Chromatin supraorganization, DNA fragmentation, and cell death in erythrocytes of the rattlesnake, Crotalus durissus terrificus (Serpentes, Viperidae), infected with the protozoan, Hepatozoon spp. (Apicomplexa, Hepatozoidae)

Abstract

Forms of the protozoan of the Hepatozoon genus are detected free in the circulation and also within some of the erythrocytes of infected snakes. In healthy snakes, DNA fragmentation and cell death usually affect a few circulating erythrocytes in agreement with the long life span expected for these cells. In the present study we investigated whether infection by Hepatozoon spp. affected the incidence of DNA fragmentation and cell death in erythrocytes from the rattlesnake, Crotalus durissus terrificus. Methods such as the kinetics of Feulgen-DNA hydrolysis, and the TUNEL and comet assays, previously used for the study of chromatin organization and DNA fragmentation in erythrocytes of healthy snakes, were used. The results indicated that Hepatozoon spp. increased the DNA fragmentation and chromatin condensation typical of cell death in circulating erythrocytes of C. d. terrificus, including cells that do not harbour the parasite. The Hepatozoon infection is thus suggested to accelerate destruction of erythrocytes in the rattlesnake, not only affecting cells harbouring the parasite, but also in those without it.

The comet assay in animal models: From bugs to whales – (Part 2 Vertebrates)

“…c c e p t e d M a n u s c r i p t protozoan (Hepatozoon spp.) a higher level of damage in the erythrocyte DNA was found compared to the erythrocyte DNA from non-infected snakes[277].…”

Feulgen‐DNA response and chromatin condensation in Malpighian tubules of Melipona rufiventris and Melipona quadrifasciata (Hymenoptera, Apoidea)

“…The finding of Feulgen-DNA values after the acid hydrolysis for 50 and 90 min generally much closer in M. quadrifasciata than in M. rufiventris indicated that DNA depurination occurs faster in M. quadrifasciata (Sandritter et al, 1965;Mello, 1979Mello, , 1983Miyamoto et al, 2005;Miyamoto and Mello, 2007). The faster depurination response in M. quadrifasciata may be promoted by its less packed chromatin states in comparison to M. rufiventris.…”

“…Acid hydrolysis was done in 4 M HCl at 25 C for 50 or 90 min followed by treatment with Schiff reagent for 40 min. Mild acid hydrolysis conditions as those used here are considered to be more suitable for the discrimination of differences in Feulgen hydrolysis kinetics provided by differential DNA depurination (ascending branch of the hydrolysis curve) or apurinic acid solubilization (descending branch of the hydrolysis curve) (Sandritter et al, 1965;Mello, 1979Mello, , 1997Mello and Vidal, 1980;Miyamoto et al, 2005;Miyamoto and Mello, 2007). The hydrolysis times were chosen after a preliminary test which revealed that at the time of 90 min a maximal depurination occurred for the DNA of both species.…”

“…The Feulgen reaction is a classic cytochemical method useful to distinguish the kinetics of DNA depurination in DNAe protein complexes differing in composition and organization in the cell nuclei as assessed microspectrophotometrically or by video image analysis (Sandritter et al, 1965: Agrell andBergqvist, 1967;Mello, 1979Mello, , 1983Mello, , 1997Mello and Vidal, 1980;Miyamoto et al, 2005;Miyamoto and Mello, 2007). When analysing the somatic interphase nuclei of M. quadrifasciata and M. rufiventris stained with the C-banding technique, it was hypothesized that the Feulgen reaction would be a better method to distinguish the patterns of composition and/or organization of the DNAeprotein complexes in interphase cells of these species presumed to differ in terms of heterochromatin content.…”