Assessing CometChip technology for DNA damage studies in non-model species: distinct UV-induced responses in turtles and mammals

Abstract

Objective: We evaluated the feasibility of using the high-throughput CometChip to assess DNA damage in non-model species. Specifically, we measured UVA-induced damage in fibroblasts from five turtle and four mammalian species with diverse life histories and cancer rates.

Results: Turtles exhibited significantly higher endogenous DNA damage than mammals but showed lower UVA-induced damage after both 2-min and 5-min exposures. At 5 min, bats exhibited the most DNA damage (21.3%), followed by mice (11.3%). Elephants showed intermediate responses (Asian: 6.49%, African: 3.58%), while all turtles remained below 3%, suggesting resilience to oxidative stress. Despite the assay's ability to detect DNA damage across species, several challenges emerged. Endogenous damage varied widely both within and between species. Differences in culture requirements between turtles and mammals limited experimental standardization. Additionally, characterizing species-specific responses is challenging, as multiple cell lines per species are often unavailable for non-model organisms, making it difficult to account for intraspecific variation. Addressing these limitations will be crucial for conducting robust comparative studies of DNA damage responses in future research.

Keywords: Aging; Cancer; Cell biology; Comet assay; Comparative oncology; Evolution; High throughput assays; Longevity; Oxidative stress.

Fig. 1

Endogenous damage at 0 min of UVA exposure. Percent damage, based on the length of the comet tail, was aggregated from individual cells analyzed across two replicate wells and three separate runs (N > 1,000 cells per species). Each data point represents the percent damage from a single cell. Mean damage values (yellow bars) represent averages across all cells, replicate wells, and runs. Tukey’s post-hoc comparisons identified a significant difference between the little brown bat and the Roti Island snake-necked turtle (p = 0.041)

Fig. 2

DNA damage induced by UVA exposure. The mean values were calculated from the aggregate of individual cells read from two wells at each time exposure in three runs (N per species > 1,000). The quantitative data point was an estimate of percent DNA damage based on the length of comet tail created by damaged DNA. The two UVA exposure times were 2 min (94.8 KJ/s/m²) and 5 min (37 KJ/s/m²). 2- and 5-min values were divided by the mean 0-min value for that species to account for endogenous damage. The ratio of induced damage from 2 to 5 min being represented by 5/2 in the graph. The Welch’s ANOVA p-values for the 2- and 5-min comparisons were significant: 2 min = 8.216e-06, 5 min = 1.044e-05, and 5/2 min = 0.056