Trypanosoma cruzi: blood parasitism kinetics and their correlation with heart parasitism intensity during long-term infection of Beagle dogs

The goals of the present study were to evaluate the kinetics of blood parasitism by examination of fresh blood, blood culture (BC) and PCR assays and their correlation with heart parasitism during two years of infection in Beagle dogs inoculated with the Be-78, Y and ABC Trypanosoma cruzi strains. Our results showed that the parasite or its kDNA is easily detected during the acute phase in all infected animals. On the other hand, a reduced number of positive tests were verified during the chronic phase of the infection. The frequency of positive tests was correlated with T. cruzi strain. The percentage of positive BC and blood PCR performed in samples from animals inoculated with Be-78 and ABC strains were similar and significantly larger in relation to animals infected with the Y strain.Comparison of the positivity of PCR tests performed using blood and heart tissue samples obtained two years after infection showed two different patterns associated with the inoculated T. cruzi strain: (1) high PCR positivity for both blood and tissue was observed in animals infected with Be-78 or ABC strains; (2) lower and higher PCR positivity for the blood and tissue, respectively, was detected in animals infected with Y strains. These data suggest that the sensitivity of BC and blood PCR was T. cruzi strain dependent and, in contrast, the heart tissue PCR revealed higher sensitivity regardless of the parasite stock.

Experimental infection of dogs with a Brazilian strain of Rickettsia rickettsii: clinical and laboratory findings

The bacterium Rickettsia rickettsii is the etiological agent of an acute, severe disease called Rocky Mountain spotted fever in the United States or Brazilian spotted fever (BSF) in Brazil. In addition to these two countries, the disease has also been reported to affect humans in Mexico, Costa Rica, Panama, Colombia and Argentina. Like humans, dogs are also susceptible to R. rickettsii infection. However, despite the wide distribution of R. rickettsii in the Western Hemisphere, reports of R. rickettsii-induced illness in dogs has been restricted to the United States. The present study evaluated the pathogenicity for dogs of a South American strain of R. rickettsii. Three groups of dogs were evaluated: group 1 (G1) was inoculated ip with R. rickettsii; group 2 (G2) was infested by R. rickettsii-infected ticks; and the control group (G3) was infested by uninfected ticks. During the study, no clinical abnormalities, Rickettsia DNA or R. rickettsii-reactive antibodies were detected in G3. In contrast, all G1 and G2 dogs developed signs of rickettsial infection, i.e., fever, lethargy, anorexia, ocular lesions, thrombocytopenia, anemia and detectable levels of Rickettsia DNA and R. rickettsii-reactive antibodies in their blood. Rickettsemia started 3-8 days after inoculation or tick infestation and lasted for 3-13 days. Our results indicate that a Brazilian strain of R. rickettsii is pathogenic for dogs, suggesting that canine clinical illness due to R. rickettsii has been unreported in Brazil and possibly in the other South American countries where BSF has been reported among humans.

Wild Triatoma infestans, a potential threat that needs to be monitored

The current persistence of Triatoma infestans, and therefore of Chagas disease transmission, in the Andean valleys of Bolivia and the Gran Chaco (precisely where wild populations of the vector are widespread), indicates a possible relationship between these two occurrences. This paper provides an overview of the current knowledge regarding wild T. infestans in Bolivia. The different morphs of the wild vector, their known distributions and some traits of their biology and ecology are presented. Particularly interesting is the considerable behavioural and chromatic plasticity that is displayed by wild T. infestans. According to the biogeographic region, different morphs of the vector occur in rupicolous habitats (common form and Mataral morph in Andean wild T. infestans) or arboreal ones ("dark morph" populations from the Chaco). The high genetic variability found at the microgeographical scale in Andean wild T. infestans favours the hypothesis that the Andes were the centre of origin and dispersal of T. infestans throughout South America. The relevant question regarding the origin of domestic populations is also addressed. Finally, current considerations of the epidemiological significance of wild T. infestans are discussed in the context of recent discoveries. Even if several observations support the epidemiological risk represented by wild T. infestans, the climatic and environmental conditions of their distribution areas would not favour the continued flow of triatomines between sylvatic refuges and domestic environments.

Rotavirus A genotype G1P[8]: a novel method to distinguish wild-type strains from the Rotarix® vaccine strain

Rotaviruses are important enteric pathogens for humans and animals. Group A rotaviruses (RV-A) are the most common agents of severe gastroenteritis in infants and young children and vaccination is the most effective method to reduce RV-A-associated diseases. G1P[8], the most prevalent RV-A genotype worldwide, is included in the RV-A vaccine Rotarix®. The discrimination between wild-type G1P[8] and vaccine G1P[8] strains is an important topic in the study of RV-A epidemiology to manage outbreaks and to define control measures for vaccinated children. In this study, we developed a novel method to segregate the wild-type and vaccine strains using restriction endonucleases. The dsRNA from the Rotarix® vaccine was sequenced and the NSP3 gene was selected as the target gene. The vaccine strain has a restriction pattern that is different than that of wild-type RV-A G1P[8] isolates after digestion with the restriction endonuclease BspHI. This pattern could be used as a marker for the differentiation of wild-type G1P[8] strains from the vaccine strain.

An experimental protocol for the establishment of dogs with long-term cellular immune reactions to Leishmania antigens

Domestic dogs are considered to be the main reservoirs of zoonotic visceral leishmaniasis. In this work, we evaluated a protocol to induce Leishmania infantum/Leishmania chagasi-specific cellular and humoral immune responses in dogs, which consisted of two injections of Leishmania promastigote lysate followed by a subcutaneous inoculation of viable promastigotes. The primary objective was to establish a canine experimental model to provide positive controls for testing immune responses to Leishmania in laboratory conditions. After inoculation of viable promastigotes, specific proliferative responses of peripheral blood mononuclear cells (PBMCs) to either Leishmania lysate or recombinant proteins, the in vitro production of interferon-γ by antigen-stimulated PBMCs and a significant increase in circulating levels of anti-Leishmania antibodies were observed. The immunized dogs also displayed positive delayed-type hypersensitivity reactions to Leishmania crude antigens and to purified recombinant proteins. An important finding that supports the suitability of the dogs as positive controls is that they remained healthy for the entire observation period, i.e., more than seven years after infection. Following the Leishmania antigen lysate injections, the infection of dogs by the subcutaneous route appears to induce a sustained cellular immune response, leading to an asymptomatic infection. This provides a useful model for both the selection of immunogenic Leishmania antigens and for immunobiological studies on their possible immunoprotective activities.

Leishmania infection in humans, dogs and sandflies in a visceral leishmaniasis endemic area in Maranhão, Brazil

Leishmania infection in humans, dogs and sandflies was examined in the endemic visceral leishmaniasis (VL) municipality of Raposa, state of Maranhão, Brazil. In this study, we examined Leishmania chagasi infection in the blood serum of both humans and Canis familiaris and the natural Leishmania sp. infection rate in the sandfly vector, Lutzomyia longipalpis. Enzyme-linked immunosorbent assay, indirect immunofluorescence reaction and polymerase chain reaction were performed to detect Leishmania infections in humans, dogs and sandflies, respectively. Overall, 186 out of 986 studied human beings were infected with L. chagasi parasites, representing an infection prevalence of 18.9%. An even higher infection rate was detected in dogs, where 66 (47.8%) out of 138 were infected. Among all Lu. longipalpis captured (n = 1,881), only 26.7% were females. The Leishmania infection frequency for the vector Lu. longipalpis was 1.56%. Remarkably, all infected sandflies were found in the peridomiciliary area. Furthermore, a high incidence of asymptomatic forms of VL in the human and canine populations was observed. The results of this study suggest autochthonous transmission of L. chagasi in this endemic area for visceral leishmaniasis because infection by Leishmania sp. was identified in all important elements of the transmission chain.

Susceptibility and resistance to deltamethrin of wild and domestic populations of Triatoma infestans (Reduviidae: Triatominae) in Bolivia: new discoveries

Bolivia is a high-endemic country for Chagas disease, for which the principal vector is Triatoma infestans (Triatominae). This is a mainly domestic species that is also found in the wild environment. Recently, an increasing number of studies have shown the importance of Triatominae resistance to insecticides, especially in Bolivia. Data regarding the susceptibility/resistance of wild and domestic populations of T. infestans to deltamethrin are presented. For the first time, domestic populations of the department of Santa Cruz were tested, showing low resistance. Although most of the wild populations were found to be susceptible to deltamethrin, three populations from three departments showed a mortality rate of less than 100%. This result is emphasised here.

Myenteric plexus is differentially affected by infection with distinct Trypanosoma cruzi strains in Beagle dogs

Chagasic megaoesophagus and megacolon are characterised by motor abnormalities related to enteric nervous system lesions and their development seems to be related to geographic distribution of distinct Trypanosoma cruzi subpopulations. Beagle dogs were infected with Y or Berenice-78 (Be-78) T. cruzi strains and necropsied during the acute or chronic phase of experimental disease for post mortem histopathological evaluation of the oesophagus and colon. Both strains infected the oesophagus and colon and caused an inflammatory response during the acute phase. In the chronic phase, inflammatory process was observed exclusively in the Be-78 infected animals, possibly due to a parasitism persistent only in this group. Myenteric denervation occurred during the acute phase of infection for both strains, but persisted chronically only in Be-78 infected animals. Glial cell involvement occurred earlier in animals infected with the Y strain, while animals infected with the Be-78 strain showed reduced glial fibrillary acidic protein immunoreactive area of enteric glial cells in the chronic phase. These results suggest that although both strains cause lesions in the digestive tract, the Y strain is associated with early control of the lesion, while the Be-78 strain results in progressive gut lesions in this model.

The TcI and TcII Trypanosoma cruzi experimental infections induce distinct immune responses and cardiac fibrosis in dogs

Trypanosoma cruzi infection may be caused by different strains with distinct discrete typing units (DTUs) that can result in variable clinical forms of chronic Chagas disease. The present study evaluates the immune response and cardiac lesions in dogs experimentally infected with different T. cruzi strains with distinct DTUs, namely, the Colombian (Col) and Y strains of TcI and TcII DTU, respectively. During infection with the Col strain, increased levels of alanine aminotransferase, erythrocytes, haematocrit and haemoglobin were observed. In addition, CD8+ T-lymphocytes isolated from the peripheral blood produced higher levels of interleukin (IL)-4. The latter suggests that during the acute phase, infection with the Col strain may remain unnoticed by circulating mononuclear cells. In the chronic phase, a significant increase in the number of inflammatory cells was detected in the right atrium. Conversely, infection with the Y strain led to leucopoenia, thrombopoenia, inversion of the ratio of CD4+/CD8+ T-lymphocytes and alterations in monocyte number. The Y strain stimulated the production of interferon-γ by CD4+ and CD8+ T-lymphocytes and IL-4 by CD8+ T-cells. In the chronic phase, significant heart inflammation and fibrosis were observed, demonstrating that strains of different DTUs interact differently with the host.

Leishmania amazonensis DNA in wild females ofLutzomyia cruzi (Diptera: Psychodidae) in the state of Mato Grosso do Sul, Brazil

Studies on natural infection by Leishmania spp of sandflies collected in endemic and nonendemic areas can provide important information on the distribution and intensity of the transmission of these parasites. This study sought to investigate the natural infection by Leishmaniain wild female sandflies. The specimens were caught in the city of Corumbá, state of Mato Grosso do Sul (Brazil) between October 2012-March 2014, and dissected to investigate flagellates and/or submitted to molecular analysis to detect Leishmania DNA. A total of 1,164 females (77.56% of which were Lutzomyia cruzi) representing 11 species were investigated using molecular analysis; 126 specimens of Lu. cruziwere dissected and also submitted to molecular analysis. The infection rate based on the presence of Leishmania DNA considering all the sandfly species analysed was 0.69%; only Leishmania (Leishmania) amazonensis was identified in Lu. cruzi by the molecular analysis. The dissections were negative for flagellates. This is the first record of the presence of L. (L.) amazonensis DNA in Lu. cruzi, and the first record of this parasite in this area. These findings point to the need for further investigation into the possible role of this sandfly as vector of this parasite.

Clostridium difficile ribotypes in humans and animals in Brazil

Clostridium difficile is an emerging enteropathogen responsible for pseudomembranous colitis in humans and diarrhoea in several domestic and wild animal species. Despite its known importance, there are few studies aboutC. difficile polymerase chain reaction (PCR) ribotypes in Brazil and the actual knowledge is restricted to studies on human isolates. The aim of the study was therefore to compare C. difficileribotypes isolated from humans and animals in Brazil. Seventy-six C. difficile strains isolated from humans (n = 25), dogs (n = 23), piglets (n = 12), foals (n = 7), calves (n = 7), one cat, and one manned wolf were distributed into 24 different PCR ribotypes. Among toxigenic strains, PCR ribotypes 014/020 and 106 were the most common, accounting for 14 (18.4%) and eight (10.5%) samples, respectively. Fourteen different PCR ribotypes were detected among human isolates, nine of them have also been identified in at least one animal species. PCR ribotype 027 was not detected, whereas 078 were found only in foals. This data suggests a high diversity of PCR ribotypes in humans and animals in Brazil and support the discussion of C. difficile as a zoonotic pathogen.

Vaccination of dogs with Trypanosoma rangeli induces antibodies against Trypanosoma cruzi in a rural area of Córdoba, Argentina

Dogs play a major role in the domestic cycle of Trypanosoma cruzi, acting as reservoirs. In a previous work we have developed a model of vaccination of dogs in captivity with nonpathogenic Trypanosoma rangeli epimastigotes, resulting in the production of protective antibodies against T. cruzi, with dramatic decrease of parasitaemia upon challenge with 100,000 virulent forms of this parasite. The aim of this work was to evaluate the immunogenicity of this vaccine in dogs living in a rural area. Domestic dogs, free from T. cruziinfection, received three immunisations with fixed T. rangeliepimastigotes. Dogs were not challenged with T. cruzi, but they were left in their environment. This immunisation induced antibodies againstT. cruzi for more than three years in dogs in their natural habitat, while control dogs remained serologically negative.

Cannibalism studies on eggs and newly hatched caterpillars in a wild population of Ascia monuste (Godart) (Lepidoptera, Pieridae)

Ascia monuste (Godart), also known as cabbage caterpillar, is considered herbivorous and specialist in Brassicaceae; however, it performs cannibalism. Experiments carried out in this work aimed to quantify the cannibalism occurrence in A. monuste, to verify whether cannibalism interferes in the species performance and to check whether cannibalism is influenced by the population size. The parameters used in order to evaluate the performance were time of development, adults weight and size, fecundity, oviposition and survival rate. Cannibalism occurred in all larval instars. There was a tendency to increase the cannibalism practice as the number of co-specific increased (potential preys). Cannibalism was more intense in larvae at the end of larval development. Generally, cannibalism did not interfere in the population performance. The most probable hypothesis for the occurrence of cannibalism in the conditions here studied would be the hypothesis of cannibalism as having the function of population control.

Influence of male nutritional conditions on the performance and alimentary selection of wild females of Anastrepha obliqua (Macquart)(Diptera, Tephritidae)

Influence of male nutritional conditions on the performance and alimentary selection of wild females of Anastrepha obliqua (Macquart) (Diptera, Tephritidae). The behavior of A. obliqua females is regulated by endogenous and exogenous factors and among these the presence of males. Experiments were carried out to investigate whether the presence of males and their nutritional condition may affect the behavior of self-selection feeding and the performance of A. obliqua females. Females were sorted in groups containing yeast-deprived females and males, and non-yeast-deprived females and males. The females were maintained apart from the males by a transparent plastic screen. Several yeast and sucrose combinations were offered to the females in a single diet block or in separate blocks. Ingestion, egg production, longevity and diet efficiency were determined. The non-yeast-deprived males positively influenced the females performance when the latter were fed with yeast and sucrose in distinct diet blocks. Performance was better in the groups without males and with yeast-deprived males where the females could not select the nutrient proportions (yeast and sucrose in a single diet block).

Selection of artificial hosts for oviposition by wild Anastrepha obliqua (Macquart) (Diptera, Tephritidae): influence of adult food and effect of experience

Several factors influence the selection of oviposition substrates by insects. The aim of the present work was to find answers to the following questions related to the oviposition behavior of Anastrepha obliqua. Can carbohydrates (glucose or sucrose) present in the adult diet have influence on the female preference for an oviposition substrate with similar composition? Can the previous experience with a host containing one of mentioned carbohydrates interfere in further selection of oviposition substrates? The results showed that the kind of carbohydrate present in the adult diet did not affect the female preference for an artificial oviposition substrate, neither when it was presented by itself nor in combination with brewer's yeast. The effect of experience in the oviposition behavior was observed when there was a previous contact with artificial oviposition substrates containing yeast and sucrose. The data are discussed in terms of the behavioral plasticity presented by this species in relation to feeding and oviposition behaviors.