Dataset Information

Image-Based Dynamic Phenotyping Reveals Genetic Determinants of Filamentation-Mediated ?-Lactam Tolerance.

ABSTRACT: Antibiotic tolerance characterized by slow killing of bacteria in response to a drug can lead to treatment failure and promote the emergence of resistance. ?-lactam antibiotics inhibit cell wall growth in bacteria and many of them cause filamentation followed by cell lysis. Hence delayed cell lysis can lead to ?-lactam tolerance. Systematic discovery of genetic factors that affect ?-lactam killing kinetics has not been performed before due to challenges in high-throughput, dynamic analysis of viability of filamented cells during bactericidal action. We implemented a high-throughput time-resolved microscopy approach in a gene deletion library of Escherichia coli to monitor the response of mutants to the ?-lactam cephalexin. Changes in frequency of lysed and intact cells due to the antibiotic action uncovered several strains with atypical lysis kinetics. Filamentation confers tolerance because antibiotic removal before lysis leads to recovery through numerous concurrent divisions of filamented cells. Filamentation-mediated tolerance was not associated with resistance, and therefore this phenotype is not discernible through most antibiotic susceptibility methods. We find that deletion of Tol-Pal proteins TolQ, TolR, or Pal but not TolA, TolB, or CpoB leads to rapid killing by ?-lactams. We also show that the timing of cell wall degradation determines the lysis and killing kinetics after ?-lactam treatment. Altogether, this study uncovers numerous genetic determinants of hitherto unappreciated filamentation-mediated ?-lactam tolerance and support the growing call for considering antibiotic tolerance in clinical evaluation of pathogens. More generally, the microscopy screening methodology described here can easily be adapted to study lysis in large numbers of strains.

SUBMITTER: Zahir T

PROVIDER: S-EPMC7082316 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Image-Based Dynamic Phenotyping Reveals Genetic Determinants of Filamentation-Mediated β-Lactam Tolerance.

Zahir Taiyeb T Wilmaerts Dorien D Franke Sabine S Weytjens Bram B Camacho Rafael R Marchal Kathleen K Hofkens Johan J Fauvart Maarten M Michiels Jan J

Frontiers in microbiology 20200313

Antibiotic tolerance characterized by slow killing of bacteria in response to a drug can lead to treatment failure and promote the emergence of resistance. β-lactam antibiotics inhibit cell wall growth in bacteria and many of them cause filamentation followed by cell lysis. Hence delayed cell lysis can lead to β-lactam tolerance. Systematic discovery of genetic factors that affect β-lactam killing kinetics has not been performed before due to challenges in high-throughput, dynamic analysis of vi ...[more]

PMID: 32231648

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Project description:BACKGROUND:Developing chilling tolerant accessions of domesticated Asian rice is a potential source of significant crop improvement. The uniquely chilling sensitive nature of the tropically originating Oryza sativa make it the most important cereal crop that can gain significantly from improved tolerance to low temperatures. However, mechanisms underlying this complex trait are not fully understood. Oryza sativa has two subspecies with different levels of chilling tolerance, JAPONICA and INDICA, providing an ideal tool to investigate mechanistic differences in the chilling stress tolerance responses within this important crop species. RESULTS:The Rice Diversity Panel 1 (RDP1) was used to investigate a core set of Oryza sativa accessions. The tools available for this panel allowed for a comprehensive analysis of two chilling tolerance traits at multiple temperatures across a 354-cultivar subset of the RDP1. Chilling tolerance trait values were distributed as mostly subpopulation specific clusters of Tolerant, Intermediate, and Sensitive accessions. Genome-wide association study (GWAS) mapping approaches using all 354 accessions yielded a total of 245 quantitative trait loci (QTL), containing 178 unique QTL covering 25% of the rice genome, while 40 QTL were identified by multiple traits. QTL mappings using subsets of rice accession clusters yielded another 255 QTL, for a total of 500 QTL. The genes within these multiple trait QTL were analyzed for Gene Ontology (GO) term and potential pathway enrichments. Terms related to "carbohydrate biosynthesis", "carbohydrate transmembrane transport", "small molecule protein modification", and "plasma membrane" were enriched from this list. Filtering was done to identify more likely candidate pathways involved in conferring chilling tolerance, resulting in enrichment of terms related to "Golgi apparatus", "stress response", "transmembrane transport", and "signal transduction". CONCLUSIONS:Taken together, these GO term clusters revealed a likely involvement of Golgi-mediated subcellular and extracellular vesicle and intracellular carbohydrate transport as a general cold stress tolerance response mechanism to achieve cell and metabolic homeostasis under chilling stress.

Dataset Information

Image-Based Dynamic Phenotyping Reveals Genetic Determinants of Filamentation-Mediated ?-Lactam Tolerance.

Publications

Image-Based Dynamic Phenotyping Reveals Genetic Determinants of Filamentation-Mediated β-Lactam Tolerance.

Similar Datasets

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