In vitro phenotypic characterization of DEL, NonDEL and HTZ strains.

A) Normalized 3’NT activity for NonDEL, HTZ and DEL strains. The 3’NT activity was measured for DEL (n = 08), HTZ (n = 03) and NonDEL (n = 11) parasites harvested at stationary phase. The original values, expressed as nmol Pi x h^-1 x 10 ^-7 cells, were normalized by the values obtained with the L. amazonensis assays. Compared to the L. amazonensis control, four NonDEL samples presented higher activity (a); three NonDEL and two HTZ strains presented lower activity (b); all DEL and one HTZ samples presented the lowest enzyme activity (c). Unpaired t-test. B) 3’NT activity average expressed as nmol Pi x h^-1 x 10 ^-7 for each genotype group composed by the strains individually presented in graph A. Kruskal-Wallis test followed by Mann-Whitney U test. C) 3’NT activity expressed as nmol Pi x h^-1 x 10 ^-7 cells in cultures from: stationary phase promastigotes = Pro; metacyclic enriched fraction of stationary phase culture = PNA−; SC = stressed culture parasites “axenic amastigotes”. D) Percentage of metacyclic at stationary phase. Dots represent the average value of two independent experiments for each strain (DEL = 07 and NonDEl = 09). Unpaired t-Test. E-F) Percentage of metacyclic at stationary phase of less-than-20 passages (<20P) culture and culture-adapted parasites - more than 30 passages (>30P). Percentage of metacyclic was determined 72 hours (late stationary phase) after an initial inoculum of 10⁶ parasites/ml. Metacyclic enrichment was obtained by Peanut agglutinin (PNA); percentage of cells from the PNA− fraction was determined in relation to the total cell count. Red = DEL; Blue = NonDEL; Grey = HTZ. Unpaired t test. E-F) Paired t-test for DEL and NonDEL groups of parasites in <20P and >30P conditions. ns = not-significant. Moderate to strong negative correlation (Pearson r = 0.64)was detected for 3’NT activity (A) and metacyclic enrichment (D) (n =14 strains; S1 Data).