Environmental dependency of ranavirus/amphibian genotypic interactions

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2013 International Symposium on Ranaviruses …

2013 International Symposium on Ranaviruses
by David Lesbarreres

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  • When assuming no environmental influences, the phenotype of infection traits in a given host-parasite interaction (such as host resistance or parasite virulence) is expected to be determined by the host and the parasite genotypesFigure I. Infection phenotype of different hypothetical combinations of two host genotypes and two parasites genotypes. The two host genotypes (A and B) are arranged along the x-axis and each line represents one parasite genotype (parasite 1 genotype, black circles; parasite 2 genotype, white circles). (a) A main effect of parasite genotype is visualized by the vertical spacing between the two lines. (b) A main effect of host genotype is indicated by the positive slope of the lines, in addition to a vertical main effect ofparasite genotype. (c,d) Host genotype by parasite genotype interactions are suggested by non-parallel lines. In (d), note that main-effect components can cumulate with an interaction.Combinations of host genotype A and B by parasite genotype interactions in two environments. In any given environment, parasite fitness is higher in hosts with an exact genetic match. A different environment (environment 1 and 2) might cause changes in host resistance for its matched parasite (GH E) or parasite fitness in its matched host (GP E). A complete reversal of host–parasite genetic compatibility (GH GP E) might also occur. Parallel lines would indicate the absence of interactions. The environment might also change the fitness of all parasites on matched hosts (E).
  • Objectives: interaction between two common North American frogs species (Lithobatespipiens and Lithobatessylvaticus) and 3 strains of their deadly pathogen Ranavirus in two temperature settings in order to assess the potential for GxGxE interactions.1) Wild type (wt) Frog Virus 3 infecting frogs, including Lithobatespipiens, and which is expected to be the most virulent. 2) An azacytidine (azaC)-resistant mutant, that is thought to be less virulent because its unmethylated genome may trigger an early innate immune response (Essani et al. 1987). 3) SsMe, isolated from a spotted salamander in Maine, USA.14 and 22 degrees
  • Tissue Culture Infectious Dose50
  • Interesting: it seems that tadpoles infected by Azac (less virulent) are growing faster than controls in LF both in cold and warm environments. May indicate 2 things:1-Infection stimulates ressource allocation to growth, reflect eventually the strategy escape instead of fight in the immune/growth trade-off, 2-that below a certain treshold of virulence, tads can allocate more ressource to growth than they do with more virulent strain such as Wt.
  • Overall seems the most virulent strain, particularly for LFOverall Azac seems to be the less virulent which was expected since this mutant lack some receptor preventing it to infect host as efficiently as Wt. SsMeV seems to have an intermediate virulence.LF are more sensitive to cold than WFs are as they suffer drastic fitness depletion for a given trait in cold but less in warm, especially when infected by Wt.Significant differences between Species are noticable (LF vs WFs), differences genotypes within species are more subtle and contingent on the trait, the strain considered and the influence of temperature. Result on growth rate tend to indicate that infection might influence resource allocation in the context of a immune function/growth trade-off. At least for Azac (milder strain) host seems to choose to escape rather than fighting the infection.
  • The analysis revealed considerable variation in life history trait in response to Temperature, Species and Virus Strain factor interactionsThis form of GHxGPxE indicates variability in Strain virulence and species susceptibility suggesting the potential for coevolution based on frequency-dependent selection illustrating a coevolutionary Rubik’s cube This study shed the light on some of the underlying mechanisms that might explain amphibian ranavirus associated die-offs variability in the field and illustrate the relevance of using integrative approaches to understand host-pathogen epidemiology and coevolution.


  • 1. Pierre Echaubard, and David Lesbarrères Ranavirus II, Knoxville, TN Environmental dependency of Ranavirus/Amphibian genotypic interactions: a coevolutionary Rubik’s Cube
  • 2. (Lambrechts et al 2005) (Salvaudon et al 2005) (Carius et al 2001) (Lambrechts et al 2006) (P1) (P2) GH x GP indicate the potential for non trivial coevolution based on Frequency dependent selection (Wolinska and King 2009) GH x GP x E Reaction norm and Phenotypic plasticity On the importance of H-P genotypic interactions in understanding coevolution
  • 3. *** L > H • Density is detrimental Pollution Competition for resources • Virulence is context dependent Visible effects in low density only a a b b a a b b a a a a a a a a Context-dependent effects Echaubard et al. 2010 PLoS One
  • 4. Investigation of GHxGPxE Hosts Parasites Environments Wt FV3 Azac Ssme Objectives
  • 5. WtFV3 AzacFV3 SsMV Control 14 °C Life history traits: -Size -Time to metamorphose -Growth rate -Mortality rate 22 °C 2 1 1 WtFV3 AzacFV3 SsMV Control Infection: 1.0x104 pfu/ml)
  • 6. Temperature (F1,369 =5.422; p=1.07x 10-7) 0 0.05 0.1 0.15 0.2 0.25 0.3 COLD WARM Weight(g),±SE COLD WARM *** TEMP effects are SPECIES specific TEMP affects Weight at metamorphosis Temp*Species*Strain (F23,3369=7.584; p=2.2x 10-16) 0 0.05 0.1 0.15 0.2 0.25 0.3 0.35 0.4 0.45 0.5 Azac Control SsMeV WtWeight(g),±SE COLD LF COLD WF1 COLD WF2 *** a a a a aa a a a b b b 0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 Azac Control SsMeV Wt Weight(g),±SE WARM LF WARM WF1 WARM WF2 a b a a a aa aa a a b *** *** *** 0 0.1 0.2 0.3 0.4 0.5 LF WF1 WF2 Weight(g),±SE COLD WARM b a a a aa *** Temp*Species (F5,387 =21.98; p=2.2x 10-16) STRAIN effects are conditional of both TEMP and SPECIES -No difference between virus Strains in cold temperature but strong effect of Wt in Warm -No difference for Wood frogs but strong effect of Wt in Leopard frogs Weight at metamorphosis
  • 7. 0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 1 2 3 4 5 6 7 8 9 10 Cumulativemortalityrate(%) Time period WARM 0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 1 2 3 4 5 6 7 8 9 10 Cumulativemortalityrate(%) Time period COLD Mortality
  • 8. 0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 1 2 3 4 5 6 7 8 9 10 Cumulativemortalityrate(%) Time period WARM 0 0.1 0.2 0.3 0.4 0.5 0.6 0.7 0.8 0.9 1 1 2 3 4 5 6 7 8 9 10 Cumulativemortalityrate(%) Time period COLD Significant Effects 1. Temperature (Warm)(t =-2.33 ; p=0.021) 2. Time(t =-3.553 ; p<0.0001) 3. TempWarm:WF1(t =-3.553 ; p<0.02) 4. TempWarm:WF2(t =2.165; p<0.03) 5. WF2:Control (t =-4.188; p= 4.29x 10-5) 6. Warm:Time (t =-2.513; p= 0.013) 7. Warm:WF1:Control (t =-2.011; p= 0.045) 8. Warm:WF2:SsMeV (t =2.034; p= 0.043) 9. Warm:WF1:Wt (t =2.253; p= 0.025) 10. Warm:Wt:Time (t =2.798; p= 0.005) 11. Warm:WF2:Wt:Time (t =-1.808; p= 0.001) -Mortality is influenced by both TEMP (1) and TIME (2) - TEMP effect is different among SPECIES (3, 4) - Mortality is dependant of STRAIN (7, 8, 9) - TIME effect is dependent of TEMP (6), SPECIES and STRAIN (10,11) Repeated measure ANOVA, Bonferroni correction applied Conclusion/Interpretation
  • 9. 2 – Infection stimulates resource allocation to growth End point Growth Rate COLD WF Azac WF ControlLF Azac LF Control Trade-off immuno/development 1 - Interaction Species*Strain Growth Rate 3 - Below a certain threshold of virulence, allocation of more resources to growth than when confronted with a more virulent strain (Wt).
  • 10. Susceptibility Azac SsMeVWt Virulence Weight Growth RateMortality rate LF FLIGHT or FIGHT? Conclusions
  • 11. The analysis revealed considerable variation in life history trait in response to: Temperature Host Species Virus Strain GHxGPxE Amphibian / Ranavirus associated die-offs variability Integrative approaches for H-P epidemiology and coevolution. Take Home
  • 12. Work in progress -Pathogens co-occurrence and environmental variability -TCID50 at different temperatures -Strain variability and distribution deletion of 757bp of coding region