Introduction
Determining the mechanisms and processes that govern the assembly and
dynamics of plant communities is of major interest in ecology
(Sutherland et al., 2013), and over the last decades, a multitude of
coexistence mechanisms have been proposed (Chesson, 2000; Wright, 2002).
For example, the classical mechanism proposed by Janzen (1970) and
Connell (1971) postulates that coexistence in plant communities may be
maintained by specialized natural enemies that cause distance- and
density-dependent mortality of early life stages if they are located
close to conspecific adults or in areas of high conspecific density,
respectively, opening space to be used by heterospecific recruitment. A
stabilizing mechanism arises if individuals of a plant species are more
frequently attacked as the species becomes more common (a rare-species
advantage; Chesson, 2000; Comita & Stump, 2020). Typically, the
Janzen-Connell hypothesis is tested by assessing survival of seeds or
seedlings located close to adults, or in areas of high conspecific
density (Comita et al., 2014), but without explicitly accounting for the
organisms mediating these interactions at plant community level. While
demographic analyses can reveal if observed patterns are consistent with
the Janzen-Connell hypothesis (e.g., Comita et al., 2014), a mechanistic
understanding requires the identification of the principal classes of
plant-associated organisms driving these patterns (Mangan et al., 2010).
From a plant perspective, interactions with plant-associated organisms
can result in a range of effects, from detrimental to beneficial
(Bronstein, 2009). Interestingly, the negative effects of antagonists
such as herbivorous insects or plant pathogens can be neutralized or
reduced by mutualists like leaf epiphytes (Bacon & White, 2016;
Pajares-Murgó et al., 2022). The balance of this counteraction varies
between plant species, and contributes to plant community dynamics and
species coexistence (Agrawal & Maron, 2022; Chomicki et al., 2019;
Johnson, 2021). Because antagonists and mutualists operate typically in
a distance- and density-dependent manner at the plant neighbourhood
scale (Bagchi et al., 2010; Bell et al., 2006; Freckleton & Lewis,
2006), we need to investigate their spatial patterns to better
understanding how they may impact plant communities (Perea et al.,
2020).
The effect of mutualists and antagonists on plant recruitment is
influenced by the seed dispersal mechanism of the species and the
emerging “seedscape” (i.e., the environment surrounding a seed that
influences its success; Beckman & Rogers, 2013), setting the template
for subsequent processes of recruitment and interactions with its
neighbours (Nathan & Muller-Landau, 2000; Wiegand et al., 2021). A key
aspect of the seedscape is the load of plant antagonists (e.g.,
herbivorous insects, pathogens) and mutualists (e.g., epiphytes)
neighbouring the location of a seed (or seedling). Following the
Janzen-Connell hypothesis, neighbourhoods in the seedscape with a high
load of antagonists, shared with the seed species, will be harmful for
the emerging recruits, while neighbourhoods with a low load of
antagonists will be beneficial or neutral. However, little is known
about how different antagonist and mutualist organisms distribute among
adult and recruit plants and how they shape the seedscape (Beckman &
Sullivan, 2023).
Different seed dispersal
mechanisms will lead to different densities and spatial patterns in the
deposition of seeds in the seedscape, with important consequences for
subsequent process of self-thinning (Schupp et al., 2010; Schupp &
Fuentes, 1995). For example, zoochorous dispersal generally transports
the seeds away from mother plants, placing them into locations where the
seeds share less antagonists with neighbouring plants compared to
locations close to mother plants. As example, frugivorous birds drop
seeds of multiple species under adult plants of other species, leading
to multispecies clumps of seeds, seedlings and saplings (Herrera, 1984;
Jordano, 2014; Perea et al., 2021). Under these circumstances it is
expected that density-responsive antagonists have only moderate effects
on the fate of the recruits. In contrast, larger proportions of seeds
located close to the parents may suffer strong effects of antagonists
(Spiegel & Nathan, 2010). If so, seeds and seedlings will show reduced
survival in areas of high share of antagonists, and the remaining
recruits that survive into the saplings stage will be located in areas
with lower load of shared antagonists.
Techniques of high-throughput sequencing are increasingly effective and
affordable, providing new exciting approaches to study
plant-microorganism interactions (Põlme et al., 2020). The availability
of such data about mutualists and antagonists associated to plant
species opens up new avenues for analysing the effect of
plant-associated organism on the dynamics and assembly of plant
communities (Pajares-Murgó et al., 2022). For example, we showed
beneficial effects of arbuscular mycorrhizal fungi in pairwise
canopy-recruit interaction networks in Mediterranean forest communities
(Garrido et al., 2023). However, limiting the analysis to pairwise
plant-plant interaction may produce biased results when recruits tend to
be surrounded by multiple plant species with whom they share antagonists
or mutualists, because not all of these organisms are host specific (Ali
& Agrawal 2012; Gilbert et al., 2015; Gilbert & Webb 2007; Parker et
al., 2015).
In this study we consider the entire plant neighbourhood that forms the
seedscape. We argue that each plant species hosts typical communities of
associated mutualists or antagonists (Gómez et al., 2010), and that the
load of mutualists or antagonists shared with its neighbours will
determine the fate of recruits in its transition from seed to sapling.
Our novel approach involves deriving for each organism type (e.g.,
herbivorous insects, pathogens or epiphytes) an index\(\delta_{\text{fi}}\) of similarity between two plant species fand i in terms of the number of shared organisms. The conspecific
index \(\delta_{\text{ff}}\) is then given by the organisms associated
only with the focal species f . Recent techniques of spatial point
pattern analysis (Perea et al., 2022; Wiegand et al., 2017) provide the
methods to take this novel neighbourhood approach. For example, the
summary functionαf ,phy (r )
determines first for each plant the mean number of organisms shared with
their neighbours before averaging over all individuals of interest. This
allows us to find out if individuals of a given species, life stage or
functional group (e.g., fleshy- or dry fruited species) share on average
more or less organisms with their neighbours than expected by a null
model that locates them randomly in the seedscape.
Our overall objective is to identify non-random spatial patterns in the
number of organisms shared by plants and their neighbours, and to
compare them to expectations under Janzen-Connell mechanisms. To this
end, we analyse data on woody plant species in two fully stem-mapped
Mediterranean forest communities with similar species composition, but
contrasting species richness and abundances, and different seed
dispersal mechanisms (i.e., fleshy fruited species vs. dry fruited
species). At each community, we collected and classified the fungi
(sequencing) and the insect species inhabiting the phyllosphere of each
of the local plant species, including leaf pathogens, sap-sucker insects
and chewer insects as antagonists, and leaf epiphytic fungi as
mutualists (see supplementary information). This unique dataset allows
us to reveal spatial patterns in the similarity of antagonist and
mutualist communities between saplings and their saplings and adult
neighbours, and adults and their adult neighbours, and whether these
patterns are conditioned by the seed dispersal mechanism. Identifying
these patterns allow us to better understand plant community assembly
during the ontogeny of the species (see Figure 1 for the conceptual
framework of this approach).
More specifically, we assess the following questions, i) Is the
host-specificity maintained across shared-organisms at species-pairwise
levels (analysis 1)? ii) Do the patterns in shared-organisms (between
sapling-sapling (analysis 2), sapling-adult (analysis 3) and adult-adult
(analysis 4) life stages) meet the contrasting expectations of
Janzen-Connell effects for dry- and fleshy-fruited species? These
analyses allow us to ask iii) Do adults conserve a spatial signature of
Janzen-Connell mechanism? and iv) Are these patterns consistent across
communities? (See Table 1 for aims, expectations and technical settings
of analyses 1 to 4).