uw medicine │ patients are first battling bugs: inroads in infectious diseases uw mini-medical...
TRANSCRIPT
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UW MEDICINE │ PATIENTS ARE FIRST
BATTLING BUGS: INROADS IN
INFECTIOUS DISEASES
UW MINI -MEDICAL SCHOOL
Brad T. Cookson M.D., Ph.D.
February 11, 2014
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“Humanity has but three great enemies: fever, famine and war; of these by far the greatest, by far the most terrible, is fever.”
Sir William Osler, 1896
FEVER: THE HOST RESPONDS
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“HOUSTON, WE’VE HAD A PROBLEM.”
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“HOUSTON, WE’VE HAD A PROBLEM.”
Four Leading Causes of Morbidity and Mortality:
• Heart Disease
• Stroke
• Cancer
• Infection
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Diagnosis. The determination of the nature of a disease. [G. a deciding] SYN: diacrisis. Diacrisis SYN: diagnosis [G. dia-, through, + krisis, a judgment]
DIAGNOSIS: KEY TO EFFECTIVE TREATMENT
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“Listen to your patient, he is telling you the diagnosis.”
Sir William Osler (1849 – 1919)
DIAGNOSIS: KEY TO EFFECTIVE TREATMENT
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Hypothesis testing:
• History• Physical Exam• (imaging)• Laboratory Testing
(Laboratory Medicine)
DIAGNOSIS: A SCIENTIFIC APPROACH
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• Chemistry• Hematology• Transfusion Medicine• Immunology• Microbiology• Virology• Medical Informatics• Molecular Diagnosis Program
(Molecular Microbiology Laboratory)
DEPARTMENT OF LABORATORY MEDICINE
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• Chemistry• Hematology• Transfusion Medicine• Immunology• Microbiology• Virology• Medical Informatics• Molecular Diagnosis Program
(Molecular Microbiology Laboratory)
DEPARTMENT OF LABORATORY MEDICINE
Google: UW Medical Laboratory Science
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Determine the nature of disease:• Predict course and potential outcome(s) of infection• Tailor therapy• Exclude non-infectious cause(s) of symptoms
IDENTIFY ETIOLOGICAL AGENTS
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Determine the nature of disease:• Predict course and potential outcome(s) of infection• Tailor therapy• Exclude non-infectious cause(s) of symptoms
IDENTIFY ETIOLOGICAL AGENTS
What you can’t see, can kill you!
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• If it grows, it can usually be identified
TRADITIONAL APPROACH
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• If it grows, it can usually be identified• Acquire patient specimen
• blood, urine, CSF• Microscopic examination
TRADITIONAL APPROACH
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• If it grows, it can usually be identified• Acquire patient specimen
• blood, urine, CSF• Microscopic examination• Isolate• Amplify
TRADITIONAL APPROACH
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ISOLATE & AMPLIFY
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ISOLATE & 109 AMPLIFICATION
(Google: Scale of universe;Powers of Ten)
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IDENTIFY ETIOLOGICAL AGENTS
Phenotype
Gene products
(physical expression of genotype)
• Identification• Antibiotic resistance
(proteins, enzymes, complex structures)
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IDENTIFY ETIOLOGICAL AGENTS
Phenotype
Gene products
Genotype
(physical expression of genotype)
• Identification• Antibiotic resistance
(proteins, enzymes, complex structures)
(blueprint for phenotype)
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• If it grows, it can usually be identified
What if…• it grows very slowly?• it does not grow in the lab?• it has disguised usual
characteristics?• it has never been seen before?
TRADITIONAL APPROACH
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• If it grows, it can usually be identified
What if…• it grows very slowly?• it does not grow in the lab?• it has disguised usual characteristics?• it has never been seen before?
• If it grows, it can usually be identified
TRADITIONAL APPROACH
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IDENTIFY ETIOLOGICAL AGENTS
Phenotype
Gene products
Genotype
(physical expression of genotype)
• Identification• Antibiotic resistance
(proteins, enzymes, complex structures)
(blueprint for phenotype)
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GENOMIC APPROACH: ISOLATE & AMPLIFY
Target: Species-identifying DNA sequences flanked by conserved primer binding sites
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GENOMIC APPROACH: ISOLATE & AMPLIFY
Target: Species-identifying DNA sequences flanked by conserved primer binding sites
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• 79-yr-old male• Soft-tissue excision• Inflammatory tissue• No microbial elements• Cultures were negative
CASE 1
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“The differential diagnosis includes:• infectious process (possible _______ infection), • inflammatory process such as rheumatoid nodule or a • neoplastic process (epithelioid carcinoma can present with foci of necrosis, however the histopathological features do not favor same).”
• PCR and DNA sequencing were performed on a PET specimen.
CASE 1
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23-yr-old man with refractory seizures
CASE 2
MRI: Vasogenic edema Gadolinium enhancing
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• Exhaustive serological testing was negative• Cultures were negative• Surgically excised lesion• Mixed inflammatory cell infiltrate without presence of microbial elements
• PCR and DNA sequencing were performed on a fresh surgical specimen.
CASE 2
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CASE 3
PCR and DNA sequencing were performed.
• Young person with AML• Lymph node biopsy• Cultures negative
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GENOMIC APPROACH: ISOLATE & AMPLIFY
Target: Species-identifying DNA sequences flanked by conserved primer binding sites
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What if…• infection occurs at a site with
normal microbiota? • more than one pathogen is
present?
GENOMIC APPROACH: ISOLATE & AMPLIFY
Target: Species-identifying DNA sequences flanked by conserved primer binding sites
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NORMAL MICROBIOTA
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POLYMICROBIAL INFECTION
Clin. Microbiol. Rev. 2012, 25(1):193.
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Dr. Toby Russell, assisted by Dr. Beverly Crusher, 2368. Genitronic replication of Worf’s new spinal column, Episode #115, Star Trek, The Next Generation
NEXT GENERATION SEQUENCING
“Sequencing the genome at 109 base pairs per second…”
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• Capture single DNA molecules• Cluster formation: amplify ~1,000 copies• Parallel DNA sequencing of clusters• Read millions of clusters per flow cell!
NGS: ISOLATE & AMPLIFY
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BACTERIAL VAGINOSIS
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CYSTIC FIBROSIS
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• 35-yr-old male with meningitis• Found to have brain abscess• CSF cultures were negative• Traditional PCR + DNA sequencing revealed polymicrobial infection
CASE
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• 35-yr-old male with meningitis• Found to have brain abscess• CSF cultures were negative• Traditional PCR + DNA sequencing revealed polymicrobial infection• Next Generation Sequencing was performed• Antibiotic regimen was optimized
CASE
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Mission
Support delivery of the best possible patient care by providing excellence in the laboratory science of diagnosing infectious diseases.
MOLECULAR MICROBIOLOGY LABORATORY
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Google: UW Molecular Microbiology