Coupling Imaging and Omics in Plankton Surveys: State-of-the-Art, Challenges, and Future Directions

Abstract

A major challenge in characterizing plankton communities is the collection, identification and quantification of samples in a time-efficient way. The classical manual microscopy counts are gradually being replaced by high throughput imaging and nucleic acid sequencing. DNA sequencing allows deep taxonomic resolution (including cryptic species) as well as high detection power (detecting rare species), while RNA provides insights on function and potential activity. However, these methods are affected by database limitations, PCR bias, and copy number variability across taxa. Recent developments in high-throughput imaging applied in situ or on collected samples (high-throughput microscopy, Underwater Vision Profiler, FlowCam, ZooScan, etc) has enabled a rapid enumeration of morphologically-distinguished plankton populations, estimates of biovolume/biomass, and provides additional valuable phenotypic information. Although machine learning classifiers generate encouraging results to classify marine plankton images in a time efficient way, there is still a need for large training datasets of manually annotated images. Here we provide workflow examples that couple nucleic acid sequencing with high-throughput imaging for a more complete and robust analysis of microbial communities. We also describe the publicly available and collaborative web application EcoTaxa, which offers tools for the rapid validation of plankton by specialists with the help of automatic recognition algorithms. Finally, we describe how the field is moving with citizen science programs, unmanned autonomous platforms with in situ sensors, and sequencing and digitalization of historical plankton samples.

Mock samples resolve biases in diversity estimates and quantitative interpretation of zooplankton metabarcoding data

“…Given the successful results, we can anticipate that metabarcoding-based protocols are close to becoming adapted as a method of choice for large-scale zooplankton monitoring programs. Although our results highlight the strengths of metabarcoding, they also underline the value of an integrated taxonomic approach by applying combined morphological and molecular analyses, which has also been recommended by previous studies (Di Capua et al, 2021;Karlusich et al, 2022;Matthews et al, 2021).…”

Unraveling Zooplankton Diversity in a Pre-Alpine Lake: A Comparative Analysis of ZooScan and DNA Metabarcoding Methods

“…On one hand, the digitization of zooplankton through different optical systems offers a means to obtain reliable data on the quality, quantity, and size (biomass) of zooplankton samples [15][16][17]. On the other hand, non-optical molecular techniques such as high-throughput sequencing (HTS) provide the opportunity for rapid and standardized analyses of zooplankton samples [18][19][20]. Such molecular methods can detect organisms below the usual thresholds of optical identification methods [4] but do not easily allow necessary zooplankton biomass estimations [18].…”

“…On the other hand, non-optical molecular techniques such as high-throughput sequencing (HTS) provide the opportunity for rapid and standardized analyses of zooplankton samples [18][19][20]. Such molecular methods can detect organisms below the usual thresholds of optical identification methods [4] but do not easily allow necessary zooplankton biomass estimations [18]. Recently, studies combined automated optical/digitization methods with molecular methods to achieve the highest level of resolution in analyzing zooplankton samples [18,21].…”

“…Such molecular methods can detect organisms below the usual thresholds of optical identification methods [4] but do not easily allow necessary zooplankton biomass estimations [18]. Recently, studies combined automated optical/digitization methods with molecular methods to achieve the highest level of resolution in analyzing zooplankton samples [18,21]. There has been ongoing discussion about the discrepancy between these two methods, and attempts are made to establish correlations between the abundance of individuals, their biomass, and their taxonomically specific sequence reads determined through molecular approaches [18,[20][21][22][23][24][25][26].…”

“…Recently, studies combined automated optical/digitization methods with molecular methods to achieve the highest level of resolution in analyzing zooplankton samples [18,21]. There has been ongoing discussion about the discrepancy between these two methods, and attempts are made to establish correlations between the abundance of individuals, their biomass, and their taxonomically specific sequence reads determined through molecular approaches [18,[20][21][22][23][24][25][26]. With the increasing use of advanced analytical techniques, two prominent methodologies, namely ZooScan and DNA metabarcoding, have emerged as potent automatized tools for investigating zooplankton diversity and abundance in lakes [15,27,28].…”