Molecular & Biochemical Parasitology 138 (2004) 183–194 DNA microarrays for comparative genomics and analysis of gene expression in Trypanosoma cruzi  Cassio Silva Baptista a , Ricardo Z.N. Vˆ encio a,b , Sarah Abdala a , Maria Paula Valadares a , Camila Martins a , Carlos Alberto de Braganc ¸a Pereira b , Bianca Zingales a,∗ a Departamento de Bioqu´ ımica, Instituto de Qu´ ımica, Universidade de S˜ ao Paulo, Avenida Professor Lineu Prestes 748, CEP 05508-000 S ˜ ao Paulo, SP, Brazil b Departamento de Estat´ ıstica, Instituto de Matem ´ atica e Estat´ ıstica, Universidade de S˜ ao Paulo, S˜ ao Paulo, SP, Brazil Received 23 September 2003; received in revised form 25 May 2004; accepted 9 June 2004 Available online 21 September 2004 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 ≤ 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 ≤ 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. © 2004 Elsevier B.V. All rights reserved. Keywords: Trypanosoma cruzi; Stocks; DNA microarrays; Comparative genomics; Gene expression; Transcript levels Abbreviations: EST, expressed sequence tag; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; rRNA, ribosomal RNA; SIRE, short inter- spersed repetitive element; TEUF, TENF, TEUQ and TENQ, EST from a non-normalised (TEUF, TEUQ) and normalised (TENF, TENQ) cDNA li- brary of CL Brener epimastigotes sequenced at FIOCRUZ (TEUF, TENF) and Instituto de Qu´ ımica (TEUQ, TENQ)  Note: Nucleotide sequence of 134 ESTs reported in this paper have been submitted to the GenBank TM database with the accession numbers CF134346–CF134365 and CF243279–CF243392. ∗ Corresponding author. Tel.: +55 11 3091 3810x217; fax: +55 11 3815 5579. E-mail address: zingales@iq.usp.br (B. Zingales). 1. Introduction The protozoon Trypanosoma cruzi is the etiological agent of Chagas disease, which affects 16–18 million people in South and Central America. T. cruzi life cycle alternates be- tween vertebrates and triatomine insects, with different de- velopmental stages in each host: epimastigotes and meta- cyclic trypomastigotes in the insect vector and intracellular amastigotes and bloodstream trypomastigotes in the mam- malian host. T. cruzi is diploid, its mode of replication is predominantly asexual and, therefore, parasite strains repre- sent independent clonal lineages [1]. The strains, also named as stocks or isolates, were shown to be divergent for various 0166-6851/$ – see front matter © 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.molbiopara.2004.06.017