Tissue distribution of a plasmid DNA encoding Hsp65 gene is dependent on the dose administered through intramuscular delivery

In order to assess a new strategy of DNA vaccine for a more complete understanding of its action in immune response, it is important to determine the in vivo biodistribution fate and antigen expression. In previous studies, our group focused on the prophylactic and therapeutic use of a plasmid DNA encoding the Mycobacterium leprae 65-kDa heat shock protein (Hsp65) and achieved an efficient immune response induction as well as protection against virulent M. tuberculosis challenge. In the present study, we examined in vivo tissue distribution of naked DNA-Hsp65 vaccine, the Hsp65 message, genome integration and methylation status of plasmid DNA. The DNA-Hsp65 was detectable in several tissue types, indicating that DNA-Hsp65 disseminates widely throughout the body. The biodistribution was dose-dependent. In contrast, RT-PCR detected the Hsp65 message for at least 15 days in muscle or liver tissue from immunized mice. We also analyzed the methylation status and integration of the injected plasmid DNA into the host cellular genome. The bacterial methylation pattern persisted for at least 6 months, indicating that the plasmid DNA-Hsp65 does not replicate in mammalian tissue, and Southern blot analysis showed that plasmid DNA was not integrated. These results have important implications for the use of DNA-Hsp65 vaccine in a clinical setting and open new perspectives for DNA vaccines and new considerations about the inoculation site and delivery system.     

Factors influencing the habitat use by ocelots in one of the last large Atlantic Forest remnants in southeastern Brazil

Ocelots (Leopardus pardalis) are widely distributed throughout the Americas, being dependent on forested areas to survive. Although ocelot ecology is broadly studied throughout the species range distribution, studies concerning factors that may affect ocelot occupancy in the Atlantic Forest are still scarce. We used camera traps to evaluate factors influencing the probabilities of detection and occupancy of ocelots in a protected area of the Atlantic Forest, the Rio Doce State Park (RDSP), southeastern Brazil. To assess ocelot occupancy and detection probabilities, we measured the distances between sampling stations and rivers, lakes, cities, pasture, and Eucalyptus plantations. In addition, we recorded the mean rainfall levels for each sampling occasion, and native grassland areas within a 500 m‐buffer around each sampling station. We found a strong and positive association between ocelot detection and the dry season, which might be due to a higher number of individuals moving through the Park during this season. Moreover, we found a strong and positive association of ocelot detection with native grassland areas around lakes, which may be related to the ocelot behavior of searching for prey in these areas. Conversely, the ocelot occupancy probability was intermediate (Ψ^ = 0.53, 95% CI = 0.36–0.69) and was not strongly associated with the evaluated covariates, which may be explained by the high‐quality of forest habitats and water resources that are homogeneously distributed within the Park. Our study indicates that the RDSP still provides a structurally suitable forest habitat for ocelots, but because of the current worrying scenario of over fragmentation, reduction of forest cover, and weakness of the protective legislation of this biome, the long‐term persistence of the species in RDSP is uncertain.     

Viability of Ascaris lumbricoides eggs eliminated after anti-helminthic therapy.

The viability of Ascaris lumbricoides eggs passed in the feces was evaluated after treatment of patients with one of the anti-helminthic drugs (thiabendazole, levamisole, cambendazole, pyrantel pamoate, mebendazole or praziquantel). For each drug, a group of 5 children was selected and their feces collected 24 h before treatment and 24, 48 and 72 h after drug administration, except for mebendazole, with the feces being collected throughout the period of treatment. After sedimentation, the total amount of eggs from each collection was transferred to tissue culture flasks containing 10 ml H2SO4 0,1N, with the addition of 3 drops of a miconazole solution, and incubated at 28 degrees C, individually, for 80 days. The flasks were maintained open and the culture were oxygenated daily by manual agitation. On the 80th day of culture, 20-days-old albino mice were inoculated with 3,200 embryonated eggs, per os. Larvae were recovered from their lungs and hearts, on the 8th day after infection, according to Baerman's method (Morais, 1948). Thiabendazole showed 100.0% ovicidal capacity as early as 48 h after treatment. Inhibition of embryonal development was observed when thiabendazole was used. This drug also had an effect on the eggs infectivity when inoculated into normal mice. No significant effect on embryonal development was observed for the other drugs tested.