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UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific
Leptospirosis Surveillance by Filtration Sampling and Next GenerationSequencing in Okinawa, Japan
CPT Sean Kluckman, P.E.
CPT Sean Kluckman, [email protected] 1 of 16
HUMANENVIRONMENT
ANIMAL
ONE HEALTH
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 2
OVERVIEW
• Public Health Command – Pacific • Leptospirosis Overview• Leptospirosis Case Study
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 308SEPT2015
Public Health Command-Pacific
Human HealthMedical Entomology
Preventive Medicine Doctors
Animal HealthVeterinary Care
Environmental HealthEngineering ServicesLaboratory Services
Leptospira Outbreak
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 408SEPT2015
Public Health Command-Pacific
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 508SEPT2015
Leptospirosis
Victoriano et al. BMC Infectious Diseases, 2009
• Multiple strains of disease-causing Leptospira that are endemic to all tropical and subtropical regions.
• Outbreaks have been linked to heavy rainfall events.
• Transmitted through contact with water, moist soil and vegetation contaminated with the urine of infected animals.
• Symptoms for humans and dogs, resulting in fever, kidney failure and can cause death if untreated.
SuspectedHost
(Mongoose)
Infected Military
Working Dog
Infected Military
Personnel
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 608SEPT2015
Leptospirosis Case Study
Trigger Event – September 2014• A long hot dry period followed by heavy rainfall
occurred prior to a water training exercise.
• Marines present with the symptoms of “Leptospirosis”.
• Loss of a Marine Military Working Dog (MWD) with presumptive Leptospirosis diagnosis.
Navy Times, 03 NOV 2014
Stripes Japan, 09 OCT 2014
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 708SEPT2015
Leptospirosis Case Study
Initial Response
United States Air Force• Collects Water/Tissue
Samples from Nuisance Wildlife (MONGOOSE)
United States Army• Analyzes Water and Tissue
Samples• Conduct Site Evaluation
United States Marine Corps• Provided medical oversight
OKINAWA
Presumptive
Samples
CAMP ZAMA
Team
Dispatc
hed
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 8
Leptospirosis Case Study
Problems Encountered
Raw water was sediment laden and clogged the instruments.
Polymerase Chain Reaction (PCR) Sequencing showed that all samples were negative for Leptospira.
Military Working Dogs do not show symptoms of being infected until the late stages of the infection.
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 908SEPT2015
Public Health Command-Pacific
Human Health
Animal Health Environmental Health
Leptospira Outbreak
Response TeamEntomologistsMicrobiologistsEngineersVeterinariansDoctors
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 1008SEPT2015
Leptospirosis Case Study
Turbidity Drinking Water
0.3 NTUs
Raw Water 800 NTUs
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 118SEPT2015
Leptospirosis Case Study
Water Source
PumpFilter
Waste
Option 1: Envirocheck Method Option 2: Innovative Disk Filter System
DNA Extraction
Water Source
Trap Filter(10/2µm pore)
Analytical Filter (0.45µm pore)
Waste
DirectDNA Extraction
Pros- Established method- Commercially available
Cons- High equipment cost- High consumable cost- Requires electricity- Clogs easily
Pros- Low cost- Light weight- No electricity- Rugged- Easy to Use
Cons- Undeveloped- Untested
100mL
140 ml syringe for delivery of 100ml of sample
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 1208SEPT2015
Leptospirosis Case Study
Next Generation Sequencing16S rDNA Amplicon Sequencing
Conventional Polymerase Chain Reaction (PCR)
• Rapid ID of specific threat• Prior sample knowledge• All Samples Negative (Possibly Due to Sensitivity)
• Rapid ID of ALL threats• No prior sample knowledge• 3 variations of pathogenic Leptospira
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 1308SEPT2015
Leptospirosis Case Study
Recommendations
Administer Leptospirosis vaccine for all Military Working Dogs.
Continue monthly sampling to try and determine a correlation between Leptospira detection and environmental conditions.
Develop a formal kit for filtration-based water sampling.
The daily number of hourly observed precipitation reports during the last 12 months (2014), color coded according to precipitation type, and stacked in order of severity. From the bottom up, the categories are thunderstorms (orange); heavy, moderate, and light snow (dark to light blue); heavy, moderate, and light rain (dark to light green); and drizzle (lightest green). Not all categories are necessarily present in this particular graph. The faint shaded areas indicate climate normals. The bar at the top of the graph is green if any precipitation was observed that day and white otherwise.
Max. Normal Precipitation
http://weatherspark.com/history/33303/2014/Okinawa-Okinawa-Prefecture-Japan
2014 Potential Predictive Environmental Cue
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 1408SEPT2015
Leptospirosis Case StudyConclusions
Worked as an integrated team to identify and provide solutions to the Leptospirosis outbreak.
Develop a water filtration device and incorporated Next Generation Sequencing to evaluate samples.
Recommendations apply to similar training environments around the Pacific.
Provided recommendations to allow training to continue.
ChemoprophylaxisVaccination of MWDsEducation and Awareness
UNCLASSIFIED
UNCLASSIFIEDPublic Health Command - Pacific AS OF: Slide 1508SEPT2015
Questions
HUMANENVIRONMENT
ANIMAL
ONE HEALTH