The Pantanal is a tropical, seasonal wetland of approximately 140,000 km 2 that is fed by tributaries of the upper Paraguay River in the centre of South America, which covers mainly Brazilian but also Paraguayan and Bolivian territories and is classified as one of the largest freshwater wetland ecosystems in the world (Alho 2005). The Brazilian Pantanal, which represents 85% of the total Pantanal area, is located within the states of Mato Grosso do Sul and Mato Grosso in Central-West Brazil, which are recognised as South and North Pantanal, respectively (Alho et al. 1988). This region is ecologically classified into sub-regions according to vegetation, flooding and physiography (Silva & Abdon 1988). The Nhecolândia sub-region that is situated in South Pantanal is one of the largest, comprising approximately one fifth of the total area, and is characterised by hundreds of shallow lakes that display varying degrees of salinity and coalescence with the system during floods (Adámoli 1982).Recent studies have demonstrated that some crocodilian species may be infected by WNV (Steinman et al. 2003, Jacobson et al. 2005 and that Alligator mississippiensis may also have a viremic load that affords the infection of blood-feeding Culicidae vectors, which suggests that in areas with high population densities of these alligators, juvenile individuals could play an important role in WNV transmission (Klenk et al. 2004). Taking these data into account, the unsuccessful efforts to detect WNV circulation in the avian hosts in Brazil and the widespread, high prevalence of the Crocodilia species, Caiman crocodilus yacare, in the Brazilian Pantanal (Campos et al. 2005), our strategy was to investigate the WNV circulation in the potential vectors, dead-end hosts and natural secondary amplifying hosts through the collection and testing of mosquitoes and serum samples of horses and wild caimans from the Nhecolândia sub-region. No morbidity was observed during the sampling period and only apparently healthy horses and caimans were sampled in the present study. MATERiALS AND METHoDSStudy area -In February 2009, mosquitoes and blood samples from horses and wild caimans were collected in six different cattle ranches of the Nhecolândia sub-region (18º20' 19º40'S and 57º14' 55º00'W) during the rainy season. Considering the free circulation of mosquitoes, horses and wild caimans in large ranches, sample collections for the study were undertaken in a 700-square-kilometre (70,000 ha) area (Figure). The collections for this study were authorised by the Brazilian Institute of Environment and Natural Resources (license IBAMA 18363-1/2009).Samples collections -Adult mosquitoes were captured at sites that were randomly selected using CDC automatic light traps and manual aspirators while landing to blood-feed on horses and research team members as routinely reported. Living mosquito specimens were transported to a field laboratory where they were immobilised by chilling and the species were identified through direct observation of the morphological ...

Serum samples collected from 547 equids in the Pantanal region of Brazil were evaluated for antibodies to Equine Infectious Anemia Virus (EIAV) by the agar gel immunodiffusion test. Risk factors associated with EIAV seropositivity were evaluated and spatial dependence investigated using a Spatial Lag Model. EIAV prevalence on farms in the Pantanal was 52.0% (13/25) with adjusted prevalence between equids of 31.5% (17.4-48.8% 95% CI). Intra-herd prevalence ranged from 5.0 to 77.0%. Statistical analysis demonstrated that farms and animals in regularly flooded areas had respectively 60 and 146 fold higher chance to be sero-positive than farms and animals located in non-flooded areas. Spatial Lag Model results were generally consistent with this conclusion although there was a negative spatial correlation between farms located within in regularly inundated regions, suggesting that other factors, such as management practices, probably play a significant role in transmission of EIAV. Equids with clinical signs were 3.74-fold more likely to be sero-positive than those without clinical signs. The results of this work reveal a high prevalence of EIAV in the Pantanal area of Brazil demonstrating that equids reared in this region are at great risk of infection.

As in humans, sub-clinical infection by arboviruses in domestic animals is common; however, its detection only

An epidemic of pneumonia with fibrinous polyserositis and multifocal arthritis emerged in captive American alligators (Alligator mississippiensis) in Florida, United States, in 1995. Mycoplasma alligatoris sp. nov. was cultured from multiple organs, peripheral blood, synovial fluid, and cerebrospinal fluid of affected alligators. In a subsequent experimental inoculation study, the Henle-Koch-Evans postulates were fulfilled for M. alligatoris as the etiological agent of fatal mycoplasmosis of alligators. That finding was remarkable because mycoplasmal disease is rarely fatal in animals. An enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies produced by alligators in response to M. alligatoris exposure was developed by using plasma obtained from naturally infected alligators during the original epidemic. The assay was validated by using plasma obtained during an experimental dose-response study and applied to analyze plasma obtained from captive and wild crocodilian species. The ELISA reliably detected alligator seroconversion (P < 0.05) beginning 6 weeks after inoculation. The ELISA also detected seroconversion (P < 0.05) in the relatively closely related broad-nosed caiman Caiman latirostris and the relatively distantly related Siamese crocodile Crocodylus siamensis following experimental inoculation with M. alligatoris. The ELISA may be used to monitor exposure to the lethal pathogen M. alligatoris among captive, repatriated, and wild crocodilian species.An epidemic of mycoplasmosis emerged in a collection of American alligators (Alligator mississippiensis) in Florida in 1995 (4, 8). Thirty-three 200-to 300-kg adult male alligators died, and 13 moribund alligators were euthanatized within 1 month of the index case. Lesions observed at necropsy ranged from mild interstitial and peribronchiolar pneumonia to fibronecrotic pneumonia. Extrapulmonary complications included pericarditis, myocarditis, and multifocal arthritis. Mycoplasma alligatoris proposed sp. nov. was isolated from multiple tissues, blood, synovial fluid, and cerebrospinal fluid of affected alligators. In a pilot experimental inoculation study (5, 6), healthy alligators were inoculated with M. alligatoris strain A21JP2 T (ATCC 700619) to reproduce the disease and fulfill the HenleKoch-Evans postulates (11) for M. alligatoris as the etiological agent of synovitis, polyserositis, and pneumonia of alligators. The results were remarkable because, except for bovine and caprine pleuropneumonia, mycoplasmal disease is rarely fatal in animals.The origin of the 1995 epidemic remains unknown, but the disease may emerge under conditions of captivity. Crocodilians are frequently sold or traded among collections, exhibits, zoos, and ranches. Many collections include multiple species of crocodilians. Some crocodilians from commercial ranches are repatriated after "head-starting" hatchlings from eggs collected in the wild, a potential vector for catastrophic infection of wild populations if lethal disease emerges in captivity. A validated ser...

Mycoplasma alligatoris was the suspected etiology of an epidemic of acute multisystemic inflammatory disease which emerged in captive American alligators (Alligator mississippiensis) in Florida (USA) in 1995. In an experimental inoculation study conducted from April through October 1999, 18 alligators were inoculated with 10 2 , 10 4 , or 10 6 colony forming units (CFU) of M. alligatoris by instillation into the glottis. As early as 1 wk post-inoculation (PI), mycoplasma were cultured from blood of three of six alligators inoculated with 10 6 CFU. Two of those died and the third was euthanatized within 4 wk PI. Necropsy gross findings included fibrinous polyserositis and polyarthritis. Histopathologic changes in affected individuals included pulmonary edema, interstitial pneumonia, pericarditis, myocarditis, meningitis, and synovitis. Mycoplasma were cultured quantitatively in high numbers from trachea, lung, coelomic cavity, liver, spleen, interior of pericardial sac, heart, blood, brain, and limb joints. In alligators inoculated with 10 6 CFU, heterophilia and moderate hyperglycemia peaked about 4 wk PI, and seroconversion occurred by 6 to 8 wk PI. Necropsy gross and histologic findings were generally unremarkable for the surviving alligators inoculated with 10 6 CFU, alligators inoculated with 10 2 or 10 4 CFU, and four uninoculated control alligators. Mycoplasma were not cultured at any time point from those alligators. The findings confirm that M. alligatoris can cause fulminant inflammatory disease and rapid death of alligators.

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Equine infectious anemia (EIA) has a worldwide distribution, and is widespread in Brazil. The Brazilian Pantanal presents with high prevalence comprising equine performance and indirectly the livestock industry, since the horses are used for cattle management. Although EIA is routinely diagnosed by the agar gel immunodiffusion test (AGID), this serological assay has some limitations, so PCR-based detection methods have the potential to overcome these limitations and act as complementary tests to those currently used. Considering the limited number of equine infectious anemia virus (EIAV) sequences which are available in public databases and the great genome variability, studies of EIAV detection and characterization molecular remain important. In this study we detected EIAV proviral DNA from 23 peripheral blood mononuclear cell (PBMCs) samples of naturally infected horses from Brazilian Pantanal using a semi-nested-PCR (sn-PCR). The serological profile of the animals was also evaluated by AGID and ELISA for gp90 and p26. Furthermore, the EIAV PCR amplified DNA was sequenced and phylogenetically analyzed. Here we describe the first EIAV sequences of the 5' LTR of the tat gene in naturally infected horses from Brazil, which presented with 91% similarity to EIAV reference sequences. The Brazilian EIAV sequences also presented variable nucleotide similarities among themselves, ranging from 93,5% to 100%. Phylogenetic analysis showed that Brazilian EIAV sequences grouped in a separate clade relative to other reference sequences. Thus this molecular detection and characterization may provide information about EIAV circulation in Brazilian territories and improve phylogenetic inferences.

The working equid population in Corumbá, Southern Pantanal, is very large and has a crucial role in the main economic activity of the State of Mato Grosso do Sul, the beef cattle industry. The aim of the present study was to estimate the prevalence of equine infectious anaemia (EIA) in working equids of ranches in the municipality of Corumbá, by the official agar gel immunodiffusion (AGID) test, and evaluate the adoption of the Programme for the Prevention and Control of Equine Infectious Anaemia proposed by Embrapa Pantanal and official entities in the 1990s. From September to November 2009, forty ranches distributed through the area of the municipality were visited, and serum samples were obtained from 721 equines and 232 mules. According to previous publications and the present data, it was concluded that the prevalence of EIA in this population has increased from 18.17% to 38.60%, which represents at this time approximately 13,000 infected animals. There was no significant difference between the apparent prevalence of equines and mules. It was also verified that the control programme was not known by the greater part of the interviewed ranch owners, managers and foremen and, in their perception, EIA is not a primary threat to address. Among the studied variables, the serologic testing practice significantly reduced the risk for the presence of EIA seropositivity, as well as the separation of riding equipment and segregation of seropositives.