Oral Presentation Australian Society for Microbiology Annual Scientific Meeting 2018

Genetic Composition and Regulatory Control of IncA/C Plasmid Conjugation (#149)

Steven J Hancock 1 , Minh Duy Phan 1 , Alvin Lo 1 , Kate M Peters 1 , Brian M Forde 1 , Zhen Luo 2 , Bostjan Kobe 1 , Ji Yang 3 , Richard A Strugnell 3 , David L Paterson 4 , Timothy R Walsh 5 , Scott A Beatson 1 , Mark A Schembri 1
  1. Australian Infectious Diseases Research Centre, School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane, QLD, Australia
  2. University of Queensland, ST LUCIA, QLD, Australia
  3. Department of Microbiology and Immunology, The University of Melbourne at the Peter Doherty Institute for Infection and Immunity, Melbourne, VIC, Australia
  4. The University of Queensland Centre for Clinical Research, Brisbane, QLD, Australia
  5. Department of Medical Microbiology and Infectious Disease, Cardiff University, Cardiff, United Kingdom

Plasmids are major vehicles for the carriage and spread of antibiotic resistance genes to all major classes of antibiotics. IncA/C plasmids are a significant group of broad-host range plasmids associated with the carriage of multiple resistance genes in enteric human pathogens, including resistance to extended spectrum cephalosporins and last-line carbapenems. Thus, IncA/C plasmids have contributed to the emergence of Carbapenem Resistant Enterobacteriaceae (CRE), an urgent threat to public health due to their extensive antibiotic resistance profile, including resistance to last line carbapenems. Despite their impact on the dissemination of antibiotic resistance genes, our current understanding of the genetics of IncA/C conjugation is limited. In this study, we utilised hyper-saturated transposon mutagenesis coupled with transposon directed insertion site sequencing (TraDIS) to determine the complement of genes required for conjugation of the prototype IncA/C plasmid pMS6198A. In total, 27 genes were identified, including all 17 predicted and known conjugation genes, two known regulatory genes (acaDC) and eight genes not previously associated with conjugation. The eight novel genes were individually mutated and the resulting mutant plasmids were examined for their ability to conjugate. Five of the eight novel genes were confirmed to significantly affect conjugation. One of these genes, which we refer to as acaB, is a novel regulator that acts at the top of the hierarchical cascade of conjugation control. Our data demonstrate that AcaB activates conjugation via upregulation of acaDC, a controlling element that in-turn directly activates the transcription of genes involved in the production of conjugative pili and DNA transfer. Overall, this work has led to the identification of novel genes associated with IncA/C conjugation, thus advancing our understanding of the spread of this important group of broad-host range multidrug resistance plasmids.