Environmental factors affecting the variation of morphological
traits in North Uist and South Uist
To quantify associations between morphological traits of resident
stickleback and different environmental factors of the loch water in
North Uist and South Uist, GLMs were fitted with a Gaussian distribution
and identity link function. PC1 and PC2 from the PCA of armour traits
were fitted as response variables, with location (North or South Uist),
pH, salinity and conductivity as predictor variables. GLMs using the
mean of each morphological trait (standard length, 1stdorsal spine length, 2nd dorsal spine length, pelvic
spine length, length of pelvis, height of pelvis and plate count) of
each population as the response variable were fitted in relation to pH
value, conductivity and salinity of loch water. A Scheffe post-hoc test
(Scheffe 1951) for multiple comparisons of mean with unequal sample
sizes was performed to observe the differences in the size of measured
armour traits among four low pH lochs in North Uist.
3. Results
3.1 Variation in
morphological traits between North Uist and South
Uist
The first two PCs from the PCA of the armour traits for the anadromous
stickleback from both North and South Uist accounted for 58% of the
total variation. All measured armour traits had positive loadings for
PC1 and PC2 except pelvic spine length and length of pelvis (Fig. 2).
Overall, anadromous fish from North and South Uist were morphologically
similar, with some exceptions. The mean height of the pelvis of South
Uist anadromous fish was significantly greater than North Uist (one-way
ANOVA: F = 5.19, df = 1, 44, p = 0.02; Table 1). The standard length of
South Uist anadromous stickleback showed significantly less variation
than North Uist fish (MSLRT = 4.15, p = 0.04) (Fig. 3).
Resident populations of stickleback from North and South Uist were
significantly smaller (smaller mean standard length) than the anadromous
populations (W = 9742, p < 0.001; Fig. 3). Mean standard
length of the North Uist resident stickleback (mean ± S.E. 36.44 ± 0.64
mm) was significantly greater than South Uist fish (34.55 ± 0.53 mm)
(one-way ANOVA: F= 5.21, df = 1, 216, p= 0.02; Fig. 3), although the
difference in length is small.
The first two principal components of a PCA of armour traits for
resident fish collected from North and South Uist explained
approximately 85% of the variance among individuals (Fig. 4A). PC1 was
strongly correlated with all measures of armour traits so that high
values of PC1 are associated with more armour. Plate count was highly
correlated with PC2 and had a stronger influence on the variation in
North Uist fish than South Uist. All measured armour traits (except
plate count) were strongly correlated with each other Pearson’s
correlation (r)>0.5 (Fig. 4B). For resident fish, despite
no significant differences in trait means, there was significantly more
variation (CV) in all traits (except standard length) in North Uist than
South Uist (Table 1).
3.2 Factors affecting the
variation of morphological traits in North Uist and South Uist
There was wide variation in many of the environmental factors including
salinity and pH across the 8 lochs of South Uist and 10 lochs of North
Uist: loch pH ranged from low (6.5, acidic) to high (~
9, alkaline) pH of water (one-way ANOVA: F = 10.28, df = 1, 216, p =
0.001) in the freshwater lochs of North Uist compared to mostly neutral
(~ 7) lochs of South Uist (Table A1 and A2).
There were significant associations between the first two PCs of armour
traits and the pH of the 18 populations of resident fish collected from
North and South Uist (Table 2). There was striking morphological
variation in fish found in acidic freshwater lochs relative to other
lochs (Fig. 4).
All morphological traits of the resident fish showed significant
associations with pH levels in both North and South Uist (Table 2).
Total plate count showed different associations with pH of water across
the two locations with significantly lower number of plates in higher pH
on South Uist, although overall across both North and South Uist the
mean plate count was highest in the high pH populations (6.58 ± 1.41)
and lowest in freshwater low pH lochs of North Uist (1.86 ± 0.27)
(F1, 17 = 11.3, p<0.001; Fig. 5A). The mean
standard length of resident fish was highest in freshwater high pH
populations (37.02 ± 0.97 mm) and lowest in freshwater low pH (33.49 ±
0.92 mm) (F1, 17 = 4.85, p = 0.044; Fig. 5B). Among the
18 populations of resident fish of both islands, the mean length of all
measured armour variables increased with increasing pH (Fig. 5C-F). One
exception to the overall trend was in Tros (pH 6.63), were armour
measurements were relatively large. Post-hoc tests (Scheffe Test)
revealed that all measured armour traits Tros were significantly larger
in length than in the three other freshwater low pH populations of North
Uist (p < 0.001).
4. Discussion
Studies of adaptive radiation have tended to concentrate on what we can
learn from adaptive radiations themselves, rather than their absence. In
this study we have compared lacustrine populations of three-spined
stickleback from isolated habitats on two neighbouring Scottish islands.
North Uist has been known for decades to support a striking example of
adaptive radiation in this species (Campbell 1985; Magalhaes et al.
2021), apparently linked to variation in the abiotic and biotic
environments that is indexed by variation in pH (Giles 1983, MacColl et
al. 2013, Magalhaes et al. 2016, Haenel et al. 2019). In contrast,
variation in stickleback on South Uist has hardly been considered,
despite apparently similar variation in aquatic environments. We have
shown that in fact South Uist lacks the striking morphological variation
of North Uist, and this lack of evolutionary diversification is well
explained by a lack of variation in the pH of water bodies.
Anadromous stickleback, that are likely to provide a good approximation
for the ancestors of Uist freshwater stickleback showed similar
variation in armour traits and size on North and South Uist, consistent
with close common ancestry in the marine environment, and supporting the
idea that differences between the two islands arise because of
differences in freshwater. In contrast, resident fish collected from
North and South Uist showed variation in all measured armour traits
within and between populations which indicate there is adaptive
radiation of phenotypic characters among populations as a result of
colonization and adaptation to the freshwater environment (Bell and
Foster 1994; Magalhaes et al. 2021). There was significantly more
variation in armour traits in resident fish of North Uist than South
Uist which demonstrates differences in evolutionary diversification of
closely-related lineages.
The pH of the lochs showed strong associations with the armour traits of
resident fish collected from North and South Uist consistent with
several previous studies of stickleback adaptive evolution in freshwater
environments (Giles 1983; Bell and Foster 1994; Spence et al. 2012;
MacColl et al. 2013; Magalhaes et al. 2016). In the present study, the
armour traits of resident stickleback showed substantial variation in
dorsal spines, pelvic spines and pelvis length in the freshwater
populations of both islands. However, North Uist stickleback exhibited
much more morphological variation than the stickleback of South Uist and
this variation was directly associated with greater diversity in pH of
the loch water on North Uist. This suggests that the differing extent of
morphological diversification between North and South Uist is a direct
consequence of differences in some aspect or aspects of environment that
are related to pH.
It is not immediately obvious why there should be less environmental
variation on South, than North, Uist. At first sight, the surface
geology of both islands, the root cause of the pH variation (Waterston
et al.1979), is rather similar, with acidic, peaty lochs in the east and
alkaline, machair lochs in the west. On North Uist, the acidic lochs
where the most extreme phenotypes have evolved are relatively large.
Adjacent, smaller lochs with similar pH have less extreme phenotypes
(ADCM, personal observations). It may be that larger stickleback
populations in larger lochs facilitate evolution. On South Uist the
eastern lochs are generally smaller, but even the larger ones (e.g.
Druidibeg) do not contain unusual stickleback. In any case, the eastern
lochs in South Uist are clearly less acidic, but again it is unclear
why. Druidibeg is rather shallow, with a largely rocky bottom, and it
may be that a shorter residence time of water in the loch, coupled with
less contact with peat, may prevent the development of more extreme
acidity. The topography of South Uist is also rather different to that
of the North. North Uist is generally low-lying, and catchments drain in
a radial pattern, meaning that there is little variation in surface
geology within catchments. In contrast, South Uist is hilly in the east
and the main catchments drain to the west across the machair. The latter
pattern results in a kind of ‘environmental flow’ (movement of water
from east to west) that may reduce variation in water chemistry between
lochs, at least the development of more alkaline conditions in machair
lochs. These linear catchments may also facilitate gene flow between
lochs on South Uist that is absent on North Uist, and this could inhibit
diversification, but the restricted environmental variance alone on
South Uist appears sufficient to explain the reduced morphological
diversification, without invoking differences in gene flow.
It is not obvious exactly why phenotypic variation should be associated
with variation in pH, although on North Uist resource availability,
competition and predation regimes covary with pH (MacColl et al. 2013;
MacColl and Aucott, 2014; Magalhaes et al. 2016). It seems most likely
that many differences in ecology between alkaline and acidic lochs are
fundamentally linked to a reduction in nutrient availability in the
latter. This is consistent with the reduced size and armour that is
apparent in acidic populations. Body size of some low pH populations was
extremely small which mirrored the previous finding of dwarfism in
stickleback in association with low pH indicative of poor resource
conditions (Giles 1983; MacColl et al. 2013). The dorsal spines and
pelvic complexes were also either rudimentary or missing in most of the
populations from low pH lochs (except Tros, see below) in North Uist.
Previously, several studies have suggested that this striking variation
in armour was due to calcium ion limitations in acidic conditions (Giles
1983; Bourgeois et al. 1994). There is no obvious correlation on North
Uist between calcium availability and armour evolution, but it is likely
that there is a more general shortage of nutrients in the acidic lochs,
which are usually oligotrophic verging on dystrophic. This suggests that
the evolution of extreme morphological variation on North Uist is driven
more by ecological constraint than by ecological opportunity.
The striking exception provided by stickleback in Loch Trosavat (Tros)
to the general pattern of relationship between armour traits and pH is
illuminating, if anecdotal. Tros is linked to the sea by a short stream,
and this means that large, migratory salmonids (Atlantic salmon,Salmo salar and sea trout, S. trutta ) and potentially
other marine fish that prey upon stickleback, occur in this loch. This
by itself could explain the more developed armour in Tros stickleback,
but it is also likely that proximity to the sea results in Tros having a
better nutrient status than most acid lochs, which could compound the
effect of predation.
To conclude, the increased evolutionary variation of North Uist
stickleback populations compared to South Uist populations can be
explained by greater environmental variation on North Uist, especially
linked to variation in the pH of loch water. Research is on-going to
understand the role of pH in the ecology and evolution of Uist
stickleback populations.
Table 1: One-way analysis of variance (ANOVA) for mean value and
modified signed-likelihood ratio test (MSLRT) for coefficients of
variation (CV) of each morphological trait [standard length,
1st dorsal spine length (DSL), 2nddorsal spine length (DSL), pelvic spine length (PSL), length of pelvis
(LPS), height of pelvis (HPS) and plate count] of stickleback sample
fish (217 residents and 45 anadromous) collected from two locations
(North Uist and South Uist, Scotland). Significant P values are
in bold.