Trypanosoma cruzi genome project: biological characteristics and molecular typing of clone CL Brener

Clone CL Brener is the reference organism used in the Trypanosoma cruzi Genome Project. CL Brener was obtained by cloning procedures from bloodstream trypomastigotes isolated from mice infected with the CL strain. The doubling time of CL Brener epimastigotes cultured at 28°C in liver infusion-tryptose (LIT) medium is 58±13 h. Differentiation to metacyclic forms is induced by incubation of epimastigotes in LIT-20% Graceʹs medium. Metacyclics give very low parasitemia in mice, contrary to what is observed for blood forms which promote 100% mortality of the animals with inocula of 5×103 parasites. CL Brener blood forms are highly susceptible to nifurtimox, benznidazole and ketoconazole. Allopurinol is inefficient in the treatment of mice experimental infection. The clone infects mammalian cultured cells and performs the complete intracellular cycle at 33 and 37°C. The molecular typing of CL Brener has been done by isoenzymatic profiles; sequencing of a 24Sα ribosomal RNA gene domain and by schizodeme, randomly amplified polymorphic DNA and DNA fingerprinting analyses. For each typing approach the patterns obtained do not change after prolonged parasite subcultivation in LIT medium (up to 100 generations). The stability of the molecular karyotype of the clone was also confirmed.