In 2014, Dr. Quave presented the keynote for the Frontiers in Science Public Lecture Series at Florida Atlantic University. Abstract: In the face of widespread multidrug resistance, new antibiotics are in high demand. In this talk, Dr. Quave discusses the ethnobotanical approach to drug discovery, and the critical role that natural products will play in the future of healthcare.

1. Anti-infective Drugs from Nature:
The next generation of antimicrobial therapy?
Cassandra L. Quave, Ph.D.
Assistant Professor
Emory University
Department of Dermatology (SOM)
Center for the Study of Human Health (ECAS)
E-mail: cquave@emory.edu
Website: http://etnobotanica.us/
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2. Clinical Relevance
• On the precipice of the post-
antibiotic era?
▫ 2M serious infections, 23k fatalities
linked to MDR infection in US
• Antibiotic pipeline nearly empty
▫ Last line therapies for Shigella and
Neisseria gonorrhoeae
▫ CDC Urgent threat level:
 CRE, Clostridium dificile, N.
gonorrhoeae
▫ CDC Serious threat level includes
drug resistant strains of:
 Acinetobacter, Camplyobacter,
VRE, MRSA, Pseudomonas
aeruginosa, Salmonella, Shigella,
Streptococcus pneumonia, TB
• We already face a high economic
burden for infectious disease (HAI’s in
US = $28.4-45 B)
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3. The Problem: Drug Resistance
Intrinsic vs. acquired
resistance: 17M new
biofilm infections per
year in US = 550k
fatalities

4. Staphylococcus aureus as a Pathogen
• Opportunistic pathogen
• Colonizes nasal passages of 30%
healthy adults in US
• Leading cause of:
▫ Bacteremia
▫ Sepsis
▫ Brain abscesses
▫ Medical device infections
▫ Skin and soft tissue infections
(SSTI)
• Commonly implicated in:
▫ Bone and joint infections
▫ Surgical site infections
▫ Pneumonia
▫ Endocarditis
• HA-MRSA vs. CA-MRSA
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5. The Current Approach
• More of the “same”
▫ bacteriostatic and
bactericidal agents
▫ Are we asking the right
questions???
• Sources of new drugs
▫ Chemical library screens
▫ Environmental samples
▫ Genome sequencing
and combinatorial
chemistry
▫ Animal proteins
▫ Plants and fungi
Natural products (or small molecules
derived from them) account for 75%
of antibacterial drugs!

6. Why look to plants?
• Natural botanical
products:
▫ Have rich structural diversity,
chirality, and extensive
functional group chemistry
 In other words, they are very
cool chemicals!
▫ Are likely produced by the
plant to fill a specific need
(i.e. defense against
pathogens)… there is
potential for efficacy against
human pathogens

7. Secondary Metabolites
• Organic compounds not
directly involved in basic survival
of the organism (growth,
development, or reproduction)
• In plants, these are used for:
▫ Defense against predation and
herbivory
▫ Competitive “warfare” with
other organisms in the
community
▫ Pollinator attractors
▫ Dispersal
▫ Responsible for plant colors,
flavors, and odors
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Tree creature: Lord of the Rings
Since plants are sessile, secondary metabolites
are critical in either attracting or deterring other
organisms as needed and can increase fitness of
the plant.

8. Deter other
plant species
from growing
nearby
Fight off
microbial
invasion/infection
Attract pollinatorsDefense against
herbivory
Secondary Metabolites

9. History of plants as a source of medicine
• Ancient tradition, dating to
prehistory
▫ Pollen samples found in cave
rich in medicinal species
 Yarrow, cornflower, hollycock,
bachelor’s button, ragwort,
grape hyacinth, and woody
horsetail
Shanidar Neanderthal cave
site, Iraq (60k-80k years b.p.)
Ephedra altissima Desf.

10. History of plants as a source of medicine
Hippocrates
Engraving by Peter Paul Reubens, 1638
Hippocrates mentions the use
of 300-400 medicinal plants in
his writings (ca. 460-370 B.C.)
Dioscorides De Materia Medica (reproduction in
Arabic - Spain, 12th-13th century)
Pedanius Dioscorides seminal work “De
Materia Medica” recorded medicinal
practices of ancient Greece and the Roman
Empire during the time of Nero (ca. 40-90
A.D.). Multiple uses of more than 600
medicinal plants are described.

11. History of plants as a source of medicine
• Many of these medical traditions persist today
worldwide in both indigenous groups and
popular culture:
▫ Ayurveda
▫ Unani
▫ Jamu
▫ Kampo
▫ Ancient Egyptian medicine
▫ Traditional Chinese Medicine
▫ Shamanism
▫ Native American traditions
▫ African traditional medicine
▫ Folk medicine
Unani Medicine
Painting by Mahaveer Swami
http://www.hinduonnet.com/TM can be found in every region of
the globe where humans live.

12. Ethnobotanical Approach to Drug Discovery
• Ethnobotany (from
ethnology, study of
culture, and botany,
study of plants) is the
scientific study of the
relationships that exist
between peoples and
plants.
• Ethnobotany is the
science of survival.
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Ethnobotanical-directed study of plants
used for infectious disease is often more
effective than a random approach.

13. The Ethnobotanical Approach

14. Study Sites in the Mediterranean

15. Mount Vulture – Basilicata, Italy

16. Southern Italy
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17. Truffle Hunting in Southern Italy
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18. Plant Collecting
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19. Plant Collecting
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20. Mountain Landscapes – NE Albania

21. Kukes District, Albania

22. 22

23. Local economy

24. Local Economy
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25. Strategy for New Drug Identification
Field-work & Lit. Review of Medicinal Plants for skin infections
Plant collection (bulk & voucher)/ Taxonomic identification
Process materials & prepare crude extracts
MIC/ MBC Cytotoxicity In vitro
bioassays
Bioassay-guided fractionation of active extracts
In vivo models Clinical trials
Isolation of active principle(s)
QSI Biofilm

26. • Layers of Consent:
▫ IRB
▫ Prior informed consent
▫ Country & community
agreements
▫ Plant permits
• Data Collection:
▫ Semi-structured
interviews
▫ Focus groups
▫ Participant-observation
• Biological Sampling:
▫ Voucher collection
▫ Bulk specimen collection
Methods: Ethnobotanical Research

27. Lulë balsami
Hypericum perforatum L., Hypericaceae
St. John’s Wort

28. Plantago spp., Plantaginaceae
Plantago lanceolata L.
Plantago major L.

29. Topical therapies
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Juglans regia L.

30. Evil-eye amulets of Albania

31. Zootherapy & Evil-eye Amulets

32. Methods: Plant Extraction
Dried 48-72 hrs
Vacuum-sealed
with silica packets
Pulverized with a
grinder

33. Methods: Plant Extraction
1:10 extraction in
95% EtOH for 72 hrs.
or boiled in water
for 30 minutes
Plant materials
separated from
extract with
vacuum
filtration
Solvent removed
under reduced
pressure with a
rotary evaporator

34. Methods: Plant Extraction
After freezing at -80°C,
extracts were
lyophilized
Dried extracts
were scraped
out and
weighed
DMSO added,
creating a stock
concentration of
10 mg/ml &
sterile filtered
(0.2 μm)

35. On Tipping the Balance…
We can not expect
antimicrobials to sterilize the
body, but to tip the balance
back in the body’s favor.
What threatens this: Drugs (including natural products) that confer
extreme selective pressure, quickly yielding resistance.
Example: Resistance to the broad spectrum antimicrobial Tea Tree Oil
(Melaleuca alternifolia Cheel, Myrtaceae) is easily acquired after repeat
exposure.
McMahon et al. 2007. Letters in Applied Microbiology 5:958-965

36. Example: Garlic
• Allium sativum L.,
Amaryllidaceae
• Inhibits communication and
toxin production in P.
aeruginosa
• Decreases hyphae formation
(budding in C. albicans)
• Improves response of C.
albicans to Amphotericin B in a
synergistic fashion
Allicin
IJAA, 33:258-63; FEMS Immun. & Med. Micro., 58: 161-8;
J. Applied Micro., 105: 2169-2177

37. Example: Cranberry
• Vaccinium macrocarpon Ait.,
Ericaceae
• >1 million women in US get UTI’s each
year
• Proanthocyanidins inhibit adhesion of
P-fimbriated E. coli to uroepithelial
cells
Howell. 2007. Mol. Nutr. Food Res. 51, 732-737

38. Why Target Pathogenesis & Virulence?
• Attacking microbial defense and offense
mechanisms makes them more vulnerable to
attack by both the host immune response and
existing antimicrobial therapies.

39. Biofilms and Intrinsic
Resistance
• 5-step Process:
1. Initial attachment
2. Irreversible attachment
3. Maturation I
4. Maturation II
5. Dispersion
• Uni- or Poly-microbial
• Heightened gene exchange
• Slow growth/metabolism
• Matrix presents a physical
barrier to host immune
response and antibiotic
therapy
James et al. (2008) Biofilms in chronic wounds. Wound Repair Regen. 16(1):37-44
Scanning electron micrograph demonstrating the presence of
mixed species biofilm in a chronic wound. Both cocci and bacilli
are seen embedded in an amorphous matrix characteristic of
biofilm formation.
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40. Models for Biofilm
Formation & Dispersal
microtiter plate
catheters in vivo
(Imaging with IVIS)
flow cells
catheters in vitro

41. Elmleaf Blackberry
• Traditional uses in S. Italy:
▫ Leaves: furuncles, abscesses,
and other skin inflammations
▫ Roots: hair loss
▫ Fruits: eaten fresh and in
marmalades
• One of 116 remedies related to
SSTIs and other topical
dermatological treatments
identified
• 168 extracts screened
• Anti-biofilm activity first identified
& published in 2008 and # 220
marked as possible lead
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Quave et al. Journal of Ethnobiology & Ethnomedicine. 2008. 4(5)
Quave et al. Journal of Ethnopharmacology. 2008. 118:418-428
Rubus ulmifolius Schott. (Rosaceae):
The source of the bioactive
composition “220D-F2”.

42. 220D-F2 is effective against all clonal lineages of S.
aureus, regardless of antibiotic resistance profile and is
nontoxic to mammalian cell lines.
Quave et al., PLoS One. 2012: 7(1)
Biofilm Prevention

43. Biofilm Inhibitor: 220D-F2
220D-F2 improves response to functionally distinct classes of antibiotics,
including daptomycin, clindamycin, vancomycin, and oxacillin.
Quave et al., PLoS One. 2012: 7(1)
Biofilm Treatment

44. Summary
• There is great promise in nature!
• Pathogenesis is an important drug target
• Our biofilm inhibitor improves antibiotic
activity
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45. Conclusions
Single drug/single
target,
Bacteriostatic
Bactericidal
New
Direction
for Drug
Discovery
Immune
adjuvants
Conjugation
Biofilms
Quorum
sensing
Efflux pumps
Synergy
Clinically
relevant
isolates
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46. Acknowledgments
• Collaborators: Dr. Andrea Pieroni (UNSIG, Italy), Dr. Brad Bennett
(FIU), Dr. Lisa Plano (Univ. of Miami), Dr. Michael Otto
(NIH/NIAID), Dr. Mark Smeltzer (UAMS), Dr. Cesar Compadre
(UAMS), Dr. Alex Horswill (Univ. of Iowa)
• All of the study participants in the Mediterranean
• Emory Mentors and Collaborators: Dr. Michelle Lampl, Dr. Dennis
Liotta, Dr. Jorge Vidal, Dr. Sharmilla Talekar, Dr. Bill Shafer, Dr.
Joanna Goldberg, Dr. John Varga, Dr. Effrosyni Seitaridou, Dr.
Maira Goytia, Dr. David Weiss, Dr. Phil Rather, Dr. Emily Weinert,
Dr. Effrosyni Seitaridou, Dr. Brian Pollack and Dr. Jack Arbiser
• Current Lab Team: Kate Nelson, Dr. James Lyles, Parth Jariwala,
Janessa Aneke, Sam Anderson, Boru Wang, Samir Hussaini,
Sandy Jiang, Michelle Paine, Matt Dorian, Sarah Meadows,
Paula Tyler, Ian Buller and Amelia Muhs
• National Institutes of Health, National Center for Complementary
and Alternative Medicine (PI: Quave)
▫ R01 AT007052
▫ F32 AT005040
▫ F31 AT004288
• Georgia Research Alliance
▫ GRA.VL13.C7 (Phase IA & IB)
• Center for the Study of Human Health
People
Funding Sources
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