Description: The aim of the WP3 “Network Integration and Improvements” is to coordinate and enhance key aspects of integration of European observing technology (and related data flows) for its use in the context of international ocean monitoring activities. One of the dimensions of the integrations is the constitution of thematic networks, that is, networks whose aim is to address specific observational challenges and thus to favor innovation, innovation that will ultimately support the Blue economy. In this context, the specific aim of Task 3.8 is to accelerate the adoption of molecular methods such as genomic, transcriptomic (and related “omics”) approaches, currently used as monitoring tools in human health, to the assessment of the state and change of marine ecosystems. It was designed to favor the increase the capacity to evaluate biological diversity and the organismal metabolic states in different environmental conditions by the development of “augmented observatories”, utilizing state-of-art methodologies in genomic-enabled research at multidisciplinary observatories at well-established marine LTERs, with main focus on a mature oceanographic observatory in Naples, NEREA. In addition, an effort is dedicated to connecting existing observatories that intend to augment their observations with molecular tools. Molecular approaches come with many different options for the protocols (size fractioning, sample collection and storage, sequencing etc). One main challenge in systematically implementing those approaches is thus their standardization across observatories. Based on a survey of existing methods and on a 3-year experience in collecting, sequencing and analyzing molecular data, this deliverable is thus dedicated to present the SOPs implemented and tested at NEREA. The SOPs consider a size fractioning of the biological material to avoid biases toward more abundant, smaller organisms such as bacteria. They cover both the highly stable DNA and the less stable RNA and they are essentially an evolution of the ones developed for the highly successful Tara Oceans Expedition and recently updated for the Expedition Mission Microbiomes, an All-Atlantic expedition organised and executed by the EU AtlantECO project. Importantly, they have only slight variations with respect the ones adopted by the network of genomic observatories EMOBON. Discussions are ongoing with EMOBON to perfectly align the protocols. The SOPs are being disseminated via the main national and international networks.

Description: The oligotrophy of the southern Adriatic Sea is characterized by seasonal stratification which enables nutrient supply to the euphotic layer. A set of interdisciplinary methods was used to elucidate the diversity and co-dependency of bacterio- and phytoplankton of the water column during the stratification period of July 2021. A total of 95 taxa were determined by microscopy: 58 diatoms, 27 dinoflagellates, 6 coccolithophores, and 4 other autotrophs, which included Chlorophyceae, Chrysophyceae, and Cryptophytes. Nanophytoplankton abundances were higher in comparison to microphytoplankton. The prokaryotic plankton community as revealed by HTS was dominated by Proteobacteria (41–73%), Bacteroidota (9.5–27%), and cyanobacteria (1–10%), while the eukaryotic plankton community was composed of parasitic Syndiniales (45–80%), Ochrophyta (2–18%), Ciliophora (2–21%), Chlorophytes (2–4%), Haptophytes (1–4%), Bacillariophyta (1–13%), Pelagophyta (0.5–12%) and Chrysophyta (0.5–3%). Flow cytometry analysis has recorded Prochlorococcus and photosynthetic picoeukaryotes as more abundant in deep chlorophyll maximum (DCM), and Synechococcus and heterotrophic bacteria as most abundant in surface and thermocline layers. Surface, thermocline, and DCM layers were distinct considering community diversity, temperature, and nutrient correlations, while extreme nutrient values at the beginning of the investigating period indicated a possible nutrient flux. Nutrient and temperature were recognized as the main environmental drivers of phytoplankton and bacterioplankton community abundance.