Leishmania, Babesia and Ehrlichia in urban pet dogs: co-infection or cross-reaction in serological methods?

INTRODUCTION: The present study was designed to assess the occurrence of co-infection or cross-reaction in the serological techniques used for detecting the anti-Leishmania spp., -Babesia canis vogeli and -Ehrlichia canis antibodies in urban dogs from an area endemic to these parasites. METHODS: The serum samples from dogs were tested for the Babesia canis vogeli strain Belo Horizonte antigen and Ehrlichia canis strain São Paulo by immunofluorescence antibody test (IFAT) and by anti-Leishmania immunoglobulin G (IgG) antibody detection to assess Leishmania infection. We used the following four commercial kits for canine visceral leishmaniasis: ELISA, IFAT, Dual Path Platform (DPP) (Bio Manguinhos(r)/FIOCRUZ/MS) and a rK39 RDT (Kalazar Detect Canine Rapid Test; Inbios). RESULTS : Of 96 serum samples submitted to serological assays, 4 (4.2%) were positive for Leishmania as determined by ELISA; 12 (12.5%), by IFAT; 14 (14.6%) by rK39 RDT; and 20 (20.8%), by DPP. Antibodies against Ehrlichia and Babesia were detected in 23/96 (23.9%) and 30/96 (31.2%) samples, respectively. No significant association was identified between the results of tests for detecting Babesia or Ehrlichia and those for detecting Leishmania (p-value>0.05). CONCLUSIONS: In the present study, we demonstrated co-infection with Ehrlichia or Babesia and Leishmania in dogs from Minas Gerais (Brazil); we also found that the serological tests that were used did not cross-react.

An atypical Toxoplasma gondii genotype in a rural Brazilian dog co-infected with Leishmania (Viannia) braziliensis

Toxoplasmosis and leishmaniasis are two worldwide zoonoses caused by the protozoan parasites Toxoplasma gondii and Leishmania spp., respectively. This report describes the clinical and laboratorial findings of a co-infection with both parasites in a 4-year-old female dog suspected of ehrlichiosis that presented anemia, thrombocytopenia, hypoalbuminemia, hyperglobulinemia, tachyzoite-like structures to the lung imprints, and polymerase chain reaction (PCR) results positive for T. gondii (kidney, lung, and liver) and Leishmania spp. Co-infection with Toxoplasma gondii and Leishmania braziliensis was confirmed by sequencing; restriction fragment length polymorphism-polymerase chain reaction (RFLP-PCR) confirmed an atypical T. gondii genotype circulating in dogs that has been reported to cause human congenital toxoplasmosis.

Polymerase chain reaction-based method for the identification of Leishmania (Viannia) braziliensis and Leishmania (Viannia) guyanensis in mucosal tissues conserved in paraffin

ABSTRACTINTRODUCTION: In the Americas, mucosal leishmaniasis is primarily associated with infection by Leishmania (Viannia) braziliensis. However, Leishmania (Viannia) guyanensis is another important cause of this disease in the Brazilian Amazon. In this study, we aimed at detecting Leishmaniadeoxyribonucleic acid (DNA) within paraffin-embedded fragments of mucosal tissues, and characterizing the infecting parasite species.METHODS: We evaluated samples collected from 114 patients treated at a reference center in the Brazilian Amazon by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analyses.RESULTS: Direct examination of biopsy imprints detected parasites in 10 of the 114 samples, while evaluation of hematoxylin and eosin-stained slides detected amastigotes in an additional 17 samples. Meanwhile, 31/114 samples (27.2%) were positive for Leishmania spp. kinetoplast deoxyribonucleic acid (kDNA) by PCR analysis. Of these, 17 (54.8%) yielded amplification of the mini-exon PCR target, thereby allowing for PCR-RFLP-based identification. Six of the samples were identified as L. (V.) braziliensis, while the remaining 11 were identified as L. (V.) guyanensis.CONCLUSIONS: The results of this study demonstrate the feasibility of applying molecular techniques for the diagnosis of human parasites within paraffin-embedded tissues. Moreover, our findings confirm that L. (V.) guyanensisis a relevant causative agent of mucosal leishmaniasis in the Brazilian Amazon.

Expression of annexin A1 in Leishmania-infected skin and its correlation with histopathological features

ABSTRACTINTRODUCTION:The aim of this study was quantify annexin A1 expression in macrophages and cluster of differentiation 4 (CD4) + and cluster of differentiation 8 (CD8)+ T cells from the skin of patients with cutaneous leishmaniasis (n=55) and correlate with histopathological aspects.METHODS:Infecting species were identified by polymerase chain reaction-restriction fragment length polymorphism, and expression of annexin A1 was analyzed by immunofluorescence.RESULTS:All patients (n = 55) were infected with Leishmania braziliensis . Annexin A1 was expressed more abundantly in CD163 + macrophages in infected skin (p < 0.0001) than in uninfected skin. In addition, macrophages in necrotic exudative reaction lesions expressed annexin A1 at higher levels than those observed in granulomatous (p < 0.01) and cellular lesions p < 0.05). This difference might be due to the need to clear both parasites and necrotic tissue from necrotic lesions. CD4 + cells in cellular lesions expressed annexin A1 more abundantly than did those in necrotic (p < 0.05) and granulomatous lesions (p < 0.01). Expression in CD8 + T cells followed the same trend. These differences might be due to the pervasiveness of lymphohistiocytic and plasmacytic infiltrate in cellular lesions.CONCLUSIONS:Annexin A1 is differentially expressed in CD163 + macrophages and T cells depending on the histopathological features of Leishmania -infected skin, which might affect cell activation.

Presence of anti-Leishmania (Viannia) braziliensis antibodies in blood donors in the West-Central region of the State of Paraná, Brazil

ABSTRACTINTRODUCTION:Serological screening in blood banks does not include all transmittable diseases. American cutaneous leishmaniasis (ACL) has a high detection rate in the municipalities of the State of Paraná.METHODS:This study analyzed the presence of anti- Leishmania braziliensisantibodies in 176 blood donors who live in these endemic areas. The variables were analyzed with the χ2 test and Stata 9.1 software. RESULTS: Twenty (11.4%) samples were positive for the presence of anti- L. braziliensisantibodies. CONCLUSIONS: The high percentage of donors with anti- Leishmania spp. antibodies indicates the need to study the risk of ACL transmission through blood donors.

Detection of Leishmania (Viannia) DNA in leucocytes from the blood of patients with cutaneous leishmaniasis

ABSTRACTINTRODUCTION:Cutaneous leishmaniasis (CL) is a serious and global public health issue, with the potential of developing a mucosal form, occurring as subclinical cases, and showing recurrence despite previous treatment.METHODS:Polymorphonuclear and mononuclear DNA obtained from 49 patients was subjected to polymerase chain reaction for detection of Leishmania (Viannia).RESULTS:DNA was detected in mononuclear cells from two patients with active primary lesions positive for CL, with infection periods of 3 and 6 months, respectively.CONCLUSIONS:The DNA of Leishmania (Viannia) indicates probable parasite dissemination possibly explaining subclinical case emergence, lesion recurrence, and mucosal lesion appearance.

Predictive factors for Leishmania infantum infection in dogs examined at a veterinary teaching hospital in Teresina, State of Piauí, Brazil

Abstract: INTRODUCTION: In Brazil, culling of seropositive dogs is one of the recommended strategies to control visceral leishmaniasis. Since infectiousness is correlated with clinical signs, control measures targeting symptomatic dogs could be more effective. METHODS: A cross-sectional study was carried out among 1,410 dogs, predictive models were developed based on clinical signs and an indirect immunofluorescence antibody test. RESULTS: The validated predictive model showed sensitivity and specificity of 86.5% and 70.0%, respectively. CONCLUSIONS: Predictive models could be used as tools to aid control programs in focusing on a smaller fraction of dogs contributing more to infection dissemination.

Intradermorreação de Montenegro em cães (Mammalia: Canidae) experimentalmente inoculados por Leishmaniaguyanensis e Leishmania braziliensis (Kinetoplastida: Trypanosomatidae), principais agentes causadores de Leishmaniose Tegumentar na Amazônia

O teste de intradermorreação de Montenegro é utilizado para detectar infecção por Leishmania em humanos. A técnica se baseia numa reação de hipersensibilidade tardia. Os antígenos de Montenegro utilizados no experimento são soluções de antígenos homólogos brutos de Leishmania (Viannia) braziliensis e L.(V)guyanensis. Este experimento demonstrou que os animais inoculados com as espécies de Leishmania inoculadas desenvolveram enduração no local do teste mais acentuada que os animais controle. Os resultados sugerem que o teste cutâneo pode vir a ser indicado como método auxiliar de diagnóstico em cães infectados por Leishmania sp.

The spiny rat Proechimys guyannensis (Rodentia: Echimydae) fails to respond to intradermal inoculation with Leishmania (Viannia) braziliensis

Leishmaniasis a disease of worldwide occurrence is caused by protozoa of the Leishmania genus. In Brazil, Leishmania (Viannia) braziliensis is the main parasite responsible for the American cutaneous leishmaniasis. Main hosts of this protozoa are small wild mammals particularly marsupials and rodents. The aim of this study was to evaluate if spiny rat Proechimys guyannensis (Rodentia: Echimydae) has role in the cycle of the American cutaneous leishmaniasis caused by L. (V.) braziliensis. Thus, promastigotes (the flagellate stage) of Leishmania (Viannia) braziliensis were used to inoculate seven spiny rats (Proechimys guyannensis). After inoculated intradermal at the ear pinna, nose and plantar pad, the rats were monitored for 180 days. Tissue samples collected at 90 and 180 days from the rats proved to be negative for the presence of genetic material from the parasite. After euthanasia, the protozoa also failed to growth in culture medium containing tissue samples collected from the rats showing that there was no infection. These results fail to prove that spiny rat has a role in the cycle of the American cutaneous leishmaniasis caused by L. (V.) braziliensis.

Bases experimentales para la identificacion de Leishmania spp. de America por morfometria de amastigotos

Estudios morfométricos sobre los amastigotos de dieciocho poblaciones de cuatro aislados pertenecientes a dos especies venezolanas de Leishmania (L. braziliensis y L. garnhami) indican que la posición del einetoplasto nose modifica en modo estadísticamente significativo cuando los parásitos son sometidos a pasajes por hamsteres y ratones,cultivos en medio NNN o por infección en un vector (Lu. townsendi). La posición del cinetoplasto, medido como la distancia entre el extremo posterior del amastigoto al cinetoplasto, dividido entre la distancia del organoide al extremo anterior de la célula, permite diferenciar a L. braziliensis de L. mexicana y L. garnhami. Los otros parámetros morfométricos no son tan confiables.

Factores dermicos que condicionan la infeccion de Lutzomyia townsendi (Ortiz, 1959) por Leishmania spp. de Venezuela

Se estudia la susceptibilidad de Lutzomyia townsendi a la infección con Leishmania spp. sobre lesiones experimentales de hamsteres. Se estudia la frecuencia y distribución de los amastigotos en la dermis, relacionándola con la profundidad que alcanza el estilete bucal del insecto. Una correlación positiva, con significante coeficiente de correlación, se establece entre la frecuencia de los parásitos a una profundidad de 100-150 nm en la dermis y el éxito de la infección de los flebótomos.

Atypical epidermal alterations in chronic Leishmania mexicana mexicana lesions fo C3H mice

C3H mice chronically infected with Leishmania m. mexicana, and in some groups treated with BCG or levamisole, presented atypical epidermal alterations, including pseudoepitheliomatous hyperplasia, hyperkeratosis and dysplasia. These alterations increased in frequency and intensity during the course of infection, but were not related to lesion size or tissue parasite load. Age matched normal, BCG and levamisole treated control mice, examined simultaneously, did not show epidermal modifications. In infected mice the dermis and hypodermis presented an inflammatory infiltrate of histiocytes, lymphocytes and plasma cells, accompanied at times by neutrophils and eosinophils, which did not vary with duration of infection.

Interaction of avirulent Leishmania species with rat peritoneal macrophages

An "in vitro" system has been developed for study of host cell-parasite interaction in visceral and cutaneous leishmaniasis. Avirulent promastigotes of L. brasiliensis and L. donovani, from strains originally isolated from human cases and mantained by serial culture in Davis' Medium were allowed to infect cultured macrophages from rat peritoneal exudate. Challenge of the macrophages by parasites took place in 199 medium, at 33ºC for L. brasiliensis and at 37ºC for L. donovani. Although the rat is resistant to infections by Leishmania spp., the promastigotes not only invaded the host cells, but transformed into amastigotes and later mutiplied, from 10 min after challenge to 24 hours later.