This study identified potential sex pheromones in the wolf spider Hogna helluo. Gas chromatography-mass spectrometry analysis found several volatile compounds that differed between male and female spiders, including octane derivatives and alcohols. Analysis of silk washings also identified two alcohols that were more abundant in female silk. These preliminary findings suggest H. helluo uses cuticular and silk pheromones for communication, similarly to other spider species. Further behavioral studies are needed to confirm the role of these compounds as sex pheromones.
More than Just Lines on a Map: Best Practices for U.S Bike Routes
Justin Stover Spring Research Presentation (2)
1. Preliminary Identification of a
Sex Pheromone in the Wolf
Spider, Hogna helluo
Justin Stover
Lou Ann Tom
Lindsay Dunkle
Matthew Persons
2. What is a Pheromone?
Chemical produced by individuals of a
species that causes a behavioral
response in other individuals of the
same species
◦ Aggregation pheromones
◦ Alarm pheromones
◦ Sex pheromones
3. Previously Characterized Spider
Pheromones
Sex pheromones common among
42,000+ spider species but
identified in only four
Photo Eric Reynolds In contrast more than 1,600 insect
Cupiennius salei pheromones have been identified
((S)-1,1'-dimethyl citrate)
(National Academy of Sciences)
Photo: Stefan Sollfors
Linyphia triangularis Agelenopsis aperta
((R)-3-hydroxybutyric acid & (8-methyl-2-nonanone)
(R)-3-[(R)-3-hydroxybutyryloxy]-butyric acid) Argiope bruennichi
(trimethyl methylcitrate)
4. Pheromones and Wolf Spider
Agroecology
High density in cropland year
round
(16 individuals/m2)
Lies at the interface of detrital and
terrestrial food webs
Poorly understood behavioral
mechanisms that allow them to
remain at high densities within
these disturbed habitats
Large potential as biocontrol
agents
5. Crop pests that reduce feeding or are
repelled by chemical cues associated
with the wolf spider Hogna helluo
Mexican Bean
Field Cricket
Beetle
Potato Beetle
Japanese Beetle
6. Sex pheromones as
biocontrol
• Attract conspecifics and facilitate rapid
colonization of recently plowed fields
7. Why Hogna helluo?: Practical
Reasons
• Large – likely produce more
pheromones than smaller spiders
• Resistant to desiccation required for
some assays
• Easy to breed and maintain in the
lab for several generations
8. Spider Collecting
• June-November 2011
• Snyder county cornfields
adjacent to SU
9. Husbandry Methods
◦ Separated by species, sex, and age
◦ Each spider collected is assigned an individual
ID
◦ Housing
16oz. White Deli dishes (Hogna helluo)
Dampened peat moss
◦ Feeding
Cricket nymphs once weekly
10. Possible Types and Sources of
Pheromones
• Volatiles from Cuticle
• Non-volatiles from Cuticle
• Volatiles from Silk
• Non-volatiles from Silk
13. Volatiles from Cuticle/Silk (CLSA)
• Closed loop stripping apparatus
• Mature female H. helluo used and compared to adult males
• Pumps ran for 4 consecutive 12 hour trails on one activated
carbon filter
• Controls ran for 12 hours
• Following trial filters were washed with dichloromethane,
centrifuged and then were analyzed by Gas Chromatography-
Mass Spectrometry
15. Gas Chromatogram of Three Filter Types
female spider
1500000
male spider
control
1000000
Abundance
500000
0
2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18
Time (min)
16. Gas Chromatograms Enlarged
1500000 1500000
Abundance
Abundance
1000000 1000000
500000 500000
0 0
2 3 4 5 6 7 8 8 9 10 11 12
Time (min) 1500000 Time (min)
Abundance
1000000
Female
Spider
Male Spider
500000
Control
0
13 14 15 16 17 18
Time (min)
17. Table of Sex-Specific Compounds from
GC
Compound Male Female
18. Table of Sex-Specific Compounds from
GC
Compound Male Female
Octane, 2,5,6-trimethyl No Yes
Eicosane, 3-methyl No Yes
2,2,7,7- No Yes
tetramethyltricyclo[6.2.1.0(1,6)]undec-4-en-3-
one
Dodecane, 4-methyl No Yes
19. Table of Sex-Specific Compounds from
GC
Compound Male Female
Octane, 2,5,6-trimethyl No Yes
Eicosane, 3-methyl No Yes
2,2,7,7- No Yes
tetramethyltricyclo[6.2.1.0(1,6)]undec-4-en-3-
one
Dodecane, 4-methyl No Yes
Ethylbenzene Yes No
Decane, 2,3,4-trimethyl Yes No
Hexane, 2,2,3-trimethyl Yes No
1-hexanol, 2-ethyl Yes No
20. Table of Sex-Specific Compounds from
GC
Compound Male Female
Octane, 2,5,6-trimethyl No Yes
Eicosane, 3-methyl No Yes
2,2,7,7- No Yes
tetramethyltricyclo[6.2.1.0(1,6)]undec-4-en-3-
one
Dodecane, 4-methyl No Yes
Ethylbenzene Yes No
Decane, 2,3,4-trimethyl Yes No
Hexane, 2,2,3-trimethyl Yes No
1-hexanol, 2-ethyl Yes No
2,2,6,6-Tetramethylheptane Yes Yes
Octane, 2,3,6,7-tetramethyl Yes Yes
1-hexanol, 2-ethyl-2-propyl Yes Yes
2,5-cyclohexadiene-1,4-dione, 2,6-bis(1,1- Yes Yes
dimethyl)
21. Volatiles from Silk
• Unmated adult females were
compared to, adult males, and
empty controls
• Spiders were placed in glass
vials and labeled
• Trials ran overnight for 12 hours
each
• Each vial was serially rinsed
with the same initial 1 mL of
dichloromethane and then
capped. The dichloromethane
from the aggregate wash of 15
vials was then filtered and
analyzed by GC/MS.
22. Gas Chromatograms of Silk with Two
Primary Compounds found and
Configuration
80000
silk
60000
Abundance
3-Penten-2-ol (12.4%)
40000
20000
0
2.5 3.0 3.5 4.0
Time (min)
5-Methyl-1-hepten-4-ol (12.7%)
23. Other Explanations for Differences
Diet-based differences
Individual Odor Signatures
Differences in metabolic processing
between the sexes
24. Future Directions
Compare to juveniles and mated
females
Confirmation
Synthesis
Behavioral Assays singly and in
combination
Field Test
Other species
25. Arachnowledgements
We would like to thank Lindsay Dunkle and Lou Ann Tom for
preparing and running the GC-MS and for analyzing the data. We
would also like to thank Thomas Duff, Rizwan Khan, Kody Hummel
and Cierra Omlor for helping to capture and maintain the spiders for
this study.
Aggregation Pheromones – Pheromone which signals members of the same species to congregate together for a specific purpose like lady bugs do in your houseAlarm Pheromones – Pheromone which warns members of the same species to either attack or run from a given threat. Honey bees do this when the hive is attacked.Sex Pheromones – Pheromone which are sent to the opposite sex of a species to indicate their maturity and willingness to select a mate.
Ubiquitous means that it is seen throughout the entire species.
Sex pheromone traps have been used to capture crop pests. Spider pheromones may be used to attract specific spiders to recolonize crops after tilling fields. Pheromones may also serve as functional kairomones—deterring some crop pests from the area. For example Mexican bean beetles, Japanese beetles, and Potato Beetles are all known to preferentially avoid feeding on plants previously walked on by spiders.
Chemosensory hairs fire when they come into contact with certain chemicals which excite them. Do not react to pressure and structure of objects, unlike tactile hairs. Each chemosensory hair is triple innervatedTactile hairs respond to pressure and movement of objects as well as structure of objects such as silk.Not entirely known: could be integrated with each other.
Tarsal organs are thought to respond to volatiles in the air and or changes in humidity in the air. Recently found to have a chemosensory function.
“Mature” just means that during an assay a male showed a courtship response to the female silk