The alkaline comet assay, or single cell gel electrophoresis, is one of the most popular methods for assessing DNA damage in human population. One of the open issues concerning this assay is the identification of those factors that can explain the large inter-individual and inter-laboratory variation. International collaborative initiatives such as the hCOMET project - a COST Action launched in 2016 - represent a valuable tool to meet this challenge. The aims of hCOMET were to establish reference values for the level of DNA damage in humans, to investigate the effect of host factors, lifestyle and exposure to genotoxic agents, and to compare different sources of assay variability. A database of 19,320 subjects was generated, pooling data from 105 studies run by 44 laboratories in 26 countries between 1999 and 2019. A mixed random effect log-linear model, in parallel with a classic meta-analysis, was applied to take into account the extensive heterogeneity of data, due to descriptor, specimen and protocol variability. As a result of this analysis interquartile intervals of DNA strand breaks (which includes alkali-labile sites) were reported for tail intensity, tail length, and tail moment (comet assay descriptors). A small variation by age was reported in some datasets, suggesting higher DNA damage in oldest age-classes, while no effect could be shown for sex or smoking habit, although the lack of data on heavy smokers has still to be considered. Finally, highly significant differences in DNA damage were found for most exposures investigated in specific studies. In conclusion, these data, which confirm that DNA damage measured by the comet assay is an excellent biomarker of exposure in several conditions, may contribute to improving the quality of study design and to the standardization of results of the comet assay in human populations.

Reports on the state of the environment in Kosovo have emphasized that river and ground water quality is affected by pollution from untreated urban water as well as the waste water from the industry. One of the main contributors to this pollution is located in Obiliq (coal power plants). Prishtina-the capital city of Kosovo-is heavily influenced too. Furthermore, the pollutants combined together with those from heavy traffic are dissolved in Prishtina runoff water, which is discharged into the creek entering the river Sitnica together with urban waste water. The available data show the complex pollution with excessive quantities of nitrites, suspended materials, organic compounds, detergents, heavy metals, polychlorinated biphenyls, etc. In this study, the cytotoxic and genotoxic potential of water samples taken at these sites was tested in primary rat hepatocytes. The results obtained indicate that water samples collected in Prishtina and Obiliq had a significant cytotoxic potential in primary rat hepatocyte cultures even when diluted to 1 %. The increased cytotoxicity, however, was not accompanied by an increased genotoxicity as measured by the percentage of micronucleated cells. Further investigations addressing the chemical composition of the samples and the identification of the toxicants responsible for the cytotoxic effects found will be carried out in a next step.

We have recently shown that inhabitants of two polluted areas in Kosovo display more DNA damage (strand breaks in blood cell DNA) than do residents of a cleaner area. Here, we present additional analyses of these data and discuss additional data sets from Kosovo. Based on our data as well as the available data from other authors, age and sex-related differences in DNA damage or in susceptibility to DNA-damaging agents in the environment should be carefully considered when designing biomonitoring studies and when carrying out statistical analysis of the data.

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