Fig 5 -

MTT assay of cloned cell lines from A10/BT (A) and A21/AT1 (B) isolates. Numbers on the x axis (-1, 0, 1, 2, 3 and 4) indicates drug concentrations used to treat cell lines at doses of 0, 0.8, 7, 125, 1000 and 1500 ng/mL, respectively.</p

MTT assay of <i>T</i>. <i>annulata</i> infected cell lines.

A.T. annulata isolates with low IC50 values (2–3 ng/mL). B. T. annulata isolates with medium IC50 values (3–7 ng/mL). C. T. annulata isolates with IC50 values over (7 ng/mL). Numbers on the x axis (-1, 0, 1, 2, 3 and 4) indicates drug concentrations used to treat cell lines at doses of 0, 0.8, 7, 125, 1000 and 1500 ng/mL, respectively.</p

Agarose gel electrophoresis of AS-PCR.

Gels showing detection of PCR amplicons of isolates with mutations V135A (A) and P253S (B), respectively. Products amplified using drug sensitive and resistance specific forward primers are given at the top and bottom of each gel, respectively. M, 100 bp molecular size marker (Thermo Scientific Corp.); lanes 1–16, products from template DNA of T. annulata samples; lanes S sensitive control, lanes R resistance control, lane X mixed control. (TIF)</p

Alignments of putative Q_o1 and Q_o2 domains of <i>Cyto b</i>.

Predicted amino acid sequences of Qo1 and Qo2 domains encoded by the Cyto b gene from the published T. annulata / C9 genome sequence and different T. annulata isolates with treatment failure history. Putative Qo1 and Qo2 domains are located between 116–144 and 238–273 amino acids of the protein around the ubiquinone binding site. Predicted amino acid substitutions associated with buparvaquone resistance are marked with ◊.</p

Fig 4 -

Alignments of TaPIN1 nucleotide (A) and predicted amino acid (B) sequences. Sequences derived from a buparvaquone resistant Tunisian isolate and various Turkish T. annulata isolates with treatment failure history were compared for mutations with the T. annulata / C9 reference genome, obtained from PiroplasmDB.org database; the systematic identifier for TaPIN1 is TA18945. Nonsynonymous substitutions detected among alignments were indicated with an asterisk (*) coloured in yellow.</p

IC₅₀ values of A10/BT and A21/AT1 clones with substitutions V135A and P253S.

IC50 values of A10/BT and A21/AT1 clones with substitutions V135A and P253S.</p

Number of alleles detected by nine micro- and mini-satellite markers before and after buparvaquone treatments.

Number of alleles detected by nine micro- and mini-satellite markers before and after buparvaquone treatments.</p

Mutations detected in the <i>Cyto b</i> gene of <i>Tancytb</i> reference gene in PiroplasmDB database and different <i>T</i>. <i>annulata</i> isolates.

Mutations detected in the Cyto b gene of Tancytb reference gene in PiroplasmDB database and different T. annulata isolates.</p

Frequency of V135A and P253S mutations in independently collected <i>T</i>. <i>annulata</i> positive blood samples and isolates.

Frequency of V135A and P253S mutations in independently collected T. annulata positive blood samples and isolates.</p

AS-PCR results of clonal cell lines.

(PDF)</p