Sensitivity of trypanosomatid protozoa to DFMO and metabolic turnover of ornithine decarboxylase.

alpha-Difluoromethylornithine (DFMO), the specific and irreversible inhibitor of ornithine decarboxylase (ODC), was able to induce the arrest of proliferation in Leishmania mexicana and ODC-transformed Trypanosoma cruzi cultures grown in a semi-defined medium essentially free of polyamines. Conversely, Crithidia fasciculata and Phytomonas 274 were not affected by the inhibitor. The drug-resistance of Crithidia and Phytomonas was neither caused by an impairment of DFMO uptake nor by a decrease of the enzyme affinity for the inhibitor. We were also able to rule out the possibility of ODC overexpression in the drug-tolerant parasites. The measurements of ODC metabolic turnover indicated that the enzymes from Crithidia and Phytomonas have a short half-life of 20-40 min, while ODC from Leishmania and transgenic Trypanosoma cruzi are rather stable with a half-life longer than 6 hours. Analyses of polyamine internal pools under different growth conditions have shown that DFMO was able to markedly decrease the levels of putrescine and spermidine in all parasites, but the depletion of spermidine was higher in trypanosomatids containing an ODC with slow turnover. Our results suggest that in these parasites cultivated in the presence of the drug, spermidine might decrease below critical levels needed to maintain trypanothione concentrations or other conditions essential for normal proliferation.

Trypanosoma cruzi epimastigotes lack ornithine decarboxylase but can express a foreign gene encoding this enzyme.

Trypanosoma cruzi, a pathogenic protozoan causing Chagas disease, lacks ornithine decarboxylase (ODC), the enzyme catalyzing the first step of polyamine biosynthetic pathway in eukaryotic cells. Our results indicate that the auxotrophy for diamines of T. cruzi epimastigotes is due to the absence of an active ODC gene in these parasites and not to the inability for the expression of this gene. The introduction of an exogenous complete coding region from Crithidia fasciculata ODC gene inserted in an expression vector specific for trypanosomatids induces the normal expression of the foreign genetic information allowing the transformed T. cruzi to overcome the exogenous polyamine requirement for growth. The enzyme expressed in the transformed parasites has shown a considerably extended metabolic stability. The loss of ODC activity in T. cruzi might be related to the parasite adaptation to the intracellular stages of its life cycle.

Paternal directional mating in two Amerindian subpopulations located at different altitudes in northwestern Argentina.

We used mitochondrial DNA (mtDNA) and Y-chromosome DNA polymorphisms to analyze the ethnic origin of maternal and paternal lineages in two Amerindian subpopulations from northwestern Argentina. One of the subpopulations was from San Salvador de Jujuy, located 1200 m above sea level. The second subpopulation inhabits the Quebrada de Humahuaca area at altitudes ranging from 2500 to 3500 m. Both subpopulations have the same ethnic background. All mtDNA haplotypes were identified as Amerindian with a frequency of 64.6% of the B form (9-bp deletion in mtDNA region V). Because all Central Andean Amerindian populations studied so far exhibit high frequencies of the B haplotype, we propose that they probably are derived from a common ancestral population that inhabited the Central Andes 6000-8000 years B.P. The presence of paternal directional mating (asymmetric contribution of one parental lineage to interethnic gene mixtures) was demonstrated by the finding of an average introgression of 40.5% Spanish Y chromosomes into our Amerindian sample. This introgression was more evident at low altitude than at high altitude, with frequencies of 64.3% in San Salvador de Jujuy (low altitude) and 27.6% in Quebrada de Humahuaca (high altitude) (p < 0.05). The San Salvador de Jujuy subpopulation also showed a significantly higher Y-chromosome gene variability than the Quebrada de Humahuaca subpopulation. These findings are in good agreement with historical reports indicating that the colonization of South America was undertaken by men who usually practiced polygamous unions with Amerindian women and that San Salvador de Jujuy was the main northwestern Argentinian region of European to Amerindian gene admixture. We found 16.7% of cases with Spanish Y chromosomes and Amerindian family names, and the same percentage with Amerindian Y chromosomes and Hispanic names. The former group probably is the result of unions between Hispanic men, who transmitted the Y chromosome, and Amerindian women, who transmitted the family name to the progeny. The latter group likely illustrates the practice of changing names from Amerindian to Hispanic during the baptism of native Americans in colonial times.