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JFB 195
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Journal of Fish Biology (2003) 63, 1–5
doi:10.1046/j.1095-8649.2003.00195.x, available online at http://www.blackwell-synergy.com
BRIEF COMMUNICATION
Sex-specific survival and parasitism in three-spined
sticklebacks: seasonal patterns revealed by molecular
analysis
K. E. A R N O L D *†, A. A D A M *, K. J. O R R *, R. G R I F F I T H S *
I. B A R B E R ‡
AND
*Division of Environmental and Evolutionary Biology, Graham Kerr Building,
University of Glasgow, Glasgow G12 8QQ, U.K. and ‡Institute of Biological Sciences,
University of Wales Aberystwyth, Aberystwyth, Ceredigion SY23 3DA, U.K.
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(Received 3 October 2002, Accepted 27 June 2003)
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A molecular method of identifying sex in three-spined sticklebacks Gasterosteus aculeatus
showed that adult males had a higher prevalence of dermal Glugea anomala cysts than adult
females, and young-of-the-year had more than adults. At the end of the breeding season, as
predicted, the adult sex ratio became female biased and there was a disproportionate increase in
# 2003 The Fisheries Society of the British Isles
G. anomala parasitism in adult males.
Key words: Glugea anomala; molecular sexing; parasites; sex ratio; three-spined stickleback.
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A sex difference in vulnerability to parasites, and therefore mortality, is thought
to be a viability cost associated with sexual selection (Moore & Wilson, 2002).
Thus, the population sex ratio is expected to vary temporally within species that
possess sexually selected traits (Andersson, 1994). This bias in sex ratio is only
expected to arise among sexually mature adults not in juveniles, so will have
important implications for population structure. Procuring information regarding temporal changes in the sex ratio of field populations, however, is often
difficult because of problems in non-destructively identifying the sex of individuals. In fishes, such problems are exacerbated in juveniles and in species that
lack sexual dimorphism, or are only morphologically distinguishable during the
breeding season. In species with chromosomal sex determination, molecular
techniques allow the unambiguous assignment of the sex of fishes regardless
of their age, reproductive status or condition. In the present study, a recently
developed molecular method (Griffiths et al., 2000) was used to determine sex
differences in microsporean parasitism, body size and survival throughout the
season in a population of three-spined sticklebacks Gasterosteus aculeatus L.
†Author to whom correspondence should be addressed. Tel.: þ44 (0) 141 3302898; fax: þ44 (0) 141 3305971;
email: [email protected]
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2003 The Fisheries Society of the British Isles
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K. E. ARNOLD ET AL.
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From March to October 2000 inclusive, monthly three-spined stickleback
samples were taken from Inverleith Pond. This is a small (c. 1 ha), shallow
(c. 1 m deep) artificial lake close to Edinburgh, U.K. (55 550 N; 03 100 W) and is
connected to another smaller pond. Three-spined sticklebacks are the only
species of fish in the lake, and they are subject to predation by gulls (Laridae).
In this population, the bulk of the diet consists of chironomids and nematodes,
but this varies with season (Tierney, 1991). On each sampling day, three types of
capture technique were employed in order to minimize sex-biased sampling.
Eight baited bottle traps were placed at random throughout the pond for
30–60 min c. every 1–2 h over a 6 h period. A 1 m hand-trawl (4 mm mesh) was also
used to collect individuals, and hand nets (2 mm mesh) were used to catch both
adults and young-of-the-year (YOY) from the periphery of the pond and
around submerged obstacles. Standard body length (LS to 01 mm) and wet
mass (to 001 g) of all fish were recorded. During the breeding season (May to
August), when sexing of adult three-spined sticklebacks can be performed
visually using cues from eye and throat colour (Wootton, 1976), the sex was
estimated based on these morphological cues at the time of capture in addition
to the molecular method. During the sampling period the study population was
infected with Glugea anomala Moniez, an obligate intracellular microsporidian
parasite, which cause visible white dermal cysts, up to 8 mm in diameter, on the
skin surface of infected fishes. Three-spined sticklebacks become infected when
spores, released from cysts of dying or dead fish, are ingested either directly, or
via contaminated invertebrates. Hatched spores infect individual host cells,
which undergo hypertrophy, forming a cyst in which the parasite reproduces
vegetatively. Infection with microsporideans such as G. anomala is associated
with severe pathology and mortality, particularly in young fishes (Canning,
1977). Because each cyst is founded by a single invading parasite, the number
of visible dermal cysts was counted to provide an index of the level of
G. anomala parasitism. Although the index is not a precise measure, because the
cysts can merge in heavy infections and may also be present internally, the
number of visible dermal cysts was regarded as an acceptable approximation of
the relative infection level. A total of 913 individuals were collected and a small
spine clipping (c. 3 mm) was taken from one of the ventral bony spines of 745
fish for molecular sexing before being returned to the pond. Small fry were
collected whole.
The results of molecular sexing of adults sampled during the breeding season
agreed 100% with estimates of their sex based on morphological cues (Griffiths
et al., 2000). A logistic regression model (GLM version 4.09) was performed to
examine the sex of individuals in relation to a number of predictors. The
following were entered into the model as factors; capture method (both types
of net or trap) and ‘age’ (YOY or adult, discriminated by body size). The month
of capture (March to October) was entered into the model as a co-variate.
Statistics were based on the w2 distribution.
Adult males had more G. anomala cysts than females (ANOVA, d.f. ¼ 1 and
592, P ¼ 002), but there was no sex difference in G. anomala cysts among YOY
(ANOVA, d.f. ¼ 1 and 149, P > 05). The adult sex difference in G. anomala
infection varied throughout the year, with males becoming relatively more
infected at the end of the year (Fig. 1; two-way ANOVA, sex month, d.f. ¼ 6
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2003 The Fisheries Society of the British Isles, Journal of Fish Biology 2003, 63, 1–5
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SEX-SPECIFIC PARASITISM
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Log10 number of cysts
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FIG. 1. Box-and-whisker plots showing the number of dermal Glugea anomala cysts (logþ1 transformed)
on (a) adult and (b) young-of-the-year Gasterosteus aculeatus caught at different times of the year.
Males (&) and females ( ) are shown separately. Horizontal bars are medians, boxes are interquartile range, whiskers show 95% CI and *, outliers.
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and 594, P ¼ 0001). YOY fish had more dermal G. anomala cysts than adults
(ANOVA d.f. ¼ 1 and 743, P < 00001). Among YOY, G. anomala parasitism
rate increased with age (two-way ANOVA, d.f. ¼ 3 and 150, P < 00001), but
there were no sex differences (Fig. 1).
Overall there was a female biased sex ratio in this population
(M : F ¼ 343 : 402; w2, P < 005). This bias, however, tended to be among adults
(M : F ¼ 277 : 324; w2, P ¼ 0055), not among YOY (M : F ¼ 66 : 78; w2, P > 03).
The GLM analyses revealed that the sex ratio varied between months (Fig. 2;
GLM Null model – scaled deviance ¼ 10281, d.f. ¼ 744, month of capture
d.f. ¼ 1, P < 0025). The sample was significantly male biased in May
(M : F ¼ 37 : 15; w2, P ¼ 0002), possibly because males were easier to catch
during the spawning season. Males are territorial and nest close to the edges
of the Inverleith Pond whereas females form shoals and move around the pond
(inshore and in the middle) (unpubl. data). The adult population was female
biased towards the end of the season (Fig. 2), as predicted in a species with
exclusive male parental care. This female bias amongst adult fish was significant
in August (M : F ¼ 62 : 93, w2, P ¼ 0013). Neither age nor capture method
influenced the sex of individuals in the model.
Adult females were longer (Fig. 2; ANOVA, d.f. ¼ 1 and 592, P < 0001) and
heavier than males (ANOVA, d.f. ¼ 1 and 592, P < 0001), but YOY were not
significantly size dimorphic (ANOVA, d.f. ¼ 1 and 149, LS: P > 05 and mass:
P > 04). Adult mass differences were at least partially due to females being
gravid from April to August, but females were also longer than males.
Amongst the three-spined sticklebacks sampled from this population, an even
sex ratio was found among YOY and adults at the beginning of the breeding
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2003 The Fisheries Society of the British Isles, Journal of Fish Biology 2003, 63, 1–5
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K. E. ARNOLD ET AL.
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10 12 14 16 18 20 22 24 26 28 30 32 34 36 38 40 42 44 46 48 50 52 54 56 58 60 62 64 66 68 70
Length (mm)
FIG. 2. Standard length frequencies ( , young-of-the-year and &, adults) and sex ratios (*, male and *,
female) for each month of capture. Mean standard lengths of males (--*--) and females (-- --)
amongst adult and YOY samples are also shown.
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season. The sex ratio became female biased during and after breeding, as
predicted from the sex differences in nuptial colouration, territorial and parental
care behaviour. Although a variety of sampling techniques was used, it is
unclear if these differences were due to a season–sex interaction in catchability,
or higher male than female mortality rates. Adult males harboured more
G. anomala cysts than females, these sex differences in infection level might be
due to the heavy demands of sexual selection (Moore & Wilson, 2002) and
fatherhood, or sex differences in the rate of encountering parasites because of
differences in ecology (Reimchen & Nosil, 2001). Parasitism rates in both adults
and YOY increased towards the end of the summer. Among adults, males were
disproportionately affected by G. anomala especially in July, just after the main
peak in paternal care.
References
Andersson, M. (1994). Sexual Selection. Princeton, NJ: Princeton University Press.
Canning, E. U. (1977). Microsporidia. In Parasitic Protozoa, Vol. IV (Kreier, J. P., ed.).
pp. 155–196. London: Academic Press.
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2003 The Fisheries Society of the British Isles, Journal of Fish Biology 2003, 63, 1–5
SEX-SPECIFIC PARASITISM
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Griffiths, R., Orr, K. J., Adam, A. & Barber, I. (2000). DNA sex identification in the
three-spined stickleback. Journal of Fish Biology 57, 1331–1334. doi: 10.1006/
jfbi.2000.1386.
Moore, S. L. & Wilson, K. (2002). Parasites as a viability cost of sexual selection in
natural populations of mammals. Science 297, 2015–2018.
Reimchen, T. E. & Nosil, P. (2001). Ecological causes of sex-biased parasitism in threespine
stickleback. Biological Journal of the Linnean Society 73, 51–63.
Tierney, J. F. (1991). Studies on the life history of Schistocephalus solidus: field observations
and laboratory experiments. PhD Thesis. University of Glasgow.
Wootton, R. J. (1976). The Biology of the Sticklebacks. London: Academic Press.
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2003 The Fisheries Society of the British Isles, Journal of Fish Biology 2003, 63, 1–5
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