DNA METABARCODIG AS A TOOL FOR ZOOPLANKTON BIODIVERSITY ASSESMENTS IN TRANSITIONAL ENVIRONMENTS

This thesis aims at evaluating the suitability of DNA metabarcoding for zooplankton biodiversity assessments. The taxonomic identifications and diversity patterns assessed with this molecular method, based on a COI marker proposed by Leray et al. (2013), were compared to the classical morphological analysis (CHAPTER A). The transitional environments Venice Lagoon and Gulf of Venice were chosen as study area, as the high spatial and temporal variability of these ecosystems is advantageous for evaluation purposes. Moreover, especially these characteristics require high sampling and identification effort, which would particularly benefit from the rapidness and cost-effectiveness of DNA metabarcoding. On one site, in comparison to the morphological method, the molecular analysis resulted in higher taxa richness (224 vs. 88 taxa), discriminating better the meroplanktonic component, morphologically identified only up to order level. In addition, DNA metabarcoding was able to detect numerous non-indigenous species (NIS), highlighting its power as an early-detection systems. On the other site, both methods revealed similar spatio-temporal distribution patterns, and the sequence abundances were significantly correlated with individual counts for various taxonomic groups. The overall results of CHAPTER A indicate that DNA metabarcoding is an efficient tool for biodiversity assessments and laid the foundation for the subsequent applications of DNA metabarcoding. This study was already published in March 2020 in Marine Environmental Research (https://doi.org/10.1016/j.marenvres.2020.104946). The second chapter (CHAPTER B), describes the application of DNA metabarcoding on the zooplankton biodiversity in the Venice Lagoon by high spatial and temporal frequency sampling, testing the advantages of this fast and reliable method in increasing the monitoring effort. This large dataset revealed the presence of monthly patterns in the zooplankton community, confirming the importance of high temporal frequency assessments to detect community changes over the year and enabling a more precise description of the zooplankton biodiversity in the Venice Lagoon. Successively, in CHAPTER C, an additional COI barcode was evaluated for zooplankton biodiversity assessment by comparing it with the COI barcode already validated in CHAPTER A. The results indicate that after a slight adjustment of the originally proposed primer sequence by Leray et al. (2013), the reverse mlCOIintR primer performs, in combination with the jdgLCO1490 forward primer, very comparably to the much more widely used primer pair mlCOIintF/HCO2198. This is true in terms of level of taxonomic resolution, species detection and their relative abundance in terms of numbers of reads. This study was submitted in April 2021 to Marine Environmental Research and is in the phase of revision. As this primer combination has the advantage of not amplifying ctenophores, it was found to be optimal for investigating on the diet of the highly invasive zooplanktivorous ctenophore Mnemiopsis leidyi, whose theme was developed in CHAPTER D. In fact, while in the past, to study the diet of Mnemiopsis, the gut content was mainly analyzed by morphological identification, this is the first study investigating the feeding preference of this species utilizing DNA metabarcoding with the benefit of overcoming the limitations in e.g., identifying partially digested prey. The comparison of the gut content with the in-situ mesozooplankton community indicated that Mnemiopsis feeds on a variety of prey, mostly coinciding with the in-situ zooplankton assemblage. However, some groups, like decapod larvae and the slow-swimming larvae of gastropods and bivalves seem to be favored. Conversely, the abundance of copepods resulted being higher in-situ than in the gut content of Mnemiopsis.