Performances of PCR tests in Sets A and B.

<p>LbX/1-6, Laboratory and test identification; Bold Type, Good Performing Methods in sets A or B; GPM, Good Performing Methods in sets A and B; Sp, 100% of specificity in all controls without <i>T. cruzi</i> DNA; Co, Coherence in PCR positive reports; DL, Detection limit in fg DNA/ul; Y, Affirmative; N, Negative; NA, Not available; ND, Not detectable<b>.</b></p

Examples of the outputs of the four best performing PCR methods.

<p>A. LbD2 ; B.LbD3, C. LbF1 and D. LbQ. The methods are described in <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0000931#s2" target="_blank">Materials and Methods</a> and <a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0000931#pntd-0000931-t001" target="_blank">Table 1</a>. 6, 15: seropositive samples; 35; seronegative sample (<a href="http://www.plosntds.org/article/info:doi/10.1371/journal.pntd.0000931#pntd-0000931-t003" target="_blank">Table 3</a>). PC: Positive control: 10 fg/µl of T.cruzi VI. NC. Negative Control: Master Mixes devoid of DNA.</p

Intra-laboratory evaluation of best performing methods in human samples.

<p>ID, sample identification number; LbX/1-6, Laboratory and test identification; % pos, Percentage of Positivity; Cons, Consensus PCR Result; pos, positive; neg, negative.</p

Concordance of PCR results reported for each clinical case of Set C.

<p>Patients 1 to 32 are seropositive and 33 to 42 seronegative. 28 kDNA tests and 13 Sat DNA tests were performed for each sample.</p><p>kDNA, minicircle DNA; Sat-DNA, satellite DNA; GPM Good performing Methods in panels A and B; ID, sample identification number; G, Gender; Ag, age in years; EN, Endemic precedence; %: Percentage of positive results; Cons, Consensus PCR result; F, female; M, male; NA, not available; 28 kDNA tests and 13 Sat DNA tests were performed for each sample.</p>1<p><i>T.cruzi</i> DTU I,</p>2<p><i>T.cruzi</i> DTU II/V/VI, NE, not endemic; Uk, Unknown; Pos, positive consensus; Ind, indeterminate consensus; Neg, negative consensus; cChHD, chronic Chagas heart disease, Mega Megacolon, CBBB, Complete Branch Bundle Blockage, HTx, Heart transplantation; Arg: Argentina; Bo: Bolivia; Br: Brazil; Par: Paraguay; BAs, Buenos Aires; Bh, Bahia; Go, Goias; MG: Minas Gerais; Sg: Santiago del Estero.</p

Intra-Laboratory Evaluation of the four Best Performing Methods in samples from Table 5.

<p>LbX/1−6, Laboratory and test identification; Se, sensitivity; Sp, specificity; Acc, accuracy; kappa, kappa index.</p

Performance of PCR tests in comparison to consensus GPM reports and serodiagnosis.

<p>LbX/1-6, Laboratory and test identification; BPM, Best Performing Methods; Consensus GPM K + S: consensus findings of GPM by kDNA and Satellite DNA PCRs; C, Conventional PCR, RT, Real Time PCR; K, kDNA; S, Satellite DNA; Se, sensitivity; Sp, specificity; Acc, accuracy; kappa, kappa index; N, negative; Y, affirmative; 25–75p, 25th-75th percentiles; Bold type, Good Performing Methods (GPM) in sets A and B.</p

Analytical Sensitivity of specific and coherent PCR tests in sets A and B.

<p>Distribution of detection limits (DL) of specific and coherent PCR tests targeted to Sat-DNA (A) and kDNA sequences (B) for detecting serial dilutions of purified DNA from 3 parasite stocks (Set A) representative of <i>T. cruzi</i> DTU I (Silvio×10), DTU IV (Can III cl1) and DTU VI (Cl Brener). C. Distribution of detection limits (DL) of specific and coherent PCR tests targeted to Sat-DNA (black bars) and kDNA sequences (white bars) carried out from human blood spiked with serial dilutions of parasite cells (Set B).</p