title: Trace incorporation of heavy water reveals slow and heterogeneous pathogen growth rates in cystic fibrosis sputum.
abstract: Effective treatment for chronic infections is undermined by a significant gap in understanding of the physiological state of pathogens at the site of infection. Chronic pulmonary infections are responsible for the morbidity and mortality of millions of immunocompromised individuals worldwide, yet drugs that are successful in laboratory culture are far less effective against pathogen populations persisting in vivo. Laboratory models, upon which preclinical development of new drugs is based, can only replicate host conditions when we understand the metabolic state of the pathogens and the degree of heterogeneity within the population. In this study, we measured the anabolic activity of the pathogen Staphylococcus aureus directly in the sputum of pediatric patients with cystic fibrosis (CF), by combining the high sensitivity of isotope ratio mass spectrometry with a heavy water labeling approach to capture the full range of in situ growth rates. Our results reveal S. aureus generation times with a median of 2.1 d, with extensive growth rate heterogeneity at the single-cell level. These growth rates are far below the detection limit of previous estimates of CF pathogen growth rates, and the rates are slowest in acutely sick patients undergoing pulmonary exacerbations; nevertheless, they are accessible to experimental replication within laboratory models. Treatment regimens that include specific antibiotics (vancomycin, piperacillin/tazobactam, tobramycin) further appear to correlate with slow growth of S. aureus on average, but follow-up longitudinal studies must be performed to determine whether this effect holds for individual patients.
authors.affiliation: Division of Geological and Planetary Sciences, California Institute of Technology, Pasadena, CA 91125; Howard Hughes Medical Institute, California Institute of Technology, Pasadena, CA 91125; dkn@caltech.edu als@gps.caltech.edu sebastian.kopf@colorado.edu.
authors.forename: Sebastian H
authors.initials: SH
authors.lastname: Kopf
authors.affiliation: Division of Geological and Planetary Sciences, California Institute of Technology, Pasadena, CA 91125; dkn@caltech.edu als@gps.caltech.edu sebastian.kopf@colorado.edu.
authors.forename: Alex L
authors.initials: AL
authors.lastname: Sessions
authors.affiliation: Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125;
authors.forename: Elise S
authors.initials: ES
authors.lastname: Cowley
authors.affiliation: Pediatric Pulmonology, Children's Hospital Los Angeles, Los Angeles, CA 90027.
authors.forename: Carmen
authors.initials: C
authors.lastname: Reyes
authors.affiliation: Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125;
authors.forename: Lindsey
authors.initials: L
authors.lastname: Van Sambeek
authors.affiliation: Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125;
authors.forename: Yang
authors.initials: Y
authors.lastname: Hu
authors.affiliation: Division of Geological and Planetary Sciences, California Institute of Technology, Pasadena, CA 91125;
authors.forename: Victoria J
authors.initials: VJ
authors.lastname: Orphan
authors.affiliation: Pediatric Pulmonology, Children's Hospital Los Angeles, Los Angeles, CA 90027.
authors.forename: Roberta
authors.initials: R
authors.lastname: Kato
authors.affiliation: Division of Geological and Planetary Sciences, California Institute of Technology, Pasadena, CA 91125; Howard Hughes Medical Institute, California Institute of Technology, Pasadena, CA 91125; Division of Biology and Biological Engineering, California Institute of Technology, Pasadena, CA 91125; dkn@caltech.edu als@gps.caltech.edu sebastian.kopf@colorado.edu.
authors.forename: Dianne K
authors.initials: DK
authors.lastname: Newman
mesh_terms: D000293:Adolescent; D000900:Anti-Bacterial Agents / Q000494:pharmacology / Q000627:therapeutic use; D002648:Child; D003550:Cystic Fibrosis / Q000188:drug therapy / Q000382:microbiology*; D017666:Deuterium Oxide / Q000378:metabolism*; D005227:Fatty Acids / Q000378:metabolism; D005260:Female; D054884:Host-Pathogen Interactions / Q000187:drug effects; D006801:Humans; D007553:Isotope Labeling; D008297:Male; D036103:Nanotechnology; D018629:Spectrometry, Mass, Secondary Ion; D013183:Sputum / Q000187:drug effects / Q000382:microbiology*; D013203:Staphylococcal Infections / Q000188:drug therapy / Q000382:microbiology; D013211:Staphylococcus aureus / Q000187:drug effects / Q000254:growth & development*; D035501:Uncertainty; D055815:Young Adult
chemical_list: D000900:Anti-Bacterial Agents; D005227:Fatty Acids; D017666:Deuterium Oxide
keywords: cystic fibrosis; hydrogen isotope labeling; infectious disease; metabolic heterogeneity; slow growth