A toxicogenomic approach for development of biomarkers of pesticide exposure

CESCHIN D.G.; MARDIROSIAN M.; PIRES N; LASCANO C; VENTURINO A

Natal

Congreso; 9th Congress of Toxicology in Developing Countries and XIX Congress of Brazilian Society of Toxicology; 2015

Soc. Bras. Toxicol. - IUTOX

Introduction: NorthernPatagonia is an irrigated area holding 95% of exportable production of appleand pear. Around 900 ton/year of pesticides have been applied since year 2000where 90% correspond to organophosphates. 50% of the pesticides applied aredispersed into the atmosphere without reaching the intended targets and endingmainly in irrigation canals, ponds, streams and rivers. Biomonitoring ofenvironmental impact of pesticides using native species is a preferred resourcefor its ecological significance. Rhinellaarenarum is a toad widelydistributed in Argentina and less in Brazil, Bolivia and Uruguay.Organophosphates produce lethal and sublethal organism effects includingpresence of abnormalities, impaired growth and biochemical effects. In thissense, enzymes and metabolites of detoxification system are widely used as biomarkers.  However, many of them are not specific and inseveral cases not responsive enough. Therefore, it is necessary to developearly, specific and sensitive ecotoxicological biomarkers. Here, we present ahigh throughput approach (RNA-Seq) to screen  new potential biomarkers in R. arenarumlarvaeexposed to chlorpyrifos (CLP) pesticide.Objective: todevelop new biomarkers for chlorpyrifos exposition.Materials andMethods:R. arenarum embryos were obtained by in vitrofertilization. Larvae (complete operculum (CO) + 11 days) were exposed tosublethal concentration of CLP (0.1 mg/L) [LC50 2.5 mg/L)]. Samples were takenat 6h and 24h to evaluate reduced glutathione (GSH) levels and the activitiesof Glutathione S transferase (GST) and Catalase (CAT). At same time, sampleswere collected for RNA purification and massive sequencing. Results anddiscussion: GSH levels and GST and CAT activities were notsignificantly affected by CLP exposition neither 6h nor 24h compared withcontrol. However, the RNA-Seq profiles showed differences between 6h vs 24h andcompared to control. There was a first wave of gene expression at 6h whereprotein of several of them could drive transcription of genes detected at 24h.As expected, detoxification and oxidative stress pathways were activated.Beside, clustering of genes using Gene Ontology classification showed hits forBiological Process category: development, metabolic and cellular process, amongothers.Conclusion: biomarkers are a biological response to toxicexposition which can be used for management and environmental protection. Here,we show that classic biomarkers were no modified when toad larvae were exposedto sublethal concentration of CLP. However, using RNA-Seq several geneactivation even at early time (6h) were detected. Thus, new molecularbiomarkers could be defined and a combination of them could bring a moresensitive and specific tool as biomarker.