Transcript Cystic fibrosis John R W Govan University of Edinburgh Medical School

Cystic fibrosis
John R W Govan
University of Edinburgh Medical School
• What is cystic fibrosis?
• Significance of respiratory infections
• Problem pathogens and new technologies
Cystic Fibrosis
• Cystic Fibrosis is the most common, life-threatening,
recessively inherited disease of Caucasian populations
• Carrier rate of 1 in 25 and incidence rate of I in 2500
live births
• UK ~ 7500 babies, children and young adults.
• Cause. Mutations in gene encoding the cystic fibrosis
transmembrane conductance regulator (CFTR) gene which
encodes an epithelial chloride channel.
• Survival. Progressive lung disease secondary to respiratory
infection is the main cause of morbidity and mortality.
CFTR, defective mucocilary clearance and salt-sensitive
antimicrobial peptides (defensins)
Bacterial Pathogens in CF
• Limited spectrum
• Age related sequence
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–
–
–
–
Haemophilus influenzae
Staphylococcus aureus
Pseudomonas aeruginosa
Burkholderia cepacia complex
S. maltophilia/A. xylosoxidans
‘Burkholderia cepacia’ – the pathogen
• Swamp foot in US marines
• Nosocomial infections due to
contaminated fluids. ICUs
• Chronic granulomatous disease.
Inherited loss of neutrophil
oxidative killing leading to fatal
lung infection
• ‘Cepacia syndrome’ in cystic
fibrosis – fatal unexpected
pneumonia with bacteraemia in
20-30% of infected patients
Edinburgh – 1986
Glass & Govan J Infect 1986; 13:157-158.
Fatal cepacia syndrome in
9-year-old CF female.
No transfer to CF sibling.
Genomovar III (B. cenocepacia)
J415, cblA and bcesm –ve
Transmission of B. cepacia
Govan et al. Lancet 1993
• Spread within and
between CF clinics
includes social contacts.
• Outcome fatal but
unpredictable
• Leper-like infection
control -- segregation and
anxiety!
• Virtually untreatable
‘Burkholderia cepacia’
how resistant?
• Biodegradation of
synthetic herbicides and
petroleum oxidants.
• Panresistance to
antibiotics. Can utilise
penicillin as nutrient!
• Resistance to natural
antimicrobial peptides –
(defensins) of amoebae,
insects, animals and
humans – explains CGD.
B. cepacia as an animal pathogen
Berriatua et al 2000. J Clin Microbiol 2001;39: 990-4
• Untreatable outbreak of
ovine mastitis
• No environmental source
identified
• Genomovar III responsible
Bacterial hybrids or new taxons?
Simpson et al. J Antimicrob Chemother1994; 34: 353-61
‘Burkholderia cepacia complex ’
• Evolving taxonomy
– At least 10 distinct genomovars recognised by
polyphasic taxonomy and RecA PCR
• All genomovars isolated from CF patients
• However, gvr II (B. multivorans) and III
(B. cenocepacia) account for 90% of isolates and
most transmission – includes the ET12 lineage
‘B. cepacia’ AMMD
Parke & Gurion-Sherman J Phyt 2002
The biopesticide issue -update
• ‘There is no evidence that CF
patients acquire B. cepacia
from the environment – CF
isolates are different’
- biopesticide developer 1999
• Not true – even clonal
- Govan, Vandamme & Balandreau
ASM News 2000;66:124
• New use rule. Fed Reg 2002
Antibiotics in trouble!
Evolution of bacterial resistance and
pathogenesis – a two way process
• Common perception:
Bacterial pathogens accumulate resistance to
antibiotics -- rendering them virtually untreatable .
MRSA and vancomycin-resistant Enterococcus faecium
• What’s also happening:
Inherently resistant environmental bacteria acquire
opportunistic virulence for plant, human and animal
hosts. Burkholderia cepacia complex
Bacteriology IDG
Collaborations
B. cepacia
B. pseudomallei
Richard Titball
Petra Oyson
Eshwar
Mahenthiralingam
Peter
Vandamme
Ty Pitt
John
Govan
Mahidol University
Sirirurg Songsivillai
C.A. Hart
B. cepacia - general features J2315 - Edinburgh ET12 isolate
Chr. 1
Chr. 2
Chr. 3
plasmid
Total
Size
3,870,082 bp
3,217,062 bp
875,977 bp
92,661 bp
8,055,782 bp
G+C content
66.7 %
67.3 %
66.9 %
62.8 %
66.9 %
Genes
3,531
2,840
773
82
7,226
Coding density
86.6 %
87.8 %
86.6 %
80.9 %
87.0 %
Av. gene length
950 bp
995 bp
982 bp
914 bp
971 bp
rRNA
4x
16S-23S-5S
1x
16S-23S-5S
1x
16S-23S-5S
0
6x
16S-23S-5S
tRNA
66
5
2
0
73
B. cepacia – chromosome 2 – LHF analysis
fatty-acid
metabolism
+ transport
chemotaxis,
helicase,
drug efflux?
phospholipases
drug efflux,
plasmid
catabolism,
conjugation
regulators
system
polysaccharide
biosynthesis
phage,
haemolysin
phage
rRNA
haemolysin
-related
protein
cell-surface
protein
B. cepacia and the neutrophil:
dose dependent and synergistic inflamation
• Lipopeptide toxin
(haemolysin) induces
apoptosis at low
concentrations and
degranulation at high
concentrations.
• B. cepacia LPS induces
potent TNF and IL-8
response and primes
neutrophil to a damaging
response to other microbes
and ‘procedures’.
Looking ahead
• Are all B. cepacia complex potentially virulent and
transmissible?
Transplantation, segregation and biopesticides issues
• The promise of genomics and proteomics
• ‘B. cepacia’ identification at local level.
False positives 10%; False negatives 30%.
Identification of B. cepacia complex
• Variable colonial
morphology
• Selective media not
selective enough: Mast
Cepacia medium or BCSA
(Henry et al 1997).
• Accuracy of commercial
identification kits?
ID of B. cepacia complex by API20NE, Vitek
NFC/GNI and BBL Crystal
• Accuracy varies from 64-92 %. API20NE most
accurate.
• None of the systems identified genomovar VI
• Only API20NE identified the Glasgow epidemic
strain (gvr II, B. multivorans) and the epidemic
ET12 lineage (gvr III, B. cenocepacia).
Some people’s idea of microbiologists?
IBCWG (http://go.to/cepacia/)
“…as a forum for clinicians and scientists
interested in advancing knowledge of B. cepacia
infection and colonisation in persons with CF
through the collegial exchange of information and
promotion of coordinated approaches to
research…”
“B. cepacia”: other bacteria
• Many other bacteria are (mis)identified as B. cepacia
• Known species:
– P. aeruginosa
– S. maltophilia
– R. pickettii
– A. xylosoxidans
– B. hinzii
– Brevundimonas spp.
– Chryseobacterium spp.
– Enterobacteriaceae