Discussion
We employed a large number of presumably unlinked neutral markers to
test for species replacement of marbled newts in Portugal. Northwards
hybrid zone movement has been suggested in this system, with the
reported T. marmoratus enclave signalling the competitive advance
of T. pygmaeus with incomplete replacement (Wielstra,Burke, Butlin, & Arntzen, 2017a). Species displacement might be driven
by the desertification of the region, conferring T. pygmaeus a
competitive advantage over its sister species. It is knownT. pygmaeus thrives in ephemeral water bodies, and environmental
modelling here suggests it is favoured by arid precipitation regimes,
whereas T. marmoratus prefers smaller and more permanent breeding
sites under more humid conditions (Espregueira Themudo & Arntzen, 2007;
Harrison & Rand, 1989). In fact, climatic simulations also suggest the
North Atlantic jet stream, associated with wetter conditions, was at its
southernmost position during the Last Glacial Maximum, and it
experienced a positive latitudinal shift after this period, parallel toT. marmoratus ’ estimated northwards retreat (Beghin et
al., 2016). Unidirectional introgression of functionally and physically
unlinked neutral markers was expected in the wake of a moving hybrid
zone (Wielstra, Burke, Butlin, & Arntzen, 2017a). Yet, we found
no evidence for an additional enclave or a genomic footprint consistent
with enclave formation. Thus, two main biogeographic scenarios of
enclave formation arise in the absence of a footprint: i) the species
could have undergone negligible introgression or ii) displacement could
have in fact occurred with introgressive hybridisation, but with the
signal subsequently lost.
Species replacement in the absence of locally extensive hybridisation
during T. pygmaeus’ expansion could be responsible for enclave
formation unaccompanied by a genomic footprint. Displacement might have
occurred in allopatry, involving an initial recession of T.
marmoratus’ range during the Pleistocene glaciations, followed by a
northwards postglacial expansion of T. pygmaeus . Under this
scenario, the species would not have been in contact across their
western ranges until meeting at the current hybrid zone. The T.
marmoratus enclave would have locally persisted during the retreat of
its main range, alike the fragmenting pattern of contracting species
ranges described by Wilson, Thomas, Fox, Roy, and Kunin (2004), withT. pygmaeus later enveloping the pocket. However, it seems
unlikely the marbled newts did not meet after the Last Glacial Maximum,
given the similarity in suitable habitats predicted by the environmental
modelling.
Alternatively, species replacement may have taken place sympatrically,
with strong reproductive isolation preventing introgressive
hybridisation. Prezygotic and postzygotic barriers could be limiting
hybridisation among the species, resulting in reproductive isolation.
Prezygotic effects, such as mating preference or genetic
incompatibility, have not been reported in Triturus species,
whereas postzygotic effects occur between crested and marbled newts,
brought by the direction of the interspecific cross (Arntzen, Jehle,
Bardakci, Burke, & Wallis, 2009). The northwards advance of the
contiguous species ranges could therefore have been driven by species
replacement without introgressive hybridisation, with the enclave likely
persisting under T. marmoratus’ presumed strong adaptation to
local conditions.
The scenario of species replacement with hybrid zone movement is derived
from previous findings supporting a moving contact zone between the
marbled newts (Espregueira Themudo & Arntzen, 2007; Espregueira
Themudo, Nieman, & Arntzen, 2012). Assuming widespread interspecific
hybridisation, the erosion of T. marmoratus’ molecular signal
would explain the absence of a genomic footprint. The time passed sinceT. marmoratus inhabited the Lisbon Peninsula, environmental
effects, and the competitive advance of T. pygmaeus might have
led to the erosion of the footprint. Additionally, strong selection
against hybrids, as suggested by the strong bimodality of the species’
contact zone with few admixed individuals in Arntzen (2018), could have
swiftly erased the footprint. Purifying selection could have operated to
maintain the species’ functional integrity, by eliminating deleterious
sequences being pulled into the receding taxon (Oleksyk, Smith, &
O’Brien, 2010). Under strong purifying selection, existing introgression
of neutral variants might not have been detected despite the large
number of markers here employed. In fact, the significant instances of
heterozygote deficit and admixture linkage disequilibrium are likely due
to a combination of incomplete lineage sorting or ancestral polymorphism
and relatedness amongst the sampled larvae, resulting from the
collection of siblings that can potentially bias the landscape genetic
structure of populations (Goldberg & Waits, 2010).
The proposed biogeographic scenarios of species replacement are subject
to the unknown past position of the species boundary. While
environmental modelling supports T. marmoratus inhabiting the
study area, it remains possible that this species did not previously
inhabit the Lisbon Peninsula. Moreover, the absence of an additional
enclave or disconnected footprints in mountainous areas, such as Serra
de Sintra, does not clarify T. marmoratus’ past range nor whether
species replacement occurred with or without hybridisation. If in the
past, the southernmost range of T. marmoratus included Caldas da
Rainha, enclave formation could have occurred via incomplete species
replacement. Thus, testing for a genomic footprint north of the enclave
could potentially elucidate among the aforementioned biogeographic
scenarios. We expect future studies on the area between the T.
marmoratus enclave and its main distribution range to unravel a genomic
footprint of species replacement with hybridisation, confirming the
dynamic nature of the hybrid zone movement between the marbled newts.
Remarkably, the enclave in Caldas da Rainha showed notable levels of
introgression, possibly signalling the beginning of its erosion, as
predicted by Espregueira Themudo & Arntzen (2007), and further
illustrating the dynamics of species replacement. The enclave is
expected to eventually disappear under T. pygmaeus’ competitive
advance, leading to the loss of T. marmoratus gene variants over
time. The expansion of T. pygmaeus is likely influenced by the
loss of breeding sites in southern Spain and Portugal, an area with
unique biodiversity patterns stricken by a decline in temporary ponds,
driven by climate warming, desertification and agricultural
intensification (Arntzen et al. , 2004; Thomas, Franco, & Hill,
2006; van de Vliet et al. , 2014). Understanding shifts in species
distributions, particularly when driven by climate change and
anthropogenic activities, therefore becomes especially relevant in
deciphering the dynamics of species replacement (Taylor, Larson, &
Harrison, 2015).