Abstract
A new methodology has been developed to assess cytochrome P4501A expression in two South Atlantic Spanish fish, guilthead seabream ( Sparus aurata) and grey mullet ( Liza aurata), used as pollution bioindicators. Degenerate oligos were used to amplify by reverse transcription and PCR (RT-PCR) specific cyp1A cDNA sequences, used subsequently to design specific primers to get the full cDNA by rapid amplification of cDNA ends. A new assay has been developed to quantitate cyp1A expression by RT-PCR in an automated DNA sequencer. The effect of β-naphthoflavone inducing biotransformation has been used to compare three distinct pollution biomarkers: EROD activity, ELISA determination of CYP1A, and 2-aminoanthracene (2-AA) activation. Immunodetection by ELISA or Western blot was inconsistent in S. aurata and L. aurata. EROD activity yielded satisfactory results; the higher induction was observed by bioactivation of 2-AA to mutagens detected with strain BA149 of Salmonella typhimurium, in agreement with the high sensitivity previously described for this biomarker. The present paper summarizes the current status of our research.
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