Efeitos da temperatura e tempo de estocagem de fezes de mazama jucunda no isolamento viabilidade e cultivo celular

Abstract

This study investigated the viability, isolation, and cultivation of exfoliated cells from the feces of Neotropical deer, proposing a non-invasive method for cellular analysis and future applications in animal conservation. Fecal samples were collected and subjected to different storage conditions, being kept at room temperature (27°C) and refrigerated (4°C) for periods of 0h, 1h, 4h, 6h, 12h, and 24h. The objective was to assess the impact of these conditions on the number and viability of exfoliated cells, considering time as a determining factor for cell integrity. After cell isolation, viability tests were performed using trypan blue and fluorescent dyes such as propidium iodide and acridine orange to distinguish viable, apoptotic, and necrotic cells. Statistical analysis was conducted using a Completely Randomized Design (CRD), evaluating the effects of time and storage condition factors on the dependent variable. Cells isolated using the Percoll gradient method were cultured in a supplemented DMEM medium. The cultivation was carried out in 96-well plates and maintained at 37°C and 5% CO₂, with media changes every two days. The results revealed that time had a significant effect on cell viability, while the storage condition factor had a smaller effect. The highest means of viable cells were observed at the initial times of 0h and 1h, with a progressive reduction over 24 hours, especially at room temperature. In contrast, samples stored under refrigerated conditions showed a milder decrease in viability. Regarding cell cultures, a high contamination rate was observed, resulting in the loss of five out of six cultures. However, in one culture, cells remained viable for up to 120 hours, demonstrating the feasibility of the method for future molecular and cell culture analyses. Thus, the proposed method could be a viable alternative for cellular studies in wild animals, avoiding invasive procedures and enabling population health monitoring. It is concluded that the isolation and cultivation of exfoliated cells from deer feces are possible and effective, but protocol improvements are necessary to increase the success rate of cell isolation and, consequently, cultures.