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Approximately 10% of the Trypanosoma cruzi genome is formed by a satellite DNA, composed by 195-bp repeats organized in 30+/-10 kb clusters in some, but not all chromosomes. Here, the satellite DNA of six representative T. cruzi strains was sequenced and used for phylogenetic inference. The results show that CL Brener contains satellite repeats from T. cruzi I and T. cruzi II strains, although type II sequences are more abundant. The presence of types I and II sequences extends previous propositions that genetic exchange between the two major T. cruzi lineages have occurred in CL Brener, although our data accommodate alternative scenarios of hybridization within T. cruzi II, as proposed by others. Altogether, present data suggest a complex origin for CL Brener. Sequence analysis of satellites isolated from chromosomal bands indicates that satellite DNA sequences are not chromosome specific. Neighbor analysis of in tandem satellite DNAs containing up to five repeats shows that each cluster contains only one type of sequence. Consequently, clusters with intercalated types I and II repeats were not found. We propose that the CL Brener genome contains large pieces of satellite DNA originated mainly from chromosomes of T. cruzi II with introgression of T. cruzi I lineage.

Pulsed-field gel electrophoresis and DNA hybridization were used to establish and compare some parameters of the molecular karyotype of nine stocks classified into Trypanosoma cruzi I and T. cruzi II groups. The isolates showed a variable number of chromosomal bands (17-22) comprised between 0.4 and 3.3 Mbp. The total number of chromosomes and the genome size were estimated based on the fluorescence intensity of SYBR Green I-stained chromosomal bands. Differences in the length of the telomeric regions among the stocks and between chromosomes of the same stock were observed. No correlation was found between the length of the telomeric region and the group to which the isolate belongs. Hybridization of 54 genetic markers revealed extensive chromosome size polymorphism. Nevertheless, the most represented pattern was the hybridization of the probes in larger chromosomes in stocks of T. cruzi II as compared to T. cruzi I. Eight putative syntenic groups, encompassing 29 non-redundant genetic markers and distributed in 11 CL Brener chromosomal bands were disclosed. The syntenic groups were conserved in all the stocks. The relative abundance of repetitive DNA sequences was determined. C6, B11/L1Tc and E12 elements presented maximum 1.7-fold variation in copy number, whereas 195-bp satellite DNA (120,000 copies in Y strain) was four- to nine-fold more abundant in T. cruzi II stocks. The novel aspects of T. cruzi karyotype here presented contribute to the comprehension of the genome organization of this parasite and will assist the assignment of scaffold to the CL Brener chromosomal bands.

Trypanosoma cruzi has about 10(5) copies of a 195 bp repeat, called satellite DNA, which correspond to about 10% of the total DNA. Since very little is known about satellite distribution and function in this and in many other organisms, we studied the genomic organization of satellite DNA in T. cruzi Y, CL Brener, and Silvio X10 cl1 strains. We found that satellite repeats are present in some, but not all, chromosomes in these strains. We also found that the copy number of the satellite repeats is distinct in each one of the strains. We have estimated that the satellite DNA is four to six times more abundant in Y and CL Brener strains as compared to Silvio X10 cl1. Despite these differences, the repeated units are organized in a similar way. Based on restriction nuclease digestion analysis, we found that satellite repeats are grouped in clusters comprising about 30+/-10 kb. The same cluster size was also found in the different chromosomes, as seen by two-dimensional pulsed field-gel electrophoresis. As evidenced by a fluorescence hybridization assay of spread DNA fibers, we additionally found that these clusters are constrained in specific regions of the chromosomes. Sequence analysis of several repeat units of satellite DNA of the three strains revealed a remarkable similarity with conserved nucleotide blocks separated by more variable regions. Such conserved distribution and common sequence may suggest that these repeated units might have a structural role in the T. cruzi chromosomes.