DNA microarrays for comparative genomics and analysis of gene expression in Trypanosoma cruzi

Abstract

Trypanosoma cruzi presents high genetic diversity and parasite isolates show remarkable differences in biological parameters. In this study, we evaluated whether DNA microarrays containing CL Brener cDNAs can be used for comparative genomics and for the analysis of gene expression in T. cruzi. We constructed a prototype microarray with 710 expression sequence tags of CL Brener and 20 sequences of T. cruzi strains. These probes represent 665 unique genes. Results from four hybridisations with genomic DNA of Silvio (T. cruzi I) and CL Brener (hybrid genotype) identified 9.3% of the probes (68/730) differentially represented in the two genomes. Data from eight hybridisations with cDNA obtained from three independent parasite harvests of Silvio and CL Brener disclosed 84 sequences of 730 (11.5%) that showed statistical significant (P < or = 0.01) changes in expression (1.6-6.5-fold). Some of the array-identified sequences were confirmed by Southern and Northern blot analysis. Only 20% of the probes with increased expression in Silvio or CL Brener presented higher hybridisation with genomic DNA of either strain. Approximately 2.5% (18/730) and 9.0% (65/730) of the probes were differentially expressed (P < or = 0.01), respectively, in epimastigotes and metacyclic trypomastigotes of two T. cruzi II strains isolated from chronic chagasic patients. Microarrays identified several sequences for which differences in gene copy number and/or in the levels of RNA transcripts were previously demonstrated by different approaches. The data indicate that DNA microarrays are a useful tool for comparative studies between strains and provide further evidence for a high level of post-transcriptional regulation of RNA abundance in T. cruzi.