biosafety and the 2014 ebola epidemic during the 2014 ebola outbreak... · organismic and...
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Nathan Yozwiak and Danny Park Broad Institute of MIT and Harvard September 10, 2014
Harvard University!Center for Systems Biology!Organismic and Evolutionary Biology!
Biosafety and the 2014 Ebola Epidemic
Graduate-level genomics education programs
Diagnostic metagenomic studies
Genomics teaching and sequencing labs
Sample collection networks
Partnered training programs
Education Research Genomics capacity
On-site training workshops
Diagnostics core facility
BioinformaticsSustainable science careers
African Center of Excellence for Genomics of Infectious Diseases
Afr
ican
Centre of Excellence for
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Redeemer s ni ersit i er
ia
Genomics of Infectious D
iseas
es
Kenema Government
Hospital
Cheikh Anta-Diop University
University of Sierra Leone
International Partners
Senegal
University of Ibadan
Redeemer’s University
Irrua Specialist Teaching Hospital
Sierra Leone
Nigeria
Sierra Leone
African Center of Excellence for Genomics of Infectious Diseases
Afr
ican
Centre of Excellence for
K^]^^f^k�l�Ngbo^klbmr%�Gb`^kbZ
Redeemer s ni ersit i er
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Genomics of Infectious D
iseas
es
Long-standing research partnerships
3,109 suspected and confirmed cases of Ebola (Aug 31 2014)
“Six months into the worst Ebola epidemic in history, the world is losing the battle to contain it.” - Dr. Joanne Liu, MSF President to the UN, Sept 2, 2014
First outbreak in West Africa, and the biggest outbreak recorded
1000 km
GuineaSierra LeoneLiberia
DRCGabon
20142007-08
1976-77
100100 1000
RC2001-051994-96
cases
Outbreak exhibits exponential growth: 35 day doubling rate
Num
ber o
f cas
es
Month of 2014
0
500
1000
1500
2000
Apr May Jun Jul Aug
Liberia
Sierra Leone
Guinea
Exponential fit R2=0.96Doubling time = 35 days
Outbreak supplies Containment and Biosafety Ebola diagnostics
Pre-Outbreak Preparedness
3000bp
100bp
Negative Patient 1 Negative Patient 2 NC
1:1 1:10 1:100 1:1K 1:1 1:10 1:100 1:1K
EBOV seed stock spike-in dilutions
A
B
3000bp
100bp76bp
1:1 1:10 1:100 1:1KEBOV seed stock dilutions
1:1 1:10 1:100 1:1K
118bp
1:1 1:10 1:100 1:1K
Kulesh assay
419bp
FiloAB assay KGH assay
118bp
Augustine Goba, Director MoHS-KGH Laboratory
May 25, 2014: First confirmed case of Ebola in Sierra Leone
For Kenema Government Hospital’s Ebola effort
1. Resupply PPE and diagnostic reagents
2. Streamline diagnostic workflow
3. Share preliminary findings with health leaders
July 2014: Outbreak Response
35
Confirmed
Suspected
Traditional
Suspected Ebola
14Confirmed Ebola
14 cases identified within first week of outbreak
Can view mutation evolution in near real-time
NSS
NC
GUINEASIERRA LEONE CLUSTER 1
SIERRA LEONE CLUSTER 2
SIERRA LEONE CLUSTER 3
NP VP35 VP40 GP VP30 VP24 L POLYMERASE
14
15
0
5
10
15
May 26 Jun 02 Jun 09 Jun 16Date in 2014
# of
new
pat
ient
s
SL1SL2SL3
15!
New mutations fall in geographic and temporal patterns
Kissi Teng
Nongowa
Kissi Tongi
Luawa
Mandu
Kpeje West
Malema
Kpeje Bongre
Jawie
Kissi Kama
Njaluahun
BoKenema
Kailahun
Kambia
Kono
SL1SL2SL3
Geneticcluster
# patients
1 32
GUINEA
LIBERIA
KissiTeng
Jawie
Mambolo
KakuaMay 31
June 15
Chi
efdo
m
?
June 17June 7
unknown
HCW-ADriver
HCW-B
GUINEA
LIBERIA
Kissi Teng
Jawie
Mambolo
Kakua
KGH
Kissi Teng
Nongowa
Kissi Tongi
Luawa
Mandu
Kpeje West
Malema
Kpeje Bongre
Jawie
Kissi Kama
Njaluahun
BoKenema
Kailahun
Kambia
Kono
SL1SL2SL3
Geneticcluster
# patients
1 32
GUINEA
LIBERIA
Intrahost variation elaborates transmission epidemiology
New mutation is rapid and random during outbreak
2014 outbreak
All of EBOV
Po
ste
rio
r p
rob
ab
ility
Subs. rate (site-1xyear -1
x10-3)
0 1 2 3 4
0.0
0.1
0.2
0.3
0.4
Faster rate of change
0
20
40
60
80
May 26 Jun 02 Jun 09 Jun 16
Cum
ulat
ive
tota
l
Patients
Date in 2014
Unique mutationsUnique lineages
Continually increasing viral diversity
NSS
136
559
55
79
75
110
p = 3.0e-07p = 1.2e-8
Betwee
n outb
reaks
With
in ou
tbrea
ks
With
in ho
sts
1.00
0.75
0.50
0.25Fr
actio
n
Enriched for non-synonymous SNPs
Largest, deepest dataset from an Ebola outbreak
Complete, reliable genomes
HIV-1
Human enterovirus A
Aphid lethal paralysis virus
GB virus C
Lassa virus
Plasmodium
Suspected EVD Patients
A
EBOV conf.
10
20
% m
alar
ia in
fect
ion
B
EBOV susp
.
Find any and all human pathogens
CommunityHospital/ Lab
Internationalcollaborators
Patient samples
Increases accuracyof diagnosis
Sample collection
Local diagnosis
Refined diagnostics:cheaper/field deployable
High throughput genomicanalysis and serology
The impact of genomic surveillance
CommunityHospital/ Lab
Internationalcollaborators
1. Lab practices 2. Lab design 3. Researcher training
BL3
BL2+
BL1
Safety consideration!(for both field and advanced labs sites)
Blood
Inactivated plasma
Nucleic acid
CommunityHospital/ Lab
Internationalcollaborators
Lab Practices
• PPE additions: Coveralls and face-shields are required at all times, in addition to the usual PPE of double gloves, booties, gowns and N95 face masks.
• All absorbent paper has been removed - except what is needed for the ELISAs.
• All surfaces are sprayed with bleach and ethanol every evening and again in the morning.
• All material is discarded into bleach – including all tips, tubes and blood collection tubes.
• A disposable glove box has been installed; confirmed (or highly suspected) EBOV samples can thus be handled in complete isolation.
• A third biosafety cabinet has been installed in the main lab so all ELISA plate washing and reading equipment can be contained.
• Standard protocols have been posted in the lab, and personnel have been trained to help maintain biosafety.
Enhanced safety strategies:
CommunityHospital/ Lab
Internationalcollaborators
Kenema Government Hospital Laboratory
Ebola ward, Kenema, Sierra Leone
Lab Design
CommunityHospital/ Lab
Internationalcollaborators
Irrua Specialist Teaching Hospital Laboratory, Nigeria
RNA Extraction&
PCR set-up
Mater Mixset-up
Sample Processing&
Inactivation
Research&
Sample Storage
Work Room
Storage Room
PCR RoomBL3 lab space
Office
Patient Sampling
Room
StaffRoom
Restroom
Restroom
Gel-electrophoresis&
Imaging Room
Converted tostorage
Gel
imag
ing
Sink Fridge
Gel running stationGel prep area
PCR machinesqPCR
AnteRoom
Sinkbenchtop
clean hoodnot vented
Frid
ge
blood chemistrymachine
buck
etce
ntrif
uge
Lab coats
SinkExtraction Area
Freezer
Vacufuge Centri-fuge
Thermomixer
Centrif.Chemicalstorage
under counter
A/C
Freezer
Mas
ter m
ix a
dditi
on Fridge
Freezer
Sam
ple
colle
ctio
n ki
t as
sem
bly
and
pick
-up
Diss
ican
t bo
x
Storage
Sink
Sink
Fridge Freezer
Freezer
LN2Freezer
Comp-uter
SampleLabel printer
Mic
rosc
ope
Mal
aria
Dia
g.
Freezer
FreezerHallway
LN2Freezer
Freezer
AnteRoomExit
ELIS
A st
atio
n
Mobile Glovebox
Incu
bato
r
Fire Exit
BL3 Sample Storage
RNA Extraction&
PCR set-up
Mater Mixset-up
Restroom
Sample Processing&
Inactivation
Research&
Sample Storage
Work Room
Storage Room
PCR RoomELISA
Office
Restroom
Patient Sampling
Room
StaffRoom
Restroom
Gel-electrophoresis&
Imaging Room
Converted tostorage
Gel
imag
ing
Sink Fridge
Gel running stationGel prep area
PCR machinesqPCRCloset
AnteRoomcoat and shoe
area
Autoclave
Sinkbenchtop
clean hoodnot vented
Biosafety HoodFr
idge
Incubator
ELISA station
blood chemistrymachine
buck
etce
ntrif
uge
Lab coats
SinkExtraction Area
Freezer
Vacufuge Centri-fuge
Thermomixer
Centrif.Chemicalstorage
under counter
A/C
Freezer
Mas
ter m
ix a
dditi
on Fridge
Freezer
Sam
ple
colle
ctio
n ki
t as
sem
bly
and
pick
-up
Diss
ican
t bo
x
Storage
Sink
Sink
Fridge Freezer
Freezer
LN2Freezer
Comp-uter
SampleLabel printer
Mic
rosc
ope
Mal
aria
Dia
g.
Water distillationmachine
Freezer
Freezer
-80 Freezer
Fire Exit
Hallway
A/C
Current Laboratory Space Proposed changes to Laboratory Space
BL3 designated spaceBL1 designated space
AnteRoomEnter
-80 Freezer
A/C A/CA/C
A/C
A/C
Biosafety Hood
RestroomRestroom
Chimney
Fire Exit
Freezer Room/Storage
Water distillationmachine
Water distillationmachine
Autoclave
Irrua, Nigeria
Lab Design
CommunityHospital/ Lab
Internationalcollaborators
Genomics Laboratory, Redeemer’s University, Nigeria
CDC Resource Needs Assessment
Lab Design
CommunityHospital/ Lab
Internationalcollaborators
Training ACEGID 2014 Summer Genomics Training - Harvard/Broad
7-week training program Focus:• Microbiology theory • Genomic diagnostic tools
• PCR, deep sequencing • Bioinformatics • Teaching pedagogy • Biosafety
CommunityHospital/ Lab
Internationalcollaborators
ACEGID 2014 Summer Genomics Training - Harvard/BroadTraining
Training-of-Trainers Courses Developed:
• Foundations of Genomic Technology • Lassa Diagnostics: Clinical
applications of molecular technology • Foundations of PCR • Intro to Molecular Cloning
Biosafety training
CommunityHospital/ Lab
Internationalcollaborators
ACEGID 2014 Summer Genomics Training - Harvard/Broad
• General lab safety at Harvard • BL3 field site training at Broad • Lecture: Ara Tahmassian, Chief Research
Compliance Officer • Glovebox tutorial • NIEDL BL-4 lab tour
Training
Expanded scientific capabilities to enable sustainable, collaborative and independent health research
African Centre of Excellence for Genomics of Infectious DiseaseK^]^^f^k�l�Ngbo^klbmr%�Gb`^kbZ
Afr
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Centre of Excellence for
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Genomics of Infectious D
iseas
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The Goals of ACEGID
Thanks to our partners
- Genomic Center for Infectious Diseases - Sequencing Platform - Technology Labs- Center for Systems Biology
- Department of Organismic and Evolutionary Biology
- Kenema Government Hospital - University of Sierra Leone - Sierra Leone Ministry of Health and Sanitation
African Center of Excellence for Genomics of Infectious Diseases
Afr
ican
Centre of Excellence for
K^]^^f^k�l�Ngbo^klbmr%�Gb`^kbZ
Redeemer s ni ersit i er
ia
Genomics of Infectious D
iseas
es
Gaps and Challenges Remaining* Potential Research Contributions
Coordination of the outbreak Single or multiple outbreaks
Financing Donations, Sustainable testing
Communication Open-access data release
Cross border collaboration Pathogen origin location
Logistics Diagnostic-Centered Response
Case management Pathogen origin date
Infection control Containment decisions
Surveillance Strain-specific diagnostics
Contact tracing Transmission chains
Community participation Improved public trust
… and Research
* As identified at the Emergency Ministerial meeting on Ebola Virus Disease Accra 2-3 Jul 2014 http://www.afro.who.int/en/media-centre/pressreleases/item/6695-health-ministers-agree-on-priority-actions-to-end-ebola-outbreak-in-west-africa.html
The Importance of Viral Diagnostic Research in Outbreak Settings
Project Goals (1) Develop African research capacity in genomics by building a critical mass of well-trained scientists;
(2) Empower African researchers to utilize genomics-based tools towards the control and elimination of infectious diseases;
(3) Create genomics curricula to support and promote cutting-edge genomics-based research; and
(4) Engage communities in prevention efforts and public health education.
!
World Bank funded Project
African Center of Excellence for Genomics of Infectious Diseases
Afr
ican
Centre of Excellence for
K^]^^f^k�l�Ngbo^klbmr%�Gb`^kbZ
Redeemer s ni ersit i er
ia
Genomics of Infectious D
iseas
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Foundational training 2-month training for ACEGID members at Harvard/Broad Institute Training program planned for June-August 2014 in Cambridge, MA, USA !
On-site Workshops Annual 2-week trainings at each of the 4 ACEGID sites for larger scientific
community !
Advanced training Annual multi-week advanced training for PIs held at Broad Institute
ACEGID Action Plan for Learning Excellence
Northwest Labs, Harvard Cambridge, Massachusetts, USA
Education
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Training%focus:%• Laboratory%procedures%• Molecular%biology%theory%• Lab%Safety%• Science%communicaAon%
Broad&Pilot&Training&Programs&
1%
Education
Year 2-4 Design and Construction of new Genomics Center at Redeemer’s 1. Genomics Teaching Labs 2. Genomic Diagnostic Research Labs 3. Core scientific production facility 4. Administration and guesthouse
Design Group
GHESKIO Tuberculosis Hospital, Port-Au-Prince, Haiti
Umubano Primary School, Kigali, Rwanda
ACEGID Infrastructure