Proteomic fingerprinting enables quantitative biodiversity assessments of species and ontogenetic stages in Calanus congeners (Copepoda, Crustacea) from the Arctic Ocean

Abstract

Species identification is pivotal in biodiversity assessments and proteomic fingerprinting by MALDI-TOF mass spectrometry has already been shown to reliably identify calanoid copepods to species level. However, MALDI-TOF data may contain more information beyond mere species identification. In this study, we investigated different ontogenetic stages (copepodids C1-C6 females) of three co-occurring Calanus species from the Arctic Fram Strait, which cannot be identified to species level based on morphological characters alone. Differentiation of the three species based on mass spectrometry data was without any error. In addition, a clear stage-specific signal was detected in all species, supported by clustering approaches as well as machine learning using Random Forest. More complex mass spectra in later ontogenetic stages as well as relative intensities of certain mass peaks were found as the main drivers of stage distinction in these species. Through a dilution series, we were able to show that this did not result from the higher amount of biomass that was used in tissue processing of the larger stages. Finally, the data were tested in a simulation for application in a real biodiversity assessment by using Random Forest for stage classification of specimens absent from the training data. This resulted in a successful stage-identification rate of almost 90%, making proteomic fingerprinting a promising tool to investigate polewards shifts of Atlantic Calanus species and, in general, to assess stage compositions in biodiversity assessments of Calanoida, which can be notoriously difficult using conventional identification methods.

Perspectives of species identification by MALDI‐TOF MS in monitoring—Stability of proteomic fingerprints in marine epipelagic copepods

“…Bearing an additional physiological signature, the fingerprint may provide information beyond the pure species name, such as on developmental stage (Karger et al, 2012; Laakmann et al, 2013; Rossel et al, 2023), gender (Lafri et al, 2016), environmental conditions (Karger et al, 2019) or feeding status (Hlavackova et al, 2019; Niare et al, 2017; Tandina et al, 2018). Successful detection of microplastics by MALDI‐TOF MS (Adhikari et al, 2022) may even open future options for simultaneous detection of contamination.…”

Potential of MALDI‐TOF MS‐based proteomic fingerprinting for species identification of Cnidaria across classes, species, regions and developmental stages

“…Our results demonstrate that intraspecific variability in scyphozoans was strongly affected by the ontogenetic stage. It is possible to differentiate within species on the level of ontogenetic stage which was previously shown on calanoid copepods (Rossel, Kaiser, et al, 2022).…”

“…Our results demonstrate that intraspecific variability in scyphozoans was strongly affected by the ontogenetic stage. It is possible to differentiate within species on the level of ontogenetic stage which was previously shown on calanoid copepods (Rossel, Kaiser, et al, 2022). Although the benthic polyp and the pelagic ephyra / medusa stages in metagenetic cnidarians represent two generations with very different morphologies, the different stages formed clear species clusters.…”

“…Recently, unsupervised methods for rapid delimitation in biodiversity assessments based on MALDI‐TOF MS data were devised (Renz et al, 2021; Rossel & Martínez Arbizu, 2020) and applied in investigations of deep‐sea biodiversity (Rossel, Uhlenkott, et al, 2022). Further studies have shown that the resolution of proteomic fingerprinting can go beyond mere species identification such as differentiation of some species on sex level (Rossel & Martínez Arbizu, 2019), developmental stage (Rossel, Kaiser, et al, 2022) and host species of important pathogen‐vector species (Niare et al, 2016, 2018).…”

Evaluating species richness using proteomic fingerprinting and DNA barcoding—a case study on meiobenthic copepods from the Clarion Clipperton Fracture Zone

“…MALDI-TOF MS could be especially attractive for quantitative assessments (specimen-by-specimen) of benthic communities as it does not require an extensive taxonomic knowledge and is considerably cheaper than DNA barcoding . Furthermore, MALDI-TOF MS data can be used not only to distinguish species, but also the developmental stages within species (Rossel et al 2022). Hence, it might be a useful, additional tool to monitor resettlement and dispersion strategies of individual taxa at impacted sites.…”