Growth curves showing total viable counts of parent MRSA and <i>P. aeruginosa</i> isolates and resistant mutants which developed following exposure to fosfomycin and tobramycin.

<p>(A) MRSA CFP13, aerobic, (B) MRSA CFP13 anaerobic, (C) <i>P. aeruginosa</i> W050 aerobic and (D) <i>P. aeruginosa</i> W050 anaerobic. Filled circle (•) Parent strain; filled square (▪) Fosfomycin resistant mutant; filled triangle (▴) Tobramycin resistant mutant.</p

Fosfomycin, tobramycin and F∶T MICs for MRSA and <i>P. aeruginosa</i> strains used in this study.

<p>Fosfomycin, tobramycin and F∶T MICs for MRSA and <i>P. aeruginosa</i> strains used in this study.</p

Fosfomycin, tobramycin and F∶T MICs for MRSA isolate CFP8 following serial passage in sub-inhibitory antibiotic concentrations under both aerobic and anaerobic conditions.

<p>Exposure to (A) Fosfomycin aerobic, (B) Fosfomycin anaerobic, (C) Tobramycin aerobic, (D) Tobramycin anaerobic, (E) F∶T aerobic, (F) F∶T anaerobic. Filled circle (•) Fosfomycin MIC; filled square (▪) Tobramycin MIC; filled triangle (▴)F∶T MIC.</p

Fosfomycin, tobramycin and F∶T MICs for <i>P. aeruginosa</i> isolate W050 following serial passage in sub-inhibitory antibiotic concentrations under both aerobic and anaerobic conditions.

<p>Exposure to (A) Fosfomycin aerobic, (B) Fosfomycin anaerobic, (C) Tobramycin aerobic, (D) Tobramycin anaerobic, (E) F∶T aerobic, (F) F∶T anaerobic. Filled circle (•) Fosfomycin MIC; filled square (▪) Tobramycin MIC; filled triangle (▴)F∶T MIC.</p

Mean^a (S.D.) frequency of spontaneous MRSA (n = 6) and <i>P. aeruginosa</i> (n = 6) mutants with increased fosfomycin (FOF), tobramycin (TOB) and F∶T MICs under aerobic and anaerobic conditions at 2×, 4× and 8× MIC.

a<p>Where the spontaneous mutation frequency was above or below the limit of detection, the limit of detection was used to calculate the mean e.g. if spontaneous mutation frequency was <4.4×10⁻⁸ then 4.4×10⁻⁸ was used.</p

Fosfomycin, tobramycin and F∶T MICs for <i>P. aeruginosa</i> isolates following serial passage in sub-inhibitory antibiotic concentrations and subsequent removal of antibiotic pressure following 12 passages.

<p>(A) AY4 FOS aerobic, (B) AY4 FOS anaerobic and (C) W050 TOB aerobic. Filled circle (•) Fosfomycin MIC; filled square (▪) Tobramycin MIC; filled triangle (▴)F∶T MIC; broken line, antibiotic pressure removed.</p

There is broad agreement between qualitative bacterial culture results and 454 pyrosequencing for dominant CF pathogens.

<p>The fraction of sequence assigned to the genus (a) <i>Pseudomonas</i> or (b) <i>Burkholderia</i> are plotted for each patient and sample timepoint as a function of the patient' s reported culture status for the recognized CF pathogens <i>P. aeruginosa</i> and <i>B. cepacia</i> complex species. Patient samples are color-coded by timepoint (green, onset of exacerbation; red, end-of-treatment for exacerbation with intravenous antibiotics; blue, clinically stable interval). Patient 19 was culture negative for both <i>P. aeruginosa</i> and <i>B. cepacia</i>.</p

Abundance taxa are highly stable during exacerbation and in response to antibiotic treatment.

<p>To determine whether the relative abundance of specific taxa changed during exacerbation or following antibiotic treatment, the normalized average sequence abundance for all detected OTUs was compared (a) between exacerbation (n = 22) and end-of-treatment timepoints (n = 22) and (b) between the exacerbation (n = 22) and stable timepoints (n = 13). For each taxon, normalized average sequence abundance values are plotted as a logarithm to the base 10 (log₁₀). Red circles indicate taxa that had significantly lower normalized average sequence abundance following antibiotic treatment at a 10% false discovery rate. (c) Comparison of overall microbial richness at all three sampling timepoints indicates a slight, but transient decrease following antibiotic treatment. By pairwise t-tests, comparisons of richness between exacerbation and end-of-treatment (p = 0.06, n = 21), exacerbation and stable (p = 0.87, n = 13) and stable and end-of-treatment (p = 0.076, n = 13) timepoints all fail to reach statistical significance at a p≤0.05 threshold.</p

CF is a polymicrobial disease.

<p>Phylogenetic tree of the 169 OTUs identified in the CF sputum dataset. Tree construction was achieved by mapping consensus sequences from each OTU to the SILVA reference tree (see Methods). Each leaf of the tree represents a consensus OTU labeled with the most closely related genus assigned by the RDP classifier.</p

Antibiotic regimens used to treat acute pulmonary exacerbations in this study¹.

1<p>Table indicates antibiotic regimens used to treat each of the 26 exacerbations that occurred during the period of study. Of the 23 patients enrolled, three patients experienced two separate exacerbations. For these three patients, the first and second exacerbations were treated with different antibiotic combinations.</p