title: Resuscitation-promoting factors reveal an occult population of tubercle Bacilli in Sputum.
abstract: BACKGROUND
  Resuscitation-promoting factors (Rpfs) are a family of secreted proteins produced by Mycobacterium tuberculosis (Mtb) that stimulate mycobacterial growth. Although mouse infection studies show that they support bacterial survival and disease reactivation, it is currently unknown whether Rpfs influence human infection. We hypothesized that tuberculous sputum might include a population of Rpf-dependent Mtb cells.
  
  
  OBJECTIVE
  To determine whether Rpf-dependent Mtb cells are present in human sputum and explore the impact of chemotherapy on this population.
  
  
  METHODS
  In tuberculous sputum samples we compared the number of cells detected by conventional agar colony-forming assay with that determined by limiting dilution, most-probable number assay in the presence or absence of Rpf preparations.
  
  
  RESULTS
  In 20 of 25 prechemotherapy samples from separate patients, 80-99.99% of the cells demonstrated by cultivation could be detected only with Rpf stimulation. Mtb cells with this phenotype were not generated on specimen storage or by inoculating sputum samples with a selection of clinical isolates; moreover, Rpf dependency was lost after primary isolation. During chemotherapy, the proportion of Rpf-dependent cells was found to increase relative to the surviving colony-forming population.
  
  
  CONCLUSIONS
  Smear-positive sputum samples are dominated by a population of Mtb cells that can be grown only in the presence of Rpfs. These intriguing proteins are therefore relevant to human infection. The Rpf-dependent population is invisible to conventional culture and is progressively enhanced in relative terms during chemotherapy, indicating a form of phenotypic resistance that may be significant for both chemotherapy and transmission.
authors.affiliation: Department of Infection, Immunity, and Inflammation, University of Leicester, Leicester, LE1 9HN UK. gvm4@le.ac.uk
authors.forename: Galina V
authors.initials: GV
authors.lastname: Mukamolova
authors.forename: Obolbek
authors.initials: O
authors.lastname: Turapov
authors.forename: Joanne
authors.initials: J
authors.lastname: Malkin
authors.forename: Gerrit
authors.initials: G
authors.lastname: Woltmann
authors.forename: Michael R
authors.initials: MR
authors.lastname: Barer
mesh_terms: D000995:Antitubercular Agents / Q000627:therapeutic use; D001426:Bacterial Proteins / Q000494:pharmacology*; D001431:Bacteriological Techniques; D015169:Colony Count, Microbial; D003114:Colony-Forming Units Assay; D016207:Cytokines / Q000494:pharmacology*; D006801:Humans; D009169:Mycobacterium tuberculosis / Q000187:drug effects*; D011379:Prognosis; D011994:Recombinant Proteins / Q000494:pharmacology*; D012293:Rifampin / Q000627:therapeutic use; D013183:Sputum / Q000382:microbiology*; D014397:Tuberculosis, Pulmonary / Q000175:diagnosis* / Q000188:drug therapy / Q000382:microbiology / Q000635:transmission
chemical_list: D000995:Antitubercular Agents; D001426:Bacterial Proteins; D016207:Cytokines; D011994:Recombinant Proteins; C113704:resuscitation-promoting factor, bacteria; D012293:Rifampin