uw medicine │ patients are first battling bugs: inroads in infectious diseases uw mini-medical...
TRANSCRIPT
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
2
“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
3
“HOUSTON, WE’VE HAD A PROBLEM.”
4
“HOUSTON, WE’VE HAD A PROBLEM.”
Four Leading Causes of Morbidity and Mortality:
• Heart Disease
• Stroke
• Cancer
• Infection
5
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
6
“Listen to your patient, he is telling you the diagnosis.”
Sir William Osler (1849 – 1919)
DIAGNOSIS: KEY TO EFFECTIVE TREATMENT
7
Hypothesis testing:
• History• Physical Exam• (imaging)• Laboratory Testing
(Laboratory Medicine)
DIAGNOSIS: A SCIENTIFIC APPROACH
8
• Chemistry• Hematology• Transfusion Medicine• Immunology• Microbiology• Virology• Medical Informatics• Molecular Diagnosis Program
(Molecular Microbiology Laboratory)
DEPARTMENT OF LABORATORY MEDICINE
9
• Chemistry• Hematology• Transfusion Medicine• Immunology• Microbiology• Virology• Medical Informatics• Molecular Diagnosis Program
(Molecular Microbiology Laboratory)
DEPARTMENT OF LABORATORY MEDICINE
Google: UW Medical Laboratory Science
10
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
11
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!
12
• If it grows, it can usually be identified
TRADITIONAL APPROACH
13
• If it grows, it can usually be identified• Acquire patient specimen
• blood, urine, CSF• Microscopic examination
TRADITIONAL APPROACH
14
• If it grows, it can usually be identified• Acquire patient specimen
• blood, urine, CSF• Microscopic examination• Isolate• Amplify
TRADITIONAL APPROACH
15
ISOLATE & AMPLIFY
16
ISOLATE & 109 AMPLIFICATION
(Google: Scale of universe;Powers of Ten)
17
IDENTIFY ETIOLOGICAL AGENTS
Phenotype
Gene products
(physical expression of genotype)
• Identification• Antibiotic resistance
(proteins, enzymes, complex structures)
18
IDENTIFY ETIOLOGICAL AGENTS
Phenotype
Gene products
Genotype
(physical expression of genotype)
• Identification• Antibiotic resistance
(proteins, enzymes, complex structures)
(blueprint for phenotype)
19
• 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
20
• 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
21
IDENTIFY ETIOLOGICAL AGENTS
Phenotype
Gene products
Genotype
(physical expression of genotype)
• Identification• Antibiotic resistance
(proteins, enzymes, complex structures)
(blueprint for phenotype)
22
GENOMIC APPROACH: ISOLATE & AMPLIFY
Target: Species-identifying DNA sequences flanked by conserved primer binding sites
23
GENOMIC APPROACH: ISOLATE & AMPLIFY
Target: Species-identifying DNA sequences flanked by conserved primer binding sites
24
• 79-yr-old male• Soft-tissue excision• Inflammatory tissue• No microbial elements• Cultures were negative
CASE 1
25
“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
26
23-yr-old man with refractory seizures
CASE 2
MRI: Vasogenic edema Gadolinium enhancing
27
• 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
28
CASE 3
PCR and DNA sequencing were performed.
• Young person with AML• Lymph node biopsy• Cultures negative
29
GENOMIC APPROACH: ISOLATE & AMPLIFY
Target: Species-identifying DNA sequences flanked by conserved primer binding sites
30
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
31
NORMAL MICROBIOTA
32
POLYMICROBIAL INFECTION
Clin. Microbiol. Rev. 2012, 25(1):193.
33
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…”
34
• 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
35
BACTERIAL VAGINOSIS
36
CYSTIC FIBROSIS
37
• 35-yr-old male with meningitis• Found to have brain abscess• CSF cultures were negative• Traditional PCR + DNA sequencing revealed polymicrobial infection
CASE
38
• 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
39
Mission
Support delivery of the best possible patient care by providing excellence in the laboratory science of diagnosing infectious diseases.
MOLECULAR MICROBIOLOGY LABORATORY
40
Google: UW Molecular Microbiology