RESUMO
Tabanids, stomoxyine flies, hippoboscids and tsetse flies are the most well-known brachyceran biting flies of livestock. Only a few other higher Diptera have developed the unique mouthparts required for blood feeding. These neglected blood feeders can also have direct effects on hosts through blood loss, and are likely to contribute to the transmission of pathogens. Musca crassirostris (Diptera: Muscidae) is one of the most abundant of the muscid flies with this haematophagous lifestyle; it is widespread in the Palaearctic, Afrotropical and Oriental regions. The present study reviews and summarizes the biology and morphology of this species, and its potential for impact on animals and humans. The study also provides a fully illustrated description of the fly to facilitate its identification, and reviews information on abundance, with a focus on recent trapping surveys in Thailand. When sampled using traps designed for other biting flies, M. crassirostris appears to be four and 45 times more abundant than stomoxyines and tabanids, respectively. High numbers of M. crassirostris in the vicinity of livestock have also been associated with outbreaks of disease, such as that of a fatal plague in bovine farms in Egypt. This calls for a reconsideration of its potential impacts on livestock economics and health, and thus the development of suitable control methods.
Assuntos
Controle de Insetos , Insetos Vetores , Características de História de Vida , Muscidae , Doenças dos Animais , Animais , Comportamento Alimentar , Insetos Vetores/anatomia & histologia , Insetos Vetores/classificação , Insetos Vetores/fisiologia , Gado , Muscidae/anatomia & histologia , Muscidae/classificação , Muscidae/fisiologia , Densidade Demográfica , TailândiaRESUMO
SUMMARY: This study investigated the molecular prevalence of Trypanosoma lewisi and T. evansi in wild rodents from Cambodia, Lao PDR and Thailand. Between 2008 and 2012, rodents (and shrews) were trapped in nine locations and 616 of these were tested using three sets of primers: TRYP1 (amplifying ITS1 of ribosomal DNA of all trypanosomes), TBR (amplifying satellite genomic DNA of Trypanozoon parasites) and LEW1 (amplifying ITS1 of ribosomal DNA of T. lewisi). Based on the size of the PCR products using TRYP1, 17% were positive for T. lewisi and 1·0% positive for Trypanozoon. Results were confirmed by sequencing PCR products and by using more specific primers (LEW1 and TBR). The specificity of TRYP1 primers, however, failed as rodent DNA was amplified in some instances, giving unexpected product sizes. Using LEW1 primers, 13·3% of the samples were confirmed positive for T. lewisi, both by PCR and sequencing. In Thailand, T. lewisi was found in Rattus tanezumi, R. exulans and Berylmys; in Lao PDR, in R. tanezumi and R. exulans, and in Cambodia in R. tanezumi, R. exulans and R. norvegicus. Using TBR, 1·3% of the samples tested positive for Trypanozoon by PCR and sequencing; T. evansi is the only species of the Trypanozoon subgenus possibly present in wild Asian rodents. These results confirmed its presence in rodents from Thailand (R. tanezumi), Lao PDR (R. tanezumi, R. nitidus) and Cambodia (R. tanezumi, Niviventer fulvescens, Maxomys surifer). Based on the information related to rodent trapping, it was found that rodent species trapped in and around human dwellings had a higher prevalence of T. lewisi infection. R. tanezumi and R. exulans, two synanthropic species, were mainly found infected in this habitat suggesting a role as a reservoir and thus a potential source of T. lewisi for human infection.
Assuntos
Doenças dos Roedores/parasitologia , Trypanosoma/classificação , Tripanossomíase/veterinária , Envelhecimento , Animais , Animais Selvagens , Sudeste Asiático , Ecossistema , Feminino , Humanos , Masculino , Modelos Biológicos , Prevalência , Doenças dos Roedores/epidemiologia , Roedores , Estações do Ano , Estudos Soroepidemiológicos , Fatores Sexuais , Tripanossomíase/epidemiologia , Tripanossomíase/parasitologiaRESUMO
Trypanosoma evansi is a blood parasite responsible for surra in mammals, with a high impact in camels and horses. The WOAH-recommended reference method for detecting immunoglobulin G directed against T. evansi is ELISA, using whole cell lysate antigens (WCLAs). WCLAs are prepared with T. evansi produced in laboratory rodents, separated from blood cells using DE-cellulose anion exchange chromatography. As parasite lysates are fragile, antigens are preserved frozen pending use. For these reasons and others, T. evansi WCLAs are not commercially available. They are produced in small quantities, in a limited number of specialized laboratories, and they require a reliable and expensive cold chain for their shipment. In this study, we assessed and validated in vitro production of T. evansi and lyophilization of WCLAs in comparison with the reference method using frozen WCLAs prepared with parasites produced in rodents. Using a set of 400 samples monthly collected from 12 naturally infected camels followed-up for 1384 days, and two batches of referenced serum samples (infected, n = 12; non-infected, n = 15), statistical studies on qualitative and semi-quantitative results of the ELISAs did not show any significant difference when comparing the four combinations of parasites produced in vivo or in vitro, and frozen or freeze-dried WCLSAs. A repeatability study (28 repeats in 9 serum samples) was fully satisfying (p-value = 0.055). With the more convenient in vitro-produced freeze-dried WCLAs it was possible to: (i) avoid the ethical concern of in vivo production, (ii) improve the standardization of antigen production, (iii) secure antigen preservation during shipment and (iv) save a considerable amount of money (DE52-cellulose and dry-ice cold chain being avoided). Additional studies with other Trypanosoma spp are required for further extending ELISA to regional laboratories in enzootic areas, especially in view of the current progress in the "Progressive Control Pathway" (PCP) for trypanosomes in Africa.
Assuntos
Doenças dos Cavalos , Trypanosoma , Tripanossomíase , Animais , Cavalos , Camelus/parasitologia , Tripanossomíase/diagnóstico , Tripanossomíase/veterinária , Tripanossomíase/parasitologia , Antígenos de Protozoários , Ensaio de Imunoadsorção Enzimática/veterinária , Ensaio de Imunoadsorção Enzimática/métodosRESUMO
Trypanosoma congolense forest-type was identified by PCR in France, in a dog returning from Senegal. This paper describes the morphological features of the parasite on Giemsa-stained smears. Slender forms and "latent bodies" represent 30.4% and 20.4%, respectively. Some rosettes have been observed (0.8%). The predominant form (48.4%) is stumpy, close to "montgomeryi-form", but it is unusually broad, with a width/length ratio (WLr) of 0.40-0.55, while that of "montgomeryi-forms" is close to 0.3. To the best of our knowledge, this is the first description of such a form of T. (Nannomonas). Also unusual, the shape of the cytoplasm appears to be tightened by an "S-" or "C-" shaped flagellum. We propose naming this peculiar morphotype "hyperpachymorph", and adding its description to that of T. congolense forest-type. Thus T. (Nannomonas) forms would include: sphaeromorph or "latent body-form" (globular), hyperleptomorph (rodhaini-form, very long and slender, with a free flagellum); leptomorph (simiae-form, slender, with a free flagellum); isomorph (congolense-form, short, generally without a free flagellum); pachymorph (montgomeryi-form, short and stout; 0.25 < WLr < 0.34, without a free flagellum), and hyperpachymorph ("hyper montgomeryi-form", short and very stout; 0.35 < WLr < 0.7, without a free flagellum).
Assuntos
Doenças do Cão/parasitologia , Trypanosoma congolense/isolamento & purificação , Tripanossomíase Africana/veterinária , Animais , DNA de Protozoário/isolamento & purificação , Doenças do Cão/tratamento farmacológico , Cães , Evolução Fatal , França , Injeções Intramusculares/veterinária , Masculino , Pentamidina/administração & dosagem , Pentamidina/uso terapêutico , Reação em Cadeia da Polimerase/veterinária , Senegal , Viagem , Tripanossomicidas/administração & dosagem , Tripanossomicidas/uso terapêutico , Trypanosoma congolense/classificação , Trypanosoma congolense/genética , Trypanosoma congolense/ultraestrutura , Tripanossomíase Africana/tratamento farmacológico , Tripanossomíase Africana/parasitologiaRESUMO
The abundance and species diversity of tabanids were evaluated by trapping of insects using Vavoua traps, during the rainy season, from October 4 to November 30, 2009, in three different habitats: primary forest, secondary forest and village, in the biosphere reserve Ipassa-IRET Makokou in Gabon. Eight species belonging to three genera of tabanids have been identified for a total of 402 specimens caught. The tabanid species numerically the most abundant were: Tabanus secedens Walker, 1854 (55.2%), Tabanus obscurehirtus Ricardo, 1908 (13.9%), Chrysops dimidiatus Wulp, 1885 (11.2%) and Chrysops silaceus Austen, 1907 (10.7%). The less abundant species were Tabanus par Walker, 1854 (3.2%), Tabanus besti arbucklei Austen, 1912 (3%), Tabanus marmorosus congoicola Bequaert, 1930 (1%) and Ancala fasciata fasciata (Fabricius, 1775) (0.5%). Specimens of the genera Tabanus and Chrysops could not be identified, these insects represented respectively 0.7% and 0.5% of the insects trapped. The highest proportion of tabanids was trapped in secondary forest (75.1%) and the lower in primary forest (4.5%).
Assuntos
Biodiversidade , Dípteros/classificação , Dípteros/crescimento & desenvolvimento , Animais , Ecossistema , Gabão , Chuva , Estações do Ano , ÁrvoresRESUMO
Animal trypanosomoses due to trypanosomes of African origin (ATAO), mainly caused by Trypanosoma congolense type Savannah (TCS), T. brucei brucei (TBB), T. vivax (TV), and T. evansi, are widespread diseases that affect domestic and wild mammals and have a huge economic impact. ATAO clinical suspicions are usually confirmed by parasitological and molecular methods, while sero-epidemiological surveys are generally carried out using the OIE-recommended ELISA method based on whole cell lysate soluble antigens (WCLSA) from purified trypanosomes; this reagent is usually stored frozen. With a view to expanding this ELISA test, we assessed, standardized, and validated the use of dehydrated rather than frozen WCLSA and serum samples. For the three ELISA assays (TV, TCS & TBB), a repeatability study revealed no significant difference between repeats. The results obtained using frozen rather than freeze-dried antigen and serum strongly correlated for Pearson's correlation values (>0.93) and Lin's measure ("very good" to "excellent"). Reproducibility was robust, with Pearson's correlation values >0.97 for inter technician effects, and 0.87 (TV) to 0.97 (TBB & TCS) for inter-laboratory tests; their combination was "very satisfactory" to "excellent" according to Lin's measure and there was no impact on qualitative test results. Dehydrated reagents offer the advantage of shipment at room temperature, allowing the secured provision of reagents to regional laboratories. Together with a compendium of standard diagnostic protocols for ATAO (/OIE), dehydrated reagents will enable the serological diagnosis of ATAO at regional level in endemic countries. This very welcome improvement in the context of the Progressive Control Pathway for trypanosomes, recently launched by African countries, will possibly be extended to Latin America in the near future.
Assuntos
Trypanosoma congolense , Trypanosoma , Tripanossomíase Africana , Animais , Ensaio de Imunoadsorção Enzimática/veterinária , Indicadores e Reagentes , Mamíferos , Reprodutibilidade dos Testes , Trypanosoma vivax , Tripanossomíase Africana/diagnóstico , Tripanossomíase Africana/veterináriaRESUMO
Fasciola gigantica is the main fasciolid species in Africa; however, F. hepatica and F. gigantica overlap in some countries. Egypt deserves mentioning because of the emerging situation of human fascioliasis in the Nile Delta area. The morphometric characteristics of fasciolid adults infecting the main livestock species present in the Nile Delta human endemic area are analyzed through a computer image analysis system (CIAS) on the basis of standardized measurements known to be useful for the differentiation of both fasciolid species. This is the first time that such a study is performed in an African country and, therefore, the results are compared to (i) F. hepatica (European Mediterranean area) and F. gigantica (Burkina Faso) standard populations, i.e. geographical areas where both species do not co-exist, and (ii) F. hepatica and F. gigantica populations from geographical areas where both species do co-exist, including the presence of intermediate forms (Iran). Results indicate the presence of F. hepatica, F. gigantica and intermediate forms (Fasciola sp.) in Egypt for the first time, and demonstrate the usefulness of CIAS for the phenotypic characterization of liver fluke adults from a concrete fascioliasis endemic area. Body roundness, body length over body width, and distance between the ventral sucker and the posterior end of the body provide useful tools for studying inter- and intraspecific morphological diversity in Fasciola adults. The application of these markers to specimens from geographical areas where F. hepatica and F. gigantica co-exist, such as in Egypt and Iran, suggest a strong population-level variation in Fasciola adult morphology.
Assuntos
Doenças dos Bovinos/parasitologia , Doenças Endêmicas , Fasciola/anatomia & histologia , Fasciola/genética , Fasciolíase/veterinária , Fenótipo , Animais , Búfalos , Bovinos , Doenças dos Bovinos/epidemiologia , Egito/epidemiologia , Fasciola/classificação , Fasciolíase/epidemiologia , Fasciolíase/parasitologia , HumanosRESUMO
The first outbreak of trypanosomosis caused by Trypanosoma evansi in camels in France was reported on a farm in the Aveyron Department. Five camels were imported from the Canary Islands to the farm in early July 2006, and trypanosomes were observed on a stained blood smear from one of them, which died in October. On further investigations, trypanosomes were observed in the blood of five camels, three of them indigenous to the farm and two that had been imported. On the basis of microscopical examination (morphological criteria and measurements) and serological results based on the card agglutination T evansi test and PCR typing, the parasites were identified as T evansi. After treatment with melarsomine, the infected camels rapidly became negative by parasitological tests and were negative two to four months later by serological tests. The parasite was probably transmitted by tabanids and Stomoxys calcitrans, which were abundant in July to September 2006. No parasites were observed in other animals on the farm or on neighbouring farms, but some of the sheep on these farms were positive by PCR or serology.
Assuntos
Camelus/parasitologia , Surtos de Doenças/veterinária , Trypanosoma/classificação , Tripanossomíase/veterinária , Animais , Arsenicais/uso terapêutico , França/epidemiologia , Insetos Vetores/parasitologia , Muscidae/parasitologia , Triazinas/uso terapêutico , Tripanossomicidas/uso terapêutico , Trypanosoma/isolamento & purificação , Tripanossomíase/tratamento farmacológico , Tripanossomíase/epidemiologia , Tripanossomíase/parasitologiaRESUMO
An epidemiological study was conducted to determine the prevalence of trypanosomosis in cattle, small ruminants and Equidae, and to identify biting flies; potential mechanical vectors of trypanosomes in the three districts of Bahir Dar Zuria, Dembia and Fogera, bordering lake Tana, Ethiopia. About 1509 cattle, 798 small ruminants and 749 Equidae were bled for the prevalence study using the buffy-coat method and the measurement of the hematocrit value. Sixty-six NGU and 20 monoconical traps were deployed for the fly survey. The results indicated the presence of trypanosomes in 6.1% (92/1509) of the cattle with a maximum during the late rainy season (9.6%) than the early dry season (3.6%) at Fogera district. Prevalence at the district level varied from 4% to 9.6%. Only one sheep (1/122) and one goat (1/676) were found positive for T. vivax-like trypanosomes and none of the Equidae was positive. All the trypanosomes encountered in cattle belong to the single species of T. vivax. The PCV was negatively associated with detection of T. vivax (21.6% in infected versus 25.4% in non-infected cattle). A total of 55,398 biting flies were caught of which 49,353 (89.08%) belong to Stomoxys, 4715 (8.51%) to horse flies and 1330 (2.4%) to Chrysops species. There was no tsetse fly. Species identification has indicated the presence of Atylotus agrestis, Chrysops streptobalia, Stomoxys calcitrans, S. nigra, S. pulla, S. pallida, S. sitiens, S. taeniata, S. uruma, Haematopota lasiops and Hippobosca variegata. The overall apparent density was 214.7flies/trap/day. Seasonal comparison showed higher fly catches in the late rainy season than the early dry season. This study indicated that T. vivax infections culminate in cattle at the same time as mechanical vectors such as Stomoxys sp. and Atylotus agrestis. Therefore, attention towards T. vivax infection in cattle is essential to control the impact of the disease on productivity. A further study on biting flies is recommended.
Assuntos
Anticorpos Antiprotozoários/sangue , Dípteros/parasitologia , Insetos Vetores/parasitologia , Trypanosoma vivax/imunologia , Tripanossomíase Africana/veterinária , Animais , Bovinos , Equidae , Etiópia/epidemiologia , Cabras , Mordeduras e Picadas de Insetos , Estações do Ano , Estudos Soroepidemiológicos , Ovinos , Tripanossomíase Africana/epidemiologia , Tripanossomíase Africana/transmissãoRESUMO
A longitudinal epidemiological survey of bovine trypanosomosis and its vectors was carried out in the Volta river basin of Northern Ghana to determine the relationship between cattle management and the incidence of bovine trypanosomosis. Two groups of sentinel cattle under different systems of management, classified as "fully-sedentary" and "partially-sedentary" (depending on the type of management) were followed over a 1-year period starting from March 2003 onwards. Cattle were screened at intervals of 3 months using the buffy coat technique (BCT). Buffy coat specimen from animals that were positive for the BCT and those that were negative, but with a packed cell volume (PCV) of less than 21% were further tested using the polymerase chain reaction (PCR). Plasma from all animals were tested for antibody using the indirect antibody enzyme-linked immunosorbent assay (ELISA). Trypanosomosis challenge was determined in tandem with the epidemiological survey with watering sites of sentinel cattle being the foci of interest. The parasitological prevalence at the start of the survey was higher in the fully-sedentary group (9%) than in the partially-sedentary group (3%). In subsequent visits, however, the parasitological incidence was consistently higher in the partially-sedentary group than in the fully-sedentary group. The mean seroprevalence (ELISA) of both groups increased from 3% in March to 54% in December. Statistical analysis of the serological results using a random effect logistic regression, showed a significant difference in incidence of bovine trypanosomosis between the two groups. There was also a significant effect of time. The influence of cattle herding on host-vector-parasite interface and its consequence on the incidence of trypanosomosis are discussed.
Assuntos
Criação de Animais Domésticos/métodos , Insetos Vetores/crescimento & desenvolvimento , Trypanosoma/isolamento & purificação , Tripanossomíase Bovina/epidemiologia , Moscas Tsé-Tsé/crescimento & desenvolvimento , Animais , Anticorpos Antiprotozoários/sangue , Bovinos , Ensaio de Imunoadsorção Enzimática/veterinária , Gana/epidemiologia , Hematócrito/veterinária , Insetos Vetores/parasitologia , Modelos Logísticos , Estudos Longitudinais , Parasitemia/parasitologia , Parasitemia/transmissão , Parasitemia/veterinária , Rios , População Rural , Estações do Ano , Estudos Soroepidemiológicos , Tripanossomíase Bovina/parasitologia , Tripanossomíase Bovina/transmissão , Moscas Tsé-Tsé/parasitologiaRESUMO
Primers hybridising with the rDNA cistron have previously been evaluated for PCR diagnosis specific for kinetoplastids, and shown to detect and differentiate the Trypanosoma brucei complex and Trypanosoma cruzi. Kin1 and Kin2 primers, amplifying internal transcribed spacer 1, were subsequently evaluated for the diagnosis of African livestock trypanosomosis. Based on the size of the PCR products obtained, Kin primers allowed detection and identification of three Trypanosoma congolense types (savannah, forest and Kenya Coast), with distinction among themselves and from the subgenus Trypanozoon (T. brucei spp., Trypanosoma evansi and Trypanosoma equiperdum), Trypanosoma vivax, Trypanosoma simiae and Trypanosoma theileri. These primers were shown to be suitable for the sensitive and type-specific diagnosis of African livestock trypanosome isolates through a single PCR even in the case of multi-taxa samples. With field samples (buffy-coat from cattle blood) sensitivity was close to the sensitivity observed in single reactions with the classical specific primers for the Trypanozoon subgenus and T. congolense-type savannah, but was lower for detection of T. vivax. Additional reaction, improvement of DNA preparation, and/or new primers design are necessary to improve the sensitivity for detection of T. vivax in field samples. However, these primers are suitable for isolate typing through a single PCR.
Assuntos
DNA de Protozoário/genética , DNA Espaçador Ribossômico/genética , Trypanosoma/genética , Tripanossomíase Bovina/diagnóstico , Animais , Burkina Faso , Bovinos , Primers do DNA , DNA de Protozoário/química , DNA Espaçador Ribossômico/química , Eletroforese em Gel de Ágar/veterinária , Reação em Cadeia da Polimerase/veterinária , Sensibilidade e Especificidade , Trypanosoma/química , Trypanosoma/classificaçãoRESUMO
Preliminary studies in French Guyana with sheep experimentally infected with a local isolate of Trypanosoma vivax tended to show poor sensitivity and/or specificity of the monoclonal antibodies used in kits for the antigen-detection (Ag) ELISA for T. vivax, T. brucei, and T. congolense. To reevaluate these kits, 4 calves were infected at ILRAD, Nairobi, Kenya, with the same isolate. Blood samples were taken daily for 51 days, and examined directly on blood smears and buffy coat, and using Ag-ELISA for the three species. For the 4 calves, on 158 tests performed over the first 51 days of infection, the percentages of positive results were 66% on buffy coat; on Ag-ELISA 3.8% for T. vivax, 4.4% for T. brucei, and 3.1% for T. congolense. Blood smears showed only T. vivax. These results confirm those previously obtained in French Guyana: the test for T. vivax (at least in the initial stage) shows a very low sensitivity, far below that of parasitological techniques, and the specificity of the T. brucei and T. congolense tests is low. Certain surprising results obtained in Africa might also be due to a poor sensitivity and specificity of the monoclonals used. As ELISA is the technic of choice for epidemiological surveys, and antigen detection a logical way to confirm whether an animal is actively infected by trypanosomes, the Ag-ELISA remains a necessary tool for epidemiological surveys of trypanosomes; new monoclonals are required to develop more specific and sensitive tests.
Assuntos
Ensaio de Imunoadsorção Enzimática/veterinária , Doenças dos Ovinos/diagnóstico , Trypanosoma brucei brucei , Trypanosoma congolense , Trypanosoma vivax , Tripanossomíase Africana/veterinária , Tripanossomíase Bovina/diagnóstico , Animais , Anticorpos Monoclonais , Bovinos , Intervalos de Confiança , Ensaio de Imunoadsorção Enzimática/métodos , Guiana Francesa , Parasitemia/diagnóstico , Parasitemia/veterinária , Kit de Reagentes para Diagnóstico/veterinária , Sensibilidade e Especificidade , Ovinos , Tripanossomíase Africana/diagnósticoRESUMO
Although Guyana, Suriname, and French Guiana share borders and climatic and geographic similarities, the countries have maintained little contact, due to language, political, and administrative differences. In 1993, two international organizations involved in the improvement of animal health, the Inter-American Institute for Cooperation on Agriculture (IICA) and CIRAD-EMVT (Centre de Cooperation Internationale en Recherche Agronomique pour le Developpement-Elevage et Medecine Veterinaire des Pays Tropicaux), jointly developed a collaborative project between the veterinary services of the three countries entitled "Hemoparasite Network for the Guianas." This project seeks to pool livestock, laboratory, and technical resources between the three countries in order to generate and exchange information on hemoparasites of livestock. A Hemoparasite Reference Laboratory for the Guianas has been created at the CIRAD-EMVT laboratory in Cayenne, French Guiana. Besides processing ruminant serum samples from the three countries, specialists from this organization conduct training in hemoparasite diagnostic techniques for laboratory personnel from Guyana and Suriname. A large-scale epidemiologic study of hemoparasites of cattle in the three countries is under way, to determine the prevalence, distribution, and clinical and economic significance of hemoparasites in the three countries, particularly Trypanosoma vivax and T. evansi. Preliminary results are presented and discussed. A Hemoparasite Information Network (TRYPNET) has been initiated, including a quarterly hemoparasite newsletter (TRYPNEWS), published in English and Spanish and disseminated to researchers in the Americas, Europe, and Africa. In 1995/96, it is proposed to expand the network's scope to include Venezuela and Brazil.
Assuntos
Doenças dos Bovinos/epidemiologia , Bases de Dados Factuais , Doenças Parasitárias em Animais , Matadouros , Anaplasmose/epidemiologia , Animais , Bovinos , Demografia , Guiana Francesa/epidemiologia , Guiana/epidemiologia , Cooperação Internacional , Doenças Parasitárias/epidemiologia , Suriname/epidemiologia , Carrapatos , Trypanosoma vivax , Tripanossomíase/epidemiologia , Tripanossomíase/veterinária , Tripanossomíase Africana/epidemiologia , Tripanossomíase Africana/veterináriaRESUMO
Polymerase chain reaction (PCR) with specific oligonucleotides for the amplification of Trypanosoma vivax DNA has been developed by Masiga et al. (1992) to detect the presence of T. vivax DNA in biting flies. The aim of this experiment was to evaluate the efficacy of this technique when applied directly on cattle serum, without DNA purification, to detect infection. The sensitivity of this PCR technique was compared with parasitological techniques, namely haematocrit centrifuge technique (HCT) and buffy coat method (BCM), and with the antigen-enzyme-linked immunosorbent assay (Ag-ELISA) for T. vivax developed by Nantulya and Lindqvist (1989). Blood and serum samples were collected from four calves experimentally infected with a stock of T. vivax from French Guyana (IL4007). During the first 51 days of infection, a total of 164 samples were collected and processed using the four tests. Mean percentages of positive results were 68% with HCT, 59% with BCM, 4% with Ag-ELISA and 64% with PCR. Parasitological and PCR techniques yielded approximately the same sensitivities. PCR was able to detect active infection in serum samples when parasitaemia was over 10(3) trypanosomes/ml. With this isolate of T. vivax the Ag-ELISA was not found to be sensitive enough to be used as a diagnostic tool. The sensitivity of this PCR technique is not greater than parasitological techniques but it allows delayed processing of the samples and gives a highly species-specific diagnosis. This simple PCR technique should be evaluated for field diagnosis because it makes retrospective epidemiological survey using serum banks possible. Moreover, it can be substituted to parasitological techniques when immediate examination is not feasible.
Assuntos
Reação em Cadeia da Polimerase , Trypanosoma vivax , Tripanossomíase Africana/veterinária , Tripanossomíase Bovina/diagnóstico , Animais , Bovinos , Ensaio de Imunoadsorção Enzimática , Estudos de Avaliação como Assunto , Reação em Cadeia da Polimerase/métodos , Tripanossomíase Africana/diagnósticoRESUMO
This paper aims to review the applications of the polymerase chain reaction (PCR) for the detection and identification of trypanosomes in animals. The diagnosis of trypanosomes, initially based on microscopic observations and the host range of the parasites, has been improved, since the 1980s, by DNA-based identification. These diagnostic techniques evolved successively through DNA probing, PCR associated to DNA probing, and currently to PCR alone. Several DNA sequences have been investigated as possible targets for diagnosis, especially multi-copy genes such as mini-exon, kinetoplastid mini-circles, etc., but the most favoured target is the nuclear satellite DNA of mini-chromosomes, which presents the advantages, and the drawbacks, of highly repetitive short sequences (120-600 bp). Several levels of specificity have been achieved from sub-genus to species, sub-species and even types. Random priming of trypanosome DNA has even allowed "isolate specific" identification. Other work based on microsatellite sequences has provided markers for population genetic studies. For regular diagnosis, the sensitivity of PCR has increased with the advancement of technologies for sample preparation, to reach a level of 1 trypanosome/ml of blood, which has brought to field samples a sensitivity two to three times higher than microscopic observation of the buffy coat. Similarly, PCR has allowed an increase in the specificity and sensitivity of diagnosis in vectors such as tsetse flies. However, because of the diversity of Trypanosoma species potentially present in a single host, PCR diagnosis carried out on host material requires several PCR reactions; for example, in cattle, up to five reactions per sample may be required. Research is now focusing on a diagnosis based on the amplification of the internal transcribed spacer-1 (ITS-1) of ribosomal DNA which presents the advantages of being a multi-copy locus (100-200), having a small size (300-800 bp), which varies from one taxon to another but is conserved in size in a given taxon. This may lead to the development of a multi-species-specific diagnostic protocol using a single PCR. By reducing the cost of the PCR diagnosis, this technique would allow a greater number of field samples to be tested in epidemiological studies and/or would increase the variety of Trypanosoma species that could be detected. Further investigations are required to develop and optimise multi-species-specific diagnostic tools for trypanosomes, which could also serve as a model for such tools in other pathogens.
Assuntos
Reação em Cadeia da Polimerase/métodos , Trypanosoma/genética , Trypanosoma/isolamento & purificação , Tripanossomíase/diagnóstico , Tripanossomíase/veterinária , Animais , DNA Espaçador Ribossômico/genética , Genes de Protozoários/genética , Genoma de Protozoário , Sensibilidade e EspecificidadeRESUMO
The diagnostic performance of a polymerase chain reaction assay (PCR) for monitoring the effectiveness of aceturate diminazene treatment was compared with those of an antibody-detection ELISA test and the buffy-coat technique using sheep experimentally infected with either savannah-type or forest-type Trypanosoma congolense or T. vivax. Within the period of infection, the PCR using specific savannah-type T. congolense primers showed a significant higher diagnostic sensitivity (p<0.05) than the buffy-coat technique. Both techniques gave closed results for detecting forest-type T. congolense or T. vivax infections. Following trypanocidal treatment, the PCR showed that specific product disappeared definitively 1 or 2 days later in animals in which a decrease of the antibody level and a significant improvement of the red packed cell volume were observed. The occurrence of relapse infection was detected by the PCR in one animal infected by T. vivax on day 19 post-treatment and confirmed by the persistence and increasing antibody level whereas the buffy-coat technique detected parasites 42 days later. Then, the PCR signals remained positive on several occasions while parasitaemia was detected only two times.The application of PCR combined with the antibody detection appeared to provide a useful tool as compared to the buffy-coat technique for monitoring the effectiveness of trypanocidal treatment.
Assuntos
Diminazena/análogos & derivados , Diminazena/uso terapêutico , Reação em Cadeia da Polimerase/veterinária , Doenças dos Ovinos/tratamento farmacológico , Tripanossomicidas/uso terapêutico , Tripanossomíase/veterinária , Animais , Anticorpos Antiprotozoários/análise , Ensaio de Imunoadsorção Enzimática/veterinária , Reação em Cadeia da Polimerase/métodos , Ovinos , Tripanossomíase/tratamento farmacológicoRESUMO
Inbred Balb/c mice were infected with three clones of Trypanosoma congolense (Sam.28.1, Dind.3.1 and K60.1A) corresponding, respectively, to the three genetically distinct types (savannah, forest and kilifi) defined within this species, for the purpose of comparing their pathogenicity for a better understanding of the epidemiology of African trypanosomosis. Another clone of savannah type, IL 3000, was also tested simultaneously to study a probable strain variation. Both the clones of savannah type were found of extreme virulence with loss of appetite, rough hair, rapid respiration, lethargy, and all mice died within a week. Parasitaemias evolved rapidly to the first peak by day 3-5 post-inoculation without any remission and the course of disease was correlated positively with the prepatent period. The clones of the forest type and the kilifi type were of low virulence with chronic infection and symptoms progressively less patent throughout the infection; only one mouse died in each experimental group.
Assuntos
Anticorpos Antiprotozoários/análise , Trypanosoma congolense/genética , Trypanosoma congolense/patogenicidade , Tripanossomíase Africana/veterinária , Animais , Anticorpos Antiprotozoários/biossíntese , Ensaio de Imunoadsorção Enzimática/veterinária , Cinética , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Parasitemia/veterinária , Análise de Sobrevida , Fatores de Tempo , Trypanosoma congolense/imunologia , Tripanossomíase Africana/epidemiologia , Tripanossomíase Africana/parasitologia , VirulênciaRESUMO
The pathology of African bovine trypanosomosis was compared in Zebu cattle subcutaneously inoculated with three clones of trypanosomes corresponding to the three genetically distinct types of Trypanosoma congolense; savannah-type, west African riverine/forest-type and kilifi-type. All inoculated animals became parasitaemic between 7 and 11 days post-infection (dpi). The savannah-type showed consistently higher levels of parasitaemia and lower packed red cell volume percentages and leukocyte counts than the other two types. The syndrome was also more severe in the savannah-type and led inexorably to death between 29 and 54 dpi while animals with the forest or the kilifi-types recovered from earlier symptoms and haematological alterations after 3 months of infection. By the end of the experiment, the animals self-cured from the forest-type infection and the kilifi-type passed under control. The results of the present study indicated clear difference in pathogenicity between the three types of T. congolense; the savannah-type was virulent while the forest-type was of low pathogenicity and the kilifi-type was non-pathogenic.
Assuntos
Trypanosoma congolense/genética , Trypanosoma congolense/patogenicidade , Tripanossomíase Bovina/parasitologia , Animais , Bovinos , Hematócrito/veterinária , Cinética , Contagem de Leucócitos/veterinária , Parasitemia/sangue , Parasitemia/parasitologia , Parasitemia/veterinária , Fatores de Tempo , Trypanosoma congolense/classificação , Tripanossomíase Africana/sangue , Tripanossomíase Africana/parasitologia , Tripanossomíase Africana/veterinária , Tripanossomíase Bovina/sangue , VirulênciaRESUMO
The epidemiology of bovine trypanosomosis was investigated in two districts (Savelugu and West Mamprusi) of Northern Ghana with different land use and environmental characteristics. The land use intensity and environmental change was suspected to be higher in the Savelugu District. A cross-sectional entomological survey conducted along the White Volta river and its tributaries confirmed the presence of only Glossina palpalis gambiensis and G. tachinoides. The challenge index as measured by the product of tsetse density and tsetse infection rate was much higher in the West Mamprusi (19.6) than in the Savelugu district (4.7). A total of 1013 cattle (508 in Savelugu and 505 in West Mamprusi) were bled from a random selection of 16 villages in the Savelugu District and 13 villages in the West Mamprusi District. Blood samples were examined for trypanosomes by the buffy coat technique (BCT). Blood samples that were positive in the BCT or negative in the BCT but with packed cell volume (PCV) values below 21 were further tested with a polymerase chain reaction for trypanosomal DNA. Plasma samples of all cattle were serologically tested with an indirect ELISA for trypanosomal antibodies. The parasitological and serological prevalence of bovine trypanosomoses was significantly higher in West Mamprusi (16 and 53%, respectively) than in Savelugu District (8 and 24%, respectively). An evaluation of animal health at the village herd level, using PCV as an index of anaemia, provided various epidemiological scenarios prevalent in the entire study area.