Discussion
This is the first report showing that soil protists present in a forest soil can ingest MPs and
store them within their food vacuoles. In addition, our observations
using fluorescence microscopy clearly showed that the majority of large
(>30 μm) phagotrophic protists, irrespective of taxon, could readily ingest MPs in
any MP addition treatment. This finding was robust because green
fluorescence (550 nm) was restricted to MP addition treatments, showing
that any background fluorescent MPs were not detectable and that the
protists’ organelles are not fluorescent at that wavelength.
Approximately 75% of protists ingested MPs in the treatment with the
lowest concentration MPs and a 10-fold increase in MP
concentration resulted in nearly all protists ingesting MPs. These
findings build on previous work (e.g., Fenchel, 1980) and can have profound ecological and environmental implications
given the important role that protists play in the soil food web (Adl &
Gupta, 2006). Motile phagotrophic protists can serve both as vectors in
moving MPs in the soil matrix and in transferring them to higher trophic
levels, thereby potentially amplifying MP pollution (Geisen et al.,
2018, 2020). Furthermore, we could observe several ciliate and
flagellate morphotypes, which suggests that the feeding behavior for MPs
may not be species specific. Conversely, we anecdotally observed that
some protist morphotypes seemed to show a greater propensity for
ingesting MPs than others as indicated by the intensity of the
fluorescence within their food vacuoles. Certainly, more experimentation
is required to understand if protists show morphotype or taxa specific
feeding behaviors regarding MP ingestion.
The results of trial 2 show that soil protists can readily ingest MPs in
as little as 24 hours after these are introduced. In addition, in the
treatment with 1% MPs (w/w) nearly all protists ingested MPs. This is
consistent with data for other soil biota such as bacterial-feeding
nematode species using similar methods with fluorescent microspheres
(Mueller et al., 2020). This technique could potentially be utilized in
future experiments to quantify MP ingestion and to source track MPs.
We detected a significant overall temporal difference in protist
abundance in both trials across treatments, which is explained by
declines after approximately a week. This was expected since microcosms
are closed systems. However, the hypothesis that the overall abundance
of phagotrophic soil protists would be reduced by the addition of MPs
particles to soil was not strongly supported by the data. Nevertheless,
MP addition had a marginally significant effect on protist abundance in
trial one. These results are interesting given that most protists were
observed carrying MPs within their food vacuoles which would indicate,
at least within short-time scales, that they may not be detrimental to
the organisms’ overall health. This is consistent with studies showing
that MPs do not seem to cause considerable mortality in earthworms at
environmentally relevant concentrations. However, mortality has been
reported at relatively high concentrations (Jacques & Prosser, 2021).
Likewise, the nematode Caenorhabditis elegans appears to be
highly susceptible to high concentrations of MPs. In contrast, other
species of nematodes may be more tolerant to MPs of similar polymer
composition to those used in this study (Mueller et al., 2020). It is
possible that the commercial microspheres used in this study are indeed
inert and their ingestion may not substantially reduce the protist’s
food intake. More research using different soils and a wider diversity
of MPs types (e.g., varying the parameters shape (Lozano et al., 2021),
polymer type (Waldman & Rillig, 2020), weathering status and additives
(Kim et al., 2020)) will be required to clarify this.
If MPs particles were in fact safe to soil protists, given their high
abundance in soils and important role as nutrient cyclers (Wood &
Bradford, 2018), perhaps they may be able to further physically
breakdown MPs. However, while they could probably use some of the
additives, debris, or biofilms in the plastic particles it is unlikely
that protists would be able to utilize the long carbon-backbone chains
that makeup most plastics. The fact is, the MPs used in this experiment,
although utilized in other MP-biota interactions research (Bringer et
al., 2020), do not represent the average MP particle found in soil (Wang
et al., 2019). Future studies could utilize other MPs commonly found in
soil environments, such as polyacrylic fibers or polyethylene fragments.
However, appropriate MP detection methods for those polymers (Shan et
al., 2018) would be required in combination with methods to extract
protists from soil and sediments (Alongi, 2018). Overall, our results
show that, in general, large phagotrophic protists appear to have the ability to
ingest MPs. More research is needed to verify to what degree MPs can in
fact affect the abundance and community composition of soil protists and
understand the effect of MPs on soil food webs.
Acknowledgements
This research was conducted in Robinson-Huron Treaty territory and the
traditional territory of the Anishnaabeg, specifically the Garden River
and Batchewana First Nations, as well as Métis People. The work was
funded through a Discovery Grant from the Natural Sciences and
Engineering Research Council of Canada (NSERC) and a Canada Research
Chair awarded to P.M. Antunes.
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