title: Penicillin induced persistence in Chlamydia trachomatis: high quality time lapse video analysis of the developmental cycle.
abstract: BACKGROUND
  Chlamydia trachomatis is a major human pathogen with a unique obligate intracellular developmental cycle that takes place inside a modified cytoplasmic structure known as an inclusion. Following entry into a cell, the infectious elementary body (EB) differentiates into a non-infectious replicative form known as a reticulate body (RB). RBs divide by binary fission and at the end of the cycle they redifferentiate into EBs. Treatment of C.trachomatis with penicillin prevents maturation of RBs which survive and enlarge to become aberrant RBs within the inclusion in a non-infective persistent state. Persistently infected individuals may be a reservoir for chlamydial infection. The C.trachomatis genome encodes the enzymes for peptidoglycan (PG) biosynthesis but a PG sacculus has never been detected. This coupled to the action of penicillin is known as the chlamydial anomaly. We have applied video microscopy and quantitative DNA assays to the chlamydial developmental cycle to assess the effects of penicillin treatment and establish a framework for investigating penicillin induced chlamydial persistence.
  
  
  RESULTS
  Addition of penicillin at the time of cell infection does not prevent uptake and the establishment of an inclusion. EB to RB transition occurs but bacterial cytokinesis is arrested by the second binary fission. RBs continue to enlarge but not divide in the presence of penicillin. The normal developmental cycle can be recovered by the removal of penicillin although the large, aberrant RBs do not revert to the normal smaller size but remain present to the completion of the developmental cycle. Chromosomal and plasmid DNA replication is unaffected by the addition of penicillin but the arrest of bacterial cytokinesis under these conditions results in RBs accumulating multiple copies of the genome.
  
  
  CONCLUSIONS
  We have applied video time lapse microscopy to the study of the chlamydial developmental cycle. Linked with accurate measures of genome replication this provides a defined framework to analyse the developmental cycle and to investigate and provide new insights into the effects of antibiotic treatments. Removal of penicillin allows recovery of the normal developmental cycle by 10-20 hrs and the process occurs by budding from aberrant RBs.
authors.affiliation: Molecular Microbiology Group, University of Southampton Medical School, Southampton General Hospital, Southampton, UK.
authors.forename: Rachel J
authors.initials: RJ
authors.lastname: Skilton
authors.forename: Lesley T
authors.initials: LT
authors.lastname: Cutcliffen
authors.forename: David
authors.initials: D
authors.lastname: Barlow
authors.forename: Yibing
authors.initials: Y
authors.lastname: Wang
authors.forename: Omar
authors.initials: O
authors.lastname: Salim
authors.forename: Paul R
authors.initials: PR
authors.lastname: Lambden
authors.forename: Ian N
authors.initials: IN
authors.lastname: Clarke
mesh_terms: D000818:Animals; D018407:Bacterial Physiological Phenomena; D002692:Chlamydia trachomatis / Q000187:drug effects / Q000378:metabolism*; D003513:Cycloheximide / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D000882:Haplorhini; D007668:Kidney / Q000166:cytology; D046529:Microscopy, Electron, Transmission / Q000379:methods; D018715:Microscopy, Video / Q000379:methods; D010406:Penicillins / Q000494:pharmacology*; D010457:Peptidoglycan / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors
chemical_list: D010406:Penicillins; D010457:Peptidoglycan; D003513:Cycloheximide