(A and B) PopMLST results from 3 of 7 patients with CF in whom PopMLST detected 2 Sa strains (see Supplemental Figure 4 for the 4 of 7 patients in whom 1 strain was detected). (A) PopMLST results from DNA pooled from 90–95 cultured Sa isolates. Blue and red bars indicate different MLST loci alleles for each patient (Supplemental Table 3). Bars for each sample show relative abundance of arc, aro, glp, gmk, pta, tpi, and yqi. A third pta allele (black bar and indicated with *) differed at a single nucleotide from the allele indicated in red, likely representing sequencing error or mutation. (B) Relative abundance of arc locus as measured by PopMLST (Pop) and by individually Sanger sequencing (Sanger) 20–30 isolates from each sample from A. (C) PopMLST performed directly on DNA isolated from 3 sputum samples, A–C, and from mixtures of these samples. Red, blue, and yellow bars indicate the abundance of MLST alleles corresponding to samples A–C, respectively; green bars indicate an allele shared between samples A and C; and orange bars indicate an allele shared between samples B and C (see Supplemental Table 1). Control experiments examining more than 100 Pa isolates cultured from sputum samples A–C showed each contained a single Pa MLST type. (D) PopMLST of DNA pooled from more than 100 Sa cultured colonies (culture) and directly from sputum (sputum). Red and blue indicate different MLST types, which were confirmed by Sanger sequencing individual isolates (Supplemental Table 1). Subject 9’s sputum contained 3 loci with an additional allele, likely indicating a third MLST type (green) that was not detected in culture. Indicated significant differences were determined by the multiple t test of minor allele abundance. Black bar indicated with * indicates the presence of a third allele, likely due to mutation or sequencing error.