Stinear Laboratory
Page Contents
Include: Research interests | Laboratory staff | Some recent publications from the group
Research interests
A major research interest of the Stinear Laboratory is the use of genomic information to understand mechanisms of bacterial pathogenesis and directly address public health concerns for the bacterial pathogens Mycobacterium ulcerans, Mycobacterium marinum, Mycobacterium tuberculosis and Staphylococcus aureus. Some background information and specific research themes we are pursuing is outlined below.
Mycobacterium ulcerans
Mycobacterium ulcerans is an environmental mycobacterium that causes Buruli ulcer, a serious skin disease characterized by chronic ulceration of subcutaneous fat that can leave victims with debilitating, life-long deformity and disability (Fig. 1). M. ulcerans is related to Mycobacterium tuberculosis and Mycobacterium leprae, the causative agents of tuberculosis and leprosy respectively. Currently, an 8-week course of antibiotics is recommended as the front line treatment, often followed by surgery and skin grafting if the damaged area is extensive. There is no vaccine against Buruli ulcer. The disease is found throughout the world, including Australia where recent outbreaks in Victoria have coincided with an ongoing epidemic across West and Central Africa. In rural Central and West Africa the prevalence of Buruli ulcer now exceeds leprosy and, in some instances, tuberculosis. In 1998 the World Health Organisation launched the Global Buruli Ulcer Initiative with the principal aim of trying to control the spread of this debilitating disease ( http://www.who.int/gtb-buruli/).
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Fig 1: A typical village in West Africa where Buruli ulcer occurs |
| Fig 2: The structure of mycolactone B |  |
M. ulcerans produces an unusual lipid toxin called mycolactone (Fig. 2). This molecule is thought to be the major determinant of virulence because injection of purified mycolactone into the skin of guinea pigs provokes the appearance of ulcers. Mycolactone has been shown to have in vitro cytotoxic and immunosuppressive properties but the precise role it plays in pathogenesis is still unclear. It is also possible that M. ulcerans may elaborate other factors that play a role in ulcer formation. In collaboration with the Institut Pasteur we determined the genome sequence of M. ulcerans ( http://genolist.pasteur.fr/BuruList/). A major finding from the genome project was the discovery of a virulence plasmid and three polyketide synthase (PKS) genes on this plasmid required for mycolactone biosynthesis. The 12-membered core of mycolactone is produced by two giant type I modular PKS, whereas its side chain is synthesized by a third PKS (Fig. 3). |
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Fig 3: Biosynthetic pathway and module and domain organisation of the two type I polyketide synthases (MlsA1 and MlsA2) required for the sythesis of the mycolactone core. A similar PKS (MlsB) produces the fatty acyl side-chain. (Click on image for High Resolution image) |
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The availability of the complete genome sequence provides an important resource for international research efforts aimed at controlling the spread of Buruli ulcer. In this respect the research within our group is focused on:
- Using genome data to identify and characterize M. ulcerans specific antigens for use in the development of a rapid diagnostic tests and future vaccine studies.
- Using genome data to identify potential determinants of virulence other than mycolactone.
- Developing systems for heterologous expression of mycolactone so that a reliable source of
mycolactone and derivatives is available for:
- determining the role of mycolactone in the bacterium
- establishing the relationship between mycolactone structure and function in the eukaryotic host
Collaborators
Dr Torsten Seemann, Victorian Bioinformatics Consortium, Monash University
(http://www.vicbioinformatics.com)
Dr Grant Jerker Jenkin, Department of Infectious Diseases, Monash Medical Centre
and Department of Microbiology, Monash University
(http://www.miin.monash.org/members/jenkin-grant.html)
Dr Kellie Tuck, School of Chemistry, Monash University
(http://www.chem.monash.edu.au/staff/tuck/)
Dr Kumar Visanathan at the Centre for Inflammatory Diseases at the Monash Institute
of Medical Research
(http://www.monashinstitute.org/cid-innate-immunity.html)
Prof. Gerd Pluschke at the Swiss Tropical Institute in Basel
(http://www.sti.ch/no_cache/about-us/staff/detail-single-all/staff/163/gerd-pluschke.html)
Assoc. Prof. Paul Johnson at the Department of Infectious Diseases, Austin Health
(http://members.ozemail.com.au/~groverjohnson/Mulcerans.html)
Dr Janet Fyfe at the Victorian Infectious Diseases Reference Laboratory
(http://www.vidrl.org.au:labsandunits:mulcerans:mulcerans_idx.htm)
Prof. Peter Leadlay, Department of Biochemistry, University of Cambridge, UK
(http://www.bio.cam.ac.uk/~pflgroup/)
Dr Hui Hong, Department of Chemistry, University of Cambridge, UK
(http://www.bio.cam.ac.uk/~pflgroup/Hui.htm)
Dr Caroline Demangel, Institut Pasteur, Paris
(
http://www.pasteur.fr/recherche/unites/Pmi/index.html)
Dr Roland Brosch, Institut Pasteur, Paris
(http://www.pasteur.fr/recherche/unites/Pmi/index.html)
Dr Laurent Marsollier, Groupe d’étude des interactions hôtes-parasites, Université d’Angers, France
Staphylococcus aureus
Staphylococcus aureus still remains a dangerous human pathogen, with the ability to cause a broad range of diseases, such as bacteremia, endocarditis, sepsis and toxic shock syndrome. Over the last 20 years there has been an increasing incidence of both hospital-acquired and community–acquired S. aureus infections. Treatment of these infections has become more difficult due to the emergence of multidrug-resistant strains. In particular, methicillin-resistant S. aureus (MRSA), which was once restricted to hospitals, is spreading rapidly through the community. Furthermore, resistance to vancomycin, the primary drug used to treat MRSA, has also recently emerged.
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Fig 4: Electron micrograph of vancomycin sensitive and vancomycin intermediate S. aureus (VSSA and VISA, respectively), showing thickening of the cell wall in the resistant isolate. |
Our research into S. aureus pathogenesis focuses on using clinically relevant strains to investigate the molecular mechanisms behind the emergence of low-level vancomycin resistance in this organism. We also have an interest in investigating the host-pathogen implications of vancomycin resistance, and the role of the Panton-Valentine leukocidin (PVL) in the pathogenesis of Australian MRSA and methicillin-sensitive S. aureus isolates. To achieve these aims we utilize a variety of molecular microbiology techniques including microarrays, whole genome sequencing, allelic exchange and protein function studies. |
|
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Fig 5: Heat map showing fold-changes in gene expression of VSSA compared with VISA. |
Collaborators
Professor John Davies at Monash University
(
http://www.med.monash.edu.au/microbiology/research/davies.html)
A/Prof. Catherine Bennett, School of Population Health, University of Melbourne
(
http://www.sph.unimelb.edu.au/about/allstaff/bennett)
Austin Health Infectious Diseases Department
Austin Health Microbiology Department
Some useful WWW links:
For more information regarding polyketide synthases visit the site of one of our collaborators:
http://www.bio.cam.ac.uk/~pflgroup/
For more information on Mycobacterium ulcerans and the disease it causes visit:
http://www.health.vic.gov.au/ideas/diseases/myco_facts.html
For more information about the Stop Buruli initiative, visit:
http://stopburuli.org/
For access to the complete M. ulcerans genome sequence visit:
http://genopole.pasteur.fr/Mulc/BuruList.html
Laboratory Staff
Head of Research Group:
Dr Tim Stinear: email: tstinear@unimelb.edu.au
Research staff (as of Apr 2009)
Dr Ben Howden (Visiting Scientist)
Mr Wei Gao (Visiting Scientist)
Jessica Porter (Research Assistant)
Sacha Pidot (PhD student)
Nicholas Tobias (PhD student)
Dr Kyra Chua (PhD student)
Some recent publications from the group
- Demangel C, Stinear TP, Cole ST. Buruli ulcer: reductive evolution enhances pathogenicity of Mycobacterium ulcerans. Nat Rev Microbiol 2009; 7(1): 50-60.
- Pidot SJ, Hong H, Seemann T, Porter JL, Yip MJ, Men A, Johnson M, Wilson P, Davies JK, Leadlay PF, Stinear TP. Deciphering the genetic basis for polyketide variation among mycobacteria producing mycolactones. BMC Genomics 2008; 9: 462.
- Lavender CJ, Stinear TP, Johnson PD, Azuolas J, Benbow ME, Wallace JR, Fyfe JA. Evaluation of VNTR typing for the identification of Mycobacterium ulcerans in environmental samples from Victoria, Australia. FEMS Micrbiol Lett 2008; 287(2): 250-255.
- Howden BP, Stinear TP, Allen DL, Johnson PD, Ward PB, Davies JK. Genomic analysis reveals a point mutation in the two-component sensor gene graS that leads to intermediate vancomycin resistance in clinical Staphylococcus aureus. Antimicrob Agents Chemother 2008; 52(10): 3755-62.
- Stragier P, Hermans K, Stinear T, Portaels F. First report of a mycolactone-producing Mycobacterium infection in fish agriculture in Belgium. FEMS Microbiol Lett 2008; 286(1): 93-5.
- Tafelmeyer P, Laurent C, Lenormand P, Rousselle JC, Marsollier L, Reysset G, Zhang R, Sickmann A, Stinear TP, Namane A, Cole ST. Comprehensive proteome analysis of Mycobacterium ulcerans and quantitative comparison of mycolactone biosynthesis. Proteomics 2008; 8(15): 3124-38.
- Hong H, Demangel C, Pidot SJ, Leadlay PF, Stinear T. Mycolactones: immunosuppressive and cytotoxic polyketides produced by aquatic mycobacteria. Nat Prod Rep 2008; 25(3): 447-54.
- Stinear TP, Seemann T, Harrison PF, Jenkin GA, Davies JK, Johnson PD, Abdellah Z, Arrowsmith C, Chillingworth T, Churcher C, Clarke K, Cronin A, Davis P, Goodhead I, Holroyd N, Jagels K, Lord A, Moule S, Mungall K, Norbertczak H, Quail MA, Rabbinowitsch E, Walker D, White B, Whitehead S, Small PL, Brosch R, Ramakrishnan L, Fischbach MA, Parkhill J, Cole ST. Insights from the complete genome sequence of Mycobacterium marinum on the evolution of Mycobacterium tuberculosis. Genome Res 2008; 18(5): 729-41.
- Stinear T, Johnson PD. First Isolation of Mycobacterium ulcerans from an Aquatic Environment: The End of a 60-Year Search? PLoS Negl Trop Dis 2008; 26;2(3): e216.
- Howden BP, Smith DJ, Mansell A, Johnson PD, Ward PB, Stinear TP, Davies JK. Different bacterial gene expression patterns and attenuated host immune responses are associated with the evolution of low-level vancomycin resistance during persistent methicillin-resistant Staphylococcus aureus bacteraemia. BMC Microbiol 2008; 27;8: 39.
- Johnson PD, Azuolas J, Lavender CJ, Wishart E, Stinear TP, Hayman JA, Brown L, Jenkin GA, Fyfe JA. Mycobacterium ulcerans in mosquitoes captured during outbreak of Buruli ulcer, southeastern Australia. Emerg Infect Dis 2007; 13(11): 1653-60.
- Rondini S, Käser M, Stinear T, Tessier M, Mangold C, Dernick G, Naegeli M, Portaels F, Certa U, Pluschke G. Ongoing genome reduction in Mycobacterium ulcerans. Emerg Infect Dis 2007; 13(7): 1008-15.
- Hong H, Stinear T, Porter J, Demangel C, Leadlay PF. A novel mycolactone toxin obtained by biosynthetic engineering. Chembiochem 2007; 23;8(17): 2043-7.
- Käser M, Rondini S, Naegeli M, Stinear T, Portaels F, Certa U, Pluschke G. Evolution of two distinct phylogenetic lineages of the emerging human pathogen Mycobacterium ulcerans. BMC Evol Biol 2007; 27;7: 177.
- Fyfe JA, Lavender CJ, Johnson PD, Globan M, Sievers A, Azuolas J, Stinear TP. Development and application of two multiplex real-time PCR assays for the detection of Mycobacterium ulcerans in clinical and environmental samples. Appl Environ Microbiol 2007; 73(15): 4733-40.
- Hilty M, Käser M, Zinsstag J, Stinear T, Pluschke G. Analysis of the Mycobacterium ulcerans genome sequence reveals new loci for variable number tandem repeats (VNTR) typing. Microbiology 2007; 153(Pt 5): 1483-7.
- Lavender CJ, Senanayake SN, Fyfe JA, Buntine JA, Globan M, Stinear TP, Hayman JA, Johnson PD. First case of Mycobacterium ulcerans disease (Bairnsdale or Buruli ulcer) acquired in New South Wales. Med J Aust 2007; 15;186(2): 62-3.
- Stinear TP, Seemann T, Pidot S, Frigui W, Reysset G, Garnier T, Meurice G, Simon D, Bouchier C, Ma L, Tichit M, Porter JL, Ryan J, Johnson PD, Davies JK, Jenkin GA, Small PL, Jones LM, Tekaia F, Laval F, Daffé M, Parkhill J, Cole ST. Reductive evolution and niche adaptation inferred from the genome of Mycobacterium ulcerans, the causative agent of Buruli ulcer. Genome Res 2007; 17(2): 192-200.
- Yip MJ, Porter JL, Fyfe JA, Lavender CJ, Portaels F, Rhodes M, Kator H, Colorni A, Jenkin GA, Stinear T. Evolution of Mycobacterium ulcerans and other mycolactone-producing mycobacteria from a common Mycobacterium marinum progenitor. J Bacteriol 2007; 189(5): 2021-9.