Developing a better definition of flow cytometric life forms under the mixoplankton paradigm - Qualification and quantification of pico- and nano-plankton
Authors: Mitra, Aditee
Type: Publication
Published: 2026-08-03

<h1>Executive Summary</h1>
<p class="BT12MFRG"><span><span>·<span>      </span></span></span><span>The dominance of smaller over larger sized organisms in coastal and open ocean waters are expected to increase with climate change. There is a concern that such an increase in smaller-sized (≤5 µm) plankton could lead to a decline in food supplies to higher trophic levels (e.g., fish) and thus have deleterious impacts on ecosystem structure, and services. </span></p>
<p class="BT12MFRG"><span><span>·<span>      </span></span></span><span>Over the last decade, marine ecology has seen the emergence of the mixoplankton paradigm which overturns our understanding of the structure and function of marine ecosystems. </span></p>
<p class="BT12MFRG"><span><span>·<span>      </span></span></span><span>Mixoplankton are single-celled chlorophyll-containing eukaryotic protist plankton which employ phototrophy <u>and</u> phagotrophy synergistically to meet their nutritional needs. From an evolutionary aspect, mixoplankton are the ancestors of protist phytoplankton. They are therefore <u>not</u> ‘plants-that-eat’, <u>nor</u> ‘animals-that-photosynthesize’. They are <u>not</u> a subset of ‘phytoplankton’, <u>nor</u> are they a subset of zooplankton. They are <u>not</u> hybrids of phytoplankton and zooplankton. Various nano-sized ‘phytoplankton’ (e.g., coccolithophores, cryptophytes, <em>Phaeocystis</em>) are actually mixoplankton, ingesting varying concentrations of picoplankton. </span></p>
<p class="BT12MFRG"><span><span>·<span>      </span></span></span><span>It is thus essential that plankton monitoring methodologies are equipped to appropriately monitor ecosystem status under the new mixoplankton paradigm. </span></p>
<p class="BT12MFRG"><span><span>·<span>      </span></span></span><span>This report reviews the suitability of extant plankton monitoring analysis tools for pico- and nano-plankton under the mixoplankton paradigm. With reference to data obtained from Scottish Coastal Observations (SCObs) and Western English Channel stations (WaMS), this report also investigates the appropriateness of usage of the current definitions of life forms from such analyses. </span></p>
<p class="BT12MFRG"><span><span>·<span>      </span></span></span><span>All the current monitoring methodologies – flow cytometry, flow-through image analysis, optical microscopy, and environmental-DNA (eDNA) metabarcoding – have significant challenges associated with them. None of these methodologies, alone, is able to deliver to the task of surveying and monitoring pico- and nano-plankton under the new paradigm. </span></p>
<p class="BT12MFRG"><span><span>·<span>      </span></span></span><span>Of the two tools specifically explored in the context of the analysis of pico- and nano- plankton, flow cytometry can provide no discrimination between mixoplankton and the non-phagotrophic phytoplankton, while eDNA analysis does not (within the data accessed from SCObs and WaMS) provide the required resolution to species level.</span></p>
<p class="BT12MFRG"><span><span>·<span>      </span></span></span><span>The fixatives used for processing of samples for flow cytometry lead to damage of various fragile and/or cryptic phyto- and mixo-plankton species. Further, flow cytometry cannot capture photosynthetic signal from acquired phototrophy in mixoplankton.</span></p>
<p class="BT12MFRG"><span><span>·<span>      </span></span></span><span>Current flow cytometry life form descriptors (e.g., pico-eukaryote, nano-eukaryote) do not differentiate between phytoplankton and mixoplankton and are thus inadequate to provide data to aid understanding of structural and dynamic changes in marine ecosystems in the light of climate change. </span></p>
<p class="BT12MFRG"><span><span>·<span>      </span></span></span><span>Future analyses require greater awareness of the need to identify mixoplankton. This requires application of optical methodologies and/or of eDNA analyses to species level. </span></p>
<p class="BT12MFRG"><span><span>·<span>      </span></span></span><span>In absence of definitive separation between phytoplankton and mixoplankton, it is important to only use the term ‘phytoplankton’ in reference to cyanobacteria (prokaryote photo-osmo-mixotrophs, e.g., <em>Synechoccocus</em>, <em>Prochlorococcus</em>) and diatoms. Other life forms should always be assumed to include mixoplankton.</span></p>
<p class="BT12MFRG"><span><span>·<span>      </span></span></span><span><span>It is recommended that photo-pigmented planktonic microbes should always by default be referenced as ‘<strong>mixoplankton+phytoplankton</strong>’.</span></span></p>