Parasites, niche modification and the host microbiome: A field survey of multiple parasites

Parasites can affect and be affected by the host's microbiome, with consequences for host susceptibility, parasite transmission, and host and parasite fitness. Yet, two aspects of the relationship between parasite infection and host microbiota remain little understood: the nature of the relationship under field conditions, and how the relationship varies among parasites. To overcome these limitations, we performed a field survey of the within‐leaf fungal community in a tall fescue population. We investigated how diversity and composition of the fungal microbiome associate with natural infection by fungal parasites with different feeding strategies. A parasite's feeding strategy affects both parasite requirements of the host environment and parasite impacts on the host environment. We hypothesized that parasites that more strongly modify niches available within a host will be associated with greater changes in microbiome diversity and composition. Parasites with a feeding strategy that creates necrotic tissue to extract resources (necrotrophs) may not only have different niche requirements, but also act as particularly strong niche modifiers. Barcoded amplicon sequencing of the fungal ITS region revealed that leaf segments symptomatic of necrotrophs had lower fungal diversity and distinct composition compared to segments that were asymptomatic or symptomatic of other parasites. There were no clear differences in fungal diversity or composition between leaf segments that were asymptomatic and segments symptomatic of other parasite feeding strategies. Our results motivate future experimental work to test how the relationship between the microbiome and parasite infection is impacted by parasite feeding strategy and highlight the potential importance of parasite traits.


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O'KEEFFE Et al. requirements of the host environment and parasite effects on the host environment. Based on the concept of niche modification (Fukami, 2015;Lewontin, 1983), we develop the hypothesis that parasites with traits that more strongly modify the environment within the host can act as niche modifiers and more strongly impact the microbiome. Parasites with different feeding strategies differ in their requirements of the host environment, and parasites with certain feeding strategies may more strongly modify the environment within the host Glazebrook, 2005).
Thus, these parasites might be associated with differences in the host microbiome. To begin to confront these ideas with data under field conditions, we conducted an observational survey of the fungal microbiome in a grass host and evaluated how microbiome diversity and composition were associated with natural infections by multiple co-occurring fungal parasites.
Microbes may interact with parasites by competing for resources, releasing antimicrobial compounds, or eliciting a host immune response (Bashey, 2015;Graham, 2008;Raaijmakers & Mazzola, 2012). Such interactions can influence host health, making the host more susceptible to, tolerant of or resistant to parasites (Arnold et al., 2003;Hayes et al., 2010). The microbiome is also dynamic, and the introduction of a parasite can lead to changes in microbiome diversity and composition (Barman et al., 2008;Jani & Briggs, 2014). The link between parasites and host microbial diversity varies, with some studies showing no relationship (Li et al., 2012;Williams et al., 2017), some studies showing a negative relationship (Jani & Briggs, 2014;Leung et al., 2018;Mosca et al., 2016;Wu et al., 2014), and still others showing a positive relationship between parasite infection and within-host microbial diversity (Lee et al., 2014). One possible source of this variation among studies is variation among the parasite species studied. Yet, there is a lack of studies that examine variation among parasite species in their associations with host microbiota (but see Aivelo & Norberg, 2018), perhaps because we have lacked a robust framework for studying multiple parasites.
A trait-based approach might provide a framework for studying multiple parasite species (Oliva aet al. 2020;Zanne et al., 2019).
As well as being evolutionarily diverse (Weinstein & Kuris, 2016), parasites vary in traits such as growth rate, generation time and feeding strategy (Leggett et al., 2017). Different parasite feeding strategies can stimulate different immune responses in a host and differently impact host performance Glazebrook, 2005). Parasites infecting plants employ three typical feeding strategies. Parasites with a biotrophic feeding strategy keep host cells alive to extract resources from them. Parasites with a necrotrophic feeding strategy kill host cells to access their resources, creating necrotic tissue while they grow within their host. Finally, hemibiotrophic parasites infect as biotrophs, then switch to become necrotrophic. Given that parasites with different feeding strategies have different requirements to extract resources from the host environment and different impacts on the host environment, they may also have different associations with the host microbiome.
When an organism modifies its environment, it can change the number and types of niches available, and in turn impact the species that can reside and colonize within the ecological community via the process of niche modification (Fukami, 2015;Lewontin, 1983). Within host individuals, symbionts may also act as niche modifiers by impacting the host environment in such a way that the host is more or less suitable to new colonizers. We hypothesized that necrotrophic parasites act as niche modifiers for the microbiome. More specifically, we hypothesized that by producing necrotic tissue from when they first infect the host, necrotrophic parasites create new niches within the host for a longer time, and thus have larger impacts on microbiome richness and composition than either hemibiotrophic parasites, which only produce necrotic tissue in later stages of infection, or biotrophic parasites, which keep host cells alive while extracting resources. Conversely, parasites with different feeding strategies may respond differently to other symbionts that modify the environment within the host.
Because feeding strategy can define how a parasite interacts with the host environment, a trait-based approach grounded in parasite feeding strategy might help explain variation in parasitemicrobiome associations.
Few studies have investigated the associations between host-associated microbiota and parasites under field conditions (but see Aivelo & Norberg, 2018;Jani & Briggs, 2014;Leung et al., 2018). The lack of field studies may result from a limited number of suitable model systems for exploring these questions in the field. The long-lived nature of some hosts, limited ability to detect diseases observationally in live hosts in the field, difficulty of excising infected tissue from animals, and ethical concerns limit the utility of many study systems for field research (Borer et al., 2011). Foliar fungal parasites are a valuable model system to investigate microbiome-parasite interactions in field settings. These parasites are often readily identifiable by external, macroscopic symptoms and morphology, which facilitates observational studies in the field (Arnold et al., 2003;Christian et al., 2015). In a rare study, conducted with inbred mice, in which parasite infection was experimentally manipulated in both the laboratory and the field, the direction and magnitude of the relationship between parasite infection and the host microbiome differed between laboratory and field conditions (Leung et al., 2018). This finding underlines the importance of investigating parasite-microbiome associations in the field.
Here, we take a trait-based approach to address variation in parasite-microbiome associations in the field. We conducted a molecular survey of the within-leaf fungal community of the grass tall fescue in a field population infected by three fungal parasites.
These three parasites each employ a different feeding strategy (biotrophic, hemibiotrophic and necrotrophic). This survey provides an initial evaluation of whether three parasites, each exhibiting a different feeding strategy, and thus a different hypothesized potential to act as a niche modifier, have different associations with the host microbiome. This field was chosen based on proximity to the laboratory (to expedite sample processing), and abundance of tall fescue (Lolium arundinaceum) and its foliar fungal parasites. This study focused on disease symptoms previously identified in another field of the Duke Forest Teaching and Research Laboratory (Halliday et al., , 2018 as caused by parasites with three different parasite feeding strategies (Table 1).

| Leaf segment collection
Leaf segments were collected over the course of two days in late September 2016 (September 26 and September 30), which is when parasites tend to peak in their abundance in this system . Leaf segments were collected at a total of 36 sampling points, spaced every 20 m along six transects; the transects were 100 m long, parallel and spaced 20 m apart ( Figure S1).
Only one leaf of each symptom was collected at each point, and therefore leaves with the same symptom were always collected at least 8 m apart. This minimum distance was selected to minimize the effect of spatial autocorrelation on the structure and composition of the microbiome Higgins et al., 2007).
To standardize the relative age of sampled leaves, we always sampled the oldest fully expanded nonsenescing leaf on the tiller.
For each leaf, we estimated the percent of leaf area infected with each parasite (infection severity) by visually comparing leaves to reference images of leaves of known infection severity (Halbritter et al., 2020;e.g., Halliday et al., 2019;Mitchell et al., 2002Mitchell et al., , 2003. At each of the 36 sampling points, we collected four whole leaves-one leaf with symptoms of a necrotrophic parasite (and no other parasites), one leaf with symptoms of a hemibiotrophic parasite (and no other symptoms), one leaf with symptoms of a biotrophic parasite (and no other parasites) and one asymptomatic leaf. While co-infection is common in this system , we avoided collecting co-infected leaves. From the four whole leaves collected at each of the 36 sampling points, we excised seven leaf segments: symptomatic and asymptomatic segments from each leaf infected with one of the three parasites, as well as an asymptomatic segment collected from the one asymptomatic leaf. Thus, we collected a total of 252 leaf segments. All segments were of approximately equal size. The two segments collected from each symptomatic leaf were spaced at least 10 cm apart within the leaf. We stored each leaf segment in an individual plastic bag that was then placed on ice.
Within 4 hr, all segments were processed back in the laboratory.
Leaf segments were washed under running deionized water for 30 s to remove fungi that were on the surface of the leaf but not attached TA B L E 1 Biology and ecology of the focal parasite feeding strategies [Colour table can be viewed at wileyonlinelibrary.com]

Feeding strategy Parasite Symptom
Necrotroph: kills the living cells of a host and then feeds on the dead matter

Rhizoctonia solani
Biotroph: feeds on living cells of a host, without killing it as part of the infection process

Puccinia coronata
Hemibiotroph: initially feeds on living host tissue without causing visible symptoms prior to switching to a necrotrophic phase

Colletotrichum cereale
to the leaf. Following surface washing, leaf segments were stored in a −80°C freezer.

| DNA extraction and sequencing
Surface-washed leaf segments were ground under liquid nitrogen with a mortar and pestle and transferred to 96-well plates for DNA extraction. DNA extraction was performed with the DNEasy PowerSoil kit according to the manufacturer's protocol (Qiagen).
We assayed the fungal communities of tall fescue leaves by sequencing the internal transcribed spacer (ITS) region. The ITS is a region of the nuclear ribosomal RNA cistron that is often used as a DNA barcode for fungi, as it has less intraspecific variation than interspecific variation (Schoch et al., 2012). We amplified the first part of the internal transcribed spacer (ITS1) with a version of the primer set ITS1F and ITS2 modified for parallel sequencing on the Illumina MiSeq platform (Smith & Peay, 2014;White, 1990

| Fungal community analysis
All statistical analyses were performed with leaf segment as the unit of observation, and all analyses were performed in the R environ- in R models and corrects Illumina-sequenced amplicon errors and infers exact amplicon sequence variants (ASVs; herein referred to as taxa); these taxa are biological variants and not sequencing noise (Callahan et al., 2016). This method can resolve biological differences at a high resolution, and the output can be directly compared among studies without the need to reprocess the pooled data. We therefore employed dada2 in this study. Quality control of sequencing reads for each leaf segment consisted of truncating reads at the first quality score of 2 (a quality score of two indicates a portion of the sequence that contains mostly low-quality reads of Q15 or less), and removing any read with ambiguous base calls or greater than two expected errors. Reads shorter than 50 bases after quality trimming were removed.

| Statistical analyses: Diversity
To compare the diversity of fungal communities among asymptomatic and symptomatic leaves of tall fescue, we quantified Hill's series of diversity. Hill's series of diversity (Hill, 1973) comprises three orders (q) of diversity that summarize information about the number and relative abundances of taxa. In Hill's series, the values of q (0, 1, 2) indicate the relative weight applied to relative abundance (Bent & Forney, 2008). We estimated fungal richness (Hills' N0, q = 0), the exponent of the Shannon index (Hill's N1, q = 1), and the inverse of the Simpson index (Hill's N2, q = 2) (Jost, 2006

2013).
To test whether fungal diversity is associated with symptom type, we used linear mixed models to explain Hill's N0, N1 and N2.
In order to meet assumptions of normality and homoscedasticity of the residuals, we log-transformed diversity. We included symptom type (seven levels: the asymptomatic and symptomatic segments from leaves with each of the three focal symptoms, plus asymptomatic segments from asymptomatic leaves) as fixed effects. Highthroughput sequencing of pooled samples can result in samples that differ in sequencing depth; we accounted for observational bias stemming from this difference by incorporating sequencing depth directly into the models as another fixed effect, following Bálint et al., (2015,2016). Another common approach to account for observational bias is to normalize the sequence read numbers via rarefaction (Gotelli & Colwell, 2001). However, rarefaction poses multiple analytical and statistical problems, such as loss of power and dependence on arbitrary thresholds. Statistical models can avoid these problems by incorporating sequencing depth into the models as the first explanatory variable, and thereby explicitly account for bias due to sequencing depth. Thus, the explicit incorporation of sequencing depth into models has been robustly tested with simulated microbiome data (Zhang et al., 2017;Luna et al., 2020), and is strongly advocated as an approach to account for differences in library size across samples (Bálint et al., 2016;Nayfach & Pollard, 2016;Weiss et al., 2017). To confirm that this method does not bias our analyses, we also normalized the sequence read numbers via rarefaction and found that the inferences remain consistent (Tables S1 and S2

| Statistical analyses: Community composition
To test the hypothesis that parasites that modify niches within their host by creating necrotic tissue alter fungal community composition, we tested whether fungal community composition was correlated with symptom type. Bray-Curtis distances were calculated among leaf segments separately and visualized using nonmet- and we report predictors that were significant regardless of order (following Vannette et al., 2016). We further investigated fungal functional composition by assigning fungal ASVs from the rarefied data to trophic mode (saprotroph, symbiotroph or pathotroph, with some taxa characterized by more than one trophic mode) where possible (Nguyen et al., 2016). When taxa could not be placed, they were labelled, "not assigned." We also investigated whether the homogeneity of the composition of the fungal leaf microbiome varied with symptom type. For each leaf segment, we quantified the distances from each measured Bray-Curtis distance to the centroid of Bray-Curtis distance for that leaf segment's symptom type. We then compared the dispersion of the measurements within each symptom type across categories using the betadisper function in the vegan package.
Within leaves symptomatic of any of the three parasite feeding strategies, we assessed if the symptomatic leaf segments differed from the asymptomatic leaf segments in the relative abundance of each taxon. We report those taxa that differed between symptom types with a false discovery rate (FDR) cutoff of 0.05, as well as their

| Statistical analyses: Interpretation
To improve statistical inference, we report our results using the language of the "statistical clarity concept," instead of emphasizing statistically significant results (Dushoff et al., 2019). This approach puts forward that the results of null hypothesis significance testing are most usefully interpreted as a guide to whether a certain effect can be seen clearly in a particular context.

| Diversity
After accounting for variation in sequencing depth, symptom type strongly and clearly predicted variation in all three numbers in Hill's series (ANOVA p < .0001). There were fewer fungal taxa (Hill's N0) and there was lower fungal diversity (Hill's N1 and N2) in leaf segments that exhibited symptoms of necrotrophic parasites compared to leaf segments that were asymptomatic or symptomatic of either hemibiotrophic or biotrophic parasites (Figure 1;

| Community composition
We analysed variation in community composition using the Bray-Curtis distance metric. The fungal community composition of leaf segments with symptoms of necrotrophic parasites differed from that of leaf segments that were asymptomatic or symptomatic of other parasites (Figure 3; type. There was no effect of symptom type on the homogeneity of fungal community composition ( Figure S2; ANOVA, p = .693).
Considering fine-scale variation in the fungal microbiome, within leaves symptomatic of any of the three parasite feeding strategies, the symptomatic leaf segments differed from the asymptomatic leaf segments in the relative abundance of multiple fungal genera ( Figures S3-S5).
At that fine scale within symptomatic leaves of any parasite feeding strategy, the symptomatic leaf segments differed from the asymptomatic leaf segments not only in fungal taxonomic composition, but also in fungal functional composition. Specifically, the trophic mode of fungi in leaf segments with symptoms of necrotrophic parasites differed from that of fungi in leaf segments that were asymptomatic or symptomatic of other parasites ( Figure 5). Of note, there were clearly fewer putative fungal saprotrophs within leaf segments with symptoms of necrotrophic parasites than segments that were asymptomatic or symptomatic of other parasites ( Figure S6A; p < .001 in all pairwise comparisons). These findings were robust to misspecification of trophic mode ( Figure S7). A higher number of F I G U R E 2 Leaf-associated fungal diversity and richness did not have a negative correlation with necrotrophic disease severity (percent leaf area exhibiting symptoms), suggesting that fungal community diversity does not change progressively as a parasite grows within a leaf. Panels show fungal diversity quantified using Hill numbers for observed species richness (N0), exponent of the Shannon index (N1) and inverse of the Simpson index (N2). Each point represents a leaf segment. Lines represent best-fit regressions between disease severity and the diversity metric [Colour figure can be viewed at wileyonlinelibrary.com] F I G U R E 3 Leaf segments with necrotrophic symptoms had foliar fungal communities that differed in taxonomic composition from asymptomatic leaf segments and leaf segments with symptoms of other parasite feeding strategies (PerMANOVA p < .001, stress = 0.18). Fungal taxonomic composition was quantified by the Bray-Curtis distance metric and is illustrated by nonmetric multidimensional scaling (NMDS). Each point represents a leaf segment. As indicated by shape and colour, each leaf segment was either asymptomatic or symptomatic of one parasite feeding strategy (necrotroph, hemibiotroph, or biotroph) F I G U R E 4 Severity of symptoms caused by necrotrophic parasites predicted fungal community composition, but only explained a modest amount of the variation, suggesting that fungal community composition does not change progressively as a parasite grows within a leaf. Fungal taxonomic composition was quantified by the Bray-Curtis distance metric and is illustrated by nonmetric multidimensional scaling (NMDS). Each point represents a leaf segment. Point size indicates percent leaf area symptomatic of necrotrophic parasites [Colour figure can be viewed at wileyonlinelibrary.com] fungal taxa associated with necrotrophic symptoms were not able to be assigned to a trophic mode compared to taxa associated with other symptom types, but this finding has weak statistical support (p = .09; Figure S6b).

| DISCUSS ION
This study used a trait-based analysis of multiple co-occurring fungal parasites in a field population of a grass host to evaluate how the host microbiome is associated with different parasite species. Lower microbiome diversity and distinct composition were associated with symptoms of a parasite with a necrotrophic feeding strategy, and not with parasites of other feeding strategies. These results are consistent with a hypothesis based on niche modification: parasites with traits that more strongly impact the host environment and available niches within the host more strongly impact the host microbiome.
In niche modification, a species changes the types of niches available within a site and, consequently, the identities of species that can colonize the community (Fukami, 2015;Lewontin, 1983). Niche modification is closely related to the concepts of niche construction (particularly with respect to evolutionary implications, Odling-Smee et al., 2003) and ecosystem engineering (Jones et al., 1994) and has been documented in numerous communities of free-living organisms (Fukami & Nakajima, 2011;Naiman et al., 2009). Among parasites infecting plant leaves, only parasites with a necrotrophic feeding strategy create necrotic host tissue throughout their entire infection process (Glazebrook, 2005;Suzuki & Sasaki, 2019). We therefore expected the necrotrophic parasite to be a particularly strong niche modifier that impacts the host environment and, consequently, the host fungal community. While our study system included only one parasite species or morphotype per feeding strategy, symptoms of a necrotrophic parasite were associated with fungal communities of lower diversity relative to asymptomatic leaves, and symptoms of two other types of parasite feeding strategies (biotrophs and hemibiotrophs) were not. Our data do not definitively establish the direction of this association, but these results are consistent with our hypotheses based on feeding strategy.
As a survey and not an experimental manipulation, this study does not establish a causative relationship between parasite infection and host microbiota. While we use our results to evaluate a causative hypothesis related to the niche modifying abilities of parasites, our results are correlative. Furthermore, while we take advantage of a naturally occurring host/multiparasite system in which parasites differ in key traits (namely, parasite feeding strategy), this same system was limited to one representative parasite per strategy. Each feeding strategy was not replicated by multiple parasites, which would be required for a more definitive test of the connection between parasite feeding strategy and variation in the host microbiome. Future studies could address these limitations with other approaches, such as longitudinal surveys across different plant/parasite systems in which microbiota are assayed before and after infection.
Despite these limitations, our hypothesis that necrotrophic parasites are particularly strong niche modifiers was supported by analysis of fungal community composition. The composition of the fungal communities of leaf segments exhibiting symptoms of necrotrophic parasites differed from that of asymptomatic leaves, and there was no clear difference in fungal composition between leaf segments exhibiting symptoms of biotrophic or hemibiotrophic parasites and that of asymptomatic leaves. We hypothesized that this F I G U R E 5 Leaf segments with necrotrophic symptoms had foliar fungal communities that differed in functional composition from asymptomatic leaf segments and leaf segments with symptoms of other parasite feeding strategies. Proportionate abundances of fungal taxa matched to putative trophic modes for parasite symptom types. Multitrophic mode groups are indicated by multiple trophic modes listed. Taxa unable to be assigned are labelled as "not assigned" shift in composition resulted from necrotrophic parasite infections making the host environment more suitable for saprobes (Suzuki & Sasaki, 2019). Inconsistent with this hypothesis, the number of putative saprotrophs was lower in leaf segments exhibiting symptoms of necrotrophic parasites than those that were asymptomatic or exhibiting symptoms of other parasites. However, a higher proportion of fungal taxa associated with necrotrophic symptoms was unable to be assigned a trophic mode. Therefore, while the number of putative saprotrophs does not support our hypothesis, the robustness of this specific test was limited by lack of definitive data on trophic mode.
On a more general level, the disproportionately high number of uncharacterized fungal taxa in leaf segments exhibiting symptoms of necrotrophic parasites suggests that necrotrophic parasites were associated with fungal communities of distinct functional composition.
Given that the region sequenced here, ITS1, cannot reliably place sequences to species (Dobon et al., 2016;Gazis et al., 2011;Lindner et al., 2011;Porras-Alfaro et al., 2014), future work that sequences the entire ITS region and other loci would be required to further characterize both the taxonomic and the functional composition of these communities.
Fungal community composition was only weakly associated with necrotrophic parasite disease severity, and fungal diversity did not have a negative correlation with disease severity. These results suggest that fungal community composition does not change and diversity does not decrease progressively as the parasite grows within a leaf. The parasite may instead disrupt the host environment, and consequently, the fungal microbiome, upon initial infection. Such microbiome disruption upon initial infection is consistent with evidence from at least one other system; in experimental inoculations of frogs with Batrachochytrium dendrobatidis, microbiome diversity declined upon infection and had no relationship with pathogen load (Jani & Briggs, 2014).
While we expected necrotrophs to act as particularly strong niche modifiers, we expected hemibiotrophs to modify their environment as well, given that they create necrotic tissue in the latter part of the infection process (Glazebrook, 2005;Suzuki & Sasaki, 2019). However, we found contrasting results between necrotrophic and hemibiotrophic parasites; the fungal communities of leaf segments exhibiting symptoms of hemibiotrophic parasites had no clear differences in diversity and composition compared to those of asymptomatic leaf segments. While both hemibiotrophs and necrotrophs ultimately require killing host cells, they differ in how they initially interact with host tissue. The initial infection by a necrotrophic parasite may be the key stage in which diversity of the microbiome declines. This is consistent with a weakly supported positive correlation between disease severity and fungal taxa diversity that we observed, as diversity was lowest when necrotrophic parasite disease severity was low (i.e., change in diversity occurred early in the infection process).
As an alternative to the niche-modification hypothesis, it is possible that parasite-microbiome associations were instead driven by variation in the fungal microbiome that altered host susceptibility to parasite infection (Croswell et al., 2009;Khosravi & Mazmanian, 2013). Specifically, if a less diverse fungal microbiome made leaf sections particularly susceptible to necrotrophic parasites, this could explain why necrotrophic parasites were associated with a less diverse microbiome in symptomatic leaf segments. Additionally, because necrotrophic parasites differ from hemibiotrophic and biotrophic parasites in how they initially interact with host tissue, this could also explain why this association was only observed for the necrotrophic parasite. However, within each leaf with necrotrophic symptoms, fungal diversity was lower and composition was distinct, on average, only in the leaf segment that was symptomatic, and not in the asymptomatic segment of that same leaf, nor in asymptomatic leaves, which suggests that this mechanism is unlikely. To clarify the cause of the parasite-microbiome associations, researchers could survey the microbiome of individuals before and after natural infection by parasites. However, such longitudinal field sampling of individuals is challenging, because assaying the fungal microbiome of leaves is destructive. Alternatively, researchers could experimentally manipulate the microbiome and subsequently monitor for parasite infections, or experimentally manipulate parasite infection then characterize the microbiome (Berg & Koskella, 2018), but such experiments are also challenging.
While we interrogated relationships between parasites and the fungal microbiome, there is growing evidence that bacterial and fungal microbiota associate with different factors (Elhady et al., 2017;Bergelson et al., 2019). For example, while we found that a biotrophic parasite had no relationship with fungal microbiome diversity, recent work investigating the bacterial microbiome of wheat found that leaves infected with a parasite infecting as a biotroph had higher bacterial diversity than uninfected leaves (Seybold et al., 2020). Such differences between the fungal and bacterial microbiome may result from many factors, including their differences in generation times and abundances within a host. For more complete understanding of parasite-microbiome associations, studies that integrate surveys of the bacterial and fungal communities will be essential (Laforest-Lapointe & Arrieta, 2018; Porras-alfaro & Bayman, 2011).
In investigations of plant, human and other animal diseases, increasing numbers of studies are characterizing how microbial communities associate with specific parasites and progression of disease (Cho & Blaser, 2012;Jani and Briggs, 2014;Jakuschkin et al., 2016;Lebreton et al., 2019). Here, we propose that functional traits can be used to explain variation among parasites in their associations with host microbiota. Trait-based approaches have played an important role in plant functional ecology (Adler et al., 2014;Cadotte, 2017