Changes in microbial (Bacteria and Archaea) plankton community structure after artificial dispersal in grazer-free microcosms

Microbes are considered to have a global distribution due to their high dispersal capabilities. However, our knowledge of the way geographically distant microbial communities assemble after dispersal in a new environment is limited. In this study, we examined whether communities would converge because similar taxa would be selected under the same environmental conditions, or would diverge because of initial community composition, after artificial dispersal. To this aim, a microcosm experiment was performed, in which the temporal changes in the composition and diversity of different prokaryoplankton assemblages from three distant geographic coastal areas (Banyuls-sur-Mer in northwest Mediterranean Sea, Pagasitikos Gulf in northeast Mediterranean and Woods Hole, MA, USA in the northwest Atlantic), were studied. Diversity was investigated using amplicon pyrosequencing of the V1-V3 hypervariable regions of the 16S rRNA. The three assemblages were grown separately in particle free and autoclaved Banyuls-sur-mer seawater at 18 °C in the dark. We found that the variability of prokaryoplankton community diversity (expressed as richness, evenness and dominance) as well as the composition were driven by patterns observed in Bacteria. Regarding community composition, similarities were found between treatments at family level. However, at the OTU level microbial communities from the three different original locations diverge rather than converge during incubation. It is suggested that slight differences in the composition of the initial prokaryoplankton communities, resulted in separate clusters the following days even when growth took place under identical abiotic conditions

Unicellular eukaryotic community response to temperature and salinity variation in mesocosm experiments

Climate change has profound impacts on marine biodiversity and biodiversity changes in turn might affect the community sensitivity to impacts of abiotic changes. We used mesocosm experiments and Next Generation Sequencing to study the response of the natural Baltic and Mediterranean unicellular eukaryotic plankton communities (control and +6oC heat shock) to subsequent salinity changes (-5 psu, +5 psu). The impact on Operational Taxonomic Unit (OTU) richness, taxonomic and functional composition and rRNA:rDNA ratios were examined. Our results showed that heat shock leads to lower OTU richness (21% fewer OTUs in the Baltic and 14% fewer in the Mediterranean) and a shift in composition towards pico- and nanophytoplankton and heterotrophic related OTUs. Heat shock also leads to increased rRNA:rDNA ratios for pico- and micrograzers. Less than 18% of shared OTUs were found among the different salinities indicating the crucial role of salinity in shaping communities. The response of rRNA:rDNA ratios varied highly after salinity changes. In both experiments the diversity decrease brought about by heat shock influenced the sensitivity to salinity changes. The heat shock either decreased or increased the sensitivity of the remaining community, depending on whether it removed the more salinity-sensitive or the salinity-tolerant taxa

Variability and Community Composition of Marine Unicellular Eukaryote Assemblages in a Eutrophic Mediterranean Urban Coastal Area with Marked Plankton Blooms and Red Tides

The Thessaloniki Bay is a eutrophic coastal area which has been characterized in recent years by frequent and intense phytoplankton blooms and red tides. The aim of the study was to investigate the underexplored diversity of marine unicellular eukaryotes in four different sampling sites in Thessaloniki Bay during a year of plankton blooms, red tides, and mucilage aggregates. High-Throughput Sequencing (HTS) was applied in extracted DNA from weekly water samples targeting the 18S rRNA gene. In almost all samples, phytoplankton blooms and/or red tides and mucilage aggregates were observed. The metabarcoding analysis has detected the known unicellular eukaryotic groups frequently observed in the Bay, dominated by Bacillariophyta and Dinoflagellata, and revealed taxonomic groups previously undetected in the study area (MALVs, MAST, and Cercozoa). The dominant OTUs were closely related to species known to participate in red tides, harmful blooms, and mucilage aggregates. Other OTUs, present also during the blooms in low abundance (number of reads), were closely related to known harmful species, suggesting the occurrence of rare taxa with potential negative impacts on human health not detectable with classical microscopy. Overall, the unicellular eukaryote assemblages showed temporal patterns rather than small-scale spatial separation responding to the variability of physical and chemical factors

Haematococcus: a successful air-dispersed colonist in ephemeral waters is rarelyfound in phytoplankton communities

In a literature search, the presence of Haematococcus in phytoplankton communities and its biogeography were investigated. Haematococcus, although showing a wide biogeographical distribution, has been rarely found in phytoplankton communities. Simultaneously, the colonization potential of air-dispersed Haematococcus in ephemeral waters and its interactions with coexisting phytoplankton taxa were examined by microscopy and molecular methods. Haematococcus was a successful colonist, appearing among the first taxa in the experimental containers. According to principal component analysis, Haematococcus growth rate was negatively correlated with the abundance and species richness of the other autotrophs. Furthermore, a negative correlation between Haematococcus and Chlamydomonas and a positive one between Haematococcus and Chlorella were found. Overall, Haematococcus appears to be an effective air-dispersed alga that can successfully colonize and establish populations in small ephemeral water bodies. However, its absence from phytoplankton in larger permanent water bodies could be related to its high light requirements, its competitive disadvantages against other algae, and the grazing pressures from predators. The results of our study suggest a life strategy based on adaptation to higher light intensities in very shallow waters compared with optical dense lakes. Therefore, ephemeral waters are the regular habitat for Haematococcus instead of being “stepping stones” for the colonization of lake phytoplankton

Review of Harmful Algal Blooms in the Coastal Mediterranean Sea, with a Focus on Greek Waters

Anthropogenic marine eutrophication has been recognized as one of the major threats to aquatic ecosystem health. In recent years, eutrophication phenomena, prompted by global warming and population increase, have stimulated the proliferation of potentially harmful algal taxa resulting in the prevalence of frequent and intense harmful algal blooms (HABs) in coastal areas. Numerous coastal areas of the Mediterranean Sea (MS) are under environmental pressures arising from human activities that are driving ecosystem degradation and resulting in the increase of the supply of nutrient inputs. In this review, we aim to present the recent situation regarding the appearance of HABs in Mediterranean coastal areas linked to anthropogenic eutrophication, to highlight the features and particularities of the MS, and to summarize the harmful phytoplankton outbreaks along the length of coastal areas of many localities. Furthermore, we focus on HABs documented in Greek coastal areas according to the causative algal species, the period of occurrence, and the induced damage in human and ecosystem health. The occurrence of eutrophication-induced HAB incidents during the past two decades is emphasized

Airborne microorganisms in urban areas: biodiversity, succession and dispersion

The present thesis aims at the microscopic and phylogenetic characterization of the airborne microorganisms of Thessaloniki’s city center, with simultaneous observations in the city of Kastoria, as well as the rural areas of Mikri Volvi and Sarti. All study areas in proximity with natural aquatic systems. At the same time, the seasonal changes in the composition and abundance and the effect of meteorological conditions in the diversity, succession and dispersal of airborne microorganisms in Thessaloniki, were studied. The ecological characteristics of the dominant microorganisms in the atmosphere and their ability to colonize newly formed aquatic systems, were examined. On this basis, an assessment of the possible effects of the airborne microorganisms that were identified on human health, was attempted. This thesis is the first study on airborne phototrophic microorganisms in urban areas in Greece and the first in international level that attempts molecular characterization of the diversity of airborne eukaryotes in general. During the study, a high diversity of airborne algae, cyanobacteria, fungi and protists was identified in Thessaloniki. The total number of taxa that were identified with microscopic analysis was 101 (59 cyanobacteria and algae taxa and 42 fungi taxa). This diversity represents a large local pool of airborne microorganisms in relation to the species pool that have been reported worldwide, supporting the hypothesis of unexplored airborne microbial diversity. A small portion of the total number of airborne microorganisms (12%) were identified in the simultaneous observation of the proximate aquatic systems of Thessaloniki Bay and the university water pool, suggesting a possible allochthonous origin from distant source pools or transport from local soil sources. The abundance of airborne microorganisms in Thessaloniki showed maximal values that are close to the highest values that have been reported from other urban areas worldwide (up to 500 cells m-3 of air for cyanobacteria and algae and 1500 cells m-3 of air for fungi). A large portion of the airborne microorganisms that were identified in Thessaloniki, that belonged mainly in the taxonomic groups of fungi, but also consisted of cyanobacteria and algae taxa as well, has been acknowledged to inflict illness when inhaled, such as asthma, allergy and respiratory illness. During the colonization experiments in Thessaloniki, 24 cyanobacteria and algae taxa and 14 protozoan taxa were identified in total. Only 30% of the total number of taxa were identified at the same time in direct air samples, suggesting that the airborne microorganisms disperse through the air with forms, such as cysts, that are not identifiable. The colonization of the experimental containers in the first two weeks of the experiments was rapid, but it quickly stabilized to a plateau. The successful colonists, that were the chlorophytes Haematococcus lacustris, Scenedesmus cf. οbliquus και Chlorella sp., the cyanobacterium Limnothrix sp. and the protozoa HNF and Amoeba sp., were also the final winners of the experimental systems, as they were found to be dominant in the final weeks of the experiments. During three seasons in Thessaloniki, the airborne colonizers formed complex food webs in the experimental containers.Η παρούσα διδακτορική διατριβή στοχεύει στον ταξινομικό (μικροσκοπική και φυλογενετική ανάλυση) και οικολογικό χαρακτηρισμό των αερομεταφερόμενων μικροοργανισμών (κυανοβακτήρια, μικροφύκη και πρωτόζωα) στο κέντρο της Θεσσαλονίκης με παράλληλες παρατηρήσεις στην Καστοριά, καθώς και στις απομακρυσμένες από αστικά κέντρα περιοχές της Μικρής Βόλβης και της Σάρτης. Διερευνήθηκαν οι χρονικές μεταβολές στη σύνθεση και αφθονία των μικροοργανισμών του αέρα, καθώς και η επίδραση των μετεωρολογικών παραγόντων στην ποικιλότητα, διαδοχή και διασπορά των αερομεταφερόμενων μικροοργανισμών στη Θεσσαλονίκη. Εξετάστηκαν τα οικολογικά χαρακτηριστικά των κυρίαρχων μικροοργανισμών στον αέρα και η ικανότητα τους να εποικίσουν υδάτινα ενδιαιτήματα. Με βάση τα παραπάνω, πραγματοποιήθηκε προσπάθεια εκτίμησης των πιθανών επιδράσεων των αερομεταφερόμενων μικροοργανισμών στην ανθρώπινη υγεία. Αυτή η διδακτορική διατριβή αποτελεί την πρώτη εργασία που αφορά σε αερομεταφερόμενους αυτότροφους και ετερότροφους μικροοργανισμούς σε αστικές περιοχές της Ελλάδας, ενώ είναι η πρώτη σε διεθνές επίπεδο που επιχειρεί μοριακό χαρακτηρισμό της ποικιλότητας αερομεταφερόμενων ευκαρυωτικών ειδών γενικότερα. Κατά τη διάρκεια της εργασίας διαπιστώθηκε ότι ο αέρας της Θεσσαλονίκης παρουσιάζει υψηλή ποικιλότητα αερομεταφερόμενων μικροφυκών, κυανοβακτηρίων, μυκήτων και πρωτόζωων, με το συνολικό αριθμό taxa που αναγνωρίστηκαν με μικροσκοπική ανάλυση να είναι 101 (59 taxa κυανοβακτηρίων και μικροφυκών και 42 taxa μυκήτων). Η ποικιλότητα αυτή αντιπροσωπεύει μία μεγάλη τοπική δεξαμενή αερομεταφερόμενων μικροοργανισμών σε σχέση με τη δεξαμενή ειδών που έχουν αναφερθεί σε παγκόσμια κλίμακα. Ένα μικρό ποσοστό (12%) του συνολικού αριθμού των αερομεταφερόμενων μικροοργανισμών καταγράφηκαν ταυτόχρονα και στα προσκείμενα υδάτινα συστημάτα του Όρμου της Θεσσαλονίκης και της υδατοσυλλογής του πανεπιστημίου. Τα αποτέλεσματα αυτά υποδηλώνουν πιθανή αλλόχθονη προέλευση του μεγαλύτερου ποσοστού των μικροοργανισμών από μακρινές υδάτινες δεξαμενές ειδών ή μεταφορά τους από χερσαίες τοπικές δεξαμενές ειδών. Η αφθονία των αερομεταφερόμενων μικροοργανισμών στη Θεσσαλονίκη παρουσίασε μέγιστα που πλησιάζουν τις ανώτερες τιμές που έχουν καταγραφεί σε αστικές περιοχές διεθνώς (μέχρι 500 κύτταρα m-3 αέρα για κυανοβακτήρια και μικροφύκη και 1500 κύτταρα m-3 αέρα για μύκητες). Ένα μεγάλο ποσοστό των μικροοργανισμών, οι οποίοι ανήκαν κυρίως στους μύκητες, αλλά συμπεριλάμβαναν και κυανοβακτήρια και μικροφύκη που αναγνωρίστηκαν στον αέρα της Θεσσαλονίκης, έχει βρεθεί να προκαλεί παθήσεις, όπως άσθμα, αλλεργίες και παθήσεις του αναπνευστικού, όταν εισπνέονται. Κατά τη διάρκεια των πειραμάτων εποικισμού υδάτινων μικροαιολικών συστημάτων στη Θεσσαλονίκη αναγνωρίστηκαν συνολικά 24 taxa κυανοβακτηρίων και μικροφυκών και 14 taxa πρωτόζωων. Μόνο το 30% του συνολικού αριθμού taxa αναγνωρίστηκαν ταυτόχρονα και με τις δειγματοληψίες αέρα, υποδηλώνοντας ότι οι μικροοργανισμοί μεταφέρονται με τον αέρα με μορφές, όπως ανθεκτικές κύστεις, που δεν αναγνωρίζονται. Ο εποικισμός των πειραματικών διατάξεων τις πρώτες δύο εβδομάδες των πειραμάτων πραγματοποιήθηκε με ταχείς ρυθμούς, ενώ στη συνέχεια ο ρυθμός εποικισμού σταθεροποιήθηκε σε πλάτωμα κορεσμού. Οι επιτυχημένοι εποικιστές, οι οποίοι συμπεριλάμβαναν τα χλωροφύκη Haematococcus lacustris, Scenedesmus cf. οbliquus και Chlorella sp., το κυανοβακτήριο Limnothrix sp. και τα πρωτόζωα HNF και Amoeba sp., ήταν και οι τελικοί νικητές των συστημάτων, καθώς βρέθηκαν να επικρατούν και στις τελικές εβδομάδες των πειραμάτων. Σε τρεις εποχές στη Θεσσαλονίκη οι αερομεταφερόμενοι εποικιστές σχημάτισαν τροφικά πλέγματα στις πειραματικές διατάξεις

Parasitic Eukaryotes in a Meso-Eutrophic Coastal System with Marked Phaeocystis globosa Blooms

This study explored the potential interactions and relations of the putative Symbiotic/Decomposer/Parasitic community (SymbDec) with other taxa and predominant environmental parameters in the eastern English Channel (EEC) over a 2.5 years period (32 sampling dates). The EEC is a meso-eutrophic coastal system portrayed by intense reoccurring patterns in plankton succession with blooms of the Haptophyte Phaeocystis globosa and abundant populations of colonial diatoms and dinoflagellate grazers before and after the bloom. The study of the V2-V3 hypervariable region of the 18S rRNA gene allowed an overview of the planktonic community, which consisted of 32 high-level taxonomic groups. Forty-two percent of the identified OTUs belonged to symbionts, decomposers or other putative parasitic taxa belonging mainly to Syndiniales (MALV—MArine ALveolates), Fungi, Cercozoa, Perkinsea, and others, most of which were plankton parasites. We examined the Local Similarity Analysis (LSA) network of lag delayed correlations on the 142 most abundant OTUs (>0.1% of the total number of reads). LSA showed that 99 OTUs had highly significant connections, involving 26 OTUs characterized as potential parasites, and 2 decomposers. The parasitic network had “small world” characteristics, rendering the assemblage more robust to abiotic variations, but more exposed to the removal of highly associated taxa. The majority of the highly connected OTUs belonged to MALV, which were mainly connected with dinoflagellates, suggesting that it might play an important role on dinoflagellate top-down control. The bloom forming P. globosa was not observed in connection with parasitic OTUs in the network. Four environmental parameters (O2, N, T, and N/P) were strongly connected with only 11 OTUs, while correlations between microbes dominated the network

Metagenomic Characterization of Bacterial Communities on Ready-to-Eat Vegetables and Effects of Household Washing on their Diversity and Composition

Ready-to-eat (RTE) leafy salad vegetables are considered foods that can be consumed immediately at the point of sale without further treatment. The aim of the study was to investigate the bacterial community composition of RTE salads at the point of consumption and the changes in bacterial diversity and composition associated with different household washing treatments. The bacterial microbiomes of rocket and spinach leaves were examined by means of 16S rRNA gene high-throughput sequencing. Overall, 886 Operational Taxonomic Units (OTUs) were detected in the salads’ leaves. Proteobacteria was the most diverse high-level taxonomic group followed by Bacteroidetes and Firmicutes. Although they were processed at the same production facilities, rocket showed different bacterial community composition than spinach salads, mainly attributed to the different contributions of Proteobacteria and Bacteroidetes to the total OTU number. The tested household decontamination treatments proved inefficient in changing the bacterial community composition in both RTE salads. Furthermore, storage duration of the salads at refrigeration temperatures affected the microbiome, by decreasing the bacterial richness and promoting the dominance of psychrotropic bacteria. Finally, both salads were found to be a reservoir of opportunistic human pathogens, while washing methods usually applied at home proved to be inefficient in their removal

Microzooplankton community associated with phytoplankton blooms in the naturally iron-fertilized Kerguelen area (Southern Ocean)

The spatial and temporal community composition of microzooplankton (dinoflagellates and ciliates) was assessed in the Kerguelen area (Southern Ocean) during the KEOPS2 cruise in early spring (October-November) 2011. This naturally iron-fertilized region was characterized by a complex mesoscale circulation resulting in a patchy distribution of phytoplankton blooms. Collectively, 97 morphospecies of dinoflagellates and ciliates belonging to 41 genera were identified by microscopy, and 202 Alveolata-related OTUs (operational taxonomical units) were retrieved with tag-pyrosequencing. Microscopy and pyrosequencing data were in accordance, in that diatom-consuming dinoflagellates were the most enhanced taxa in the blooms. Dinoflagellates also showed significant positive relationships with phytoplankton pigments, while no major differences were found in the ciliate abundances inside and outside the blooms. Cluster analysis showed clear differences in the phytoplankton and microzooplankton community structures between the iron-fertilized and HNLC (high nutrient low chlorophyll) waters, and between the blooms, concerning their location and the fertilization mechanisms. These results were combined with the rates of primary production and mesozooplankton consumption determined for the study area. The potential role of dinoflagellates and ciliates as phytoplankton consumers and as prey for mesozooplankton was then evaluated. Overall, heterotrophic dinoflagellates were probably the most important group of phytoplankton grazers, and a potential food source for copepods