Landscape genetics across the Andes mountains: Environmental
variation drives genetic divergence in the leaf-cutting ant Atta
cephalotes
Vanessa Muñoz-Valencia1, James
Montoya-Lerma1, Perttu Seppä2 &
Fernando Diaz3
1 Group of Agroecosystem Ecology and Natural Habitats,
Department of Biology, Faculty of Natural Science, Universidad del
Valle, Cali, Colombia.
2 Organismal and Evolutionary Biology Research
Programme, Faculty of Biological and Environmental Sciences, University
of Helsinki, Finland.
3 Biology Department, Colgate University, Hamilton,
NY, USA.
Corresponding authors:
Vanessa Muñoz-Valencia,
vanem28@gmail.com; James
Montoya-Lerma,
james.montoya@correounivalle.edu.co;
Fernando Diaz,
ferdiazfer@gmail.com
ABSTRACT
Distinguishing among the mechanisms underlying the spatial distribution
of genetic variation resulting from the environmental or physical
barriers from those arising due to simple geographic distance is
challenging in complex landscapes. The Andean uplift represents one of
the most heterogeneous habitats where these questions remain unexplored
since multiple mechanisms may interact, confounding their relative
roles. We explore this broad question in the leaf-cutting ant Atta
cephalotes , a species that is distributed across the Andes mountains,
using nuclear microsatellite markers and mtCOI gene sequences. We
investigate spatial genetic divergence across the western range of the
northern Andes in Colombia by testing the relative role of alternative
scenarios of population divergence, including isolation by geographic
distance (IBD), climatic conditions (IBE), and the physical barriers
presented by the Andes mountains (IBB). Our results reveal substantial
genetic differentiation among A. cephalotes populations for both
types of markers, but only nuclear divergence followed a hierarchical
pattern with multiple models of genetic divergence imposed by the
western range. Model selection showed that the IBD, IBE (temperature and
precipitation), and IBB (Andes mountains) models, often proposed as
individual drivers of genetic divergence, interact and explain up to
33% of the genetic divergence in A. cephalotes . The IBE model
remained significant after accounting for IBD, suggesting that
environmental factors play a more prominent role than with IBB. These
factors, in combination with the idiosyncratic dispersal patterns of
ants, appear to determine the hierarchical patterns of gene flow. This
study enriches our understanding of the forces shaping population
divergence in complex habitat landscapes.
KEYWORDS
Andean uplift, western mountain range, spatial genetic structure,
isolation by distance, isolation by barrier, model selection, isolation
by environment.
INTRODUCTION
Population divergence can be determined by the spatial distribution of
individuals, where geographic proximity modulates genetic similarity,
leading to a pattern of isolation by distance – IBD (Lee &
Mitchell-Olds, 2011; Shafer & Wolf, 2013; Slatkin, 1993; Wright, 1943).
However, populations in complex landscapes are often exposed to
environmental variation and physical barriers that can also contribute
to genetic divergence (Manel & Holderegger, 2013; Manel, Schwartz,
Luikart, & Taberlet, 2003; Shafer & Wolf, 2013). Populations could
adapt to their local environment, maximizing the fitness of individuals
under local conditions while decreasing the fitness of immigrants from
alternative environments (Carro, Quintela, Ruiz, & Barreiro, 2019;
Sobel, 2014; Wang & Bradburd, 2014; Wang, Glor, & Losos, 2013). This
adaptive reduction in gene flow can produce a pattern of isolation by
environment – IBE (Sexton, Hangartner, & Hoffmann, 2014; Shafer &
Wolf, 2013; Wang & Bradburd, 2014). Alternatively, gene flow could be
restricted by allopatric scenarios of genetic differentiation mediated
by physical barriers in the landscape, generating patterns of isolation
by barrier – IBB (De Queiroz, Torrente-Vilara, Quilodran, da Costa
Doria, & Montoya-Burgos, 2017; Haffer, 2008; Rull, 2011;
Turchetto-Zolet, Pinheiro, Salgueiro, & Palma-Silva, 2013). With
increasing landscape complexity, gene flow is likely to be influenced by
a combination of geographical and ecological factors in which these
isolating mechanisms are not mutually exclusive (Crispo, Bentzen,
Reznick, Kinnison, & Hendry, 2006; Edwards, Keogh, & Knowles, 2012;
Noguerales, Cordero, & Ortego, 2016; Wang et al., 2013).
The Andean mountain ranges not only represent one of the most unexplored
environments, but also offer a great complexity of landscapes, promoting
the diversification of a wide range of taxa (Salgado-Roa et al., 2018).
Across these mountains, restricted gene flow has been reported in
multiple organisms, including birds (Cadena, Pedraza, & Brumfield,
2016), plants (Lagomarsino, Condamine, Antonelli, Mulch, & Davis, 2016;
Luebert & Weigend, 2014; Pérez-Escobar et al., 2017), mammals
(Antonelli et al., 2009; Hoorn et al., 2010), insects (Antonelli et al.,
2009; De-Silva et al., 2017; Hoorn et al., 2010), and other arthropods
(Salgado-Roa et al., 2018). However, organisms from contrasting
populations in these habitats are often exposed to a combination of
distance, environmental, and physical barriers to dispersal, challenging
the investigation of the relative roles of different isolating
mechanisms (James, Coltman, Murray, Hamelin, & Sperling, 2011;
Meirmans, 2015; Noguerales et al., 2016). It remains unclear how the
Andean uplift has modulated patterns of gene flow and the evolution of
several of the most ecologically important groups in the Neotropics,
including social insects. For example, although the entire evolution of
Neotropical ants occurs across the Andes (Mueller et al., 2017), the
interplay between their population structure and environmental variation
relative to the effect of these mountains on isolated populations
remains largely unknown.
The leaf-cutting ant A. cephalotes is a major urban and
agricultural pest in the Neotropics, colonizing a wide spectrum of
environments (Della Lucia, Gandra, & Guedes, 2014; Fernández,
Castro-Huertas, & Serna, 2015; Hölldobler & Wilson, 2011). In
Colombia, its distribution overlaps with the maximum complexity of the
Andean uplift, ranging from 0 to 2100 m.a.s.l. (Fernández et al., 2015),
with a vertical thermal gradient of 0.6 °C/100 m (Hermelin, 2015). The
northern section of these mountains in Colombia splits into three main
branches: the western, central, and eastern ranges. Populations ofA. cephalotes are separated by these mountains while
simultaneously being exposed to complex combinations of topographical
and environmental variation (Kattan, Franco, Rojas, & Morales, 2004;
Pérez-Escobar et al., 2017; Salgado-Roa et al., 2018). Such conditions
provide a tremendous climatic spectrum for local adaptation, with an
interplay between population dynamics and species-specific dispersal
patterns (Hakala, Seppä, & Helanterä, 2019). Evolution under such
environmental heterogeneity could act to shape patterns of gene flow (De
Queiroz et al., 2017; Lee & Mitchell-Olds, 2011; Noguerales et al.,
2016; Wang et al., 2013), which often produces more complex scenarios
than genetic divergence due to IBD alone (Slatkin, 1993; Wright, 1943).
For example, we recently found that the eastern range of the Andes in
Colombia plays a major role as a geographic barrier to historical gene
flow, restricting the dispersion of A. cephalotes from north to
south (Muñoz-Valencia, Vélez-Matínez, Montoya-Lerma, & Díaz, 2021).
Although this initial study demonstrates the significant influence of
the Andes on population divergence in the leaf-cutting ant at the
phylogeographic scale, the role of local adaptation occurring at more
regional scales across the Andes remains untested.
This study focuses on a finer and more complex geographic distribution
scale of A. cephalotes : that of the western range of the Andean
uplift in Colombia. We use a landscape genetic approach to investigate
the role of geographic features and environmental variation in the
definition of patterns of spatial genetic structure in A.
cephalotes . Using nuclear (microsatellites) and mitochondrial
(mtCOI ) markers, we test the relative roles played by geographic
distance, climate variation, and a major dispersal barrier (the western
range) in modulating patterns of gene flow. As a monogynous
(single-queen) species, A. cephalotes presumably undertakes
long-distance nuptial flights that can potentially overcome isolating
barriers (Cherrett, 1968; Helms, 2018; Moser, 1967). Our results
demonstrate that gene flow is limited by a complex interaction of the
three isolating mechanisms (IBD, IBE, and IBB) rather than IBD alone,
while IBE appears to play a stronger role than IBB. Investigating the
spatial genetic structure of a species in an exceptionally heterogeneous
environment helps to elucidate the evolution and diversity of this
ecologically dominant group of ants in the Neotropics.
MATERIALS AND METHODS
Sampling
Ant sampling was conducted in the Colombian Pacific and Andean regions,
which are separated by the western mountain range of the Colombian Andes
(Figure 1). The Pacific region is classified as a tropical rainforest
with an extremely humid climate and an annual average temperature of 27
°C. The Andean region is further divided into two groups: Andean 1 (800
- 1050 m.a.s.l.) and Andean 2 (1300 - 2200 m.a.s.l.), with highly
variable climatic conditions. The inner valleys in Andean 1 are
climatically classified as tropical savanna and tend to be dry, with an
annual temperature of 25 °C, while
the range summits in Andean 2 are more humid, with a temperate climate,
tropical monsoons, and an annual temperature of 21 °C (Supplementary
table ST1) (Chen & Chen, 2013; Hernández-Camacho, 1992; Kattan et al.,
2004; Köppen, 1884; Peel, Finlayson, & McMahon, 2007).
Environmental variation across the three regions was characterized by
differences in temperature, humidity, and precipitation, measured as
five-year averages of the annual temperature (°C), relative humidity
(%), and precipitation (mm), respectively, for each location (IDEAM,
2019). In addition, a climate classification was represented using four
categories (1 to 4) of different climatic conditions mediated by the
tropical Andes. Variation in topography was estimated by elevation above
sea level. A dummy variable was used to evaluate the Andean uplift as a
major geographic barrier to gene flow. The code 0 was used for
populations from the western side of the western range (Pacific region),
and 1 for populations from the eastern side of these mountains
(Supplementary table ST1).
Worker ants from nine to twenty nests in ten locations (total of 153
nests) were sampled in the period 2017-2018 (Table 1). The distance
between nests in each location was at least 1.5 km, ensuring that the
sampled nests were independent colonies. Three, two, and five locations
were sampled from the Pacific, Andean 1, and Andean 2 regions,
respectively (Table 1).
Molecular methods
DNA extraction and PCR
amplification of microsatellite
markers
Total DNA extraction was carried out for five workers from each nest
(total 765 workers) using TNES lysis buffer (Tris 50mM, NaCl 0.4M, EDTA
100mM, SDS 0.5%), pH 7.5, and chloroform:isoamyl alcohol (24:1),
following Wasko et al. (2003), with minor modifications as described by
Muñoz-Valencia et al. (2020).
Thirteen microsatellite loci developed for A. cephalotes(Muñoz-Valencia et al., 2020) were used (Supplementary Table ST2). PCR
reactions were carried out following Muñoz-Valencia et al. (2020) in a
10 µL volume containing 10 ng of DNA, 1 X Phusion Flash PCR Master Mix
(Thermo Fisher Scientific), and 2 µM of each labeled primer. The thermal
profile was: 98 °C for 1 min, followed by 34 cycles of 98 °C for 1 s,
annealing temperature for 15 s, and 72 °C for 20 s, followed by a final
extension step at 72 °C for 1 min. The fluorescent amplified fragments
were visualized using an automated DNA sequencer ABI 3130 Genetic
Analyzer (Applied Biosystems), and allele sizes were estimated using
GeneMapper version 4.0 (Thermo Fisher Scientific).
Sequencing of the mtCOI
gene
A 368 bp fragment of the mitochondrial cytochrome oxidase subunit I gene
(mtCOI ) was sequenced from one sample per nest, obtaining a total
of 146 sequences after discarding failed amplifications and sequencing
(GenBank accession numbers: MW245066 - MW245211). PCR amplification was
carried out using the universal primers Ben and Jerry, following
Kronauer et al. (2004) and Simon et al. (1994). PCR reactions were
performed in a 10 µl volume, containing 10 ng of DNA, 1X GoTaq® Master
Mix (PROMEGA), and 2 µM of each primer. The thermal cycling profile was
94 °C for 2 min followed by 30 cycles of 94 °C for 1 min, 58 °C for 1
min, and 72 °C for 1 min, with a final extension step at 72 °C for 10
min. PCR products were confirmed by electrophoresis on 1 % agarose
gels. All PCR products were sequenced by Psomagen, Maryland, USA.
Population genetics analyses