2013 International Symposium on Ranaviruses

1. Ecology and Epidemiology of Ranaviruses:
Mechanisms Contributing to Outbreaks
University of Tennessee
Center for Wildlife Health
Department of Forestry, Wildlife and Fisheries
Matthew J. Gray
M. Niemiller

2. Presentation Contributors
M. Brand, University of Tennessee
R. Brenes, University of Tennessee
J. Chaney, University of Tennessee
J. Earl, NSF NIMBioS
N. Hilzinger, University of Tennessee
J. Hoverman, Purdue University
R. Huether, University of Tennessee
A. Kouba, Memphis Zoo
D. Miller, University of Tennessee
P. Reilly, University of Tennessee
S. Roon, Oregon State University
B. Sutton, Clemson University
J. Tucker, Humboldt University
T. Waltzek, University of Florida
B. Wilkes, University of Tennessee
Unpublished Data

3. Ranavirus Ecology
Gray et al. (2009)
Gray et al.
(2009)

4. Global Distribution of
Ranavirus Cases: Amphibians
All Latitudes, All Elevations
14 Families: Alytidae, Ranidae, Hylidae, Bufonidae, Leptodactylidae, Dendrobatidae,
Discoglossidae, Myobatrachidae, Rhacophoridae, Scaphiopodidae,
Ambystomatidae, Salamandridae, Hynobiidae, Cryptobranchidae
5 Continents: 1992
Miller et al.
(2011)
>70 Species

5. Reported Ranavirus Cases in
North America: Amphibians
>30 States & 5
Provinces;
46 Species
Families
Bufonidae
Hylidae
Ranidae
Scaphiopodidae
Ambystomatidae
Cryptobranchidae
Plethodontidae
Salamandridae
Norman Wells,
NWT
Uncommon
Lithobates sylvaticus
2011

6. Cases of FV3-like Ranaviral
Disease in Reptiles
Over >95% homology with
1000-bp region of MCP
Gopherus polyphemus, Testudo hermanni, Terrapene carolina
carolina, Trionyx sinensis, Uroplatus fimbriatus, and
Chondropython viridis
(Westhouse et al. 1996; Marschang et al. 1999, 2005; Hyatt et al. 2002; DeVoe et al. 2004;
Huang et al. 2009; Allender et al. 2006, 2011; Johnson et al. 2007, 2008, 2011)
At least 14 reptile species Marschang (2011)

7. Cases of Ranaviral Disease
in Fishes
Ictalurus melas, I. nebuosa, Silurus glanis, Psetta maxima, Sander
lucioperca, Perca fluviatilis, P. flavescens, Oncorhynchus mykiss,
Pomoxis nigromaculatus, Gambusia affinis, Epinephelus tauvina
Journal of Fish Diseases
33:95-122
At least 20 fish speciesEHNV, ECV LMBV, SGIV

8. Species Challenges
FV3-like Ranaviruses

9. Single-species FV3-like Challenges
Amphibians
Percentmortality
0
10
20
30
40
50
60
70
80
90
100
FV3
RI
Hoverman et al. (2011): 19 Species Tested
High Moderate
Low

10. Single-species FV3-like Challenges
Amphibians
Ambystomatidae Ranidae
Brenes (2013)
15 Additional
Species

11. Life History and Phylogeny
Amphibians
P = 0.354 • Fast development hatching time*
• Low aquatic index
• Breeding habitat (temporal)*
• Breeding time (spring)
• Distance: Population & Isolate
Brenes (2013)
All Three Isolates
Smoky Mountains Isolate
No Phylogenetic Signal
35 spp
Co-evolution
High Correlation
(r = 0.88) between
Infection Prevalence
and Percent
Mortality

12. Single-species FV3-like Challenges
Chelonians
Mississippi
Map Turtle
Control Turtle Fish Amph
Soft-shelled Turtle
Brenes (2013)
Terrapene carolina, T. ornata, Elseya latisternum, Emydura krefftii , Trachemys scripta
Water bath exposure sufficient for transmission
with some species.
Greatest infection and morbidity with IP
infection or oral inoculation.
Ariel (1997), Johnson et al. (2007), Allender (2012), Gray et al. (unpubl. data)

13. Single-species
FV3-like & ATV Challenges
Fishes
Amelurus melas, Esox luscious,
Sander lucioperca,
Micropterus salmoides
Cyprinus carpio, Carassius auratus,
Lepomis cyanellus
Scaphirhynchus albus
No Transmission:
Low Transmission:
High Mortality:
Jancovich et al. (2001), Bang Jensen et al. (2011a)
Gobbo et al. (2010), Bang Jensen et al. (2009, 2011b), Picco et al. (2010)
Waltzek et al. (unpubl. data)

14. Single-species FV3-like Challenges
Fishes
Channel catfish
Control Turtle Fish Amph
Mosquito fish
Control Turtle Fish Amph
No Transmission: tilapia, bluegill and fathead minnow
Brenes (2013)

15. Reservoirs or Amplification Hosts?
FV3-like Ranaviruses
Low Mortality
(Subclinical)
Low Mortality
(Subclinical)
Low – High Mortality
(Subclinical & Clinical)
Reservoir
Reservoir or
Amplification
Reservoir

16. Can Interclass Transmission Occur?
Bandin & Dopazo (2011)

17. Evidence from the Wild
13 February
2012
26 of 31 Box
Turtles Die
from
Ranaviral
Disease
Larval
anurans and
salamanders
dead too
Farnsworth
and Seigel,
Towson U.
2008 – 2011
North Branch
Stream Valley
State Park

18. Evidence of Interclass Transmission
Bayley et al. (2013)
Pike-perch Iridovirus Common Frog
Tadpoles
Frog Virus 3
Pike
Pike-perch
Black Bullhead
Bang Jensen 2009, 2011;
Gobbo et al. 2010

19. Pallid
Transmission to Anurans
Gray et al. (unpubl. data)
85% 80% 95%
(35 – 70%) (5%) (95%)
5%
Gray Bull Wood

20. Transmission to Turtles
35%
45%
5%
Brenes Presentation
Gray et al. (unpubl. data)

21. Superspreaders
and Amplifying Species
Paull et al. (2012)
Frontiers in
Ecology and the
Environment
10:75-82
2012
Superspreading Individuals Amplification Species
Disease Hotspots
Susceptibility Contact RateShedding Rate Contact
Host Community Contact RatePersistence Dispersal
•Green et al. (2002)
•Petranka et al.
(2003)
•Harp and
Petranka (2006)
•Gahl and
Calhoun (2010)
•Uyeharaet al.
(2010)
•Brunner et al.
(2011)

22. Ranavirus Superspreaders
Reilly, Gray, & Miller (unpubl. data)
6 hrs cohabitation
3-day 103
PFU/mL
n = 10 tadpoles/tub
20/80 Rule: Superspreading

23. Exposure Order Matters
Brenes (2013)
Only Wood Frogs
Only Chorus Frogs
Only Spotted Salamanders
Control
n = 5 pools/trt
10 larvae/spp
Inoculated in Lab
103
PFU/mL FV3
60 days
Exposure
Treatments
Design
Wood Frogs 100%
43%
12%
Chorus Frogs
Spotted Salam
72%
3%
Wood Frogs
Spotted Salam
24%
18%
Chorus Frogs
Wood Frogs
Chorus Frogs 44%
Spotted Salam 6%
52%
16%
40%
Appalachian Community
Ecosystem Effects?
Disease-induced Trophic Cascades

24. Community Composition Matters
Brenes (2013)
Only Gopher Frogs
Only Chorus Frogs
Only Southern Toad
Control
n = 5 pools/trt
10 larvae/spp
Inoculated in Lab
103
PFU/mL FV3
60 days
Exposure
Treatments
Design
Gopher Frogs 100%
52%
34%
Chorus Frogs
Southern Toad
70%
58%
Gopher Frog
Southern Toad
32%
80%
Chorus Frogs
Gopher Frog
Chorus Frogs 78%
Southern Toad 76%
62%
62%
68%
Gulf Coastal Plain,
USA

25. Evidence of
Environmental Persistence
(Nazir et al. 2012)
•Soil: 13-22 d
•Soil: 30-48 d
(1) FV3, FV3-like
•PW (unsterile): 22-34 d
•PW (unsterile): 58-72 d
20 C =
4 C =
(T-90 Values)

26. Impacts of Stressors
Gray et al. (2009)

27. *
**
**
**
χ2
3 = 40.1 ; p<0.001
Hatchling – 3X > Embryo
Larval – 4X > Embryo
Metamorph – 5X > Embryo
Impacts of Development
Across Seven Species
ML Estimate:
Egg membrane
may act as a
protective barrier
Haislip et al. (2011)

28. Tree frog Chorus frog Wood frog Green frog
b
c
b
c
a
b
b
b
b
Kerby et al. (2011)
Anax increased
susceptibility to ATV
(A. tigrinum)

29. Competing Temperature Hypotheses
• Virus Replication Hypothesis
– Ranavirus replication increases with temperature
up to 32 C
– Caveat: Immune function in ectotherms also
increases with temperature
• Temperature Induced Stress Hypothesis
– Early Spring Breeding Species:
• Stressed by Warm Temp
– Summer Breeding Species:
• Stressed by Cold Temp
High Pathogenicity at Higher Temperatures
Pathogenicity is Species-specific and Related to Typical Water
Temperature Experienced During Tadpole Development
Bayley et al. (2013)

30. Impacts of Genetic Isolation
Pearman and
Garner (2005)

31. Factors Contributing to Emergence
Other Possible Stressors: Pesticide Mixtures, Nitrogenous Waste,
Endocrine Disruptors, Acidification, Global Warming, Heavy Metals
Pathogen Pollution:
Anthropogenic introduction of novel strains to naïve populations
(Cunningham et al. 2003)
•Fishing Bait
•Ranaculture Facilities
•Biological Supply Companies
•International Food & Pet Trade
•Contaminated FomitesPicco et al. (2007) Schloegel et al. (2009)
Anthropogenic Stressors:
1) Herbicide (Atrazine)
Forson & Storfer (2006); Gray et al. (2007); Greer et
al. (2008); Kerby et al. (2011)
ATV SusceptibilityA. tigrinum
2) Cattle Land Use: Prevalence Green Frogs and Tiger
Salamanders
Insecticide (Carbaryl)

32. Percentmortality
0
10
20
30
40
50
60
70
80
90
100
FV3
RI
Ranaculture isolate 2X more lethal than FV3
Risk of Pathogen Pollution
Majji et al. (2006), Storfer et al. (2007), Mazzoni et al. (2009), Hoverman et al. (2011)

33. Are Ranaviruses
Capable of Causing
Local Extirpations and
Species Declines?
0
50
100
150
200
250
1960
1963
1966
1969
1972
1975
1978
1981
1984
1987
1990
1993
1996
NumberofPopulations
Collins & Crump
(2009)
Muths et al.
(2006)

34. Traditional Theory
(Anderson and May 1979)
Extirpation is possible if:
Frequency Dependent
All Conditions
Met with
Ranavirus-
Host System

35. Evidence of Declines
Dr. Amber Teacher
Southeastern England
Animal
Conservation
13:514-522
1996/97 and 2008
Ranavirus (+)
populations
81% Median
Reduction A. Teacher
A. Teacher
Teacher et al.
2010
81%

36. Evidence of Re-occuring Die-offs
Dr. Jim Petranka
Tulula Wetland Complex, NC
Rescue Effect
Biological Conservation
138:371-380
Wetlands
23:278-2901998-2006
Recruitment at most
wetlands failed due
to ranavirus
Persistence Possible
from Source
Populations

37. Extinction Probability in 1000 years
Earl and Gray (unpubl. data)
Haislip et al. (2011)
Keith Berven

38. Female Population Size
Earl and Gray (unpubl. data)
Death of
Pre-metamorphic Stages
Matters!

39. Evidence of Rare Species Effects
Sutton, Gray, Miller & Kouba
Endangered Dusky Gopher Frog

40. Evidence of Rare Species Effects
Chaney, Gray, Miller & Kouba
Threatened Boreal Toad
Tadpoles Metamorphs
2 – 5 d
5 – 7 d

41. Commonality of Being Uncommon
Southeastern United States
Federally Listed:
Species of Concern:
Rana capito sevosa, Ambystoma cingulatum,
Phaeognathus hubrichti, Ambystoma bishopi
113 Species and 25 Genera Total
1) Alabama = 14 species (11 genera)
2) Arkansas = 25 species (12 genera)
3) Florida = 19 species (12 genera)
4) Georgia = 22 species (15 genera)
5) Kentucky = 22 species (11 genera)
6) Louisiana = 15 species (10 genera)
7) Mississippi = 18 species (12 genera)
8) North Carolina = 41 species (15 genera)
9) South Carolina = 19 species (13 genera)
10) Tennessee = 26 species (14 genera)
50% U.S.
If uncommon species are highly susceptible,
ranaviruses could have a significant impact
on amphibian communities.

42. Take Home Messages
Should we be Concerned?
•Ranavirus Die-offs have Global Distribution
•Ranavirus Prevalence can be High
•Ranaviruses Infect Multiple Amphibian Species with
Different Susceptibilities
•Community Composition Matters
•Interclass Transmission is Possible – Abundant Reservoirs
•Ranavirus Persistence is Long
•High Transmission: Breeding and for Schooling Spp.
•Anthropogenic Stressors and Pathogen Pollution
contribute to Ranavirus Emergence
Epidemiological Theory AND Initial Simulations Supports
the Premise that Ranaviruses Could Cause Local
Population Extirpations and Contribute to Species Declines

43. Host-Pathogen Community
Aeromonas
hydrophila
Ranaviruses
Batrachochytrium
dendrobatidis
Alveolates
Complex Epidemiology
Spatially Structured Breeding Sites
I(t)ijkl S(t)ijkl
IM(t)ijkl
EM(t)ijkl
i = species
j = age class
k = pathogen
l = wetland
P(Nt)il < 0
S(t|k)ijklI(t|k)ijkl
Metacommunity of
Hosts & Pathogens

44. Questions??
mgray11@utk.edu
865-974-2740
Photo:
N. Wheelwright