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Trypanosoma cruzi is currently classified into 2 major phylogenetic lineages, T. cruzi I and II, that correlate with the formerly described zymodeme 1 and 2, respectively. Another isoenzymic group (zymodeme 3-Z3) was also described. In this study, we analysed the genetic diversity among Z3 isolates of the Brazilian Amazon by restriction fragment length polymorphism of the intergenic transcribed spacers (ITSs) of the ribosomal RNA cistron and the size of the divergent domain D7 of the 24Salpha rRNA gene. DNAs from 12 T. cruzi Z3 isolates obtained from humans (2), Panstrongylus geniculatus (1), and Rhodnius brethesi (9) were submitted to PCR amplification of the ITSs plus the 5.8S rDNA. The PCR products were digested with 4 distinct endonucleases and the profiles analysed by a numerical methodology. The phenetic dendrogram revealed a clear dichotomy in the Z3 group, defining 2 groups that were named Z3-A and Z3-B. Dimorphism was also found in the band sizes of the amplified D7 divergent domain of the 24Salpha rDNA, which showed a perfect correlation with the ITSs clustering. The organization of the ribosomal cistron was investigated by Southern blotting and shown to be conserved in the genome of the 2 Z3 groups. This study shows that the rDNA cistron allows the definition of 2 distinct subclusters in Z3 isolates.

Introduction: Elimination of malaria in areas of interrupted transmission warrants careful case assessment to avoid the reintroduction of this disease. Occasional malaria cases are reported among visitors of the Atlantic Forest area of Brazil, while data on residents of this area are scarce. Methods: A sectional study was carried out to examine 324 individuals living in a municipality where autochthonous cases were detected. Results: Asymptomatic Plasmodium infections were detected in 2.8% of the individuals by polymerase chain reaction (PCR), with one case of P. falciparum (0.3%), two cases of P. vivax (0.6%), and six cases of P. malariae (1.9%). The thick blood smears were negative in all individuals. Serological tests performed in 314 subjects were reactive in 11.1%, with 3.5% for P. falciparum and 7.7% for P. vivax. A subsample of 42 reactive individuals for any Plasmodium species showed P. malariae in 30.9% of specimens. Individuals who entered the Atlantic Forest region were 2.7 times more likely to exhibit reactive serology for P. vivax compared with individuals who did not enter this region (p<0.05). Children <15 years had a higher chance of reactive serology for P. falciparum and P. vivax than individuals ≥15 years of age (p<0.05). Individuals living in the Paraiso district had a higher chance of reactive serology for P. vivax compared to other districts (p<0.05). No associations were found between sex, past exposure to malaria, or serological response to antibodies of any Plasmodium species. Conclusions: The implications of these results for the elimination of malaria were discussed.

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