Ecological Entomology (2011), DOI: 10.1111/j.1365-2311.2011.01268.x
SHORT COMMUNICATION
Stable isotopes reveal different patterns of inter-crop
dispersal in two ladybeetle species
K A T H E R I N E J . F O R B E S 1 and C L A U D I O G R A T T O N 1,2
1
Department of Zoology,
University of Wisconsin, Madison, Wisconsin, U.S.A. and Department of Entomology, University of Wisconsin, Madison,
Wisconsin, U.S.A.
2
Abstract. 1. Dispersal plays a key role in structuring the local population densities of
many insect species, yet the movement patterns across the landscape of most species
are poorly understood. By measuring the stable isotope of carbon (δ 13 C) from multiple
tissues, a novel approach applied to field-collected insects, we were able to infer
differences in movement patterns of two species of mobile generalist insect predators.
2. Coccinella septempunctata L (7-spot ladybeetle) and Harmonia axyridis Pallas
(multicoloured Asian ladybeetle) were collected in agricultural habitats in 2003 and
2004, and were assayed for δ 13 C in the elytra (slow turnover) and fat/reproductive
tissues (fast turnover). δ 13 C values were used to infer diet use of C3 versus C4 crops.
3. Coccinella septempunctata was relatively more faithful to a particular habitat
and tended to stay in alfalfa and soybean (C3-based photosynthetic crops) over long
periods during the summer. This contrasts with H. axyridis which showed isotopic
evidence consistent with frequent late-season movement between C3 and C4 crops
such as corn in the landscape.
4. These differing patterns suggest that in the late summer season H. axyridis
individuals traverse the environment more extensively and utilise broadly dispersed
aphid resources, whereas C. septempunctata adults are more specialised on alfalfa and
soybean crops.
Key words. Dispersal, diet-switching, inter-crop movement, stable isotopes.
Introduction
The extent and dynamics of movement of organisms between
different habitat patches within the landscape are recognised as
key to determining the abundance and persistence of populations and for driving inter-specific interactions such as competition and predation (Hanski, 1999). There is a growing body
of empirical research that documents the ability of species to
move across the landscape and the consequences of this movement for the dynamics of populations and communities at a
landscape scale (Gurr et al., 2004). However, inferring movement patterns has been especially challenging for small-bodied
animals highlighting the need to develop new and complementary approaches to study dispersal.
Ladybeetles (Coleoptera: Coccinellidae) are important predators in many agricultural landscapes. In the present study,
Correspondence: Claudio Gratton, 237 Russell Labs, 1630 Linden
Drive, Madison, WI 53706, U.S.A. E-mail: [email protected]
© 2011 The Authors
Ecological Entomology © 2011 The Royal Entomological Society
we compared the late-season movements of two common
aphidophagous ladybeetle predators in the upper Midwest,
Coccinella septempunctata L., the seven-spot ladybeetle, and
Harmonia axyridis Pallas, the multicolored Asian ladybeetle. From observations of ladybeetle prevalence in the field
(Forbes, 2008) and building on what is known in the literature
(Evans & Toler, 2007), we hypothesised that C. septempunctata
and H. axyridis have different spatial patterns of host use. In
particular, C. septempunctata was hypothesised to have higher
residence times in alfalfa where pea aphids, a preferred prey,
are present in the landscape throughout the summer (Gross
et al., 2005). In contrast, H. axyridis, an arboreal predator common in row crops, consumes a wide range of prey (LaMana &
Miller, 1996), and exhibits rapid density-dependent responses
to aphids at small spatial scales (Donaldson et al., 2007). As
such, we hypothesised that H. axyridis would have a greater
propensity for inter-crop movement than C. septempunctata.
We utilised stable isotopes of C (13 C/12 C or δ 13 C) to infer
spatial patterns of feeding by ladybeetles. As plants with a
1
2 Katherine J. Forbes and Claudio Gratton
C3-photosynthetic pathway such as soybean and alfalfa have
δ 13 C ≈ −28‰ whereas C4-photosynthetic plants such as corn
have δ 13 C values around −13‰ (Fry, 2006) this facilitates
the interpretation of naturally occurring isotope ratios in consumers. As differences in δ 13 C propagate through food chains,
it is possible to infer the ultimate C resource (i.e. resource
‘end-members’) utilised by predators. In a novel application
of this technique, we also took isotope measurements from
multiple tissues with slow and fast isotopic turnover rates from
single individuals (e.g. forewings and fatbody/reproductive tissues, respectively) to discriminate consumers that had recently
switched between two isotopically distinct resources from
those that have been using one resource for a long period
of time (Podlesak et al., 2005; Gratton & Forbes, 2006). We
used this approach to test the hypotheses that C. septempunctata would show isotopic evidence of habitat fidelity whereas
H. axyridis would have an isotopically more variable diet, suggesting greater intercrop movement.
Methods
Ladybeetle sampling
Agriculture is an important feature of southern Wisconsin
landscapes with corn, soybean, and alfalfa averaging 22%,
7%, and 7% of the landcover, respectively. Perennial habitats are characterised by open grasslands/pasture (13%) and
woodlands (29%) (USDA NASS, 2007). Between late July and
mid-September 2003 we sampled ladybeetles in five alfalfa
fields, two soybean fields, and two corn fields at irregular
2- to 14-day intervals at the University of Wisconsin’s Arlington Agricultural Research Station, Columbia County, Wisconsin (43◦ 18 N, 89◦ 21 W). In 2004, ladybeetles were collected
along a 300-km south-north transect across the southern twothirds of Wisconsin. From 29 August to 1 September we only
sampled alfalfa fields (n = 11) selecting fields with alfalfa
25–60 cm in height. All stages of larval and adult C. septempunctata and H. axyridis were collected either by hand or
by sweep netting. Samples were frozen before preparation for
isotope analysis.
(C4 plants) or soybean/alfalfa (C3 plants), under the assumption that larval dispersal between crops is unlikely (Appendix
SI). Larval isotope values were not distinguishable from aphid
values collected in the same habitats (data not shown). Soybean
and alfalfa are both C3 plants and as their δ 13 C values overlap, we were not able to isotopically distinguish between them.
Thus in 2003 beetles were pooled across these two habitats for
analysis.
For a given isotopic value of a tissue, we used inverse regression (Draper & Smith, 1998, Appendix SII) to estimate the
mean proportion (and variation) of the diet that came from
C3-based resources given the mean and standard deviation of
the two diet end-members. By calculating the mixing model
estimate of diet use from two different tissues in the same
individual, we were able to asses diet switching. For example,
overlapping estimated ranges of a diet source would indicate
tissues are at equilibrium with the two diet resources – a pattern which would result if ladybeetles had been feeding on
a single resource (or a mixture) for an extended period of
time. On the other hand, non-overlapping estimates is taken
as evidence for a recent switch in diets as the tissue with a
slower turnover has not yet equilibrated to the novel resource
(Podlesak et al., 2005; Gratton & Forbes, 2006).
From the results of the two-tissue mixing models we classified individual beetles into one of five categories. The first three
categories are individuals that did not recently switch diets and
for which mixing models suggested that both tissues were at
equilibrium with resources: (i) beetles with a a long-term C3
diet (>90% C3 diet); (ii) ladybeetles with a long-term diet with
<10% composed of C3 resources (i.e. >90% C4 resource), and
(iii) beetles with diets estimated to be between 10% and 90%
of C3 resources. The last group is beetles not in categories
(i) or (ii), but which have δ 13 C values of both tissues which
give diet estimates that are statistically indistinguishable. The
final two groups (iv) and (v) contain beetles where the δ 13 C
of two tissues indicate different diet composition, suggesting a
recent diet switch. The frequency of H. axyridis and C. septempunctata in different isotopic categories was analysed using a
two-tailed Fisher’s exact test.
Results and discussion
Isotope analyses
Stable isotope analyses were performed separately on the
elytra (forewings) and the reproductive and fat tissues (combined) of each beetle. Tissue samples (0.5–1.0 mg) were analysed for stable isotopes of carbon at the Colorado Plateau
Stable Isotope Lab (Northern Arizona University, Flagstaff,
AZ, U.S.A.). Ratios of 13 C/12 C were expressed relative to a
known standard (VPDB) in per mil (‰) notation, with a measurement error on duplicate samples approximately ±0.15‰.
Linear mixing models were used to infer the proportion of
the diet attributable to C3-based resources (Phillips, 2001).
To estimate the isotopic composition of mixing model endmembers (i.e. homogenous isotopic resources), we used the
isotopic values of larval beetles collected within either corn
Coccinella septempunctata and H. axyridis, two important
ladybeetle predators in agricultural landscapes, have significantly different patterns of late-season inter-crop movement in
southern Wisconsin. Coccinella septempunctata collected from
all crops in 2003 were dominated by individuals showing evidence of C3-based diets (soy, alfalfa or other habitats with
C3-based resources such as forb-dominated areas) regardless of
the crop in which they were collected (Fig. 1a, Table 1). Coccinella septempunctata was more likely to show a C3 isotope
signature than H. axyridis (Region i vs. ii–v, Fisher’s exact
test, P = 0.002). In addition, the population of H. axyridis was
more heterogeneous in δ 13 C isotope values (Fig. 1b) compared
with C. septempunctata, suggesting beetles fed in both C3 and
C4 habitats (Table 1). Harmonia axyridis was also more likely
to show an intermediate diet than C. septempunctata (Region
© 2011 The Authors
Ecological Entomology © 2011 The Royal Entomological Society, Ecological Entomology, doi: 10.1111/j.1365-2311.2011.01268.x
6
13
25
44
24
25
0
0
0
5
8
16
0
8
16
7
4
36
17
15
32
36
0
12
0
0
28
5
0
24
83
77
24
48
88
12
1c
1c
C. septempunctata
H. axyridis
1b
H. axyridis
2004
1a
C. septempunctata
Corn (C4)
Soy/Alfalfa (C3)
Corn (C4)
Soy/Alfalfa (C3)
Alfalfa (C3)
Alfalfa (C3)
N
V (C4 → C3)
IV (C3 → C4)
III (mix)
II (C4)
I (C3)
Collected in:
2003
iii–v vs. i–ii, Fisher’s exact test, P = 0.02, Table 1) indicative
of a recent movement into a crop after having fed on a crop of
a different isotopic composition. In 2004, a similar pattern of
isotope patterns was observed: C. septempunctata had evidence
of feeding mostly within C3-based habitats (88% beetles)
Figure
Fig. 1. δ 13 C (‰) values of the elytra and reproductive and fat tissues
from adult Coccinella septempunctata (a, n = 19) and Harmonia
axyridis (b, n = 69) collected at the Arlington Agricultural Research
Station (Columbia Co., WI) in summer 2003. Ladybeetles were
collected in soybean (dark grey circles), alfalfa (light grey circles), and
corn (open squares). In 2004 (c) collections of adult C. septempunctata
(gray, n = 24) and H. axyridis (open, n = 25) were exclusively from
alfalfa. Region ‘i’ corresponds to beetles consuming a >90% C3
diet (soybean and/or alfalfa), ‘ii’ are beetles consuming >90% C4
(corn) diet. Ladybeetles in region ‘iii’ show evidence of a mixed diet
(10–90% diet of C3). Ladybeetles in region ‘iv’ show evidence of
a C3 → C4 switch in diet, whereas ladybeetles in region ‘v’ show
evidence of a C4 → C3 switch in diet.
Species
(c)
Year
(b)
Percent of beetles in Region:
(a)
Table 1. Percentage of ladybeetles collected in either corn (C4) or soy/alfalfa crops (C3) showing evidence of their long-term feeding in a C3 crop (I), C4 crop (II), a long-term mixture of the
diet from the two crop types (III) or a recent transition from C4 to C3 habitats (IV) or C3 to C4 habitats (V) based on δ 13 C analysis of fast turnover tissues (fat body/reproductive tissues) and
slow turnover tissues (elytra) depicted in Fig. 1. N is total number of beetles collected in a particular habitat.
Differential dispersal of two coccinellids 3
© 2011 The Authors
Ecological Entomology © 2011 The Royal Entomological Society, Ecological Entomology, doi: 10.1111/j.1365-2311.2011.01268.x
4 Katherine J. Forbes and Claudio Gratton
compared with only 12% of H. axyridis. Harmonia axyridis
also had more heterogeneous isotopic values than C. septempunctata (difference between species, Fisher’s exact, P <
0.0001, Fig. 1c, Table 1) with almost two-third of beetles intermediate between C3 and C4 resources compared with only
12% of C. septempunctata.
These differences in the isotope patterns of C. septempunctata and H. axyridis imply differential movement of these
predators that may be the result of multiple species-specific
traits. Harmonia axyridis is known to readily disperse throughout fields, finding and consuming aphids in isolated patches
more quickly than some other species (With et al., 2002).
Harmonia axyridis’ varied isotopic diet suggests frequent intercrop movement as crop types are isotopically homogenous and
distinct from one another. Donaldson et al. (2007) found that
H. axyridis rapidly aggregates to high densities of soybean
aphid prey at two spatial scales but does not readily locate lowdensity aphid patches, suggesting that this predator is highly
mobile within soybean fields. In contrast, C. septempunctata is
more likely to stay near aphids it encounters, even at low density, and is more likely to remain in alfalfa in general (Forbes,
2008; Harmon et al., 2009). Coccinella septempunctata is frequently described as exhibiting a strong tendency to engage in
area-restricted searches for prey, meaning it is likely to forage
in a relatively small area provided it is encountering aphids
(Ives et al., 1993). In spite of its ability to consume a wide
range of prey, C. septempunctata may also be relatively specialised on alfalfa as a habitat to the extent that it will consume
non-aphid prey when pea aphid, Acyrthosiphon pisum (Harris),
densities are low (Evans & Toler, 2007). Although aphids are
the most likely prey for both species, the recent laboratory
finding that H. axyridis readily feeds on corn seedlings as a
larva (Moser et al., 2008) may be another avenue by which
δ 13 C can be enriched in adults.
Dispersal as a response to resource availability is an important determinant of species’ abundance and distribution in the
landscape. The success of biological control programmes often
implicitly relies on the ability of predators to move rapidly
across the landscape in response to changes in prey availability. Using a novel application of a stable isotope approach to
infer inter-crop movement, the present study found that for
two important predators of key pests of agricultural crops in
the upper Midwest, the propensity to move across the agricultural landscape varies widely. It is possible that differences in
dispersal, the mechanisms for which are not understood, could
also result in differential abilities of the ladybeetles to suppress
aphid prey at landscape scales. Differences in the likelihood of
dispersing across an agricultural landscape matrix may make
these beetles differentially sensitive to changes in landscape
structure and composition (Gardiner et al., 2009) or to habitat
manipulations intended to enhance their abundance in agricultural habitats (Gurr et al., 2004).
this paper. KJF was supported by an NSF pre-doctoral fellowship. This study was supported in part by a National Research
Initiative of the USDA Cooperative State Research, Education
and Extension Service (CSREES), Grant 2004-35302-14726,
the University of Wisconsin College of Agriculture and Life
Sciences Hatch Funding to CG (WIS01285), and by National
Science Foundation grant DEB-0108300 to A. R. Ives.
Supporting Information
Additional Supporting Information may be found in the online
version of this article under the DOI reference:
10.1111/j.1365-2311.2011.01268.x
Appendix SI. Average δ 13 C (‰) (SD) of Harmonia
axyridis and Coccinella septempunctata larvae collected in
soybean, alfalfa, and corn in 2003, and alfalfa in 2004 used
as isotopic diet end-members in mixing model analyses. Larvae of both species within a crop and year were combined
to get a single estimate of beetles feeding on an isotopically
homogeneous diet.
Appendix SII. The proportion of a diet coming from endmember a from a single measurement of δ 13 C is estimated
using an inverse regression approach described in Forbes
(2008). The procedure requires estimating the proportion of
resource a in the diet, given an isotopic value (Y) of tissue i, where i = elytra or fat/reproductive tissue. Figure A1.A
depicts the linear relationship in this two source mixing model.
The circles with error bars represent isotope values (mean and
their SE, estimated from nia individuals) of a consumer’s tissues when fed isotopically homogeneous resource a, (‘1’) or
b (‘0’). These values were derived from laboratory feeding
experiments and field measurements (Appendix SI). The variation around the estimate of the line (dotted lines) is given
by the variation in the end-member values (here shown as
±SEM) which determines the level of variation in the regression relationship. The width of the regression uncertainty was
arbitrarily set with an α-level of 0.05. We can use this linear
relationship to estimate the mean fraction of diet a, x, and the
confidence range (grey rectangle) of an assayed tissue with an
isotopic value of tissue i, yi , using inverse regression (Draper
& Smith, 1998) (Fig. A1.B). Performing this procedure for
two tissues in the same individual will give two estimates of
x. If the estimates are not statistically distinguishable, then tissues are at isotopic equilibrium with the resources, while if the
estimates differ (do not overlap), there is evidence of a recent
change in diet which has resulted in different isotope values
in the two tissues.
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Acknowledgements
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© 2011 The Authors
Ecological Entomology © 2011 The Royal Entomological Society, Ecological Entomology, doi: 10.1111/j.1365-2311.2011.01268.x