Animals of both groups were immunosuppressed with dexamethasone and received enrofloxacin (7.5 mg/kg) whenever mean values of parasitemia for A. marginale were ≥ 5% per group. Cattle of group T02 were also treated with isometamidium chloride (0.5 mg/kg) on D25. On D17 and D22 to D28 of the study, there was a higher (P ≤ 0.05) A. marginale parasitemia in animals of T02 than in those of T01. Animals of T01 required one enrofloxacin treatment to decrease A. marginale parasitemia, while those from T02 needed five treatments. From D5 to D37 of study, the mean values of PCV for calves from T02 were lower (P ≤ 0.05) than that for calves from T01. In conclusion, bovines co-infected T. vivax needed four more treatments with enrofloxacin to reduce A. marginale parasitemia and keep PCV values within reference standards.

Follow-up of dairy cattle naturally infected by Trypanosoma vivax after treatment with isometamidium chloride.

Trypanosoma vivax infections cause nonspecific clinical signs in cattle associated with aparasitemic intervals, making disease diagnosis a challenge. In Brazil, diminazene aceturate and isometamidium chloride (ISM) are available to treat bovine trypanosomosis. The objective of this study was to follow-up, by molecular and serological techniques, dairy cattle naturally infected by T. vivax after ISM treatment. Thirty cattle naturally infected with T. vivax received two applications of ISM, at a dosage of 1.0 mg/kg intramuscularly, on days 0 and 150. For T. vivax diagnosis, EDTA-blood and serum samples were evaluated on 0, 7, 15, 30, 60, 90, 120, 150, 180, 210, and 240 days after treatment PCR, Loop-mediated isothermal amplification (LAMP) and ELISA. Animals with persistent detection of T. vivax DNA by both PCR and LAMP were found and continuous detection of anti-T. vivax IgG antibodies by ELISA, suggesting the presence of T. vivax resistance to ISM. The combination of LAMP and ELISA tests can prevent misdiagnosis of the parasite clearance in treated cattle, contributing to better disease control. This is the first experiment that demonstrates the persistence infection of T. vivax under ISM treatment in a natural infected herd and evidence of ISM chemotherapy-resistant T. vivax in Brazil.

Follow-up of dairy cattle naturally infected by Trypanosoma vivax after treatment with isometamidium chloride / Acompanhamento de bovinos leiteiros naturalmente infectados com Trypanosoma vivax após tratamento com cloreto de isometamidium

Abstract Trypanosoma vivax infections cause nonspecific clinical signs in cattle associated with aparasitemic intervals, making disease diagnosis a challenge. In Brazil, diminazene aceturate and isometamidium chloride (ISM) are available to treat bovine trypanosomosis. The objective of this study was to follow-up, by molecular and serological techniques, dairy cattle naturally infected by T. vivax after ISM treatment. Thirty cattle naturally infected with T. vivax received two applications of ISM, at a dosage of 1.0 mg/kg intramuscularly, on days 0 and 150. For T. vivax diagnosis, EDTA-blood and serum samples were evaluated on 0, 7, 15, 30, 60, 90, 120, 150, 180, 210, and 240 days after treatment PCR, Loop-mediated isothermal amplification (LAMP) and ELISA. Animals with persistent detection of T. vivax DNA by both PCR and LAMP were found and continuous detection of anti-T. vivax IgG antibodies by ELISA, suggesting the presence of T. vivax resistance to ISM. The combination of LAMP and ELISA tests can prevent misdiagnosis of the parasite clearance in treated cattle, contributing to better disease control. This is the first experiment that demonstrates the persistence infection of T. vivax under ISM treatment in a natural infected herd and evidence of ISM chemotherapy-resistant T. vivax in Brazil.

Resumo Em bovinos, infecções por Trypanosoma vivax geram sinais clínicos inespecíficos que, associados a intervalos aparasitêmicos, faz com que o diagnóstico da enfermidade seja desafiador. No Brasil, somente aceturato de diaminazeno e cloridrato de isometamidum (ISM) estão disponíveis para o tratamento da tripanossomose bovina. Este trabalho teve como objetivo acompanhar bovinos leiteiros naturalmente infectados por T. vivax, após o tratamento com ISM por meio de técnicas moleculares e sorológica. Foram utilizados 30 bovinos naturalmente infectados com T. vivax, sendo estes tratados com duas aplicações de ISM, na dosagem de 1,0 mg/kg por via intramuscular profunda, nos dias 0 e 150. Foram avaliadas, para diagnóstico de T. vivax, amostras de sangue acrescido de EDTA e soro, colhidas nos 0, 7, 15, 30, 60, 90, 120, 150, 180, 210 e 240 dias após os tratamentos pela reação em cadeia da polimerase (PCR), amplificação circular isotérmica do DNA (LAMP) e ensaio de imunoabsorção enzimático (ELISA). Verificou-se a presença de animais com persistência na detecção de DNA de T. vivax pela PCR e LAMP, bem como detecção contínua de anticorpos IgG anti-T. vivax pelo método de ELISA, sugerindo a presença de resistência de T. vivax ao ISM. A combinação dos testes LAMP e ELISA pode evitar falsos diagnósticos da eliminação do parasita nos bovinos tratados, contribuindo para um melhor controle da doença. Este é o primeiro experimento que demonstra infecção persistente do T. vivax em rebanho naturalmente infectado, tratado com ISM, e evidencia possível resistência ao quimioterápico no Brasil.

Epidemiological and molecular identification of Trypanosoma vivax diagnosed in cattle during outbreaks in central Brazil.

Bovine trypanosomosis has been spreading in Brazil. In the present study, we evaluated the spatial distribution, prevalence and risk factors of this disease in the state of Goiás, Brazil, and performed both molecular and phylogenetical analyses of Trypanosoma vivax. A total of 4049 blood samples were collected from cattle for a period of 2 years. The parasitological diagnosis was performed using the Woo method and a questionnaire was administered to the farmers to document risk factors associated with the disease in the herd. Positive samples were DNA sequenced and compared to GenBank codes. The prevalence of T. vivax was 8.84%, occurring on 24 ranches only in dairy cattle and mainly in the central and southern portions of the state. The acquisition of new animals infected with T. vivax and the administration of exogenous oxytocin to cows using the same syringe and needle were the main associated factors (P ≤ 0.05). After an outbreak, milk production decreased by 39.62%. The presence of biting flies (tabanids, Haematobia irritans and Stomoxys calcitrans) was not a risk factor (P > 0.05) for the occurrence of T. vivax. The epidemiological data demonstrate the importance of restricting the practice of auctions as well as eliminating the use of exogenous oxytocin in animals during milking. The samples tested by polymerase chain reaction were positive for T. vivax and were genetically homologous with T. vivax found in different states of Brazil and west Africa based on the 18S rRNA gene.

Comparison of therapeutic efficacy of different drugs against Trypanosoma vivax on experimentally infected cattle.

In this study, we evaluated the therapeutic efficacy of diminazene diaceturate at a dose of 7 mg/kg (DA), imidocarb dipropionate at 4.8 mg/kg (IMD), isometamidium chloride at 0.5 and 1.0 mg/kg (ISM 0.5 and ISM 1.0) and combinations applied through different methods to treat Trypanosoma vivax in experimentally infected calves. Thirty male Girolando calves were kept indoors and infected intravenously with T. vivax trypomastigotes (approximately 1 × 106). On D-1, the calves were randomized based on the quantity of infecting parasites per animal, yielding six groups of five animals each: G1: positive control group without treatment; G2 animals treated with DA on Day 0 intramuscularly (IM); G3 animals treated with IMD on Day 0 and D + 14 subcutaneously; G4 animals treated with ISM 0.5 on Day 0 IM; G5 animals treated with ISM 1.0 on Day 0 IM; G6 animals received DA on Day 0 and ISM 1.0 on D + 14, both IM. Throughout 180 days, blood samples were collected for the evaluation of T. vivax using the Woo, Brener and PCR methods. The results indicated that the treatment protocols with DA and/or ISM 0.5 and ISM 1.0 had high efficacy (100 %) against T. vivax. Interestingly, cattle that received ISM remained free of parasites until D + 180. In contrast, animals treated with IMD had relapsed T. vivax detected on the 10th and 14th days post-treatment (DPT). Cattle that received ISM 1.0 did not exhibit relapsed T. vivax in the blood, even after reinfection performed on the 50th DPT. However, treatment with DA on Day 0 failed to prevent a new infection of T. vivax on the 50th DPT. The animals that received ISM 1.0 had a transient decrease in packed cell volume similar to that found in the control group. The reappearance of T. vivax in herds in Brazil treated with DA likely occurred due to the short half-life of the drug and not necessarily due to T. vivax resistance to DA.

Infection capacity of Trypanosoma vivax experimentally inoculated through different routes in bovines with latent Anaplasma marginale.

The aim of the present study was to compare the infection capacity of Trypanosoma vivax experimentally inoculated through different routes in calves naturally infected with latent Anaplasma marginale. On Day 0 of the study, 25 calves (breed: Girolando) were divided into five groups. The first four groups of five calves each received approximately 1 × 106 trypomastigotes of T. vivax through the intradermal, subcutaneous, intramuscular and intravenous routes. Another five animals remained unaffected to serve as A. marginale naturally infected controls. The study of T. vivax was performed on all calves from D+1 to D+30 using the Woo, Brener and blood smear techniques. PCR was performed on Days +1, +3, +4, +5, +28, +29 and + 30. The results indicated that T. vivax was capable of infecting and developing the disease in the calves independent of the inoculation route. A positive correlation was found between T. vivax and rectal temperature (P ≤ 0.05) and a negative correlation was seen between the protozoan and globular volume (P ≤ 0.05). Latent A. marginale in the calves acted as co-infection for T. vivax. Jaundice was seen only in calves with a high parasitemia by A. marginale. Therefore, in areas with the confirmed presence of T. vivax in bovines, this protozoan should be included in the complex denominated "Bovine Parasitic Sadness", which currently encompasses only Anaplasma and Babesia.

Comparison of conventional and molecular techniques for Trypanosoma vivax diagnosis in experimentally infected cattle.

Livestock infections by Trypanosoma vivax have been occurring with increasing frequency, mainly due to the presence of animals with subclinical infections and without apparent parasitaemia, making diagnosis challenging. The aim of the present study was to evaluate several techniques used for T. vivax diagnosis in order to assess the best way of using them during the course of the disease. Molecular methods demonstrated higher rates of detection than parasitological methods, detecting 33 of the 54 (61.1%) known positive samples, while the hematocrit centrifugation technique (best parasitological test) detected only 44.4%. The serological methods, IFAT and ELISA, detected seropositivity in 51 of the 54 (94.4%) and 49 of the 54 (90.7%) known positive samples, respectively. Despite being highly sensitive, the latter only demonstrates exposure to the infectious agent and does not indicate whether the infection is active. The present study was the first to use the qPCR for a South American isolate, improving disease detection and quantification. Furthermore, the analyses revealed that the patent phase of the disease may extend up to 42 days, longer than previously reported. The combination of several diagnostic techniques can lower the frequency of false negative results and contributes toward better disease control.

Comparison of conventional and molecular techniques for Trypanosoma vivax diagnosis in experimentally infected cattle / Comparação de técnicas convencionais e moleculares para diagnóstico em bovinos experimentalmente infectados por Trypanosoma vivax

Abstract Livestock infections by Trypanosoma vivax have been occurring with increasing frequency, mainly due to the presence of animals with subclinical infections and without apparent parasitaemia, making diagnosis challenging. The aim of the present study was to evaluate several techniques used for T. vivax diagnosis in order to assess the best way of using them during the course of the disease. Molecular methods demonstrated higher rates of detection than parasitological methods, detecting 33 of the 54 (61.1%) known positive samples, while the hematocrit centrifugation technique (best parasitological test) detected only 44.4%. The serological methods, IFAT and ELISA, detected seropositivity in 51 of the 54 (94.4%) and 49 of the 54 (90.7%) known positive samples, respectively. Despite being highly sensitive, the latter only demonstrates exposure to the infectious agent and does not indicate whether the infection is active. The present study was the first to use the qPCR for a South American isolate, improving disease detection and quantification. Furthermore, the analyses revealed that the patent phase of the disease may extend up to 42 days, longer than previously reported. The combination of several diagnostic techniques can lower the frequency of false negative results and contributes toward better disease control.

Resumo Infecções por Trypanosoma vivax têm ocorrido com frequência crescente em animais de produção, principalmente pela aquisição de animais com infecções subclínicas e sem aparente parasitemia, o que dificulta o diagnóstico. O objetivo do presente estudo foi avaliar várias técnicas empregadas para o diagnóstico de T. vivax, a fim de verificar a melhor maneira de utilizá-las durante o curso da doença. Os métodos moleculares demonstraram maiores taxas de detecção que os métodos parasitológicos, detectando 33 das 54 (61,1%) amostras sabidamente positivas, enquanto a técnica de hemoconcentração (melhor teste parasitológico) detectou apenas 44,4%. Os métodos sorológicos, RIFI e ELISA, detectaram soropositividade em 51 das 54 (94,4%) e 49 das 54 (90,7%) amostras sabidamente positivas, respectivamente. Apesar de serem altamente sensíveis, estes testes apenas demonstram a exposição ao agente infeccioso, e não indicam se a infecção permanece ativa. O presente estudo foi o primeiro a utilizar a qPCR para um isolado sul-americano, melhorando sua detecção e quantificação. Além disso, as análises revelaram que a fase patente da doença pode se estender por até 42 dias após a infecção, sendo maior que anteriormente relatado. A combinação de várias técnicas de diagnóstico pode evitar a frequência de resultados falso-negativos e contribuir para um melhor controle da doença.

A comparative in silico linear B-cell epitope prediction and characterization for South American and African Trypanosoma vivax strains.

Trypanosoma vivax is a parasite widespread across Africa and South America. Immunological methods using recombinant antigens have been developed aiming at specific and sensitive detection of infections caused by T. vivax. Here, we sequenced for the first time the transcriptome of a virulent T. vivax strain (Lins), isolated from an outbreak of severe disease in South America (Brazil) and performed a computational integrated analysis of genome, transcriptome and in silico predictions to identify and characterize putative linear B-cell epitopes from African and South American T. vivax. A total of 2278, 3936 and 4062 linear B-cell epitopes were respectively characterized for the transcriptomes of T. vivax LIEM-176 (Venezuela), T. vivax IL1392 (Nigeria) and T. vivax Lins (Brazil) and 4684 for the genome of T. vivax Y486 (Nigeria). The results presented are a valuable theoretical source that may pave the way for highly sensitive and specific diagnostic tools.

Evaluation of clinical signs, parasitemia, hematologic and biochemical changes in cattle experimentally infected with Trypanosoma vivax.

Infections by Trypanosoma vivax cause great losses to livestock in Africa and Central and South Americas. Outbreaks due this parasite have been occurred with increasing frequency in Brazil. Knowledge of changes caused by T. vivax during the course of this disease can be of great diagnostic value. Thus, clinical signs, parasitemia, hematologic and biochemical changes of cattle experimentally infected by this hemoparasite were evaluated. Two distinct phases were verified during the infection - an acute phase where circulating parasites were seen and then a chronic phase where fluctuations in parasitemia were detected including aparasitemic periods. A constant reduction in erythrocytes, hemoglobin and packed cell volume (PVC) were observed. White blood cells (WBC) showed pronounced changes such as severe neutropenia and lymphopenia during the acute phase of the illness. Decreases in cholesterol, albumin, aspartate aminotransferase (AST), lactate dehydrogenase (LDH), and increases in glucose, globulin, protein, and alkaline phosphatase (ALP) were observed. The "Lins" isolate of T. vivax showed pathogenicity for cattle, and intense parasitemia was detected in the early stages of infection. Circulating parasites were detected for about two months. The most evident laboratory abnormalities were found in WBC parameters, including thrombocytopenia.

Evaluation of clinical signs, parasitemia, hematologic and biochemical changes in cattle experimentally infected with Trypanosoma vivax / Avaliação dos sinais clínicos, parasitemia e alterações hematológicas e bioquímicas de bovinos experimentalmente infectados peloTrypanosoma vivax

Abstract Infections by Trypanosoma vivax cause great losses to livestock in Africa and Central and South Americas. Outbreaks due this parasite have been occurred with increasing frequency in Brazil. Knowledge of changes caused byT. vivax during the course of this disease can be of great diagnostic value. Thus, clinical signs, parasitemia, hematologic and biochemical changes of cattle experimentally infected by this hemoparasite were evaluated. Two distinct phases were verified during the infection – an acute phase where circulating parasites were seen and then a chronic phase where fluctuations in parasitemia were detected including aparasitemic periods. A constant reduction in erythrocytes, hemoglobin and packed cell volume (PVC) were observed. White blood cells (WBC) showed pronounced changes such as severe neutropenia and lymphopenia during the acute phase of the illness. Decreases in cholesterol, albumin, aspartate aminotransferase (AST), lactate dehydrogenase (LDH), and increases in glucose, globulin, protein, and alkaline phosphatase (ALP) were observed. The “Lins” isolate of T. vivax showed pathogenicity for cattle, and intense parasitemia was detected in the early stages of infection. Circulating parasites were detected for about two months. The most evident laboratory abnormalities were found in WBC parameters, including thrombocytopenia.

Resumo Infecções pelo Trypanosoma vivax causam grandes prejuízos à pecuária na África e Américas Central e do Sul. Surtos devido a este protozoário têm ocorrido com frequência cada vez maior no Brasil. O conhecimento das alterações provocadas pelo T. vivax durante a evolução desta enfermidade podem ser de grande valia para o auxílio no diagnóstico. Para tanto foram estudados os sinais clínicos, parasitemia, alterações hematológicas e bioquímicas em bovinos experimentalmente infectados por este hemoparasito. Foram verificadas duas fases distintas durante a infecção, uma aguda onde parasitos circulantes foram vistos durante todo o período, e posteriormente uma crônica, onde foram detectadas flutuações na parasitemia, com períodos aparasitêmicos. Foi verificada constante diminuição da contagem global de eritrócitos, teor de hemoglobina e volume globular (VG). O leucograma revelou leucopenia por neutropenia e linfopenia durante a fase aguda da enfermidade. Foram observados diminuição do colesterol, albumina, aspartato aminotransferase (AST), lactato desidrogenase (LDH) e aumento da glicose, globulinas, proteínas e fosfatase alcalina (FA). O isolado “Lins” de T. vivax apresentou patogenicidade para bovinos, verificando-se parasitemia intensa nos estágios iniciais da infecção, sendo detectados parasitas circulantes por aproximadamente dois meses. As alterações laboratoriais mais evidentes foram encontradas nos parâmetros do leucograma, ainda destacando-se um quadro de trombocitopenia.

Detection of Trypanosoma vivax using PCR and LAMP during aparasitemic periods.

Trypanosoma vivax affects cattle herds in Africa and Americas and has been spreading rapidly in Brazil, through introduction of animals with subclinical infections and without apparent parasitemia, which makes its diagnosis challenging. PCR and LAMP are effective in detecting the presence of T. vivax DNA in situations of low parasitemia. LAMP is simpler and faster technique than PCR, and can be performed in the field, with limited resources. In this study, the capacities of conventional PCR and LAMP for detecting T. vivax in bovine blood samples classified as aparasitemic were evaluated. The capacity of conventional PCR (56.25%) for detecting positive samples was lower than that of LAMP (93.73%). This may influence the choice of screening tests for cattle herds infected with T. vivax.