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1.
PLoS Negl Trop Dis ; 14(8): e0008489, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32853247

RESUMO

Piroplasmosis treatment has been based on the use of imidocarb dipropionate or diminazene aceturate (DA), however, their toxic effects. Therefore, the discovery of new drug molecules and targets is urgently needed. Cryptolepine (CRY) is a pharmacologically active plant alkaloid; it has significant potential as an antiprotozoal and antibacterial under different in vitro and in vivo conditions. The fluorescence assay was used for evaluating the inhibitory effect of CRY on four Babesia species and Theileria equi in vitro, and on the multiplication of B. microti in mice. The toxicity assay was evaluated on Madin-Darby bovine kidney (MDBK), mouse embryonic fibroblast (NIH/3T3), and human foreskin fibroblast (HFF) cell lines. The half-maximal inhibitory concentration (IC50) values of CRY on Babesia bovis, B. bigemina, B. divergens, B. caballi, and T. equi were 1740 ± 0.377, 1400 ± 0.6, 790 ± 0.32, 600 ± 0.53, and 730 ± 0.025 nM, respectively. The toxicity assay on MDBK, NIH/3T3, and HFF cell lines showed that CRY affected the viability of cells with a half-maximum effective concentration (EC50) of 86.67 ± 4.43, 95.29 ± 2.7, and higher than 100 µM, respectively. In mice experiments, CRY at a concentration of 5 mg/kg effectively inhibited the growth of B. microti, while CRY-atovaquone (AQ) and CRY-DA combinations showed higher chemotherapeutic effects than CRY alone. Our results showed that CRY has the potential to be an alternative remedy for treating piroplasmosis.


Assuntos
Anti-Infecciosos/farmacologia , Babesia/efeitos dos fármacos , Babesiose/tratamento farmacológico , Alcaloides Indólicos/farmacologia , Quinolinas/farmacologia , Theileria/efeitos dos fármacos , Animais , Anti-Infecciosos/administração & dosagem , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Humanos , Camundongos
2.
Rev Bras Parasitol Vet ; 29(3): e012420, 2020 Aug 03.
Artigo em Inglês | MEDLINE | ID: mdl-32756775

RESUMO

Piroplasm species were analyzed by molecular tools in total 31 blood samples from positive dogs, previously checked by stained slides, stored until DNA extraction between 2016 to 2018 in the laboratory Clinical Analyzes in Niterói, Rio de Janeiro. The piroplasms were identified by PCR, targeting the 18S rRNA gene and sequencing. From the total number of samples only 24 (77.4%) were positive and show adequate nucleotide sequences for interpretation with identity between 93%-100% with Babesia vogeli in compared to the sequences isolated of infected dogs from other states in Brazil deposited on GenBank. Most of dogs infected with B. vogeli had anemia (62.5%) and thrombocytopenia (95.8%). The findings of this study are compatible with previous reports in the literature and highlight B. vogeli as the most incriminated species in canine piroplasmosis in Brazil, and thrombocytopenia the hematological alteration most frequently identified in this infection. It is important to note that this is the first study involving the molecular characterization of piroplasms in the metropolitan region of Rio de Janeiro, based on PCR followed by sequencing.


Assuntos
Babesia , Babesiose , Sangue , Doenças do Cão , Manejo de Espécimes , Animais , Babesia/genética , Babesiose/sangue , Sangue/parasitologia , Análise Química do Sangue , Brasil , Doenças do Cão/sangue , Doenças do Cão/parasitologia , Cães , RNA Ribossômico 18S/genética , Manejo de Espécimes/veterinária
3.
Pol Merkur Lekarski ; 48(285): 170-173, 2020 Jun 17.
Artigo em Inglês | MEDLINE | ID: mdl-32564041

RESUMO

Lifelong withdrawal from the donor population of those who have been diagnosed with babesiosis must be used for transmission prevention. AIM: The aim of the study was a detection of Babesia antibodies level with the usage of experimental Babesia divergens whole-cell slide antigen and commercial B. microti immunofluorescence assay substrate slide (Fuller Laboratories, USA). MATERIALS AND METHODS: Experimental B. divergens whole-cell slide antigen in addition to commercial B. microti IFA substrate slide was used to create a diagnostic kit for serum Babesia antibodies level detecting, as well as for a babesiosis serodiagnosis clinical trial of different origins blood samples (patients with Lyme disease, rheumatoid arthritis and toxoplasmosis; human blood donors; cattle). RESULTS: Antibodies to B. divergens (5.4%) and B. microti (2.3%) were detected with higher (p <0.05) frequency at Lyme disease patients (16.7%) than at blood donors (1.7%). Diagnostically significant IgG titres (= 1:128) were found in 13.3% of blood samples from Lyme disease patients and 1.7% from blood donors. Specific IgM were also found in 13.3% blood samples from Lyme disease patients. Among blood samples from Lyme disease patients, in which diagnostically significant titres of Babesia antibodies were detected (16.7%), 60% of them were represented by IgG and IgM (rA= 0.63), and in 40% only one of them reached diagnostically significant titre. Conclusions. Advantages of babesiosis IFA diagnostics. CONCLUSIONS: Advantages of babesiosis IFA diagnostics are combined with its significant disadvantages (principle of evaluation, low sensitivity in the initial period of the disease, probability of false positives, absence of validated test systems and research protocols for B. divergens and B. divergens-like species).


Assuntos
Babesia microti , Babesia , Babesiose , Doença de Lyme , Animais , Babesia/isolamento & purificação , Babesiose/diagnóstico , Bovinos , Humanos , Imunoensaio , Doença de Lyme/diagnóstico
4.
Rev Bras Parasitol Vet ; 29(1): e017119, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32294720

RESUMO

The present study aimed to characterize the importance of the Anaplasma marginale, Babesia bovis and Babesia bigemina in the genesis of cattle tick fever (CTF) among dairy calves in the northwest of Minas Gerais, Brazil. Blood samples from 300 calves were collected, followed by DNA extraction and nested PCR using oligonucleotide primers to amplify fragments of the semi-nested for the msp5 gene (A. marginale), sbp-4 (B. bovis) and rap-1a (B. bigemina) Among the examined calves, the prevalence of A. marginale was 55.6% (n=167/300), B. bovis was 4.0% (n=12/300) and B. bigemina was 15.3% (n=46/300), by PCR techniques. Parasitic forms of A. marginale and B. bigemina were found in 36,3% and 2,6% of the blood smears while B. bovis was not detected. There was a statistical difference between the positivity of infected animals in the age groups 1 (10-70 days) and (>70-300 days) for A. marginale and B. bigemina. A total of 15 calves with the classic symptoms of disease were examined, and the samples obtained were confirmed as a simple infection by A. marginale through semi-nested PCR. These results confirm bovine anaplasmosis as the primary cause of CTF among the calves of dairy cattle within the studied area.


Assuntos
Anaplasma marginale/genética , Anaplasmose/parasitologia , Babesia/genética , Babesiose/parasitologia , Doenças dos Bovinos/parasitologia , Carrapatos/parasitologia , Anaplasmose/diagnóstico , Anaplasmose/epidemiologia , Animais , Babesiose/diagnóstico , Babesiose/epidemiologia , Brasil , Bovinos , Doenças dos Bovinos/diagnóstico , Doenças dos Bovinos/epidemiologia , Feminino , Masculino , Filogenia , Reação em Cadeia da Polimerase/veterinária
5.
Parasite Immunol ; 42(5): e12706, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-32119124

RESUMO

To provide useful information based on the macropathology, histopathology and immunohistochemical investigation in the spleens of dogs with Babesia rossi infection. Control spleens were collected from four healthy dogs euthanized for welfare reasons. Nine dogs that died naturally because of a mono-infection with Babesia rossi were selected for the diseased group. One haematoxylin-and-eosin-stained section of splenic tissue from each of the infected and control dogs was examined under the light microscope. Immunohistochemical markers were applied to characterize different immunocyte populations. The application of analytic software enabled semi-quantitative comparison of leucocyte subpopulations. Routine splenic histopathology revealed diffuse intermingling of white and red pulp from infected dogs with a clear loss of distinction between these zones. Immunohistochemistry revealed an increase in the proportion of tissue resident and bone marrow origin macrophages in the infected spleens. Apart from a few remnant lymphocytes within the peri-arteriolar lymphatic sheaths and follicles, the majority of the immunocytes redistributed to the red pulp, supporting the observation of white and red pulp intermingling. The majority of our findings are in agreement with histomorphological descriptions of the spleen in a variety of noncanid mammalian hosts with lethal malaria or babesiosis.


Assuntos
Babesia/fisiologia , Babesiose/patologia , Doenças do Cão/patologia , Baço/patologia , Animais , Babesiose/imunologia , Babesiose/parasitologia , Doenças do Cão/imunologia , Doenças do Cão/parasitologia , Cães , Leucócitos/imunologia , Leucócitos/parasitologia , Linfócitos/imunologia , Linfócitos/parasitologia , Baço/imunologia , Baço/parasitologia
6.
Parasitol Res ; 119(3): 1173-1176, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32140779

RESUMO

Babesia is tick-transmitted protozoan parasites that infect mammalian hosts and have a major impact on farm and pet health-associated costs worldwide. This study aimed to test the prevalence of Babesia spp. infection in a small cohort of dogs at a veterinary hospital and to perform molecular characterization of the Babesia species causing the infection. For the PCR assay, 5 mL of blood was collected by venipuncture of the cephalic or radial veins in 300 dogs of different ages, sex, and breeds, which were presented to the veterinary hospital of the Federal University of Uberlândia between March 2015 and April 2016. In addition, a drop of blood was collected from the marginal blood vessels of the ear of dogs included in this study. Ninety-two (30.67%) were positive for Babesia spp., as determined by microscopic observation of the blood smear, revealing the presence of intra-erythrocyte merozoites. For molecular characterization by PCR, 17 samples were chosen from dogs who were tested positive for Babesia spp. by blood smears. Among them, B. vogeli was found to infect all 17 dogs, as determined by 99-100% sequence identity (closest GenBank match KT246307) using primers PIRO A/PIRO B. Our results indicate that the species observed in these dogs was B. vogeli.


Assuntos
Babesiose/parasitologia , Doenças do Cão/parasitologia , Animais , Babesia/genética , Brasil/epidemiologia , Primers do DNA , Cães , Feminino , Masculino , Reação em Cadeia da Polimerase/veterinária , Prevalência , Carrapatos/parasitologia
7.
Exp Parasitol ; 212: 107870, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32142733

RESUMO

Piroplasmosis is a serious debilitating and sometimes fatal disease. Phylogenetic relationships within piroplasmida are complex and remain unclear. In the study, we assessed the relative resolution capabilities of the DNA sequences of the nuclear genes 40S ribosomal protein S5 (RPS5) and mitochondrial DNA Cytochrome c oxidase subunit III (cox3) gene in the phylogeny of Babesia and Theileria species isolates. We demonstrated that by using the cox3 gene can recover a better supported species tree for some Theileria species than when using the nuclear RPS5 gene alone, it tends to intra-specific diversity and considerable inter-specific difference. Additionally, the combined DNA sequences of the nuclear RPS5 and cox3 gene improved the inference of evolutionary relationships among Babesia and Theileria species. The mitochondrial cox3 gene outperforms nuclear RPS5 gene and yields better resolution on the intra-specific diversity of Babesia and Theileria species. However, the combined RPS5 nuclear DNA and cox3 DNA tree had more advantage in the phylogeny of Babesia and Theileria species than that of single gene alone.


Assuntos
Babesia/classificação , Complexo IV da Cadeia de Transporte de Elétrons/genética , Filogenia , Proteínas Ribossômicas/genética , Theileria/classificação , Animais , Babesia/genética , Sequência de Bases , Biodiversidade , Bovinos , DNA Mitocondrial/fisiologia , DNA de Protozoário/fisiologia , Marcadores Genéticos , Alinhamento de Sequência , Ovinos , Organismos Livres de Patógenos Específicos , Theileria/genética
8.
Acta Trop ; 205: 105388, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32035054

RESUMO

Ticks and tick-borne pathogens constitute a great threat to livestock production and are a potential health hazard to humans. Grasscutters (Thryonomys swinderianus) are widely hunted for meat in Ghana and many other West and Central African countries. However, tick-borne zoonotic risks posed by wild grasscutters have not been assessed. The objective of this study was to investigate bacterial and protozoan pathogens in ticks infecting wild grasscutters. A total of 81 ticks were collected from three hunted grasscutters purchased from Kantamanto, the central bushmeat market in Accra. Ticks were identified as Ixodes aulacodi and Rhipicephalus sp. based on morphological keys, which were further confirmed by sequencing mitochondrial 16S ribosomal DNA (rDNA) and cytochrome oxidase I (COI) genes of specimens. Protozoan infections were tested by PCR amplifying 18S rDNA of Babesia/Theileria/Hepatozoon, while bacterial infections were evaluated by PCRs or real-time PCRs targeting Anaplasmataceae, Borrelia, spotted fever group rickettsiae, chlamydiae and Candidatus Midichloria mitochondrii. The results of PCR screening showed that 35.5% (27 out of 76) of I. aulacodi were positive for parasite infections. Sequencing analysis of the amplified products gave one identical sequence showing similarity with Babesia spp. reported from Africa. The Ca. M. mitochondrii endosymbiont was present in 85.5% (65 out of 76) of I. aulacodi but not in the five Rhipicephalus ticks. Two Anaplasmataceae bacteria genetically related to Ehrlichia muris and Anaplasma phagocytophilum were also detected in two I. aulacodi. None of the ticks were positive for Borrelia spp., spotted fever group rickettsiae and chlamydiae. Since I. aulacodi on wild grasscutters are potential carriers of tick-borne pathogens, some of which could be of zoonotic potential, rigorous tick control and pathogen analyses should be instituted especially when wild caught grasscutters are being used as foundation stock for breeding.


Assuntos
Babesia/isolamento & purificação , Bactérias/isolamento & purificação , Ixodes/microbiologia , Ixodes/parasitologia , Roedores/parasitologia , Theileria/isolamento & purificação , Animais , Feminino , Gana , Humanos , Masculino , Doenças Transmitidas por Carrapatos/parasitologia
9.
Parasitol Res ; 119(4): 1423-1427, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32107621

RESUMO

We report two cases of bovine babesiosis caused by Babesia divergens in a region of central Bosnia and Herzegovina. The cases were detected in June 2017 and July 2018 from two small backyard farms. Routine clinical assessments, including physical examination and haematology, revealed lethargy, fever, anaemia, leukopenia and haemoglobinuria in the affected animals. Serum alterations included an elevation of aspartate aminotransferase and a decrease of serum phosphate or hypophosphatemia. Thrombocytopenia was detected in the first clinical case. Microscopic examination of blood smears revealed intracytoplasmic protozoan parasites from the genus Babesia. Molecular screening of both animals confirmed the presence of Babesia divergens, the causative agent of bovine babesiosis. B. divergens DNA was also detected in two engorged female Ixodes ricinus ticks removed from these animals. In addition, Mycoplasma wenyonii DNA was identified by molecular screening in the animal examined in June 2017, and in I. ricinus ticks feeding on this animal. This study provides molecular confirmation of B. divergens as a cause of piroplasmosis in cattle in South-East Europe. The detection of M. wenyonii DNA ain I. ricinus also provides the first evidence of this bacterium in ticks in Europe.


Assuntos
Babesia/genética , Babesiose/diagnóstico , Doenças dos Bovinos/microbiologia , Doenças dos Bovinos/parasitologia , Infecções por Mycoplasma/veterinária , Mycoplasma/genética , Animais , Aspartato Aminotransferases/sangue , Babesia/isolamento & purificação , Babesiose/parasitologia , Bósnia e Herzegóvina , Bovinos , DNA Bacteriano/genética , DNA Bacteriano/isolamento & purificação , DNA de Protozoário/genética , DNA de Protozoário/isolamento & purificação , Europa (Continente) , Fazendas , Feminino , Hipofosfatemia/sangue , Ixodes/microbiologia , Ixodes/parasitologia , Mycoplasma/isolamento & purificação , Infecções por Mycoplasma/diagnóstico , Trombocitopenia/sangue
10.
PLoS One ; 15(2): e0228996, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32053698

RESUMO

BACKGROUND: The plenteous resistance to and undesirable consequences of the existing antipiroplasmic therapies have emphasized the urgent need for new chemotherapeutics and drug targets for both prophylaxis and chemotherapy. Hydroxyurea (HYD) is an antineoplastic agent with antitrypanosomal activity. Eflornithine (α-difluoro-methyl ornithine, DFMO) is the best choice therapy for the treatment of late-stage Gambian human African trypanosomiasis. METHODS: In this study, the inhibitory and combination efficacy of HYD and DFMO with existing babesicidal drugs (diminazene aceturate (DA), atovaquone (ATV), and clofazimine (CLF)) deoxyribonucleotide in vitro against the multiplication of Babesia and Theileria. As well as, their chemotherapeutic effects were assessed on B. microti strain that infects rodents. The Cell Counting Kits-8 (CCK-8) test was used to examine their cytotoxicity on human foreskin fibroblast (HFF), mouse embryonic fibroblast (NIH/3T3), and Madin-Darby bovine kidney (MDBK) cells. FINDINGS: HYD and DFMO suppressed the multiplication of all tested species (B. bigemina, B. bovis, B. caballi, B. divergens, and T. equi) in a dose-related manner. HFF, NIH/3T3, or MDBK cell viability was not influenced by DFMO at 1000 µM, while HYD affected the MDBK cell viability at EC50 value of 887.5±14.4 µM. The in vitro combination treatments of DFMO and HYD with CLF, DA, and ATV exhibited synergistic and additive efficacy toward all tested species. The in vivo experiment revealed that HYD and DFMO oral administration at 100 and 50 mg/kg inhibited B. microti multiplication in mice by 60.1% and 78.2%, respectively. HYD-DA and DFMO-DA combined treatments showed higher chemotherapeutic efficacy than their monotherapies. CONCLUSION: These results indicate the prospects of HYD and DFMO as drug candidates for piroplasmosis treatment, when combined mainly with DA, ATV, and CLF. Therefore, further studies are needed to combine HYD or DFMO with either ATV or CLF and examine their impact on B. microti infection in mice.


Assuntos
Babesia/efeitos dos fármacos , Eflornitina/efeitos adversos , Eflornitina/farmacologia , Hidroxiureia/efeitos adversos , Hidroxiureia/farmacologia , Theileria/efeitos dos fármacos , Animais , Antineoplásicos , Antiprotozoários/administração & dosagem , Antiprotozoários/farmacologia , Atovaquona/efeitos adversos , Atovaquona/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Clofazimina/efeitos adversos , Clofazimina/farmacologia , Diminazena/efeitos adversos , Diminazena/análogos & derivados , Diminazena/farmacologia , Cães , Prepúcio do Pênis/citologia , Humanos , Masculino , Camundongos , Células NIH 3T3
11.
Rev Bras Parasitol Vet ; 29(1): e018019, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32049147

RESUMO

The aim of the present study was to detect Cercopithifilaria bainae and other tick-borne pathogens and to perform molecular characterization of the tick Rhipicephalus sanguineus s.l. collected from dogs. Ticks (n = 432, including 8 larvae, 59 nymphs, and 365 adults) were sampled from domiciled dogs (n = 73) living in Campo Grande, Mato Grosso do Sul (Midwest Brazil). All ticks were morphologically identified as R. sanguineus. Genomic DNA was extracted in pools (three to five ticks per animal) and was used for definition of R. sanguineus haplotypes (based on 16S rRNA analysis) and pathogen identification (Cercopithifilaria sp., Ehrlichia canis, Anaplasma platys, Hepatozoon canis, Babesia vogeli and Rickettsia spp.). Rhipicephal us sanguineus specimens were identified as haplotypes A and B. DNA of Cercopithifilaria bainae (43.83%; 32/73), Ehrlichia canis (24.65%; 18/73), Anaplasma platys (19.17%; 14/73), and Hepatozoon canis (5.47%; 4/73) was detected. The identity of pathogens was confirmed by DNA sequence analysis. The present study confirms the presence of haplotypes A and B of R. sanguineus in the state of Mato Grosso do Sul and its importance as a vector of several pathogens of veterinary concern. Finally, this is the first report to identify C. bainae in ticks in the Midwestern region of Brazil.


Assuntos
Vetores Aracnídeos/parasitologia , Cães/parasitologia , Rhipicephalus sanguineus/parasitologia , Anaplasma/genética , Anaplasma/isolamento & purificação , Animais , Babesia/genética , Babesia/isolamento & purificação , Brasil , Ehrlichia canis/genética , Ehrlichia canis/isolamento & purificação , Eucoccidiida/genética , Eucoccidiida/isolamento & purificação , RNA Ribossômico 16S/genética , Rickettsia/genética , Rickettsia/isolamento & purificação
12.
Parasitol Res ; 119(3): 1117-1123, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32100102

RESUMO

Reported fatal cases of bovine babesiosis (syn.: piroplasmosis, red water fever) in cattle were analyzed to identify spatial and temporal clusters of their incidence in the Austrian province of Styria. Data were collected within a governmental babesiosis compensation program. Diagnosis was performed using a standardized necropsy protocol. Between 1998 and 2016, a total of 1257 cases of fatal babesiosis were registered and compensated. Within the study interval, annual numbers of fatal babesiosis differed significantly among municipalities. Spatiotemporal analysis covering the entire study period revealed one high-risk cluster in the western and central northern region of Styria and a low-risk cluster in the southeastern part of Styria. Annual temporal analysis demonstrated that cases accumulated in June. Annual spatial analysis revealed consistently that cases mainly occurred in the western and central northern regions, whereas they occurred rarely in the southeastern regions. These results should increase awareness and facilitate protective actions against ticks during certain time periods and geographic areas.


Assuntos
Babesiose/epidemiologia , Babesiose/mortalidade , Doenças dos Bovinos/epidemiologia , Doenças dos Bovinos/mortalidade , Animais , Áustria/epidemiologia , Babesia , Bovinos , Incidência
13.
Parasitol Res ; 119(4): 1259-1269, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32060726

RESUMO

To reveal the genetic diversity of Babesia microti and Theileria orientalis in Southwest China, we conducted a molecular survey of piroplasms in hard ticks in a China-Myanmar border county. Host infesting and questing ticks were collected from Tengchong County in 2013 and 2014. Piroplasm infection in ticks was detected by PCR, and then, phylogenetic analysis was conducted to study the genetic diversity of the pathogens identified in ticks. All in all, six piroplasm species comprising of B. microti; B. orientalis; a novel Babesia species designated Babesia sp. Tengchong, China; T. orientalis; T. luwenshuni; and an as yet undescribed piroplasmid species referred to as Piroplasmid sp. Tengchong, China, have been identified after screening goat- and cattle-attached ticks. In addition, B. bigemina has been identified by screening questing ticks. Phylogenetic analysis based on the 18S rRNA and partial ß-tubulin gene revealed two novel potentially zoonotic genotypes designated B. microti Tengchong-Type A and B. The T. orientalis genotypes identified in the present study represent the seven known genotypes 1-5, 7, and N3 as revealed by phylogenetic analysis of 18S rRNA and MPSP genes. Importantly, an additional genotype designated N4 has also been identified in this study, which brings the number of recognized T. orientalis genotypes to a total of twelve. Thus, besides the two novel species, Babesia sp. Tengchong, China, closely related to Babesia species isolated from yak and Piroplasmid sp. Tengchong, China, our study demonstrates that additional novel B. microti and T. orientalis genotypes exist in Southwest China.


Assuntos
Babesia microti/genética , Babesia/genética , Ixodidae/parasitologia , Theileria/genética , Animais , Babesia/classificação , Babesia/isolamento & purificação , Babesia microti/classificação , Babesia microti/isolamento & purificação , Bovinos , China , Genótipo , Mianmar , Filogenia , Reação em Cadeia da Polimerase , RNA Ribossômico 18S , Theileria/classificação , Theileria/isolamento & purificação
14.
Parasitol Int ; 75: 102054, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31927139

RESUMO

In the current study, we evaluated the usefulness of a SYBR Green I (SG I) fluorescence assay for evaluation of the inhibitory effects of antibabesial drugs against the in vitro growth of Babesia gibsoni. Linearity and high-throughput screening (HTS) assays exhibited the validity of the SG I fluorescence assay for B. gibsoni parasite when performed at low hematocrits (HCTs) (2.5% and 5%) without daily changing of the medium. Interestingly, 5% HCT showed the highest value of the signal/noise ratio. Of note, there were no significant differences (P > .05) in the IC50s of the commonly used antibabesial drugs (diminazene aceturate and/or imidocarb dipropionate) that calculated by either the SG I fluorescence assay with and without daily medium changing or by the fluorescence and microscopy methods at 2.5% and 5% HCTs. Such results confirmed that both HCTs are valid for mass drug screening against the in vitro growth of B. gibsoni. While the results of the HTS assay add merit to the assay when performed at 5% HCT especially when incubating the plates for 2 h in a dark after adding lysis buffer with SG I stain. Next, nine different drugs were screened to confirm the assay's usefulness. MMV396693, pyronaridine tetraphosphate and nerolidol drugs exhibited the highest effectiveness against the in vitro growth of B. gibsoni, next to diminazene aceturate. In summary, SG I fluorescence assay with 5% HCT without daily changing of the medium for B. gibsoni offers a novel approach for the large-scale screening of huge chemical libraries in in vitro cultures.


Assuntos
Antiprotozoários/farmacologia , Babesia/efeitos dos fármacos , Babesiose/tratamento farmacológico , Ensaios de Triagem em Larga Escala/métodos , Antiprotozoários/análise , Fluorescência , Técnicas In Vitro
15.
J Parasitol ; 106(1): 30-37, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31971489

RESUMO

Ixodes scapularis is currently known to transmit 7 pathogens responsible for Lyme disease, anaplasmosis, babesiosis, tick-borne relapsing fever, ehrlichiosis, and Powassan encephalitis. Ixodes scapularis can also be colonized by endosymbiotic bacteria including those in the genus of Rickettsia. We screened 459 I. scapularis ticks submitted to the Connecticut Agricultural Experiment Station Tick Testing Laboratory with the objectives to (1) examine differences in infection prevalence of Borrelia burgdorferi, Anaplasma phagocytophilum, Babesia microti, and Borrelia miyamotoi, (2) evaluate whether prevalence of co-infections occur at the same frequency that would be expected based on single infection, and (3) determine the presence of rickettsial endosymbionts in I. scapularis. The prevalence of infection in I. scapularis was highest with Bo. burgdorferi sensu lato (nymph = 45.8%; female = 47.0%), followed by A. phagocytophilum (nymph = 4.0%; female = 6.9%), Ba. microti (nymph = 5.7%; female = 4.7%), and Bo. miyamotoi (nymph = 0%; female = 7.3%). We also identified rickettsial endosymbionts in 93.3% of I. scapularis. Nymphs were significantly more likely to be infected with Bo. burgdorferi if they were infected with Ba. microti, whereas adult females were significantly more likely to be infected with Bo. burgdorferi if they were infected with A. phagocytophilum. Our study suggests that the infection prevalence of Bo. burgdorferi is not independent of other co-circulating pathogens and that there is a substantially higher infection of Bo. miyamotoi in I. scapularis females compared with nymphs in this study. High prevalence of infection and co-infection with multiple pathogens in I. scapularis highlights the public health consequences in Connecticut, a state endemic for Lyme and other tick-borne diseases.


Assuntos
Vetores Aracnídeos/microbiologia , Ixodes/microbiologia , Rickettsia/fisiologia , Simbiose , Doenças Transmitidas por Carrapatos/transmissão , Anaplasma phagocytophilum/fisiologia , Animais , Babesia/fisiologia , Babesia microti/fisiologia , Borrelia burgdorferi/fisiologia , Connecticut/epidemiologia , Feminino , Ninfa/parasitologia , Prevalência , Doenças Transmitidas por Carrapatos/epidemiologia
16.
J Vet Med Sci ; 82(3): 286-293, 2020 Mar 05.
Artigo em Inglês | MEDLINE | ID: mdl-31969541

RESUMO

In Sudan, donkeys are important animals, providing transportation and income possibilities. However, the prevalence of parasites in donkeys in Sudan has not been thoroughly characterized. Accordingly, in this study, we aimed to detect selected hemoprotozoan parasites in donkeys in West Omdurman, Khartoum State, Sudan, wherein people depend mainly on donkeys for their daily life. In total, 198 blood samples collected from donkeys in a local market in West Omdurman, were screened using serological and molecular diagnostic techniques. Serologically, 52 (26.3%), 56 (28.3%), and 19 (9.6%) samples were positive for trypanosomosis using Card Agglutination Test for Trypanosoma evansi, Trypanosoma evansi crude antigen -based enzyme-linked immuno sorbent assay (ELISA) and recombinant Trypanosoma evansi GM6-4r-based ELISA, respectively. ELISA for equine piroplasmosis revealed 156 (78.8%) and 10 (5.1%)Theileria equi- and Babesia caballi-positive samples, respectively. PCR detected Trypanosoma congolense, subgenus Trypanozoon, Theileria equi, and Babesia caballi in 18 (9.1%), 77 (38.9%), 18 (9.1%), and 8 (4%) samples, respectively. Of the 77 Trypanozoon-positive samples, 35 (45.5%) were confirmed as Trypanosoma evansi type A. To our knowledge, this is the first report of detection of Trypanosoma congolense in donkeys outside of tsetse-infested areas in Sudan.


Assuntos
Babesiose/epidemiologia , Equidae/parasitologia , Theileriose/epidemiologia , Trypanosoma congolense/isolamento & purificação , Tripanossomíase Africana/veterinária , Animais , Anticorpos Antiprotozoários/sangue , Babesia/classificação , Babesiose/sangue , Ensaio de Imunoadsorção Enzimática/veterinária , Equidae/sangue , Sudão/epidemiologia , Theileria/classificação , Theileriose/sangue , Tripanossomíase Africana/sangue , Tripanossomíase Africana/epidemiologia
17.
Parasitol Res ; 119(2): 687-694, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31897793

RESUMO

Wild rodents, as natural reservoir hosts carrying various species of pathogens, play an important role in the evolution and emergence of zoonotic diseases. In this study, protist parasites, namely Babesia sp., Trypanosoma sp. and Hepatozoon sp. were studied in rodent populations in Lithuania. Two hundred forty rodent specimens of seven species were analysed by a combined approach using polymerase chain reaction (PCR)-based techniques and traditional microscopic examination. The total prevalence of blood parasites reached 35% in rodent communities. The prevalence of Hepatozoon sp. reached the highest value (32%), followed by Trypanosoma sp. (5%) and Babesia sp. (3%). Myodes glareolus and Microtus agrestis were the most heavily infected rodent species. Comparison of microscopy and PCR-based methods showed that the two approaches might give different results and thus can lead to an underestimation of the actual prevalence and abundance of parasites. In our study, PCR-based assays were more sensitive and robust than traditional microscopy. However, precise molecular results for the estimation of the prevalence of Babesia sp. and Hepatozoon sp. were achieved only by using several sets of primers. To avoid inaccurate results, the improvement and detailed description of molecular and microscopy protocols are required.


Assuntos
Arvicolinae/parasitologia , Babesia/isolamento & purificação , Eucoccidiida/isolamento & purificação , Trypanosoma/isolamento & purificação , Animais , Lituânia , Microscopia , Reação em Cadeia da Polimerase
18.
Vet Parasitol ; 278: 109034, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31991285

RESUMO

Ovine babesiosis is an endemic tick-borne disease of small ruminants in the Middle East, European, and some African and Asian countries, including Turkey. This study assessed whether the endemic status of this disease was stable or instable, which is important for disease control efforts. For this aim, 4115 sheep blood samples were collected from 81 cities in the seven geographical regions of Turkey. The diagnosis of Babesia ovis was made using microscopic and serological techniques. Thin blood smears were prepared from anticoagulated venous blood. Serum samples were screened for specific antibodies using an enzyme-linked immunosorbent assay (ELISA) and indirect fluorescent antibody test (IFAT). Recombinant Babesia ovis secreted antigen 1 (rBoSA1) was used in the ELISA. The antigen slides used in the IFAT were prepared from the B. ovis-infected blood at a high level of parasitemia (5 %). The animals were divided into three groups according to their age: group I (one to six months), group II (6-12 months), and group III (older than one year). The endemic status of B. ovis was determined according to the inoculation rate (h value) calculations. Babesia spp. merozoites were observed in 40 (0.97 %) of the slides. Seropositivity rates were 29.89 % (1230/4115) and 49.16 % (2023/4115) by the ELISA and IFAT, respectively. According to the IFAT results, 31.7 %, 33.6 %, and 52.8 % of the animals were seropositive in groups I, II, and III, respectively. The inoculation rates of the animals indicated that the endemic status of ovine babesiosis was mostly instable throughout the country. Endemic stability was found only in group I from four regions (Central Anatolia, Eastern Anatolia, Aegean, and Mediterranean). Based on these results, the risk of clinical infection due to tick infestation was high when the maternal immunity and non-specific age resistance weakens or disappears. Thus, vaccination is needed to protect sheep against B. ovis infections in Turkey.


Assuntos
Babesiose/epidemiologia , Doenças dos Ovinos/epidemiologia , Doenças Transmitidas por Carrapatos/veterinária , Animais , Anticorpos Antiprotozoários/sangue , Babesia , Ensaio de Imunoadsorção Enzimática/veterinária , Técnica Indireta de Fluorescência para Anticorpo/veterinária , Estudos Soroepidemiológicos , Ovinos , Doenças dos Ovinos/parasitologia , Doenças Transmitidas por Carrapatos/epidemiologia , Turquia/epidemiologia
19.
PLoS One ; 15(1): e0227234, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31923195

RESUMO

Hemangiosarcoma (HSA), a locally invasive and highly metastatic endothelial cell neoplasm, accounts for two-thirds of all cardiac and splenic neoplasms in dogs. Bartonella spp. infection has been reported in association with neoplastic and non-neoplastic vasoproliferative lesions in animals and humans. The objective of this study was to determine the prevalence of Bartonella spp. in conjunction with two other hemotropic pathogens, Babesia spp. and hemotropic Mycoplasma spp., in tissues and blood samples from 110 dogs with histopathologically diagnosed HSA from throughout the United States. This was a retrospective, observational study using clinical specimens from 110 dogs with HSA banked by the biospecimen repository of the Canine Comparative Oncology and Genomics Consortium. Samples provided for this study from each dog included: fresh frozen HSA tumor tissue (available from n = 100 of the 110 dogs), fresh frozen non-tumor tissue (n = 104), and whole blood and serum samples (n = 108 and 107 respectively). Blood and tissues were tested by qPCR for Bartonella, hemotropic Mycoplasma, and Babesia spp. DNA; serum was tested for Bartonella spp. antibodies. Bartonella spp. DNA was amplified and sequenced from 73% of dogs with HSA (80/110). In contrast, hemotropic Mycoplasma spp. DNA was amplified from a significantly smaller proportion (5%, p<0.0001) and Babesia spp. DNA was not amplified from any dog. Of the 100 HSA tumor samples submitted, 34% were Bartonella PCR positive (32% of splenic tumors, 57% of cardiac tumors, and 17% of other tumor locations). Of 104 non-tumor tissues, 63% were Bartonella PCR positive (56% of spleen samples, 93% of cardiac samples, and 63% of skin/subcutaneous samples). Of dogs with Bartonella positive HSA tumor, 76% were also positive in non-tumor tissue. Bartonella spp. DNA was not PCR amplified from whole blood. This study documented a high prevalence of Bartonella spp. DNA in dogs with HSA from geographically diverse regions of the United States. While 73% of all tissue samples from these dogs were PCR positive for Bartonella DNA, none of the blood samples were, indicating that whole blood samples do not reflect tissue presence of this pathogen. Future studies are needed to further investigate the role of Bartonella spp. in the development of HSA.


Assuntos
Babesia/genética , Babesiose/epidemiologia , Infecções por Bartonella/epidemiologia , Infecções por Bartonella/veterinária , Bartonella/genética , Doenças do Cão/epidemiologia , Hemangiossarcoma/epidemiologia , Hemangiossarcoma/veterinária , Infecções por Mycoplasma/epidemiologia , Infecções por Mycoplasma/veterinária , Mycoplasma/genética , Animais , Babesiose/parasitologia , Infecções por Bartonella/microbiologia , DNA Bacteriano/genética , Doenças do Cão/microbiologia , Doenças do Cão/parasitologia , Cães , Feminino , Hemangiossarcoma/microbiologia , Hemangiossarcoma/parasitologia , Masculino , Reação em Cadeia da Polimerase , Prevalência , Estudos Retrospectivos , Estados Unidos/epidemiologia
20.
Transfusion ; 60(2): 317-325, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31885102

RESUMO

BACKGROUND: Transfusion-transmitted Babesia microti is well recognized in the Northeast and upper Midwestern United States. Blood donation screening in Babesia-endemic states has occurred under investigational protocols prior to US Food and Drug Administration-licensed test availability. Here, we provide a prospective screening summary of nucleic acid testing (NAT) as part of a multicenter Babesia pivotal trial followed by extended investigational use. METHODS: From June 2017 to February 2018, 176,928 donation samples were tested with Procleix Babesia Assay (Grifols Diagnostic Solutions), a blood screening NAT for Babesia species ribosomal RNA detection using whole blood samples. During the pivotal trial, donations were collected in 11 endemic states plus Washington, DC, and Florida (nonendemic). Whole blood lysate samples were either tested in pools of 16 or individually. Reactive samples were confirmed by Babesia microti antibody and polymerase chain reaction (PCR) testing. If unconfirmed, further testing used a second PCR assay capable of detecting multiple Babesia species. Follow-up samples were also tested. Extended investigational testing followed pivotal trial completion. RESULTS: The pivotal trial identified 61 confirmed positives (176,608 donations): 35 (57%) PCR positive, 59 (97%) antibody positive, and two (3%) NAT positive/antibody negative, for a total yield of one positive per 2895 donations, including one Florida resident; others were from seven endemic states. During extended investigational testing of 496,270 donations in endemic states through January 2019, 211 (1:2351) repeat reactive donations were identified. CONCLUSIONS: Babesia was detected in donors from multiple US states, including one previously not associated with positive blood donors. This study supports the use of the Procleix Babesia Assay using individual testing or pools of up to 16.


Assuntos
Babesia/patogenicidade , Doadores de Sangue/estatística & dados numéricos , Programas de Rastreamento/métodos , Transcrição Genética/genética , Idoso , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase
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