Plague: Out of the Foothills

1. Plague: Out of the Foothills Rebecca E. Colman1, Robert J. Brinkerhoff2, Adina Doyle1, Chris Ray3, Paul Keim1, Sharon K. Collinge3, and David M. Wagner1 1Northern Arizona University, Flagstaff, AZ; 2Yale University, New Haven, CT; 3University of Colorado at Boulder, Boulder, CO

2. Background • Caused by the bacterium Yersinia pestis • Obligate pathogen • Cycles between rodents and their associated fleas • Endemic in several foci around the world

3. Plague in North America • Entered ~ 1900 at a California shipping port • San Francisco area, maybe L.A. • Characterized by large die-offs in rodent populations • Increased human risk • Mechanism for maintenance unknown • Several theories but how it is sustained in North America is not clear

4. So what is the reservoir? • NSF-NIH: Landscape Effects on Disease Dynamics in Prairie Dogs; 2002-2007; University of Colorado; $1,832,715 • NSF-NIH: Ecological Drivers of Rodent-borne Disease Outbreaks: Trophic Cascades and Dispersal Waves; 2003-2009; University of New Mexico; $1,746,268 • NSF-NIH: Plague As a Model for Low Prevalence/ epizootic Disease Dynamics; 2003-2009; Colorado State University; $1,281,000 • TOTAL = $4.8 million

5. Plague in the US • Was introduced into the US ~1900 and rapidly spread to the current distribution • Has become endemic in the Southwest Cully and Williams 2001

6. Cycle: Enzootic / Epizootic (Enzootic) (Epizootic)

7. Study System: Boulder Colorado Colorado Sites with Y. pestis positive samples from June to September 2005

8. Study Questions Does plague come out of the Foothills “reservoir” hosts and infect the prairie dogs in the grasslands? Or is there another local reservoir that spills over into the prairie dog colonies?

9. Hypothetical Molecular Trees Out of the Foothills hypothesis Other local reservoir hypothesis A F B E C D D C E B F A

10. Collection Methods • Rodents were trapped and combed for fleas • DNA from individual fleas was extracted • Extractions were screened with 2 PCR targets • Plasmid target (pla) and chromosome target • 78 samples were used for high resolution genetic analysis • 150 samples were used for more ancestral genetic structure

11. Different Molecular Markers Fast Slow Mutation Rate 10-3 10-9 10-10 10-4 10-5 10-6 10-7 10-8 Complex VNTRs SNPs SNR / VNTRs

12. MLVA Insertion Deletion Multi-Locus Variable number tandem repeat Analysis

13. MLVA Examines 43 regions of Y. pestis genome with varying mutation rates Provide discrimination even among local outbreak samples

14. MLVA Results • Shows population structure by site • Interesting 5A dispersal event • Reservoir sites are more genetically similar to each other than to the prairie dog sites except for 5A

15. MLVA Results

16. SNPs • Single Nucleotide Polymorphisms • Slow mutating markers compared to MLVA, therefore they are more stable markers • Show more ancestral (historical) population structure

17. SNP Results Northern AZ Colorado • Whole Genome Sequence comparisons • SNPs are extremely rare Y. pestis • Show historical divergence • Samples fell out into two SNP groups

18. SNP and MLVA combined

19. Conclusion • Geographic proximity does not equal genetic similarity

20. Implications • Reservoir sites are distinct from each other based on MLVA • Suggests plague is cycling between epizootics with no real gene flow • Two SNP groups seen in one study season • Suggests separate maintenance and dispersal of this disease on small spatial scale

21. Implications • SNP group 2.2 (blue) – plague comes out of reservoirs (in the foothills) and infects prairie dog colonies (5A) • However the other 2 prairie dog sites (MK and CR) are distinct and must have come from another introduction event • Maybe out of the grassland habitat?

22. Acknowledgements A collaboration with Sharon Collinge’s group at UC Boulder Work supported by NIH grant #1R15-AI070183-01

23. Questions ?