Abstract
Environmental gradients can influence morpho-physiological and
life-history differences in natural populations. It is unclear, however,
to what extent such gradients can also modulate phenotypic differences
in other organismal characteristics such as the structure and function
of host-associated microbial communities. In this work, we addressed
this question by assessing intra-specific variation in the diversity,
structure and function of environmental-associated (sediment and water)
and animal-associated (skin and gut) microbiota along an environmental
gradient of pollution in one of the most urbanized coastal areas in the
world. Using the tropical sea cucumber Holothuria leucospilota ,
we tested the interplay between deterministic (e.g., environmental/host
filtering) and stochastic (e.g., random microbial dispersal) processes
underpinning host-microbiome interactions and microbial assemblages.
Overall, our results indicate that microbial communities are complex and
vary in structure and function between the environment and the animal
hosts. However, these differences are modulated by the level of
pollution across the gradient with marked clines in alpha and beta
diversity. Yet, such clines and overall differences showed opposite
directions when comparing environmental- and animal-associated microbial
communities. In the sea cucumbers, intrinsic characteristics (e.g., body
compartments, biochemistry composition, immune systems), may underpin
the observed intra-individual differences in the associated microbiomes,
and their divergence from the environmental source. Such regulation
favours specific microbial functional pathways that may play an
important role in the survival and physiology of the animal host,
particularly in high polluted areas. These findings suggest that the
interplay between both, environmental and host filtering underpins
microbial community assembly in H. leucospilota along the
pollution gradient in Hong Kong.
Keywords: Microbial community, holothurian, nutrient pollution,
environmental gradient, host-microbiome
Introduction
Metazoans harbour diverse and dynamic microbial communities (microbiome)
that play essential roles in the ecology and function of their animal
host (Apprill, 2017). Symbiotic microorganisms influence host metabolic
processes (Egerton et al., 2018; Hakim et al., 2016; Hanning &
Diaz-Sanchez, 2015), development (Carrier & Reitzel, 2018), immune
responses (de Araujo et al., 2019; Krediet et al., 2013), reproduction
(Brucker & Bordenstein, 2013; Sharon et al., 2010), behaviour (Sherwin
et al., 2019) and survival (Kešnerová et al., 2020). However, microbial
communities and the interactions with their hosts are not static and can
vary across time (e.g., host’s development or seasonality, Kešnerová et
al., 2020; Shoemaker & Moisander, 2017) and space (e.g., host’s body
plan or geography, Griffiths et al., 2019; McKnight et al., 2020). Thus,
to understand the ecological dynamics of animal hosts in a changing
world, it is becoming more important to understand the mechanisms and
drivers underpinning the origin and regulation (functional and
structural) of their associated microbial communities (Kohl, 2020).
Host-associated microbiomes are shaped by a diversity of evolutionary
and ecological processes that can be explored through the framework used
in community ecology. Under this framework, the formation of microbial
communities is seen as the outcome of selective processes in which a
larger species pool is subjected to a set of biotic and abiotic filters
(Hardy et al., 2012). For instance, habitat/environmental filtering is
usually considered one of the dominant forces in structuring communities
consisting of habitat-specialized species (Cavender-Bares et al., 2009).
In this case, the habitat/environment is part of the selective processes
as it plays a dual role acting as a source for the hosts’ microbiome and
also influencing the composition and dynamics of the established
microbial community (Johnke et al., 2020). As a result, host intra- and
interspecific variation in microbiome composition is expected to be
lower for organisms inhabiting similar environments in comparison to
their counterparts living in more divergent conditions (Johnke et al.,
2020). Hosts, however, can also influence associated microbiomes in
several ways, such as through selective feeding or filtering (Gao et
al., 2014; Theis et al., 2016). This may result from evolutionary
processes that produce patterns in which closely related host species
harbour similar microbiota even if they inhabit different environments
(San Juan et al., 2020; Sanders et al., 2015). In any case,
environmental and host filtering are aligned with the fundamental idea
that deterministic processes dictate the microbial community assembly in
natural populations (Näpflin & Schmid-Hempel, 2018; J. Wang et al.,
2013; Weigel, 2020; Yan et al., 2016).
Previous studies have also suggested that stochastic processes can
influence microbial community assembly (Oliphant et al., 2019; Zhou et
al., 2013). These processes include random dispersal potential and
colonization chance of microbes (Zhou & Ning, 2017), random arrival
sequence to host from the environment during dispersal (Burns et al.,
2017), changes in community abundance and survival due to random
speciation, extinction and ecological drift (Chase & Myers, 2011).
Under this paradigm, species present in the microbial community would be
independent and unpredictable, with no specific occurrence pattern or
relation with the respective niche (Rosindell et al., 2012). This
provides us with a wider perspective when studying processes
underpinning microbial community assembly. What is worth mentioning is
that deterministic and stochastic processes are not mutually exclusive
(Bordenstein & Theis, 2015). Both processes work together in
structuring microbial communities although a stronger contribution of a
particular process during the assembly may occur depending on the
investigated model and ecosystem (Yan et al., 2016).
In marine environments, the interplay between deterministic and
stochastic factors is an important driver of inter- and intra-specific
differences in organismal characteristics across heterogeneous
seascapes. In these systems, gradients of environmental conditions are
known to influence geographic variation in attributes such as physiology
(Gaitán-Espitia et al., 2014; Gaitán-Espitia, Villanueva, et al., 2017),
life-history (Pecuchet et al., 2018), zonation (Mangialajo et al.,
2012), behaviour (Cornwell et al., 2019), intra- and inter-specific
genetic diversity (Zhong et al., 2024), stress tolerance and phenotypic
plasticity (Gaitán-Espitia, Bacigalupe, et al., 2017) in natural
populations. As such, it would be expected that environmental gradients
would also influence variation and phenotypic differences in other
organismal characteristics, including the structure and function of
host-associated microbial communities. In fact, gradients in sea surface
temperature and salinity are known to influence symbiotic microbial
assemblages in benthic marine species (Capistrant-Fossa et al., 2021;
Ketchum et al., 2021; Osman et al., 2020) and coastal ecosystems
(Bolhuis et al., 2013; Z. Wang et al., 2019; Zäncker et al., 2018; G.
Zhang et al., 2021). While these natural environmental gradients may
have a fundamental role in modulating ecological patterns of marine
microbiomes, anthropogenic-mediated gradients (e.g., nutrient pollution)
can potentially induce drastic changes in these patterns by altering
ecological dynamics, the origin, and regulation of host-associated
microbial communities (Degregori et al., 2021; Milan et al., 2018;
Stevick et al., 2021; Ziegler et al., 2019).
In this study, we aimed to assess to what extent deterministic and/or
stochastic processes along an anthropogenic-mediated gradient of
pollution influence intra-specific variation in diversity, structure,
and function of host-associated microbial communities in marine
organisms. Here, our model system was Holothuria leucospilota, a
tropical sea cucumber that dominates shallow waters in Hong Kong and the
Indo-Pacific region. H. leucospilota is a deposit-feeding species
that assimilates organic matter from surface sediments (including
bacteria, benthic phytoplankton, meiofauna, and organic detritus), which
serves as a constant environmental source for microbe acquisition (Gao
et al., 2014; Pagán-Jiménez et al., 2019). However, H.
leucospilota is characterised by the capacity to secrete secondary
metabolites from the skin, gonads, and guts, which have antibacterial
and antifouling properties (Darya et al., 2020), potentially allowing
them to regulate the influence of environmental microbial reservoirs.
Based on 16S amplicon sequencing, we examined both sea cucumber (skin
and intestine) and environment (sediment and water) microbiomes along a
pollution gradient in Hong Kong, one of the busiest ports and highly
urbanized areas in the world. If environmental filtering is the main
driver of microbial community assemblies along the pollution gradient,
then similar host and environmental microbiomes will be observed within
sites whilst higher microbiome variation would be expected across the
gradient. In this context, microbial dispersal patterns and ecological
drift might also play an important role in shaping inter-individual
microbiome variation within sites (Stothart et al., 2021). Conversely,
if host filtering and specificity are the main drivers of microbial
community assembly in H. leucospilota , then low inter-individual
variation in sea cucumber microbiome composition would be expected along
the pollution gradient independently of the variation in the
environmental microbial communities. However, intra-individual variation
would be expected as sea cucumber skin and guts have different
antibacterial properties (Darya et al., 2020).