Design and use of a new primer pair for the characterization of the cyanobacteria Synechococcus and Prochlorococcus communities targeting petB gene through metabarcoding approaches.

During the last years, the application of next-generation sequencing (NGS) technologies to search for specific genetic markers has become a crucial method for the characterization of microbial communities. Illumina MiSeq, likely the most widespread NGS platform for metabarcoding experiments and taxonomic classification, allows processing shorter reads than the classical SANGER sequencing method and therefore requires specific primer pairs that produce shorter amplicons. Specifically, for the analysis of the commonly studied Prochlorococcus and Synechococcus communities, the petB marker gene has recently stood out as able to provide deep coverage to determine the microdiversity of the community. However, current petB primer set produce a 597 bp amplicon that is not suitable for MiSeq chemistry. Here, we designed and tested a petB primer pair that targets both Prochlorococcus and Synechococcus communities producing an appropriate amplicon to be used with state-of-the-art Illumina MiSeq. This new primer set allows the classification of both groups to a low taxonomic level and is therefore suitable for high throughput experiments using MiSeq technologies, therefore constituting a useful, novel tool to facilitate further studies on Prochlorococcus and Synechococcus communities. •This work describes the de novo design of a Prochlorococcus and Synechococcus-specific petB primer pair, allowing the characterization of both populations to a low taxonomic level.•This primer pair is suitable for widespread Illumina MiSeq sequencing technologies.•petB was confirmed as an adequate target for the characterization of both picocyanobacteria.

Picocyanobacteria Community and Cyanophage Infection Responses to Nutrient Enrichment in a Mesocosms Experiment in Oligotrophic Waters.

Prochlorococcus and Synechococcus are pico-sized cyanobacteria that play a fundamental role in oceanic primary production, being particularly important in warm, nutrient-poor waters. Their potential response to nutrient enrichment is expected to be contrasting and to differ from larger phytoplankton species. Here, we used a metagenomic approach to characterize the responses to nutrient enrichment in the community of picocyanobacteria and to analyze the cyanophage response during a mesocosms experiment in the oligotrophic Red Sea. Natural picoplankton community was dominated by Synechococcus clade II, with marginal presence of Prochlorococcus (0.3% bacterial reads). Increased nutrient input triggered a fast Synechococcus bloom, with clade II being the dominant, with no response of Prochlorococcus growth. The largest bloom developed in the mesocosms receiving a single initial input of nutrients, instead of daily additions. The relative abundances of cyanophage sequences in cellular metagenomes increased during the experiment from 12.6% of total virus reads up to 40% in the treatment with the largest Synechococcus bloom. The subsequent collapse of the bloom pointed to a cyanophage infection on Synechococcus that reduced its competitive capacity, and was then followed by a diatom bloom. The cyanophage attack appears to have preferentially affected the most abundant Synechococcus clade II, increasing the evenness within the host population. Our results highlight the relevance of host-phage interactions on determining population dynamics and diversity of Synechococcus populations.

Warming and CO2 Enhance Arctic Heterotrophic Microbial Activity.

Ocean acidification and warming are two main consequences of climate change that can directly affect biological and ecosystem processes in marine habitats. The Arctic Ocean is the region of the world experiencing climate change at the steepest rate compared with other latitudes. Since marine planktonic microorganisms play a key role in the biogeochemical cycles in the ocean it is crucial to simultaneously evaluate the effect of warming and increasing CO2 on marine microbial communities. In 20 L experimental microcosms filled with water from a high-Arctic fjord (Svalbard), we examined changes in phototrophic and heterotrophic microbial abundances and processes [bacterial production (BP) and mortality], and viral activity (lytic and lysogenic) in relation to warming and elevated CO2. The summer microbial plankton community living at 1.4°C in situ temperature, was exposed to increased CO2 concentrations (135-2,318 µatm) in three controlled temperature treatments (1, 6, and 10°C) at the UNIS installations in Longyearbyen (Svalbard), in summer 2010. Results showed that chlorophyll a concentration decreased at increasing temperatures, while BP significantly increased with pCO2 at 6 and 10°C. Lytic viral production was not affected by changes in pCO2 and temperature, while lysogeny increased significantly at increasing levels of pCO2, especially at 10°C (R 2 = 0.858, p = 0.02). Moreover, protistan grazing rates showed a positive interaction between pCO2 and temperature. The averaged percentage of bacteria grazed per day was higher (19.56 ± 2.77% d-1) than the averaged percentage of lysed bacteria by virus (7.18 ± 1.50% d-1) for all treatments. Furthermore, the relationship among microbial abundances and processes showed that BP was significantly related to phototrophic pico/nanoflagellate abundance in the 1°C and the 6°C treatments, and BP triggered viral activity, mainly lysogeny at 6 and 10°C, while bacterial mortality rates was significantly related to bacterial abundances at 6°C. Consequently, our experimental results suggested that future increases in water temperature and pCO2 in Arctic waters will produce a decrease of phytoplankton biomass, enhancement of BP and changes in the carbon fluxes within the microbial food web. All these heterotrophic processes will contribute to weakening the CO2 sink capacity of the Arctic plankton community.