Bacterial whole genome sequencing in clinical microbiology, infection control and public health Julian Parkhill FIS, Birmingham, November 2013
Falling costs of genomics 2003 Cost/genome Throughput 60,000 1 genome / 20 days 2013 30 3,000 genomes / 10 days 40 20 genomes / 1 day Disclaimer: Illustrative figures only. Other sequencing machines are available
Genomics in public health microbiology Rapid high throughput sequencing will benefit in 3 areas: Epidemiology: Phenotyping: Diagnosis Rapid, unambiguous, high-resolution typing Single technique to replace multiple organism-specific schemes Common digital data can be shared globally Phylogenetic analysis allows temporal and directional inference (rather than simple same/different cutoffs) Can be used now Drug resistance/sensitivity Virulence, Transmissibility Will require further research and continuous updates Possible now in limited circumstances Widespread use will require further falls in cost and throughput
Staphylococcus aureus Widespread Gram +ve bacteria Natural biota of the skin ~40% carriage Versatile pathogen associated with a wide range of diseases Minor wound infections Food poisoning Toxic shock syndrome Endocarditis Haemolytic pneumonia Complex pathology Drug resistance Methicillin-resistant S. aureus (MRSA) Vancomycin insensitive/resistant Image kindly provided by Sharon Peacock, Cambridge University
Staphylococcus aureus population structure by MLST CC1 CC51 CC8 CC22 CC39 CC45 CC5 CC30 CC25 CC12 CC15 CC9
Methicillin-resistant Staphylococcus aureus DoH reduction targets Identification of EMRSA15+16 Livermore (2012) Int J Antimicrob Agents. 39:283-94
Addenbrooke s Special Care Baby Unit outbreak Harris et al. 2013 Lancet Infect Dis. 13:130
SCBU outbreak Harris et al. 2013 Lancet Infect Dis. 13:130
SCBU outbreak Harris et al. 2013 Lancet Infect Dis. 13:130
SCBU outbreak Harris et al. 2013 Lancet Infect Dis. 13:130
SCBU outbreak Harris et al. 2013 Lancet Infect Dis. 13:130
SCBU outbreak Harris et al. 2013 Lancet Infect Dis. 13:130
SCBU outbreak Harris et al. 2013 Lancet Infect Dis. 13:130
SCBU outbreak Harris et al. 2013 Lancet Infect Dis. 13:130
SCBU outbreak Harris et al. 2013 Lancet Infect Dis. 13:130
SCBU outbreak: consequences Harris et al. 2013 Lancet Infect Dis. 13:130
Mycobacterium abscessus in Cystic Fibrosis Prior to 1990 M abscessus was rarely isolated from CF sputum. Now one of the top NTMs infecting CF patients Estimated to affect 5 10% of CF population Naturally high antibiotic resistance resistant to all frontline TB antibiotics (isoniazid, rifampin, pyrazinamide and ethambutol) Associated with decline in lung function (only less than 10% have no clinical deterioration) Thought to be environmentally acquired clinical guidelines based on this Tested in Papworth - Sequenced 168 isolates collected from 31 CF patients Bryant et al. 2013 Lancet 381:1551
Mycobacterium abscessus phylogeny 2. Deep divergences between different 4. Loosely patient isolates Clustered isolates 1. 3. Three Tightly subspecies Clustered isolates M.a. massiliense Maximum likelihood tree of 200 isolates M.a. abscessus M.a. bolletii Bryant et al. 2013 Lancet 381:1551
Pairwise similarities Pairwise SNP distance between all isolates of same subspecies Bryant et al. 2013 Lancet 381:1551
Epidemiology of possible patient contact Bryant et al. 2013 Lancet 381:1551
Dating of possible transmission events Bryant et al. 2013 Lancet 381:1551
Transmission of drug resistance Bryant et al. 2013 Lancet 381:1551
Pairwise similarities Pairwise SNP distance between all isolates of same subspecies Bryant et al. 2013 Lancet 381:1551
Epidemiology of possible patient contact Bryant et al. 2013 Lancet 381:1551
Dating of possible transmission events Bryant et al. 2013 Lancet 381:1551
M. a. massiliense circulating clone? Bryant et al. 2013 Lancet 381:1551
Mycobacterium abscessus transmission routes 1. Independent acquisition from the environment 2. Between patient transmission 3. Dominant circulating clones Bryant et al. 2013 Lancet 381:1551
Mycobacterium abscessus in Cystic Fibrosis Most typing techniques allow only identical/not-identical discrimination Use of whole genome sequencing and phylogenetic reconstruction allows dating of relationships between isolates Combining this with epidemiological information gives unambiguous demonstration of transmission events Accurate dating of deeper relationships allows identification of circulating (emerging) clones Bryant et al. 2013 Lancet 381:1551
WTSI: Gordon Dougan Josephine Bryant Simon Harris Nick Thomson Stephen Bentley Matthew Holden Nick Croucher Ankur Mutreja Thomas Connor Cambridge: Sharon Peacock Estee Torok Nicholas Brown Claudio Köser Matt Ellington Ed Cartwright Amanda Ogilvy-Stuart M. abscessus: Andres Floto Sharon Peacock Dorothy M Grogono Daniel Greaves Juliet Foweraker Iain Roddick Thomas Inns, Mark Reacher Charles Haworth Martin Curran
Resistance prediction Current accuracy Antibiotic class False negative False positive Gentamicin 0.39% 0.20% Penicillin 0.0% 0.0% Oxacillin 2.36% 0.0% Fusidic acid 4.43% 0.10% Teicoplanin 1.38% 0.0% Vancomycin 0.20% 0.0% Erythromycin 3.05% 0.49% Mupirocin 1.38% 0.0% Linezolid 0.0% 0.0% Fluoroquinolone 0.99% 0.49% Rifampicin 0.39% 0.0% Tetracycline 0.89% 0.10% Minocycline 0.0% 0.0% Trimethoprim 2.86% 0.20%
M. a. massiliense adaption? Mammalian cell entry cluster Bryant et al. 2013 Lancet 381:1551
Origin of ST22 (EMRSA15) 193 isolates, 15 countries, 20 years Holden et al. 2013 Genome Res.
Origin of ST22 (EMRSA15) Fluoroquinolone resistance Holden et al. 2013 Genome Res.
Origin of ST22 (EMRSA15) Holden et al. 2013 Genome Res.
Origin of ST22 (EMRSA15) What was happening in the West Midlands in the mid 80s? 1986