Piroplasmid Infections Among Domestic Dogs in the Mountain City of Rio de Janeiro, Brazil.

PURPOSE: The aim of the present study was to analyze the frequency of the piroplasmids in blood from dogs and ticks recovered from these animals in Teresópolis city, located in the mountain region of Rio de Janeiro state, Brazil. In addition to the clinical and hematological profile. METHODS: A total of 400 dogs attended in a veterinary clinic in this city between 2020 and 2021 were included. The blood was collected from the dogs, along with ticks and information on these dogs was obtained through a questionnaire applied to the owners. Thin-smear analyses and complete blood counts were performed. All forms characteristic of piroplasmids were measured and classified morphologically. The blood was also subjected to PCR assays based on the genes 18S rRNA and hsp70. In addition, the ixodid ticks were classified morphologically and subjected to PCR for piroplasmids research. The amplified products were sent for gene sequencing. RESULTS: Piroplasmids were detected in 2.3% of the dogs. The variables statistically associated with infections in these animals were hemorrhage/bleeding, jaundice, anisocytosis, activated monocytes and macroplatelets (p ≤ 0.05). Piriform, ring-shaped, oval and aberrant structures were viewed in erythrocytes, neutrophils and monocytes, with lengths greater than and less than 2.5 µm. The nine positive samples from these dogs were characterized as due to Rangelia vitalii. However, one sequence from B. vogeli was detected in a single adult specimen of R. sanguineus. CONCLUSION: Although circulation of two species of piroplasmids potentially infective for domestic dogs has been observed in the mountain city of Rio de Janeiro, infection due to R. vitalii was mostly seen in the dogs of the present study.

First report of canine rangeliosis in domestic dogs from different regions of Santa Catarina State, Brazil.

Rangeliosis is the disease caused by Rangelia vitalii, a parasite reported in dogs from southeastern and southern Brazil, Uruguay, Paraguay, and Argentina. This protozoan is transmitted by the ixodid Amblyomma aureolatum, and infects erythrocytes, leukocytes, and vascular endothelial cells of the host. The common clinical signs, such as prostration, fever, anemia, thrombocytopenia, anorexia, weight loss, and dehydration, are also found in other infections, like canine babesiosis and ehrlichiosis. The similar clinical presentation with other diseases, as well as the indistinguishable morphology with intraerythrocytic Babesia canis, complicates the disease diagnostic. In the present study, blood samples from dogs presenting clinical signs compatible with hemoparasitosis were investigated for rangeliosis. The dogs were treated at veterinary clinics in the cities of Blumenau and Lages, in the State of Santa Catarina, Brazil. Blood samples from 17 dogs were analyzed by PCR. The samples were screened by a conventional piroplasma-PCR and the positives confirmed by a specific R. vitalii-qPCR. Two animals (2/17; 11.8%) were positive for R. vitalii, one from Blumenau and the other from Lages. Both animals presented unspecific signs of hemoparasitosis, such as apathy, anemia, and anorexia. The results indicate the necessity of molecular assays for the proper identification of the hemoparasite, and to investigate the real prevalence of rangeliosis in the State of Santa Catarina.

Rangelia vitalii in free-living crab-eating foxes (Cerdocyon thous) in Uruguay.

Rangelia vitalii is a protozoan parasite that causes a hemorrhagic and hemolytic disease in dogs known as rangeliosis. Current reports of the disease are concentrated in the southern and southeastern regions of Brazil, as well as in Uruguay, Argentina, and Paraguay, and mainly concern domestic dogs. South American wild canids, such as the crab-eating fox (Cerdocyon thous), the pampas fox (Lycalopex gymnocercus), and the maned wolf (Chrysocyon brachyurus) may also be affected, although existing reports are restricted to Brazil. The present study aimed to detect R. vitalii parasitism in the Uruguayan wild fox population. DNA extracted from the blood and/or spleen samples of road-killed C. thous and L. gymnocercus found in northern Uruguay were subjected to polymerase chain reaction (PCR) to amplify a 551-bp fragment of the Rangelia 18S rRNA gene. A total of 62 wild canids, including 38 C. thous and 24L. gymnocercus, were analyzed. Five crab-eating fox samples (13.2%) were positive for R. vitalii, with 99.5-100% identity between the sequences. All samples from pampas fox tested negative for R. vitalii. When compared with the R. vitalii sequences available in GenBank, a similarity of 98.9-100% was revealed. Molecular analysis results suggest that R. vitalii is circulating in the crab-eating fox population in Uruguay; however, its veterinary relevance for these foxes remains unknown.

Use of molecular tools for the diagnosis of rangeliosis by Rangelia vitalii in Argentina: A case report.

Vector-borne pathogens are responsible for serious emerging diseases and Rangelia vitalii, the etiologic agent of canine rangeliosis, is one of the most pathogenic tick-borne pathogens for dogs in South America. This protozoan is transmitted by the Amblyomma aureolatum tick bite and the clinical features associated to the disease are fever, hemolytic anemia, jaundice, hepatosplenomegaly and bleeding from natural orifices, mainly from the ear egde. The reports of canine rangeliosis in Argentina are scarce. In the present study we report the detection of Rangelia vitalii in a naturally infected dog from Gualeguay, Entre Ríos, Argentina with history of tick infestation and clinical signs compatible with rangeliosis. An initial blood sample was positive to piroplasmids by blood smear examination and the molecular amplification of a fragment of the 18SrRNA gene. Sequencing of the fragment confirmed the pathogen identity. After treatment with imidocarb dipropionate, the clinical signs remitted and the blood smear tested negative.

Implications of domestic dogs in the epidemiology of Rickettsia parkeri strain Atlantic rainforest and Rangelia vitalii in Southeastern Brazil / Implicações dos cães domésticos na epidemiologia da Rickettsia parkeri cepa Mata Atlântica e da Rangelia vitalii no Sudeste do Brasil

Abstract This study aimed to evaluate the occurrence of diseases transmitted by Amblyomma ovale in 61 dogs monitored for three years through collections of ticks and blood, interviews, telemetry and camera traps in three areas of Serra do Mar State Park, Brazil. Blood samples were used to investigate infection by Rangelia vitalii by real-time TaqMan PCR and Rickettsia parkeri by IIFA. The collected ticks were submitted to conventional PCR to investigate the presence of R. parkeri . These data were compared with the monitoring results and interviews with the owners. Dogs considered as companion presented a risk of infection by R. parkeri strain Mata Atlantica 5.4 times higher than those not considered as companion (p = 0.009). Dogs that had at least one A. ovale collected during the campaigns had a 10 times higher risk of infection by R. parkeri strain Mata Atlantica than those who did not (p = 0.009). One dog positive for R. vitalii by real-time TaqMan PCR was parasitized by A. ovale frequently during monitoring. Sequenced ompaA - positive DNA samples had 100% identity of R. parkeri strain Mata Atlantica clone As106. From the findings, it is urgent to control domestic dogs around rainforests to reduce zoonoses transmission.

Resumo A ocorrência de doenças transmitidas por Amblyomma ovale em 61 cães monitorados por três anos através de coletas de carrapatos, sangue, entrevistas, telemetria e armadilhas fotográficas foi avaliada em três áreas do Parque Estadual da Serra do Mar - SP. Amostras de sangue foram utilizadas para investigação de Rangelia vitalii através de PCR TaqMan em tempo real e Rickettsia parkeri através da RIFI. Carrapatos coletados foram submetidos à PCR convencional para investigação de R. parkeri . Estes dados foram comparados considerando os resultados do monitoramento e entrevistas. Cães de companhia apresentaram risco de infecção pela R. parkeri cepa Mata Atlântica 5,4 vezes maior que os não considerados como de companhia (p = 0,009). Cães que tiveram pelo menos um A. ovale coletado apresentaram risco de infecção por R. parkeri cepa Mata Atlântica 10 vezes maior do que aqueles que não tiveram (p = 0,009). Um cão positivo para R. vitalii através de PCR TaqMan em tempo real foi parasitado por A. ovale durante o monitoramento. Amostras positivas para o gene ompaA possuíam 100% de identidade do clone As106 de R. parkeri cepa de Mata Atlântica. Assim, é urgente o controle de cães na Mata Atlântica para redução dos riscos de zoonoses.

Presentation of hemolytic and hemorrhagic rangeliosis in Cerdocyon thous.

Rangeliosis, caused by protozoan Rangelia vitalii, is transmitted by the tick Amblyomma aureolatum. The disease is characterized by hemolytic and hemorrhagic disorder and has been described in dogs and other wild canids. The aim of this study was to compare clinicopathological findings and laboratory results of a Rangelia infection in a crab-eating fox (Cerdocyon thous) with those of canine rangeliosis. The zoo of Universidade de Caxias do Sul, received a crab-eating fox with marked jaundice in mucous membranes, dark-colored stools and neurological signs. The animal underwent an ear tip smear examination and blood collection for complete blood counts, serum biochemistry and PCR. Free-living and intraerythrocytic pyriform structures consistent with R. vitalii were found in the blood smear of the ear tip. The erythrogram revealed normocytic normochromic anemia, moderate macrocytosis, polychromasia and metarubricytosis. The leukogram revealed leukocytosis with neutrophilia and monocytosis, as well as severe thrombocytopenia. Serum biochemistry showed hypoproteinemia, hypoalbuminemia and elevated levels of urea and creatinine. The treatment was performed with imidocarb hydrochloride and dexamethasone, however 24 h after initiation of treatment the animal died. Macroscopic examination revealed jaundice, subcutaneous edema, enlarged superficial lymph nodes, splenomegaly, and hemorrhage of internal organs. Histological sections of the cerebellum, lung, pancreas, intestine and heart were consistent with R. vitalii infection of the vascular endothelium. Pathological and hematological findings were similar to those found in infected dogs, with clinical presentation characterized by hemolytic anemia and hemorrhage. The description of this case showed that C. thous does not only serve as reservoir of R. vitalii but may also develop disease.

Molecular detection of vector borne pathogens in anemic and thrombocytopenic dogs in southern Brazil / Detecção molecular de patógenos transmitidos por vetores em cães anemicos e trombocitopenicos no Sul do Brasil

Abstract Arthropod-borne pathogens are medically important because of their ability to cause diseases in their hosts. The purpose of this study was to detect the occurrence of Ehrlichia spp., piroplasmids and Hepatozoon spp. in dogs with anemia and thrombocytopenia in southern Brazil. EDTA-whole blood was collected from 75 domestic dogs presenting anemia or/and thrombocytopenia from Guarapuava, state of Paraná, Brazil. DNA samples were subjected to conventional PCR assays for Ehrlichia spp. (dsb), piroplasmids (18S rRNA) and Hepatozoon spp. (18S rRNA), followed by sequencing and phylogenetic analyses. Among the 75 dogs, one (1.33%) was positive for Hepatozoon sp. and six (8%) were positive for piroplasmids in 18S rRNA cPCR assays. None of the dogs showed positive results in Ehrlichia spp.-cPCR targeting dsb gene. The phylogenetic analyses revealed that three piroplasm sequences were clustered with Rangellia vitalii, while one sequence was grouped with B. vogeli. The only sequence obtained from Hepatozoon spp.-PCR protocol was pooled with H. canis. Therefore, there is urgent need for differential molecular diagnosis of the two piroplasm species cited as etiological agents in clinical cases of canine hemoparasitic diseases, given the higher pathogenic potential of R. vitalii than of B. vogeli.

Resumo Agentes transmitidos por artrópodes têm grande importância na medicina veterinária devido à sua capacidade de causar doenças graves em seus hospedeiros. O presente estudo objetivou investigar a ocorrência de três patógenos transmitidos por vetores, Ehrlichia canis, Rangelia vitalii e Hepatozoon canis, em cães na região sul do Brasil. Foram coletadas amostras de sangue total de 75 cães domésticos que apresentavam anemia e/ou trombocitopenia, em Guarapuava, Paraná, Brasil. As amostras de DNA foram submetidas à técnica de PCR convencional para E. canis (dsb), piroplasmídeos (18S rRNA) e Hepatozoon spp. (18S rRNA), seguida de sequenciamento e análises filogenéticas. Das 75 amostras, uma (1,33%) foi positiva para Hepatozoon spp. e seis (8%) foram positivas para Babesia spp. Nenhuma amostra mostrou resultados positivos para Ehrlichia spp. utilizando a detecção pelo gene dsb. As análises filogenéticas revelaram que três sequências obtidas foram agrupadas no mesmo clado que R. vitalii , enquanto uma foi agrupada juntamente com B. vogeli. A única sequência obtida pelo protocolo de PCR para Hepatozoon spp. foi agrupada juntamente com H. canis. Assim, é justificada necessidade de diferenciação das espécies de piroplasmas, através do diagnóstico molecular, como agentes etiológicos nos casos clínicos de hemoparasitose canina, considerando o potencial patogênico de R. vitalii quando comparado à B. vogeli.

Clinical and hematological evaluation of Rangelia vitalii-naturally infected dogs in southeastern Brazil / Avaliação clínica e hematológica de cães naturalmente infectados por Rangelia vitalii no sudeste do Brasil

Abstract Rangelia vitalii, a tick-borne piroplasm that infects dogs, has been recently molecularly characterized in Brazil, Uruguay and Argentina. Studies on molecular characterization of these piroplasms in different Brazilian regions are scarce. The aim of this study was to evaluate clinical and hematological changes in dogs caused by R. vitalii in the mountainous region of the state of Rio de Janeiro. Blood samples from 36 dogs were evaluated for piroplasms and hematological disorders using light microscopy and molecular analysis. Blood samples from all the animals included in this study were confirmed to be positive for R. vitalii through genetic sequencing. Clinical signspresented by 24 of the 36 dogs of the study were evaluated during appointments or hospitalization within private practice. The most frequent clinical disorders in these dogs that were naturally infected with R. vitalii were fever, spontaneous cutaneous bleeding and diarrhea. Normochromic non-regenerative anemia and thrombocytopenia were the most common hematological disorders in these R. vitalii-positive dogs and therefore should be considered in hematological evaluations on suspected cases.

Resumo Rangelia vitalii, um piroplasma transmitido por carrapatos que infecta cães, foi sendo recentemente caracterizado molecularmente no Brasil, Uruguai e Argentina. Nas diferentes regiões brasileiras são escassos os estudos acerca da caracterização molecular destes piroplasmídeos. O objetivo deste estudo foi avaliar as alterações clínicas e hematológicas em cães causadas por R. vitalii na região serrana do Rio de Janeiro. Amostras de sangue total de 36 cães foram examinadas quanto à presença de piroplasmas pela microscopia de luz, alterações hematológicas e análise molecular. Todos os cães do presente estudo foram positivos para R. vitalii através do sequenciamento genético. Dos 36 animais positivos para R. vitalii, 24 foram avaliados clinicamente. As alterações mais frequentemente observadas foram febre, sangramento cutâneo espontâneo e diarréia. Anemia normocítica normocrômica arregenerativa e trombocitopenia foram as alterações hematológicas mais observadas em cães positivos para R. vitalii, devendo ser consideradas na avaliação hematológica de cães suspeitos.

Rangeliosis: histopathological analysis, hematology and molecular detection of canine Rangelia vitalii in Rio Grande do Sul, Brazil / Rangeliose: achados anatomopatológicos, hematológicos e detecção molecular de Rangelia vitalii em cães no Rio Grande do Sul, Brasil

ABSTRACT: In the period from January 2004 to December 2015, 56 dogs were diagnosed with rangeliosis in the Setor de Patologia Veterinária at Universidade Federal do Rio Grande do Sul (SPV-UFRGS). The main hematological abnormalities were thrombocytopenia and anemia. The affected dogs showed signs of apathy, anorexia, fetid and bloody diarrhea, vomiting, and dehydration. At necropsy, the main changes were jaundice, splenomegaly, hepatomegaly, and lymphadenomegaly. Histological analyses revealed parasitophorous vacuoles of Rangelia vitalii in cytoplasmic endothelial cells, mainly in the heart, kidneys, lymph nodes, intestines, and pancreas. Inflammation characterized by mononuclear cells was predominant in the analysis, and most was due to the presence of plasma cells. Other lesion types observed were lymphoid hyperplasia, extramedullary hematopoiesis, erythrophagocytosis, and erythroid lineage hyperplasia in bone marrow. Of the total number of animals, 49 were diagnosed using necropsy and histological analysis, and seven were diagnosed using a molecular analysis (i.e., PCR and genetic sequencing of blood samples). This paper presented a different method of diagnosing rangeliosis in canines. This approach involved histological methods including the quantification and determination of the intensity and distribution of the infectious agent in different organs.

RESUMO: No período de janeiro de 2004 a dezembro de 2015, 56 caninos domésticos obtiveram o diagnóstico de rangeliose no Setor de Patologia Veterinária da Universidade Federal do Rio Grande do Sul (UFRGS). Os cães apresentaram sinais de apatia, anorexia, diarreia fétida e sanguinolenta, êmese e desidratação. As principais alterações hematológicas foram trombocitopenia e anemia. Na necropsia as principais alterações foram icterícia, esplenomegalia, hepatomegalia e linfadenomegalia. Na análise histológica observou-se vacúolos parasitóforos de Rangelia vitalii no citoplasma de células endoteliais, principalmente no coração, rins, linfonodos, intestinos e pâncreas. A inflamação mononuclear foi predominante na análise, sendo que a maioria deu-se pela presença de plasmócitos. Entre outras lesões frequentemente observadas estão hiperplasia linfoide, hematopoiese extramedular e eritrofagocitose, e hiperplasia de linhagem eritroide em medula óssea. Do total, 49 cães foram diagnosticados através de necropsia e análise histológica, e 7 animais através de análise molecular da PCR e sequenciamento genético de amostras de sangue. Este trabalho apresenta um diagnóstico diferencial de rangeliose em caninos, através do método histológico de quantificação e determinação de intensidade e distribuição do agente em diferentes órgãos.

Rangelia vitalii in a free-ranging maned wolf (Chrysocyon brachyurus) and co-infections.

An adult free-ranged female maned wolf was rescued from a periurban area subject to anthropogenic disturbances in the Minas Gerais, Brazil. The animal presented poor body condition and anemia. The clinical condition rapidly deteriorated culminating in dead and a necropsy was performed. The main gross lesions were marked anemia and blood content in the intestines accompanied by many types of parasites. The protozoa Rangelia vitalii was identified by histopathological analysis predominantly within the cytoplasm of endothelial cells of capillaries of the small intestine. The lymph nodes, spleen, bone marrow, dermis, lungs and kidney had similar protozoal forms but with mild or moderate intensity. Rangelia vitalii was confirmed by molecular assays. Hepatozoon sp., Leishmania sp., and Entamoeba spp., apparently not related to the clinical signs were also detected. The myriad parasites found in the intestines included nematodes (Ancylostoma caninum, A. braziliensis,, Molineus sp., Pterygodermatites sp., and Trichuris sp.), cestodes (Spirometra sp.) and (acanthocephalans. To our knowledge, R. vitalii was identified in C. brachyurus for the first time. These findings emphasize the fragility of Brazilian ecosystems, especially in disturbed areas, reinforcing the necessity of efforts to preserve these areas and wild carnivores, some of which are threatened with extinction, such as the maned wolf.

Molecular detection of Rangelia vitalii in domestic dogs from Uruguay.

The piroplasm Rangelia vitalii is the etiological agent of canine rangeliosis, a severe disease affecting domestic dogs in South America. Two domestic dogs from two different Departments (Salto and Treinta y Tres) of Uruguay presented with clinical signs such as apathy, anorexia, pale mucous membranes, jaundice, and hemorrhagic manifestations, suggestive of a canine vector-borne disease. Molecular analysis, based on PCR and DNA sequencing of portions of the 18S rRNA gene, revealed that both dogs were infected by R. vitalii. Two consensus sequences, one from Salto and one from Treinta y Tres, differed from each other by only 1 nucleotide (99.8% similarity) and were 99.8-100% identical to corresponding sequences of R. vitalii from Brazil and Argentina available in GenBank. Through phylogenetic analysis inferred by the 18S rRNA gene, the two Uruguayan sequences of R. vitalii were aligned with the corresponding sequences from 7 other R. vitalii sequences available in GenBank (5 from Brazil and, 2 from Argentina) under high bootstrap support. The two dogs of the present study were negative for Ehrlichia canis according to the E. canis-specific real-time PCR assay. Our findings not only confirm the occurrence of R. vitalii in Uruguay but also provide the southernmost record of this re-emerging agent. The only previous report of R. vitalii in Uruguay dated from 1976, a period when molecular analyses were not available. We provide the first molecular detection of R. vitalii in Uruguay. Currently, canine rangeliosis is confirmed to occur in Brazil, Argentina, and Uruguay.

First report of Rangelia vitalii infection (canine rangeliosis) in Argentina.

A 12-year old mixed breed neutered bitch from Misiones, Argentina, was presented with a history of fever and epistaxis. Blood, bone marrow, and lymph node samples were collected for hematology and cytology. Mild regenerative anemia was recorded and large, round, poorly stained piroplasms (>2.5 µm) were found within erythrocytes in blood and lymph node smears. Nested PCR-RFLP on blood and bone marrow samples was positive for piroplasm DNA. The 18S rRNA gene of piroplasms was targeted. A restriction pattern of a previously unreported piroplasm was observed. The PCR product was sequenced, and the sequence obtained had 99% identity with the Rangelia vitalii sequences from Brazil when compared by BLAST analysis. Further characterization of the detected piroplasm consisted of nearly full-length sequencing (1668 bp) of the 18S rRNA gene of this organism. Those sequences were deposited in GenBank. A phylogenetic analysis indicated that they clustered together with R. vitalii from Brazil but separately from large Babesia species of dogs such as Babesia canis, and from species of Theileria of dogs as well. This is the first report of R. vitalii infection in Argentina, and the first case of canine rangeliosis diagnosed outside Brazil.

Canine rangeliosis due to Rangelia vitalii: from first report in Brazil in 1910 to current day - a review.

Canine rangeliosis (popular names: "nambi-uvú", i.e. ``bleeding ears''; "peste de sangue", i.e. ``bleeding plague''; and "febre amarela dos cães", i.e. ``yellow fever of dogs'') is a tick-borne haemolytic and haemorrhagic disease caused by the protozoan parasite Rangelia vitalii which infects erythrocytes, leukocytes, and endothelial cells of blood capillaries. Rangelia vitalii was first reported as a novel piroplasm of dogs in 1910 in Brazil, a discovery that was met with skepticism at that time. Canine rangeliosis has been diagnosed in domestic dogs not only in Brazil but also in other South American countries (Argentina and Uruguay). Rangelia vitalii infection has also been found incidentally in Brazil in wild dogs (Cerdocyon thous, the crab-eating fox). Despite the fact that researchers in the early 1900s suggested that R. vitalii was a hitherto unidentified piroplasm that would be transmitted by the tick Amblyomma aureolatum, it was not until 2012 that these hypotheses were actually confirmed by PCR and transmission studies. Molecular studies have shown that R. vitalii is related to the Babesia sensu strictu clade, but genetically different from other morphologically similar species of Babesia that infect dogs. Another difference between Babesia spp. and R. vitalii is the ability of R. vitalii to invade endothelial cells, erythrocytes, and leukocytes. Experimental infection in dogs has successfully reproduced the clinical picture and pathology of the natural disease. In this article, epidemiology, clinical signs, laboratory findings, pathogenetic mechanisms including oxidative stress and immune response, necropsy findings, microscopic lesions, diagnosis, and treatment of canine rangeliosis are reviewed. What is currently known about this protozoal disease since its first report over a century ago is presented herein.

Natural infection of the wild canid, Cerdocyon thous, with the piroplasmid Rangelia vitalii in Brazil.

Canine rangeliosis, caused by the piroplasmid protozoon Rangelia vitalii, is currently recognized as a reemerging disease that affects domestic dogs in Brazil. In the present study, piroplasmid infection was searched in wild canids (20 Cerdocyon thous and 4 Lycalopex gymnocercus) in Brazil. Molecular analysis, based on PCR and DNA sequencing of a portion of the 18S rRNA gene, revealed that 30% (6/20) C. thous were infected by R. vitalii. Blood and bone marrow samples from one of the R. vitalii-infected C. thous were inoculated into a domestic dog, which developed clinical rangeliosis that was confirmed by molecular tests. However, the C. thous donor showed no clinical, hematological or biochemical alterations, even though its R. vitalii infection status was confirmed for at least 80 days. These observations suggest that R. vitalii is not as highly pathogenic for C. thous as it is for domestic dogs. Phylogenetic analysis inferred by the 18S rRNA gene placed R. vitalii embedded in the clade 'Babesia sensu stricto', consisting of a number of species that represent truly the genus Babesia. It is proposed that the species R. vitalii should be transferred to the genus Babesia. The present study expands our knowledge on the natural history of R. vitalii, suggesting that it might have a natural cycle involving the wild canid C. thous. Further studies are needed to confirm that C. thous is a natural reservoir of R. vitalii in Brazil.

Increased cytokine and nitric oxide levels in serum of dogs experimentally infected with Rangelia vitalii.

This study aimed to measure the levels of interferon-gamma (IFN-γ), tumor necrosis factor-alpha (TNF-α), interleukin 1 (IL-1), interleukin 6 (IL-6), and nitrite/nitrate (NO x ) in serum of dogs experimentally infected with Rangelia vitalii. Twelve female mongrel dogs were divided into 2 groups; group A (uninfected controls) composed by healthy dogs (n=5) and group B consisting of dogs inoculated with R. vitalii (n=7). Animals were monitored by blood smear examinations, which showed intraerythrocytic forms of the parasite on day 5 post-infection (PI). Blood samples were collected through the jugular vein on days 0, 10, and 20 PI to determine the serum levels of IFN-γ, TNF-α, IL-1, IL-6, and NO x . Cytokines were assessed by ELISA quantitative sandwich technique, and NO x was measured by the modified Griess method. Cytokine levels (IFN-γ, TNF-α, IL-1, and IL-6) were increased (P<0.01) in serum of infected animals. Serum levels of NO x were also increased on days 10 PI (P<0.01) and 20 PI (P<0.05) in infected animals. Therefore, the infection with R. vitalii causes an increase in proinflammatory cytokines and nitric oxide content. These alterations may be associated with host immune protection against the parasite.

Increased Cytokine and Nitric Oxide Levels in Serum of Dogs Experimentally Infected with Rangelia vitalii

This study aimed to measure the levels of interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), interleukin 1 (IL-1), interleukin 6 (IL-6), and nitrite/nitrate (NOx) in serum of dogs experimentally infected with Rangelia vitalii. Twelve female mongrel dogs were divided into 2 groups; group A (uninfected controls) composed by healthy dogs (n=5) and group B consisting of dogs inoculated with R. vitalii (n=7). Animals were monitored by blood smear examinations, which showed intraerythrocytic forms of the parasite on day 5 post-infection (PI). Blood samples were collected through the jugular vein on days 0, 10, and 20 PI to determine the serum levels of IFN-gamma, TNF-alpha, IL-1, IL-6, and NOx. Cytokines were assessed by ELISA quantitative sandwich technique, and NOx was measured by the modified Griess method. Cytokine levels (IFN-gamma, TNF-alpha, IL-1, and IL-6) were increased (P<0.01) in serum of infected animals. Serum levels of NOx were also increased on days 10 PI (P<0.01) and 20 PI (P<0.05) in infected animals. Therefore, the infection with R. vitalii causes an increase in proinflammatory cytokines and nitric oxide content. These alterations may be associated with host immune protection against the parasite.

Detection and molecular characterization of piroplasms species from naturally infected dogs in southeast Brazil / Detecção e caracterização molecular de piroplasmas em cães naturalmente infectados no Sudeste do Brasil

Rangelia vitalii is a protozoon described from dogs in the south and southeast regions of Brazil. It is phylogenetically related to Babesia spp. that infects dogs, but data on this enigmatic parasite is still limited. The aim of this work was to detect piroplasm species in dogs in the state of Rio de Janeiro, Brazil, by 18S rRNA gene-based PCR assay, restriction fragment length polymorphism (RFLP) and sequence analyses. Of 103 dogs examined, seven (6.8%) were positive for Babesia spp. by PCR. The amplified products were digested by restriction enzymes to differentiate the Babesia species, and one sample was identified as Babesia vogeli. The pattern observed for the other six amplification products did not match with pattern described for large Babesia infecting dogs. Sequencing analysis confirmed these six samples as R. vitalii, with high homologies (99-100%) with a sequence from south Brazil. This study confirms the presence of Babesia vogeli and Rangelia vitalii circulate in domestic dogs in Teresópolis, Rio de Janeiro, Brazil.

Rangelia vitalii é um protozoário que infecta cães e foi descrito nas regiões Sul e Sudeste do Brasil. R. vitalii é filogeneticamente próxima à Babesia spp., mas dados deste misterioso parasito ainda são escassos. O objetivo deste trabalho foi detectar a presença de piroplasmas em cães naturalmente infectados no estado do Rio de Janeiro, através da amplificação do gene 18S rRNA pela PCR, clivagem com enzimas de restrição (RFLP) e caracterização genética através do sequenciamento. De 103 cães, sete (6,8%) foram positivos para Babesia spp. pela PCR. Os produtos amplificados foram digeridos por enzimas de restrição para a diferenciação das espécies de Babesia e uma amostra foi identificada como Babesia vogeli. O padrão de amplificação observado nas outras seis amostras não correspondeu ao padrão descrito para babesias que infectam cães. O sequenciamento das seis amostras confirmou ser uma espécie geneticamente idêntica a R. vitalii apresentando grande homologia (99-100%) com a sequência do sul do Brasil. Este estudo confirma a presença de Babesia vogeli e Rangelia vitalii infectando cães em Teresópolis, Rio de Janeiro, Brasil.

Patogênese e achados clínicos, hematológicos e anatomopatológicos da infecção por Rangelia vitalii em 35 cães (1985-2009) / Pathogenesis, clinical, hematological, and pathological aspects of Rangelia vitalii infection in 35 dogs (1985-2009)

A patogênese e os achados clínicos, hematológicos e anatomopatológicos da infecção natural pelo protozoário Rangelia vitalii (rangeliose canina) foram estudados em 35 cães que morreram em consequência dessa condição. Os resultados obtidos permitem o seguinte conjunto de conclusões: (1) causa doença hemolítica exclusivamente extravascular e de origem imunomediada; (2) cursa invariavelmente com algum grau de hemorragia à necropsia, mas nem sempre clinicamente perceptível; (3) os principais sinais que devem chamar a atenção para a suspeita clínica são anemia, icterícia e esplenomegalia; (4) o principal achado hematológico e que deve chamar a atenção para a suspeita clínica é a ocorrência de anemia com sinais de intensa regeneração eritroide; (5) os três principais diagnósticos diferenciais são leptospirose, babesiose e erliquiose monocitotrópica aguda; (6) a principal lesão observada é uma associação de hiperplasia linfoide com inflamação mononuclear, predominantemente plasmocitária, mas por vezes granulomatosa; (7) outras lesões frequentes são secundárias à marcada anemia regenerativa; (8) o agente etiológico pode ser facilmente encontrado, pois ocorre em grande quantidade na maioria dos tecidos, principalmente nos linfonodos, no baço, na medula óssea, no coração e nas tonsilas.

The pathogenesis, clinical, hematological and pathological features of the natural infection by the protozoan organism Rangelia vitalii (canine rangeliosis) was studied in 35 dogs that died due this condition. The results allow for the following set of conclusions on canine rangeliosis: (1) causes an exclusively extravascular immune mediated hemolysis; (2) is invariably associated with some degree of hemorrhage observed at necropsy, but no always clinically apparent; (3) the clinical signs that are the hallmark of the disease are anemia, icterus and splenomegaly; (4) the main hematological aspect that establishes a clinical suspect is the development of anemia with signs of intense erythroid regeneration; (5) the three main differential diagnosis are leptospirosis, babesiosis and e acute monocytotropic ehrlichiosis; (6) the main observed histopathological lesion is an association of lymphoid hyperplasia with mononuclear inflammatory reaction, predominantly plasmacytic, but occasionally granulomatous; (7) other frequently found lesions are secondary to a marked regenerative anemia; (8) large numbers of the etiologic agent can be easily demonstrate in most tissues, mainly in lymph nodes, spleen, bone marrow, heart, and tonsils.