1) The study aims to assess how predation risk influences the defensive chemical compounds in striped skunk spray. Specifically, it will compare the amounts of trans-2-butene-thiol and trans-2-butenyl thioacetate, two abundant noxious chemicals, in skunk populations facing different predation pressures. 2) Gas chromatography-mass spectrometry will be used to identify and quantify the two target chemicals in spray samples collected from skunks in areas of high and low mammalian and avian predation risk. 3) Three hypotheses are that skunks in riskier areas will have more variable spray potency, juveniles will have stronger spray, and bolder

1. Victoria Luce and Theodore Stankowich
California State University Long Beach, 1250 Bellflower Blvd., Long Beach, CA 90804
Abstract
Aposematism, in which an antipredator adaptation is advertised through warning
signals, has been shown to occur throughout the animal kingdom. While the ability
of striped skunks to deter predators using black and white fur to advertise their
noxious spray is well known, few studies have assessed what molecules make up
the spray, and none have attempted to assess how ecological factors can influence
the spray strength.
The goals of this study are to assess the validity of aposematic black and white
stripes pelage and the influence of predation risk on striped skunk defenses by
measuring the amounts of trans-2-butene-thiol and trans-2-butenyl thioacetate, the
first and third most abundant noxious chemicals of skunk spray, using gas
chromatography and mass spectrometry (GCMS).
References
• Fuller TK, Kuehn DW. 1985. J Mammal:813-815.
• Maan ME, Cummings ME. 2012. Am Nat 179:E1-E14.
• Medill S, Renard A, Larivière S. 2011. Ethol Ecol Evol 23:41-48.
• Stankowich T. 2012. Adapt Behav 20:32-43.
• Stankowich T, Haverkamp PJ, Caro T. 2014. Evolution 68:1415-1425.
• Wood WF. 1990. J Chem Ecol 16:2057-2065.
Methods
Sample Locations
 Using predation heat maps, we can locate skunk populations with various
predation risks to collect samples from
 Photographs of skunks that I will collect samples from will be used to test for
stripe honesty
Collection
Sample Analysis
Preliminary Results
Gas Chromatography & Mass Spectrometry
 Trans-2-butene-thiol and trans-2-butenyl thioacetate have been able to be
identified in preliminary trials
 Our next step is to be able to quantify these two chemicals by comparing them
with a chemical standard
Statistical Analysis
Hypothesis 1: Levene’s Test of Variance
 Compare the variation of the chemicals between populations
 Prediction 1: Skunks in areas with low mammalian predation will have less
variation in their chemical proportions
Hypothesis 2: T-Test & Regression Analysis
 Test if there is significant difference in chemical compound proportions
between yearling and adult skunks
 Prediction 2: Juvenile skunks will have a greater proportion of 2-butenethiol
and butenyl thioacetate than adults
Hypothesis 3: ANOVA & Regression Analysis
 Test if the relationship between the chemical proportions of thiols and
thioacetates to the deviation from the mode of the whiteness index
 Prediction 3: Skunks that deviate away from the whiteness index mode will
have less noxious chemicals in their secretions
Acknowledgements
Thank you to Varenka Lorenzi and Rick Gossett from IIRMES for allowing us to use their equipment.
We would also like to thank Dr. William Wood (CSU Humboldt) for his methodology assistance. A
huge thank you to everyone in the Stankowich lab.
Introduction
Predation Risk
 Prey evolve particularly in response to the predator that they encounter most
often in their natural environment (Stankowich, 2012)
 Scent-based predators are more sensitive to striped skunk spray than other
predators (Stankowich et. al, 2014)
 Hypothesis 1: Skunks evolve more noxious anal gland secretions when
predation risk from scent-sensitive predators and predation risk in general is
high.
Development
 Younger skunks are predated upon more due to their smaller stature in
comparison with adults (Fuller and Kuehn, 1985)
 As a response, young skunks exhibit more aggressive behaviors than adults,
including an increase in their willingness to spray (Medill et. al, 2011)
 Hypothesis 2: Spray produced by young skunks are more noxious in order to
increase their chances of survival.
Honesty
Predation Heat Maps (Stankowich et al., 2014)
The Characteristics of Aposematism and Noxious Spray
in the Striped Skunk (Mephitis mephitis)
High Risk
Low Risk
 I will travel to each location with various predation risks and collect skunk
spray samples to analyze
 A local population in Frank G. Bonelli Park in San Dimas, CA will be used to
collect juvenile and adult skunk samples (IUCAC Protocol 334, DFW Permit
#13392)
Abundance(approx.ng/mL)
(1) 2-Butene-thiol
(2) 3-Methyl-1-butanethiol
(3) 2-Butenyl thioacetate
Peak 1 Peak 2 Peak 3
 Gas Spectrometry and Mass Spectroscopy (GCMS), will allow us to identify
and quantify trans-2-butene-thiol and trans-2-butenyl thioacetate in our
samples (Wood,1990).
Avian Risk Heat Map Mammalian Risk Heat Map
Gas Chromatography Mass Spectrometry
Abundance(approx.ng/mL)
2-Butene-thiol 3-Methyl-1-butanethiol 2-Butenyl thioacetate
Whiteness Index: 0.44
Whiteness Index: 0.26
 Images of skunk pelts will be analyzed using a computer software to provide
a Whiteness Index score, a measure of how much white the skunks have
(Ferguson, unpublished).
 Aposematism is an honest signal of an antipredator
defense of a prey item towards a predator using bright or
bold coloration
 Poison dart frogs with bolder colors signal their level of
unpalability to birds (Maan & Cummings 2009)
 Hypothesis 3: Black and white coloration patterns are
honest signals of the potency of noxious chemicals in the
skunk secretions.
m/zTime (minutes)