Experimental data for Pseudomonas aeruginosa from experimental evolution under different bottleneck sizes and antibiotic selection pressures

We here combined evolution experiments with genomic and genetic analyses
to assess whether bottleneck size and antibiotic-induced selection
influences the evolutionary path to resistance in pathogenic Pseudomonas
aeruginosa, one of the most problematic opportunistic human pathogens. Two
sets of evolution experiments were performed across 16 transfers, in which
either the aminoglycoside gentamicin or the fluoroquinolone ciprofloxacin
were used as antibiotic. The evolutionary response was studied using
counts of bacterial cells at the end of each transfer period (i.e., yield)
or by calculating the growth rate from regular optical density measurement
during the respective transfer periods. For the populations at the end of
the evolution experiments, we also determined their antibiotic resistance
with the help of standardized dose response curves. Moreover, we performed
whole genome sequencing to assess the frequency of variants, which emerged
and spread during the evolution experiments. We further focused on
variants in two specific genes, which were selectively favoured in the
gentamicin evolution experiments, and assessed their relative fitness
using competition experiments. We found that antibiotic resistance is
favoured under high antibiotic selection and weak bottlenecks, but also
under low antibiotic selection and severe bottlenecks. We found that the
absence of high resistance under low selection and weak bottlenecks is
caused by the spread of low-resistance variants with high competitive
fitness under these conditions. We conclude that bottlenecks in
combination with drug-induced selection are currently neglected key
determinants of pathogen evolution and antibiotic treatment outcome.