WHY IS SNAKE VENOM COMPOSITION SO VARIABLE?

Wolfgang WüsterBangor University, School of Biological Sciences, Bangor LL57 2UW, UK

What is snake venom?

Complex cocktails of numerous bioactive proteins

Variation in venom composition Occurs at all

taxonomic levels Within individual in

time Between individuals

within populations Between populations Between species &

above Can result in

profound clinical differences and treatment difficulties

Symptoms after Asiatic cobra bites

% with neurotoxicity

% with necrosis

Ratio neurotoxicity

: necrosis Luzon (Naja philippinensis)

97 %

7 %

14:1

NW Malaysia – (Naja kaouthia)

12 %

44 %

1:4

What causes variation in venom composition?

Long-standing controversy

Function of venom: overpowering and killing prey, ?defence, ?digestion

What causes variation in venom composition?

Long-standing controversy

Function of venom: overpowering and killing prey, ?defence, ?digestion

What causes variation in venom composition?

Long-standing controversy

Function of venom: overpowering and killing prey

Potential causes Selection for different prey

Evolutionary history

, ?defence, ?digestion

Electrophoretic profiles of individual Calloselasma rhodostoma

Daltry, Wüster & Thorpe, 1996

Association between venom composition and causal hypotheses in Calloselasma rhodostoma

Geographic distance (gene

flow)

Phylogeny Diet

Generalised composition No No YES

Daltry, Wüster & Thorpe, 1996

Variability of venom composition: Selection pressure or “Overkill”?

Variability of venom composition: Selection pressure or “Overkill”?

“Overkill-Hypothesis”: Snakes have far more venom than

required for killing their prey Hence no selective pressure for greater

lethality BUT: Lethality only known in white

mice!!! Some prey known to be resistant against

snake venoms

Crotalus oreganus - Pacific rattlesnakeSpermophilus beecheyi – California ground squirrel

Resistance against venom

Example: Pacific rattlesnake (Crotalus oreganus ) and ground squirrel (Spermophilus beecheyi)

Squirrels from rattlesnake areas display increased resistance to venom

Example: Crotalus oreganus – Pacific Rattlesnake

Venom quantity: ~ 90 mg dry venom LD50 in mice: 2.84 mg/kg Median lethal potential: 31.7 kg of mice• LD50 in ground squirrels: ~ 40 mg/kg =>

2.25 kg• => per 250g ground squirrel: ~ 10 mg

=> 9 ground squirrels

BUT:• Venom needs to be lethal in minutes, not

24 hrs• Reserves?

Overkill in snakes?

Sea kraitLaticauda

Moray eelGymonothorax sp.

Sensitivity of prey and non-prey moray eels to Laticauda sea snake venoms

SympatricGymnothorax

AllopatricGymnothorax

survived 45-75mg/kg

died after 0.1mg/kg

Heatwole & Poran, 1995

Evidence of the energetic cost of venom:

Increased metabolic rate of snakes after venom extraction

O2 consumption increased by ~ 11% after venom

extraction in 3 pitviper species (McCue, 2006)

Venom metering in snakes

Loss of venomous function in snakes feeding on undefended prey

Why fine-tune venom?

0

2

4

6

8

10

12

14

16

18

20

Medium Snakes Large Snakes

Small prey

Large prey

mg venom injected

Hayes et al., 1995

Venom metering in Crotalus oreganus

E.g., Aipysurus eydouxii Feeds exclusively on fish

eggs Venom of very low

toxicity Toxins non-functional Venom delivery

apparatus atrophied

Venom loss in snakes feeding on undefended prey

Saw-scaled Vipers (Echis) as a model system for studying snake venom variation

One of the main causes of snakebite mortality worldwide

Documented variability in venom composition, incompatible antivenoms

% fatality rates after Echis ocellatus bites

0

510

1520

25

E.ocellatusantivenom

E. carinatusantivenom

? ?? ?

Echis pyramidum

Echis coloratus

Echis carinatus

Echis ocellatus

Pook et al. 2009

Diet and venom activity in saw-scaled vipers (Echis)

T. Mazuch

Do saw-scaled viper venoms show evidence of adaptive evolution?

T. Mazuch Diet recorded from museum specimens

Diet of the four Echis species groups

E. coloratus

E. carinatus E. ocellatus

E. pyramidum

Arthropods

Vertebrates

Diet recorded from museum specimens

LD50 of 4 representative venoms estimated for Scorpio maurus

Functional significance: time to incapacitation and death after realistic venom dose

Do saw-scaled viper venoms show evidence of adaptive evolution?

T. Mazuch Diet recorded from museum specimens

LD50 of 4 Echis venoms on Scorpio maurus

µg

ve

no

m /

g s

corp

ion

n/s P < 0.001 P < 0.001 P < 0.05

Arthropod Scorpion feeding lethality

– –

++ ++

+ +

++ ++

– –

Origin ofarthropoddiet + arthropod-lethalvenom

Loss of arthropod diet + arthropod-lethal venom

Echis carinatus

Echis pyramidum

Bitis arietans

Echis coloratus

Echis ocellatus

0.92

1.00

1.00

Repeated co-evolution of diet and specific venom lethality => evidence of selection

Functional significance of specific venom lethality

More rapid prey death? Reduction of metabolic cost of venom?

Test: Injection of biologically realistic amounts of venom

(3 mg/g = 25-575 x LD50)

Record time to loss of coordination and death

E. p. leakeyi E. c. sochureki E. ocellatus E. coloratus B. arietans

Tim

e (

min

ute

s)

0

20

40

60

80

Tim

e (

min

ute

s)

Death

Loss of coordination

Staged encounters

Are these experiments biologically realistic?

Vipers and scorpions placed together Video recording Time 1st bite – beginning of feeding

Staged encounters

Results:

3 successful trials with E. c. sochureki

In all cases 1-2 further bites

Time bite - feeding: 40-48 Min.

Functional significance of venom variation in Echis

Greater lethality ≠ more rapid prey death!

Functional significance: Venom economy => reduced metabolic

expense? Facilitation of scorpion feeding for

juveniles?

How do they do it? The genetics of venom adaptation

Venoms consist of several multigene toxin families

How does gene diversity correlate with diet?

PIII/PIV SVMP

sub-classes

Serine proteases

Casewell et al., in prep.

Induce haemorrhage, inhibit platelet aggregation

Procoagulant, fibrinolytic, inhibit platelet aggregation

=> Link between diet, toxin gene diversity and toxin function

Conclusions: why is venom so variable?

Evidence of natural selection on venom composition – specific lethality to prey

Resistance in some prey => predator-prey arms races

Looking beyond lethality: economics of venom

Genetic mechanism in saw-scaled vipers: Importance of diversification of toxin

families

=> Extreme variation in venom composition at all levels

Cheers!

Thank you for your attention !