Transcript Bad hygiene or bad bug???

Bad hygiene or bad bug???
A case-based bioinformatics
exercise for biology/microbiology
students
In this activity students will:
Investigate a current epidemiological case about an
alarming outbreak of Clostridium difficile in Canada and
U.S. Using web-based bioinformatics tools, students will
explore the following questions:
• What makes C. difficile pathogenic?
• What might cause increased pathogenicity in C.
difficile?
• How does C. difficile regulate genes involved in
pathogenicity?
• Can a genetic mutation affect its pathogenic ability?
• What types of mutations would you predict?
• Can you test your predictions with information about
DNA sequence???
Potential audiences:
Biology course: 1st year non-majors and
above
Microbiology: allied health, general
microbiology, etc.
Problem space:
• Background information and websites about opportinists
pathogens of GI tract, exotoxins, C. difficile.
• Tutorial information about operon model in bacterial
chromosomes
• Sequences of C. difficile toxins A and B and regulator genes
from patients involved in outbreak as well as control (historical)
samples. (We will be including comprehensive sample set from
MacDonald study identifying outbreak origins and toxin
differences).
• Links to appropriate bioinformatics tools.
• CaseIt activities simulating researchers’ approach to studying
the outbreak
Sample Investigative Activity:
Identification of tcdC gene variant as
putative reason for high pathogenicity
Once students have identified tcdC gene
mutation as a causative factor, they can
compare sequences among patients.
How similar are gene sequences among
patients in Quebec, UK, and Europe?
Steps in bioinformatic analysis
• Obtain sequences
– What sequences?
• Provide data set, or
• Student-driven GenBank search
• Upload into Workbench
• ClustalW alignment and tree
• Case It! activities interwoven
Alignment of tcdC negative regulator genes
Deletion
Mutation?
tcdC negative regulator genes
Italy
U.K.
Quebec
Published PCR analysis of toxA and toxB genes from C. difficile
Case It! simulation of PCR analysis
PFGE analysis of C. difficile strains
PFGE and Southern blot of C. perfringens
Case It! simultation of PFGE analysis and Southern blot
Case It! simultation of PFGE analysis
Case It! simultation of PFGE analysis
Summary
• Student engaged in investigation of a
current medical issue
• Motivation to use a variety of
bioinformatics tools
• Adaptable for different types of
courses/student populations
• Integration with labs or lab simulation
References:
• Eggerston, Laura. 2006. Quebec strain of C. difficile in
7 provinces. CMAJ 174 (5). doi:10.1503/cmaj.060105.
• Pechine S, Janoir C, Collignon A. 2005. Quebec strain
of C. difficile in 7 provinces.
J Clin Microbiol. 2005 Oct;43(10):5018-25.
• Rupnik et. Al. 2005. Revised nomenclature of
Clostridium difficile toxins and associated genes. J Med
Microbiol 54 (2005), 113-117.
• Loo et. al., A Predominantly Clonal Multi-Institutional
Outbreak of Clostridium difficile–Associated Diarrhea
with High Morbidity and Mortality. Volume 353:24422449, Number 23
• Cohen, et. al. 2000. Analysis of the Pathogenicity
Locus in Clostridium difficile Strains. The Journal of
Infectious Diseases, volume 181 (2000), pages 659–
663