Generation times data in Blastocystis by qPCR and by microscopy

Generation time (Tg) data of Blastocystis cultures in Barret’s and in Pavlova’s media during 48 h., from IBS patients and from asymptomatic carriers, measured by microscopy and by quantitative PCR are shown in present table. The generation times of Blastocystis isolates, were calculated according to Zhang et al. [17], with the following equation: Tg=(T2-T1)/(log2(n2/n1)), where Tg denotes the generation time, n1 represents the number of cultured parasitic organisms at the initial time (T1), and n2 represents the number of parasitic cells at subsequent time (T2). Thus, (T2-T1) = 48 hours of in vitro culture. Besides, for to absolute quantification by qPCR, it was necessary to consider i) the size of the Blastocystis genome ~ 18.8Mbp[36]; ii) 1pg of DNA ~ 978Mbp[37] and iii) the concentration of DNA control was 160ng/µL; thus, by cross multiplications, the number of copies of the genetic marker amplified were estimated.