Growth rate assays reveal fitness consequences of β-lactamases

Abstract

Antibiotic resistance is a powerful model for studying evolutionary biology and population genetics. For the purpose of these evolutionary studies, fitness data have been approximated through susceptibility testing methods designed for clinical use in providing appropriate antibiotic therapies. An alternative approach for measuring fitness of microbes has experienced growing popularity: growth rates are a highly sensitive approach for measuring the fitness effects of antibiotics and resistance genes, but they differ from susceptibility testing in that a single concentration of antibiotic is used for the assay. Here we show that despite this key difference, the results of growth rates correlate well with clinical determination of resistance by minimum inhibitory concentration (MIC), while providing the sensitivity required for direct input as fitness values into mathematical models. This means that growth rates at a single sublethal inhibitory concentration can help us understand the fitness effects that result in clinical antibiotic resistance. By measuring the growth rates of sequenced clinical isolates obtained from Dignity Health Mercy Medical Center, we detected the fitness effects of individual resistance genes on bacteria exposed to different antibiotics. In our study, the CTX-M-15 gene conferred the highest fitness in assays with cephalosporins. These results show the strong fitness benefit conferred by CTX-M-15.

Case Studies in the Assessment of Microbial Fitness: Seemingly Subtle Changes Can Have Major Effects on Phenotypic Outcomes

“…In this approach, growth parameters, such as the maximum growth rate (V max ), carrying capacity (K), and area under the curve (AUC), are calculated from absorbance values and used as proxies for fitness (Kugelberg et al 2005;Hansen et al 2007;Hegreness et al 2008;Paulander et al 2009;McDonald et al 2012; Handling Editor: Kerry Geiler-Samerotte. Ketola and Saarinen 2015;Kang et al 2019;Santiago et al 2020;Frey et al 2021). Populations with increased V max , K, or AUC values are considered to be more fit relative to those with lower values.…”

“…The first method uses data obtained from growth curves as an indirect measure of fitness and is one of the most commonly used methods outside of the experimental evolution field (Warringer et al 2003 ; Kugelberg et al 2005 ; Paulander et al 2009 ; Łapińska et al 2022 ). In this approach, growth parameters, such as the maximum growth rate (V max ), carrying capacity (K), and area under the curve (AUC), are calculated from absorbance values and used as proxies for fitness (Kugelberg et al 2005 ; Hansen et al 2007 ; Hegreness et al 2008 ; Paulander et al 2009 ; McDonald et al 2012 ; Ketola and Saarinen 2015 ; Kang et al 2019 ; Santiago et al 2020 ; Frey et al 2021 ). Populations with increased V max , K, or AUC values are considered to be more fit relative to those with lower values.…”

Distribution of β-Lactamase Genes in Clinical Isolates from California Central Valley Hospital Deviates from the United States Nationwide Trends

“…This result indicates that either the genetic background of K. pneumoniae eliminates the incompatibilities of these genes, or that the antibiotic exposures of these pathogens is different from E. coli. Additionally, at DHMMC, there appears to be strong selection for bla CTX-M , which may be displacing bla TEM likely due to antagonism between these genes [ 22 ].…”

Symmetry-related residues as promising hotspots for the evolution of de novo oligomeric enzymes

“…2c), indicating that symmetry-related residues exert greater degrees of perturbation to the protein than proximal residues. The distribution of tness effects [37][38][39] was also depicted by box charts (Fig. 2c inset), where the mutations at symmetry-related residues exhibit much more outspread cell-growth rates than those of proximal residues.…”