1. Evolution and Ecology of Pathogens Martin Polz Civil & Environmental Engineering Massachusetts Institute of Technology

2. Outline Emergence of pathogens Emergence of pathogens Global importance of microorganisms Global importance of microorganisms What are pathogens? What are pathogens? Evolution of pathogenesis Evolution of pathogenesis Re-emergence of pathogens Re-emergence of pathogens Antibiotic resistance Antibiotic resistance Cholera Cholera Conclusions Conclusions

3. Germs, germs everywhere Even on a little pear, Germs germs all around, Even on the dirty ground. Germs, germs make me sick, Especially on a candy stick. Germs, germs are so small, Even on a bouncy ball. Candice's Germ Poem

4. Microbial communities drive biogeochemical cycles…

5. 100 human activities 140 biological fixation 200 denitrification SOIL ATMOSPHERE OCEANS 15 biological fixation 140 denitrification 1200 internal cycling 8000 internal cycling 10 burial 36 river flow <3 fixation in lightening groundwater Example: The Nitrogen Cycle

6. ? human activities 140 biological fixation 200 denitrification SOIL ATMOSPHERE OCEANS 15 biological fixation 140 denitrification 1200 internal cycling 8000 internal cycling ? burial ? river flow <3 fixation in lightening groundwater Nitrogen Cycle Without Microbes All processes slow. Would life be possible?

7. Microbes Bacteria Fungi Protists Viruses

8. Bacteria Small Efficient Biochemically diverse Fast growth

9. Marine Freshwater Sediments Subsurface sediments (0-3,000 m) Animal guts Cells/ ml or g x10 6 Total cells x10 26 0.5 1.0 4,600 0.34-200 1-10 5 1,000 1.5 170 38,000 0.0004 (Whitman et al. 1998) Bacteria are everywhere

10. Global bacterial biomass (Pg of C) Soil and Aquatic PlantsBacteria Subsurface Terrestrial Marine 560 1.8 26 2.2 22-215 303 Microbial biomass rivals plant biomass but has higher turnover

11. How many bacterial species are there? Wilson 1988 Total number species: ~ 1.4 million Bacteria: ~3,500 Hammond 1995 Total number species: ~ 11 million Bacteria: ~10 million

12. The great plate count anomaly microbial community plating DAPI stained marine water sample < 1% of observable bacteria grow on standard culture media

13. Genetic diversity Total nucleic acids 16S ribosomal RNA genes Sequences Diversity and evolutionary relationships Identification and quantification

15. Molecular approach: great diversity of microbes pathogens only a minor component of microbial diversity allows understanding of evolution of pathogenesis

16. Emergence of pathogens

17. What is a pathogen? An evolutionary view. Example: Escherichia coli (E. coli) Normally a harmless gut bacterium but… Eterotoxigenic strains Enteropathogenic strains Enteroinvasive strains Enterohemorrhagic strains Enteroaggregative strains Uropathogenic strains

18. Genome analysis provides answer Strains closely related Genome structure similar But…. Insertions of ‘foreign’ DNA = pathogenicity islands Comparative analysis:

19. Comparison harmless and pathogenic E. coli strains AB AB C C E. coli K12 E. coli O157:H7 Foreign DNA = locus of enterocyte effacement Responsible for pathogenicity: allows attachment and toxin productions A harmless bacterium has become a pathogen by ‘stealing’ DNA from another bacterium!

20. Mechanisms of gene transfer: 1 Transformation: uptake of DNA from environment 2 Transduction: DNA transfer by viruses 3 Conjugation: plasmid transfer between bacterial cells 1 2 3 Can all transfer genes from other bacteria that can become incorporated into genome

21. Fate of transferred genes: RecA system = recombination into genome dependent on sequence similarity % sequence difference recombination rate

22. How often does gene transfer happen? Gene transfer is rare e.g., transduction by viruses insert foreign DNA every 10 8 virus infections But…. Microbes have very large populations e.g., gene transfer in marine environment ~20 million billion times per second! Genes must be advantageous to recipient….

23. Ecology of pathogenesis Bacteria grow fast High population densities Great competition for resources Pathogen = normal bacterium that has gained access to a new resource through new genes --> Competitive advantage

24. Re-emergence of pathogens

25. Example 1: Antibiotics Antibiotics - natural warfare common resource species 1species 2

26. Example 1: Antibiotics Antibiotics - natural warfare common resource species 1species 2 antibiotic

27. Example 1: Antibiotics Antibiotics - natural warfare common resource species 1species 2 antibiotic

28. Example 1: Antibiotics Antibiotics - natural warfare common resource species 1

29. Antibiotic resistance Bacteria have evolved resistance genes to antibiotics Located on plasmids

30. Plasmid encoded resistance is easily transferred between species because plasmids are mobile Occurrence usually low unless selection through widespread antibiotic use

31. Antibiotics overuse creates ‘Superbugs’ 50 million tons antibiotics per year ‘Superbugs’ resistant to most antibiotics Example: Tuberculosis 2.5 million deaths Mycobacterium tuberculosis increasingly resistant

32. Example 2: Cholera and climate Vibrio cholerae and other vibrios ubiquitous in marine, coastal waters Genetically similar non-pathogenic and pathogenic strains co-exist V.cholera

33. Vibrio species identified as agents of human disease

34. Seasonal cholera in Calcutta (Sharma, 1998)

35. Vibrio infections linked to El Nino Dhaka, Bangladesh Cholera cases Seasonality Removed (Pascual, 2000)

36. Attachment to algae and zooplankton? Temperature dependent growth? Possible reasons for seasonality Algal growth = vibrio growth? Temperature rise = vibrio growth? Links to global warming and/or pollution

37. Conclusions

38. Re-Emergence is an evolutionary/ecological phenomenon Microbial communities extremely diverse Large numbers of individuals Potential for gene transfer Pathogenesis arises via gene transfer Result: harmless bacterial species becomes pathogen because it gains competitive advantage Ecological factors (resistance, alternate hosts, climate) may trigger increased incidence of pathogenesis

39. Outlook for the future Need to understand environmental context of pathogenesis Need to understand gene transfer rates and diversity of co-occurring genomes

40. Thanks to: Silvia Acinas Dan Distel Dana Hunt Vanja Klepac Luisa Marcelino Chanathip Pharino Ramahi Sarma-Rupavtarm Janelle Thompson NSF, NIH, Seagrant, DOE - Genomes to Life