Evaluation of homologous, heterologous, and affinity conjugates for the serodiagnosis of Toxoplasma gondii and Neospora caninum in maned wolves (Chrysocyon brachyurus)

Use of serological tests in the diagnosis of infectious diseases in wild animals has several limitations, primarily the difficulty of obtaining species-specific reagents. Wild canids, such as maned wolves (Chrysocyon brachyurus), are highly predisposed to infection by Toxoplasma gondii and, to a lesser extent, to Neospora caninum. The aim of the present study was to evaluate homologous, heterologous, and affinity conjugates in enzyme-linked immunosorbent assays (ELISAs) and indirect fluorescent antibody tests (IFATs) for detecting immunoglobulin (Ig) G antibodies against T. gondii and N. caninum in maned wolves. Serum samples were obtained from 59 captive animals in Brazil and tested by ELISA for T. gondii serology and IFAT for N. caninum serology using 3 different enzymatic and fluorescent conjugates: homologous (guinea pig anti-maned wolf IgG-peroxidase and -fluorescein isothiocyanate [FITC]), heterologous (rabbit anti-dog IgG-peroxidase and -FITC), and affinity (protein A-peroxidase and -FITC). Seropositivity to T. gondii was comparable among the homologous (69.5%), heterologous (74.6%), and affinity (71.2%) enzymatic conjugates. A significant positive correlation was found between the antibody levels determined by the 3 enzymatic conjugates. The highest mean antibody levels (ELISA index = 4.5) were observed with the protein A-peroxidase conjugate. The same seropositivity to N. caninum (8.5%) was found with the homologous and heterologous fluorescent conjugates, but protein A-FITC was not able to detect or confirm any positive samples with homologous or heterologous conjugates. Our results demonstrate that homologous, heterologous, and affinity conjugates might be used in ELISA for serological assays of T. gondii in wild canids, whereas for N. caninum infection, only the homologous or heterologous fluorescent conjugates have been shown to be useful. © American Society of Parasitologists 2005.

First telemetry study of bush dogs: Home range, activity and habitat selection

Context The bush dog (Speothos venaticus) is difficult to observe, capture, and study. To date, indirect evidence and opportunistic field observations have been the primary sources of information about the species' ecology. Field data are urgently needed to clarify the species' ecological requirements, behaviour and movement patterns. Aims The present study uses 13 months of telemetry data from a group of bush dogs to begin to address questions about area requirements, habitat preferences and movement patterns of this difficult-to-study species. Methods We tracked a group of bush dogs (two adults, one juvenile, four young) in an area of intact and altered Cerrado (woodlandsavanna biome) in eastern Mato Grosso, Brazil (Nova Xavantina District). Key results The group had a total home range of 140km2 (fixed kernel 95%), with smaller seasonal 'subareas' (areas used for 12 months before moving to another area, with repetition of some areas over time) and demonstrated a preference for native habitats. Conclusions The bush dog's home range is greater than that of other canids of the same size, even correcting for group size. Patterns of seasonal movement are also different from what has been observed in other South American canids. Implications From our observations in the Brazilian savanna, bush dogs need large tracks of native habitat for their long-term persistence. Although the present study is based on a single pack, it is highly relevant for bush dog conservation because it provides novel information on the species' spatial requirements and habitat preferences. © 2012 CSIRO.

Improvement in clinical signs and cellular immunity of dogs with visceral leishmaniasis using the immunomodulator P-MAPA

This study investigated the immunotherapeutic potential of the protein aggregate magnesium-ammonium phospholinoleate-palmitoleate anhydride immuno-modulator (P-MAPA) on canine visceral leishmaniasis. Twenty mongrel dogs presenting clinical symptoms compatible with leishmaniasis and diagnosis confirmed by the detection of anti-leishmania antibodies were studied. Ten dogs received 15 doses of the immunomodulator (2.0mg/kg) intramuscularly, and 10 received saline as a placebo. Skin and peripheral blood samples were collected following administration of the immunomodulator. The groups were followed to observe for clinical signals of remission; parasite load in the skin biopsies using real-time PCR, the cytokines IL-2, IL-10 and IFN-γ in the supernatant of peripheral blood mononuclear cells stimulated in vitro with either total promastigote antigen or phytohemagglutinin measured by capture ELISA, and changes in CD4+ and CD8+ T cell subpopulations evaluated by flow cytometry. Comparison between the groups showed that treatment with the immunomodulator promoted improvement in clinical signs and a significant reduction in parasite load in the skin. In peripheral blood mononuclear cell cultures, supernatants showed a decrease in IL-10 levels and an increase in IL-2 and IFN-γ. An increase in CD8+ T cells was observed in peripheral blood. In addition, the in vitro leishmanicidal action of P-MAPA was investigated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and no leishmanicidal activity was detected. These findings suggest that P-MAPA has potential as an immunotherapeutic drug in canine visceral leishmaniasis, since it assists in reestablishing partial immunocompetence of infected dogs. © 2013 Elsevier B.V.

Influência do estágio reprodutivo e suplementação do meio de cultivo com progesterona e/ou soro de cadela em estro, nas taxas de maturação in vitro de oócitos de fêmeas caninas

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Detecção sorológica e caracterização moleculares de agentes anaplasmataceae, micoplasmas hemotróficos, piroplasmas e Hepatozoon sp. em carnívoros selvagens mantidos em cativeiro no Brasil

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Descrição do plexo braquial do cachorro-do-mato (Cerdocyon thous Linnaeus, 1766)

The Cerdocyon thous is a canid that has a wide distribution in South America and, besides some general aspects, its morphology is little known in the literature, especially regarding the nervous system. With the aim of elucidating the anatomical composition of brachial plexus, we studied three male specimens from Paragominas-PA, donated to the Morphological Laboratory of Animal Research (LaPMA), Federal Rural University of Amazonia (UFRA), after death by trampling. The animals were fixed in an aqueous solution of 10% formaldehyde for bilateral dissection of the origin of the brachial plexus. The brachial plexus of C. thous is derived from the last three cervical nerves and the first thoracic nerve (C6-T1). The main nerves that compose it, with their respective origins were the suprascapular nerve, subscapular nerve and musculocutaneous nerve (C6-C7), axillary nerve (C7-C8), radial nerve (C7-T1 and C7-C8), median nerve, ulnar nerve, thoracodorsal and thoracic lateral nerve (C8-T1). We conclude that the brachial plexus of C. thous is similar to that described for the domestic dogs, showing small differences in the composition of some nerves.

Gastrintestinal helminths Of Cerdocyon thous (Linnaeus, 1766-Smith, 1839) from the caatinga area of the Paraiba State, Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Investigating South American biogeographic history using patterns of skull shape variation on Cerdocyon thous (Mammalia: Canidae)

Patterns of geographic variation of the canid Cerdocyon thous have historically been obscured by its remarkable intraspecific morphological variability. The observed distribution is highly associated with phytophysiognomy, a feature considered highly dynamic along geological time. In the present study, we tested whether vegetation distribution during the Holocene Glacial Maximum of South America (HGM) explains the patterns of morphological variation within Cerdocyon thous. The species was divided in groups according to paleohabitats that could support their presence during the HGM, and then tested for differences in skull morphometrics. The results obtained demonstrate that the climatic changes during the HGM influenced the population structure of this species, resulting in the establishment of geographical groups with different degrees of morphological cohesion. Higher morphological cohesion found in the Northern group might be explained by the marked discontinuity between its geographical range and the rest of the species`distribution. The Eastern and Southern morphological divergence is less striking and, although this could be related to past vegetation distribution, the disappearance of those barriers leads to a population structure that could be slowly breaking down. (C) 2009 The Linnean Society of London, Biological Journal of the Linnean Society, 2009, 98, 77-84.

Pathogens of Wild Maned Wolves (Chrysocyon brachyurus) in Brazil

The maned wolf, Chrysocyon brachyurus, is an endangered Neotropical canid that survives at low population densities. Diseases are a potential threat for its conservation but to date have been poorly studied. We performed clinical evaluations and investigated the presence of infectious diseases through serology and coprologic tests on maned wolves from Galheiro Natural Private Reserve, Perdizes City, Minas Gerais State, southeastern Brazil. Fifteen wolves were captured between 2003 and 2008. We found high prevalences of antibody to canine distemper virus (CDV; 13/14), canine parvovirus (CPV; 4/14), canine adenovirus type 2 (13/14), canine coronavirus (5/11), canine parainfluenza virus (5/5), and Toxoplasma gondii (6/8), along with Ancylostomidae eggs in all feces samples. Antibodies against Leishmania sp. were found in one of 10 maned wolves, and all samples were negative for Neospora caniman. Evidence of high exposure to these viral agents was also observed in unvaccinated domestic clogs from neighboring farms. High prevalence of viral agents and parasites such as CDV, CPV, and Ancylostomidae indicates that this population faces considerable risk of outbreaks and chronic debilitating parasites. This is the first report of exposure to canine parainfluenza virus in Neotropical free-ranging wild canids. Our findings highlight that canine pathogens pose a serious hazard to the viability of maned wolves and other wild carnivore populations in the area and emphasize the need for monitoring and protecting wildlife health in remaining fragments of the Cerrado biome.

Ecological genetics and conservation genomics of wolf (Canis lupus)

In the present work, we apply both traditional and Next Generation Sequencing (NGS) tools to investigate some of the most important adaptive traits of wolves (Canis lupus). In the first part, we analyze the variability of three Major Histocompatibility Complex (MHC) class II genes in the Italian wolf population, also studying their possible role in mating choice and their influence on fitness traits. In the second section, as part of a larger canid genome project, we will exploit NGS data to investigate the transcript-level differences between the wolf and the dog genome that can be correlated to domestication.

Vaccines against neosporosis: what can we learn from the past studies?

Neospora caninum is an intracellular apicomplexan parasite, which is a leading cause of abortion in cattle; thus neosporosis represents an important veterinary health problem and is of high economic significance. The parasite can infect cattle via trans-placental transmission from an infected cow to its fetus (vertical transmission), or through the oral route via ingestion of food or water contaminated with oocysts that were previously shed with the feces of a canid definitive host (horizontal transmission). Although vaccination was considered a rational strategy to prevent bovine neosporosis, the only commercialized vaccine (Neoguard®) produced ambiguous results with relatively low efficacy, and was recently removed from the market. Therefore, there is a need to develop an efficient vaccine capable of preventing both, the horizontal transmission through infected food or water to a naïve animal as well as the vertical transmission from infected but clinically asymptomatic dams to the fetus. Different vaccine strategies have been investigated, including the use of live attenuated vaccines, killed parasite lysates, total antigens or antigen fractions from killed parasites, and subunit vaccines. The vast majority of experimental studies were performed in mice, and to a certain extent in gerbils, but there is also a large number of investigations that were conducted in cattle and sheep. However, it is difficult to directly compare these studies due to the high variability of the parameters employed. In this review, we will summarize the recent advances made in vaccine development against N. caninum in cattle and in mice and highlight the most important factors, which are likely to influence the degree of protection mediated by vaccination.

Assessing the taxonomic status of dingoes Canis familiaris dingo for conservation

1. The conservation status of the dingo Canis familiaris dingo is threatened by hybridization with the domestic dog C. familiaris familiaris. A practical method that can estimate the different levels of hybridization in the field is urgently required so that animals below a specific threshold of dingo ancestry (e.g. 1/4 or 1/2 dingoes) can reliably be identified and removed from dingo populations. 2. Skull morphology has been traditionally used to assess dingo purity, but this method does not discriminate between the different levels of dingo ancestry in hybrids. Furthermore, measurements can only be reliably taken from the skulls of dead animals. 3. Methods based on the analysis of variation in DNA are able to discriminate between the different levels of hybridization, but the validity of this method has been questioned because the materials currently used as a reference for dingoes are from captive animals of unproven genetic purity. The use of pre-European materials would improve the accuracy of this method, but suitable material has not been found in sufficient quantity to develop a reliable reference population. Furthermore, current methods based on DNA are impractical for the field-based discrimination of hybrids because samples require laboratory analysis. 4. Coat colour has also been used to estimate the extent of hybridization and is possibly the most practical method to apply in the field. However, this method may not be as powerful as genetic or morphological analyses because some hybrids (e.g. Australian cattle dog x dingo) are similar to dingoes in coat colour and body form. This problem may be alleviated by using additional visual characteristics such as the presence/absence of ticking and white markings.

When wolves cry: long distance calling by wild maned wolves

The maned wolf (Chrysocyon brachyurus Illiger 1815) is the biggest canid in South America and it is considered a “near threatened” species by IUCN. Because of its nocturnal, territorial and solitary habits, there are still many understudied aspects of their behavior in natural environments, including acoustic communication. In its vocal repertoire, the wolf presents a longdistance call named “roar-bark” which, according to literature, functions for spacing maintenance between individuals and/or communication between members of the reproductive pair inside the territory. In this context, this study aimed: 1) to compare four methods for detecting maned wolf’s roar-barks in recordings made in a natural environment, in order to elect the most efficient one for our project; 2) to understand the night emission pattern of these vocalizations, verifying possible weather and moon phases influences in roarbark’s emission rates; and 3) to test Passive Acoustic Monitoring as a tool to identify the presence of maned wolves in a natural environment. The study area was the Serra da Canastra National Park (Minas Gerais, Brazil), where autonomous recorders were used for sound acquisition, recording all night (from 06pm to 06am) during five days in December/2013 and every day from April to July/2014. Roar-barks’ detection methods were tested and compared regarding time needed to analyze files, number of false positives and number of correctly identified calls. The mixed method (XBAT + manual) was the most efficient one, finding 100% of vocalizations in almost half of the time the manual method did, being chosen for our data analysis. By studying roarbarks’ temporal variation we verified that the wolves vocalize more in the early hours of the evening, suggesting an important social function for those calls at the beginning of its period of most intense activity. Average wind speed negatively influenced vocalization rate, which may indicate lower sound reception of recorders or a change in behavioral patterns of wolves in high speed wind conditions. A better understanding of seasonal variation of maned wolves’ vocal activity is required, but our study already shows that it is possible to detect behavioral patterns of wild animals only by sound, validating PAM as a tool in this species’ conservation.

Exploring the social behaviour of domestic dogs (Canis familiaris) in a public off-leash dog park

The growing popularity of dog parks has created an opportunity to learn more about interactions between companion dogs. Dog-dog behaviour in a public off-leash dog park was described and analyzed using a motivationally-neutral approach. I observed focal dogs from park entry for 400 s and constructed activity time budgets (percentages of time spent with dogs, humans, etc.); rates of socially-relevant dog behaviours (e.g., snout-muzzle contact, physical contact) were also calculated. On average, focal dogs spent 50% of their time alone, nearly 40% with other dogs and 11% in other activities; time with dogs decreased and time alone increased over the first six minutes. Some behaviours were very frequent (i.e., more than 90% of focal dogs initiated and received snout-muzzle contact to the anogenital and head areas, while others were rare (i.e., 9% and 12% of focal dogs initiated and received lunge approaches, respectively). Dog density and focal dog age, sex, neuter status, and size were found to influence some behavioural variables. Future studies should continue to investigate the diverse range of canid behaviours and factors that influence social behaviours in dog park settings.

Morfometria dos forames mandibular, mentual e infraorbital de Lobo-guará Chrysocyon brachyurus, Illiger, 1815 aplicada a bloqueios anestésicos

CHAPTER II - The Chrysocyon brachyurus is the biggest South American canid which has a high frequency of dental injuries, both in wild and captivity. Thus, veterinary procedures are necessary to preserve the feeding capacity of hundreds of captive specimens worldwide. The aim of this study was to study the mandibular morphometry of maned wolf with emphasis on the establishment of anatomic references to the anesthetic blockage of the inferior alveolar and mental nerves. Therefore, 15 measurements in 22 hemimandibles of C. brachyurus adults were taken. For extra-oral technique of blockage of the inferior alveolar nerve at the level of the mandibular foramen, it is stated that the needle should be advanced, close to the medial surface of the mandibular ramus, by 11.4 mm perpendiculary from the palpable concavity. Alternatively, the needle can be introduced for 30.4 mm from the angular process at 20-25 degrees angle with the ventral margin. For blocking only the mentual nerve, it is recommended the introduction of needle for 10 mm, close to the lateral aspect of the mandibular body, at the level of the lower first premolar. The mandibular foramen showed similars position, size and symmetry in the maned wolfs specimens examined. The comparison of the data with those available for other carnivores reflects the necessity for determining these anatomical references specifically for each species.