Animal Cytogenomics – Jiří Rubeš

• Comparative cytogenomics in mammals
• Oncogenetics and biodosimetry in oncology
• Effects of the air pollution on sperm quality

• To obtain new information on evolutionary karyotype changes in Ruminantia and Carnivora using comparative FISH and optical mapping
• To reveal genetic changes causative of canine cutaneous mast cell tumours using cytogenetic analysis, and whole exome and targeted sequencing
• To evaluate the FISH method in biodosimetry of professionally exposed medical staff and in cancer patients; comparative analysis of the frequency of stable chromosomal aberrations in differentially treated cancer patients, and possibilities of the secondary malignity risk assessment
• To analyse the effects of air pollution on sperm quality and on the epigenetic modification of sperm DNA

Individual species differ in chromosome size and morphology. Species evolution is often accompanied by characteristic karyotype changes. The group of cytogenomics is focused on comparative genomics in representatives of the orders Artodactyla and Carnivora. Evolutionarily conserved chromosome regions and their structural rearrangements, and the location and sequence of the repetitive DNA regions are analysed within the individual taxonomic groups. We use the comparative FISH methods with whole chromosome painting and region specific probes, including satellite DNA, which are prepared in our laboratory using laser microdissection and flow cytometry. The optical mapping using an Irys Instrument will be employed for the comparative analysis of related species. The aim of this research is to obtain new information on the evolutionary genome changes at the level of chromosomes and sequences in the related species and to analyse the data in the context of the taxon phylogeny.

Mast cell tumours are counted among the most frequent skin cancers in dogs. However, the genetic cause of the mast cell tumour formation has not yet been elucidated, so the prognosis assessment and effective treatment are still challenging. Our group performs a detailed cytogenetic and molecular genetic analysis of canine cutaneous mast cell tumours using the FISH method with whole chromosome painting and BAC probes, and whole genome sequencing (WES). Subsequently, the targeted sequencing and mutation analysis of selected genes will be performed. The aim of this study is to reveal the genetic cause of the canine mast cell tumour formation and propose reliable prognostic markers and new potential treatment targets.

Cytostatics and ionizing radiation have been found to have carcinogenic, mutagenic and teratogenic activity and so the medical staff handling these substances, as well as the treated patients, are potentially at increased risk of cancer or fertility problems. Chromosome aberrations in lymphocytes are used as biomarkers of the exposure to genotoxic agents. However, the commonly used conventional cytogenetic testing reflects mainly the short-term exposure to genotoxic agents and is greatly influenced by confounding factors, such as viral infection, life style etc. The test inaccuracy may lead to unnecessary withdrawal of employees from working process or may underestimate the risk of malignancy. In our laboratory we use the FISH method for the detection of stable chromosome aberrations in lymphocytes, which is suitable for the cumulative (long term) exposure assessments. The aim of the study is to evaluate the FISH method for the use in biodosimetry of employees at risk and to assess whether or not it has the potential to identify cancer patients at increased risk of second malignancies.

In developed industrial countries, up to 30% of couples are affected by infertility. The polluted environment, mainly air pollution, is a known factor affecting reproductive functions in both men and women. Our research is focused on the analysis of the effects of air pollution on sperm quality in men living in the Ostrava (CZ) industrial region. The basic functional and morphological parameters of their ejaculatse and genetic and epigenetic changes of sperm DNA, mainly chromatin integrity and condensation defects and changes in the sperm DNA methylation are evaluated in relation to the level of air pollution in the region. Light and fluorescence microscopy, CASA (Computer Assisted Semen Analysis) system for sperm motility analysis, flow cytometry (MoFlo XDP Cell Sorter) for the sperm chromatin integrity analysis and sperm DNA sequencing after bisulfite conversion for methylation analysis are used. The aim of the study is to reveal if the air pollution in the Ostrava region is associated with a decline in the overall sperm quality and with the epigenetic changes in sperm DNA in men living in this area.