title: Cell-associated bacteria in the human lung microbiome.
abstract: BACKGROUND
  Recent studies have revealed that bronchoalveolar lavage (BAL) fluid contains previously unappreciated communities of bacteria. In vitro and in vivo studies have shown that host inflammatory signals prompt bacteria to disperse from cell-associated biofilms and adopt a virulent free-living phenotype. The proportion of the lung microbiota that is cell-associated is unknown.
  
  
  RESULTS
  Forty-six BAL specimens were obtained from lung transplant recipients and divided into two aliquots: 'whole BAL' and 'acellular BAL,' the latter processed with a low-speed, short-duration centrifugation step. Both aliquots were analyzed via bacterial 16S rRNA gene pyrosequencing. The BAL specimens represented a wide spectrum of lung health, ranging from healthy and asymptomatic to acutely infected. Bacterial signal was detected in 52% of acellular BAL aliquots, fewer than were detected in whole BAL (96%, p ≤ 0.0001). Detection of bacteria in acellular BAL was associated with indices of acute infection [BAL neutrophilia, high total bacterial (16S) DNA, low community diversity, p < 0.01 for all] and, independently, with low relative abundance of specific taxonomic groups (p < 0.05). When whole and acellular aliquots from the same bronchoscopy were directly compared, acellular BAL contained fewer bacterial species (p < 0.05); whole and acellular BAL similarity was positively associated with evidence of infection and negatively associated with relative abundance of several prominent taxa (p < 0.001). Acellular BAL contained decreased relative abundance of Prevotella spp. (p < 0.05) and Pseudomonas fluorescens (p < 0.05).
  
  
  CONCLUSIONS
  We present a novel methodological and analytical approach to the localization of lung microbiota and show that prominent members of the lung microbiome are cell-associated, potentially via biofilms, cell adhesion, or intracellularity.
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
authors.forename: Robert P
authors.initials: RP
authors.lastname: Dickson
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
authors.forename: John R
authors.initials: JR
authors.lastname: Erb-Downward
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
authors.forename: Hallie C
authors.initials: HC
authors.lastname: Prescott
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
authors.forename: Fernando J
authors.initials: FJ
authors.lastname: Martinez
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA ; Pulmonary and Critical Care Medicine Section, Medical Service, VA Ann Arbor Healthcare System, Ann Arbor, MI 48105, USA.
authors.forename: Jeffrey L
authors.initials: JL
authors.lastname: Curtis
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
authors.forename: Vibha N
authors.initials: VN
authors.lastname: Lama
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Department of Internal Medicine, University of Michigan Medical School, Ann Arbor, MI 48109, USA ; Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
authors.forename: Gary B
authors.initials: GB
authors.lastname: Huffnagle
keywords: 16S; Bronchoalveolar lavage; Lung microbiome; Pneumonia; Pyrosequencing