Water chemistry variation promotes adaptive radiation in three-spined stickleback (Gasterosteus aculeatus)
Mahmuda Begum1, Victoria Nolan1 and Andrew D. C. MacColl1
School of Life Sciences, University of Nottingham, Nottingham NG7 2RD, United Kingdom.
Corresponding author
Mahmuda Begum PhD fellow
School of Life Sciences,
University of Nottingham,
Nottingham NG7 2RD,
The United Kingdom
E-mail: mahmudabegum.bcsir@gmail.com
Abstract
The context and cause of adaptive radiations has been widely described and explored but why rapid evolutionary diversification does not occur in related evolutionary lineages has yet to be understood. One possible answer to this is simply that evolutionary diversification is provoked by environmental diversity, and that some lineages do not encounter the necessary environmental diversity. Three-spined stickleback on the Scottish island of North Uist show enormous diversification, which seems to be associated with the diversity of aquatic habitats. Stickleback on the neighbouring island of South Uist have not been reported to show the same level of evolutionary diversity, despite levels of environmental variation that we might expect to be similar to North Uist. In this study, we compared patterns of morphological and environmental diversity on North and South Uist. Ancestral anadromous stickleback from both islands exhibited similar morphology including size and bony ‘armour’. Resident stickleback showed significant variation in armour traits in relation to pH of water. However, North Uist stickleback exhibited greater diversity of morphological traits than South Uist and this was associated with greater diversity in pH of the waters of lochs on North Uist. Highly acidic and highly alkaline freshwater habitats are missing, or uncommon, on South Uist. Thus, pH appears to act as a causal factor driving the evolutionary diversification of stickleback in local adaptation in North and South Uist. This is consistent with diversification being more associated with ecological constraint than ecological opportunity.
Keywords : Stickleback, morphometric study, North Uist, South Uist.
Introduction
Studies of adaptive radiation have tended to concentrate, for obvious reasons, on lineages and locations in which it is known to have occurred. The circumstances associated with a failure to radiate are less well explored, yet there is much that might be learned from them about the conditions that favour divergence. Differences in ecological conditions among local populations are generally assumed to be the reason for adaptive divergence leading to adaptive radiation, reproductive isolation and speciation (Schluter 1996; Schluter 2000; MacColl 2011). In particular, according to the ecological theory of adaptive radiation (Schluter, 2000), ‘ecological opportunity’, the diversity of available resources not used by other taxa, is central to explaining why adaptive radiations occur. Lineages often diversify when they colonise habitats where there is little competition for constraining resources (Simpson 1953, Schluter 2000). The concentration on biotic interactions, especially competition for food, that has followed from the idea of ecological opportunity, has meant that the role of abiotic environmental variation has been less well explored. Here, we scrutinise diversification of three-spined stickleback (Gasterosteus aculeatus ) in lochs on two neighbouring Scottish islands, for clues about the causes of adaptive radiation.
The three-spined stickleback (hereafter ‘stickleback’) has served as a model organism for the study of adaptive radiation due in part to parallel diversifications of freshwater populations from marine ancestors, that have occurred in the last 10,000 years ago (Bell and Foster 1994; Schluter 2000; Jones et al 2012; Magalhaes et al. 2021). Freshwater populations have evolved conspicuous differences in morphology, physiology and behaviour (Bell and Foster 1994; Ostlund-Nilsson et al. 2006). The divergence of morphological traits through major changes in the bony armour that have repeatedly evolved in various locations, are common components of adaptive radiation of stickleback (Colosimo et al. 2004; Chan et al. 2010). Ancestral marine anadromous sticklebacks are heavily armoured with a continuous row of 30-36 bony lateral plates running from head to tail on each side (known as a complete morph), and have dorsal spines, a well-developed pelvic girdle and further spines attached to the girdle (Colosimo et al. 2005; Barrett et al. 2008). On the other hand, derived freshwater (live in freshwater year round) and saltwater-resident (inhabit coastal saltwater all year round without migration to the open sea) stickleback exhibit substantial reduction in the total plate number with either a discontinuous row of 9-28 plates (partial morph) or with 0-9 lateral plates at the anterior end (low morph), and reductions in the size of spines and girdle (Colosimo et al. 2004; Colosimo et al. 2005). In addition, other phenotypic traits such as body size and shape of freshwater and saltwater-resident stickleback show morphological transformation from the ancestral anadromous form (Moodie and Reimchen 1976; Schluter 1993; Bell et al. 1993; Schluter et al. 2004; Shapiro et al. 2004; Ravinet et al. 2015).
To date, several abiotic and biotic environmental factors have been considered as causes for these major phenotypic changes including variations in salinity, temperature, nutrient and calcium availability, stream gradient, predators and parasites (Giles 1983; Bergstrom 2002; Barrett et al. 2009; Myhre and Klepaker 2009; Marchinko 2009; Wark and Peichel 2010). For example, predation-driven selection has been reported to influence the evolution of bony armour structures such as the lateral plates and spines within marine and freshwater stickleback populations (Marchinko 2009). Moreover, other factors such as reduced nutrients or salinity, and calcium ion deficiency have also been reported to have an association with the loss of lateral plates in freshwater populations (Giles 1983; Barrett et al. 2009; Myhre and Klepaker 2009).
The neighbouring islands of North and South Uist, in the Scottish Western Isles, appear to have much in common from the perspective of aquatic habitats and species. Both have large numbers of small to medium-sized shallow lochs, that occur over a variety of surface geology, from peat and bedrock on their eastern sides, to shell-rich machair sand on their western sides. Although fish, including three-spined stickleback, populations have been surveyed on both islands (Campbell and Williamson, 1979; Campbell, 1985), detailed studies of phenotypic variation in the stickleback have only taken place on North Uist (Giles 1983; MacColl et al. 2013; Magalhaes et al. 2016; Magalhaes et al. 2021) apparently because of a lack of such variation on South Uist (Campbell 1985).
North Uist comprises a mosaic of interconnected freshwater and brackish water lochs and lagoons which are known to have exceptional variation in their water chemistry, with high pH (~8) in the western machair lochs in the west and low pH (~6) in the east. These are associated with variation in the concentrations of alkaline metals (sodium, potassium, magnesium, calcium etc.) (Waterston et al.1979). This variation has been shown to correlate with the evolution of high diversity of stickleback populations across the island (Giles 1983; MacColl et al. 2013; Magalhaes et al. 2016; Haenel et al. 2019) and offers a unique opportunity to study the variation in phenotypes in relation to adaptation under different environmental conditions.
In contrast, environmental conditions and phenotypic variation have been poorly explored in South Uist, and there have been no reports of unusual morphological variation in stickleback. While different in topography (it has higher hills and less low-lying ground), it appears to have similar variation in aquatic environments, with both saline and freshwater lochs, and both western, machair lochs and eastern lochs on peat or bedrock. This begs the question why, on two such closely neighbouring islands, where ancestral variation is likely to be shared, and environmental variation is similar, there appears to have been little or no diversification of stickleback on one, while the other exhibits some of the most dramatic variation known in the species. Here we make a direct comparison of the morphological and environmental variation in the two islands, and relate the former to the latter, to understand what factors may promote or inhibit adaptive radiation at local scales. We hypothesised that morphological traits of three-spined stickleback would show variation between South Uist and North Uist due to differences in the environmental diversity of the lochs.
Materials and methods
2.1 Study area
Two neighbouring islands with apparently similar aquatic environments were selected to make a comparative study of morphological variation in three-spined stickleback. North Uist (57°31’12”N; 7°27’42”W) is in the centre of the Western Isles of Scotland and is approximately 303 km2in total area (Fig. 1B). It comprises a mosaic of peat bogs, heathland and low hills which makes it different from South Uist. South Uist (57°13’54”N; 7°02’38”W) is the second largest island of the Outer Hebrides (Fig. 1C). It is around 320 km2 in total area and differs greatly between its west and east sides.
2.2 Sample collection
Stickleback samples were collected from 10 lochs on North Uist and 8 lochs on South Uist between the dates of 06th – 16th May 2019 (Fig. 1). Locations on both islands were selected to maximise likely variation in surface geology (sand versus peat or bedrock), and hence water chemistry, using Google Earth. Stickleback were caught in unbaited Gee’s Minnow Traps (Gee traps, Dynamic Aqua, Vancouver, Canada), set overnight (approximately 16 hrs.) at all sites. The geolocation of each sample site (GPSmap 60CSx, Garmin, UK) and water quality parameters including temperature, absolute conductivity, salinity and pH (multi-parameter probe - Multi340/set, WTW, Germany) were recorded from all sites (Table A1 - A2). Lochs were grouped based on the pH of water: high pH >7.5, neutral pH = >7.0 - 7.4, low pH< 7.0 (Table A1 - A2).
Across all ten sampling sites of North Uist, 135 stickleback were haphazardly selected (Fig. 1B). From South Uist, 128 fish were collected in the same way from 11 sites (3 sites from loch Aroa) in eight lochs (Fig. 1C). Of the total 263 stickleback, 45 were anadromous fish sampled from both islands to estimate the ancestral state of stickleback in the Uist lochs. Fish were euthanized and preserved in 70% ethanol for morphometric study.
2.3.Body armour and spine data collection for morphometric analysis
To collect data on external bony skeletal structures (armour), ethanol-preserved samples were stained with Alizarin Red solution following a standard staining method (Peichel et al. 2001). Samples were stored in 40% isopropyl alcohol (propan-2-ol). After confirming the appearance of bony parts, a digital photograph of the right side of every fish was captured using a digital SLR camera (Nikon D5200) with 60 mm macro lens, digital ring flash and a tripod (to set a fixed distance). All photographs included graph paper as a scale (cm) and the measurements of standard length (from the tip of the snout to the end of caudal peduncle), first and second dorsal spine, pelvic spine (from insertion point to the tip), pelvis height and pelvis length were recorded using tpsDig2 version 2.31 (Rohlf 2010) (Fig. Appendix, A1). The total number of lateral plates was also counted from the right side of the stained photograph.
2.4 Data analysis
Data were collated in Excel (Microsoft) and statistical analyses were conducted using R, version 3.6.3 (R Core Team 2020).