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Temporal Variation in Venom Yield of the Australian Funnel-Web Spider Atrax
sutherlandi (Hexathelidae: Atracinae)
Author(s): Mark K. L. Wong , James D. Woodman and David M. Rowell
Source: Arachnology, 17(1):7-9.
Published By: British Arachnological Society
DOI: http://dx.doi.org/10.13156/arac.2006.17.1.7
URL: http://www.bioone.org/doi/full/10.13156/arac.2006.17.1.7
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Arachnology (2016) 17 (1), 7–9 7
Temporal variation in venom yield of the
Australian funnel-web spider Atrax sutherlandi
(Hexathelidae: Atracinae)
Mark K. L. Wong
Research School of Biology,
College of Medicine, Biology and Environment,
Australian National University,
Canberra, ACT 2601, Australia
email: markwong.research@outlook.com
James D. Woodman
Australian Plague Locust Commission,
Australian Government Department of Agriculture,
GPO Box 858,
Canberra, ACT 2601, Australia
David M. Rowell
Research School of Biology,
College of Medicine, Biology and Environment,
Australian National University,
Canberra, ACT 2601, Australia
Summary
Temporal variation in the venom yield of spiders is a relatively
poorly understood phenomenon. We investigated temporal
variation in venom yield of the Australian funnel-web spider
Atrax sutherlandi Gray 2010 (Hexathelidae: Atracinae). The
venom yield of spiders collected and milked in winter was
62.9% higher than those collected and milked in autumn, despite
all undergoing acclimatization (45 days in darkness at 10°C and
100%RH) before milking. Our ndings highlight the potential
effects of seasonality on spider venoms and lay the groundwork
for future studies to investigate the evolutionary and ecological
correlates of this phenomenon further.
Introduction
Intraspecic variation in the quantity of arachnid venoms
(i.e. venom yield) may be associated with ontogenic, sexual,
diet and temporal factors (de Oliveira et al. 1999; Herzig,
Ward & dos Santos 2004; Herzig 2010). Of these, data on
temporal variation in venom yield is the least well docu-
mented, and generally based on anecdotal evidence from
opportunistic observation (e.g. Bücherl 1953; Wiener 1956;
Wiener 1959; Schenberg & Pereira-Lima 1966) with little
discussion of underlying ecological and biological mecha-
nisms.
The neurotoxic venoms of the Australian funnel-web
spiders (Hexathelidae: Atracinae) have been subject to
extensive toxicological research, with the majority of
work focusing on the notorious Sydney funnel-web Atrax
robustus. The venom yield of this species has been observed
to vary signicantly by season, with the highest yield
reported in spring (Wiener 1959). The present study focused
on a recently described species of the genus, Atrax suth-
erlandi Gray, 2010, which is distributed across temperate
forests in most of south-eastern New South Wales. This
fossorial spider constructs permanent silk-lined burrows (c.
10–30 cm deep) in the soil beneath fallen logs and rocks.
Females appear rarely to leave the immediate vicinity of
their burrows (pers. obs.), in contrast to the vagrant males,
which have been collected from pitfall traps and temporary
refuges built under rocks in summer months (unpublished
data). During winter, the spiders appear largely inactive and
retreat deeper into their burrows (pers. obs.). In this study,
we investigate temporal variation in venom yield of A. suth-
erlandi by comparing the venom yield of spiders collected
and milked in two different seasons.
Methods
In 2014, adult females were collected from Tallaganda
forest (35°15′S–36°15′S 149°28′E–149°37′E) in two sepa-
rate batches: 23 specimens on 2 April (mid-autumn) and 22
specimens on 2 July (mid-winter). Only large spiders from
well-established burrows were collected to avoid including
pre-reproductive males (which cannot be distinguished
from females on external morphology alone). Following
collection, the spiders were housed individually in 70 ml
plastic specimen jars at 10°C and 100% RH in constant
darkness for 45 days, in preparation for venom milking. The
dark setup was intended to approximate light availability
within a burrow in the eld, and also acclimatize all spiders
to the same conditions. Spiders were fed successfully once
every three days with a medium-sized (~15 mm) Tenebrio
molitor larva for the rst 39 days, but not fed in the last six,
to allow replenishment of venom reserves in the absence of
prey capture prior to milking. No individuals moulted while
in captivity.
After the 45-day acclimatization period, venom was
milked from each specimen in a 5-minute sitting daily
(between 0900 and 1600 hrs) over three consecutive days
(autumn batch milked 17–19 May; winter batch milked
16–18 August). Pilot tests showed this regimen depleted
the venom stores of 97% of subjects. During each sitting,
the spider was repeatedly provoked by touching its front
legs with metal forceps, causing it to produce venom on
the tips of its fangs, which was milked via capillary action
using a 32 mm (5 µL) microcapillary tube (Model: P1799,
Drummond Scientic, USA). The tubes were photographed
with a digital microscope (Model: P-400Rv, Nikon, Japan)
immediately after every sitting, and the length (mm) of each
tube occupied by venom was measured and converted to
volume (µL) via image analysis using the software ImageJ
(Abràmoff, Magalhães & Ram 2004). The absolute yield
(Ya) of each spider was the total venom quantity collected
over the three-day period.
Statistical analyses were performed using SPSS 22.0
(IBM Corporation 2013). To derive a size-corrected measure
of yield, carapace width (CW) of each specimen was meas-
ured as an indicator of body size (Hagstrum 1971), then Ya
was regressed against CW, CW2 and CW3 and the resultant
coefcients of determination (r2) were scrutinized to deter-
mine the dimension of body size (i.e. width, surface area or
volume) that was most strongly associated with yield. This
measurement was then utilized for subsequent correction of
absolute yield for body size, and comparison of size-cor-
rected yield between the autumn and winter batches.
8 Temporal variation in venom yield of Atrax
Results
Among CW, CW2 and CW3, CW3 showed the strongest
relationship with absolute yield (Ya) (r2 = 0.407, p < 0.0001)
(Fig. 1). Size-corrected yield (Ys) was calculated using the
equation: Ys (µL mm-3) = Ya (µL) / CW3 (mm3)
There was no signicant difference (p = 0.634) in the
mean size (CW) of spiders between the batches from autumn
(6.76 ± 0.94 mm) and winter (6.71 ± 1.00 mm). However,
there was a signicant difference in the mean size-corrected
yield (Ys) of autumn (0.0447 ± 0.023 µL mm-3) and winter
(0.0728 ± 0.0259 µL mm-3) batches, t (47) = -3.44, p =
0.001, d = -1.15 (Fig. 2).
Discussion
In our assessment of the relationship between venom
yield and body size in A. sutherlandi, we found absolute
yield to be most strongly associated with the volumetric
dimension of body size (indicated by CW3). This may be
explained by the changing volume of the venom glands
with overall body size inuencing venom production and
venom-holding capacity (Herzig, Ward & dos Santos 2004).
Similar relationships have been demonstrated in other
spiders including A. robustus (e.g. Wiener 1959; Vapenik
& Nentwig 2000). The linearity of the relationship between
venom yield and overall body size (Fig. 1) is consistent
with results from other mygalomorphs (Herzig 2010) but
contrasts with exponential modelling reported for araneo-
morphs (e.g. Malli, Vapeli & Nentwig 1993; Herzig, Ward
& dos Santos 2004). Herzig (2010) attributed such disparity
in size-yield relationships between the infraorders to differ-
ences in (i) venom gland morphology, whereby mygalo-
morph venom glands are restricted to the basal part of the
chelicerae compared to araneomorph venom glands that
extend into the prosoma (Foelix 2011), and (ii) moulting,
whereby unlike adult mygalomorphs that may increase
venom yield with moulting and growth, araneomorphs
generally do not moult after reaching adulthood (Foelix
2011, but see exceptions in Kuntner et al. 2012). Hence, the
exponential increase in their venom yield may have devel-
oped to ensure that the spiders reach high levels of venom
production after their nal moult as subadults.
Despite undergoing a 45-day acclimatization period
prior to milking, the mean venom yield from A. sutherlandi
spiders collected and milked in winter was 62.9% greater
than that of spiders collected and milked in autumn (Fig.
2). This nding presents evidence for intraspecic temporal
variation in venom yield for a second species of Australian
funnel-web spider, in addition to seasonal variation in
venom yield previously reported in A. robustus (Wiener
1959). The magnitude of temporal increase in A. suther-
landi venom yield is also noteworthy; in comparison, the
maximum difference between yields of two seasons in A.
robustus was 45%, between spring and autumn (Wiener
1959). Apart from the Atracinae, temporal effects have also
been observed in the venom yield of araneomorphs such as
the theridiid Latrodectus hasseltii, and ctenids Phoneutria
fera and Phoneutria nigriventer (Bücherl 1953; Wiener
1956; Schenberg & Pereira-Lima 1966). While it is possible
that temporal variation may occur in the venom production
of other spiders, considering how infrequently such varia-
tion is reported, further investigation is required to substan-
tiate and qualify this phenomenon in nature.
In addition to being scarce, most accounts on temporal
variation in spider venom yield are derived from incidental
observations made during pharmacological or biochemical
studies (e.g. Schenberg & Pereira-Lima 1966). Hence, the
evolutionary and ecological underpinnings for such varia-
tion remain unknown. While it may be intuitive to attribute
the observed variation in A. sutherlandi venom yield to
plastic responses to seasonality, a common precursor to
seasonal polyphenism is the perception of seasonal climatic
changes (Azel 2002). However, the acclimatization period
(45 days) and controlled microclimatic conditions (temper-
ature, humidity, darkness) used are likely to have precluded
the spiders from detecting any climatic shifts following
collection from the eld. Nevertheless, since the batches
were collected in mid-autumn and mid-winter, we are unable
to reject the possibility that the spiders had already adjusted
their venom production in response to perceived climatic
changes before capture, and maintained these adjustments
throughout the acclimatization period.
The observed temporal variation in venom yield is not
likely to be an artefact of yield differences at the initial time
of capture, since a 45-day period is almost certainly suf-
cient for complete regeneration of venom stores to maximal
capacity; Perret (1977) reported that, in other mygalo-
morphs, regeneration of 50% of original venom supply is
achieved within 2–3 days. Furthermore, we observed that,
during feeding in the laboratory, like other mygalomorphs,
A. sutherlandi from both batches primarily immobilized the
relatively soft-bodied T. molitor larva by physical manip-
ulation (mashing) with their strong chelicerae, as opposed
to relying on fang-piercing alone (Wigger, Kuhn-Nentwig
& Nentwig 2002). It is thus possible that venom reserves
were well conserved during predation over the 45 days; see
discussion on the venom optimization hypothesis (Wigger,
Kuhn-Nentwig & Nentwig 2002; Morgenstern & King
2013). In addition, even if some venom was utilized for
prey submission during the 45-day period, this should not
Fig. 1: Relationship between absolute yield (Ya) and carapace volume
(CW3), which showed the strongest relationship with absolute yield
(r2 = 0.407).
M. K. L. Wong, J. D. Woodman & D. M. Rowell 9
have affected the observed temporal variation in yield, since
the feeding and milking were performed at the same time
intervals in both batches.
Perret (1977) also observed the occurrence of moulting
in his mygalomorphs to vary with season. Although venom
yield of A. robustus and Coremiocnemis tropix have been
shown to decrease prior to moulting (Wiener 1959; Herzig
2010), none of the A. sutherlandi used in our study moulted
during the 45 day acclimatization period nor in the 30 days
following milking (after which the spiders were preserved
for size measurement). Furthermore, as adult araneomorphs
generally do not moult, seasonal variation in moulting is
unlikely to account for temporal variation in venom yield of
Latrodectus and Phoneutria (Bücherl 1953; Wiener 1956;
Schenberg & Pereira-Lima 1966).
It is unclear why such a large disparity (62.9%) exists
between venom yields of A. sutherlandi milked in winter
compared to autumn. One explanation may be that the
spiders possess mechanisms to increase venom accumula-
tion during winter as a means of capitalizing on their relative
inactivity (pers. obs.) in preparation for increased feeding in
the coming spring – potentially on prey items that are more
difcult to immobilize compared to T. molitor. For example,
the remains of formicids and coleopterans have been found
at A. sutherlandi burrow entrances on many occasions (pers.
obs.). We propose that future longitudinal comparisons of
venom yield and metabolic expenditure as well as prey type
and consumption quantity across the seasons would appro-
priately test this postulation. Indeed, the Atracinae would be
ideal study candidates for such pursuit, given the relative
ease of venom collection and their extensive (>15 years)
lifespans (King, Tedford & Maggio 2002; Morgenstern &
King 2013).
Acknowledgements
This study was supported by the following grants
awarded to the rst author: The National Geographic Soci-
ety’s Young Explorer’s Grant (9633-14), The Australasian
Wildlife Management’s Student Travel Award, and The
Australian Wildlife Society’s University Student Grant.
The authors would also like to thank the two anonymous
reviewers for their comments on the manuscript.
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Fig. 2: Temporal variation in venom yields for A. sutherlandi collected and
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and associated standard errors. Mean size-corrected yield (Ys) for
the winter batch was 62.9% higher than that of the autumn batch.
