Environmental Microbiology Reports

Macroalgae are found in a variety of marine vegetation ecosystems around the world, contributing significantly to global net primary production. In particular, the sea lettuce species, i.e., members of the genus Ulva (Chlorophyta), are located in many ecological niches and are characterized by excellent adaptability to environmental changes but depend on essential associated bacteria, which release algal growth and morphogenesis-promoting-factors (AGMPFs). Our work investigated the hypothesis that bacteria need to be stress-adapted to provide sufficient amounts of AGMPFs for the growth and morphogenesis of Ulva throughout its life cycle, even under severe environmental conditions. Our study thus aimed to understand which bacteria contribute to overcoming a variety of stressors in polar regions. Green macroalgae were collected from Potter Cove, King George Island (Isla 25 de Mayo), Antarctica, to study the associated microbiome and, subsequently, to identify AGMPFs releasing bacteria. Therefore, microbiome analysis was combined with morphogenetic bioassays and chemical analysis, identifying bacteria essential for algal growth under Antarctic conditions. Hereby, axenic cultures of a Mediterranean Ulva compressa (cultivar Ulva mutabilis), previously developed as a model system for bacteria-induced algal growth and morphogenesis, were inoculated with freshly isolated and cultivable Antarctic bacteria to determine their morphogenetic activity. The exploratory microbiome investigation identified numerous cold-adapted AGMPF-producing bacteria. Unlike the reference bacterial strains isolated from the Mediterranean Sea, the cold-adapted isolates Maribacter sp. BPC-D8 and Sulfitobacter sp. BPC-C4, released sufficient amounts of AGMPFs, such as thallusin, necessary for algal morphogenesis even at 2{degrees}C. Our results illustrate the role of chemical mediators provided by bacteria in cross-kingdom interactions under cold conditions within aquatic systems. The newly isolated bacteria will enable further functional studies to understand the resilience of the holobiont Ulva and might applied in algal aquaculture even under adverse conditions. The study highlights the importance of ecophysiological assays in microbiome analysis.

- Pico-phytoplankton have ample scope to react to environmental change. But we know little about the underlying physiological mechanisms that govern how evolutionary history may affect short-term responses to environmental change. - We investigated growth rates and carbon uptake related traits (i.e. fitness proxies) in different temperatures and at different times during the microbial growth curve of eight novel strains of Ostreococcus sp. (ca. 1-2{micro}m). The strains were isolated from two distinct regions of the Baltic Sea differing in salinity and temperature from North-East (Bornholm Basin) to South-West (Kiel area). - Strains from the warmer, more variable Kiel area had higher growth rates in general and showed more variable growth rates compared to strains from the colder and less variable Bornholm Basin. - In addition, growth was maintained in early stages of the growth curve by organic carbon acquisition and the increase in growth over time and with temperature was associated with an increase in inorganic carbon acquisition (net primary productivity). - Based on the differences between net primary productivity and potential growth on organic carbon, we postulate a shift in carbon acquisition between inorganic and organic sources in Ostreococcus sp. with potential implications on ecological dynamics within microbial communities.

Steinernema species are soil-dwelling and insect-parasitic nematodes that associate with symbiotic Xenorhabdus bacteria. During the infective juvenile (IJ) stage, Steinernema nematodes package species-specific Xenorhabdus bacteria in the anterior intestinal pockets. The nematodes can survive in the soil for months as they seek insect prey. The mechanisms of how these nematodes associate with environmental microbes other than their Xenorhabdus symbionts is barely known. Here, we report a new mechanism of E. coli Nissle (EcN) association with the nematode Steinernema hermaphroditum. We show that EcN cells are enclosed and lysed in at least four pairs of coelomocytes, suggesting these immune cells respond to bacterial invasion. During the IJ stage of nematode development, EcN cells localize to posterior intestinal vacuoles and enter the inter-cuticular space, where they proliferate, aggregate, then lyse. EcN cells expressed proteins in the cell lysates were maintained in the nematode cuticle over eight weeks in non-sterile soil. These observations suggest sequential steps of EcN colonization in the host nematodes involving an immune response that is distinctive from interactions with mutualistic symbiont. Our work establishes a novel framework of nematode-bacteria interaction with potential applications in environmental bioengineering.

Carbon monoxide (CO) degrading microorganisms are present in volcanic deposits throughout succession, with vegetation and soil influencing the communities present. The carboxydovores are a subset of CO degraders that use CO only as an energy source, raising the question of how the physiological and metabolic features of the carboxydovores can make these bacteria more competitive in harsh volcanic ecosystems. An enrichment strategy was modified, which enabled the isolation of two carboxydovore representatives from genera that were abundant in the native soils, Cupriavidus sp. CV2T (92.3% ANI vs. Cupriavidus basilensis DSM 11853) and a putative strain of Paraburkholderia terrae (Pb. terrae COX) (96.42% ANI vs. Pb. terrae KU-64T). These isolates oxidise CO across a very broad range of concentrations, and genome sequence analysis indicated that they use form-I carbon monoxide dehydrogenase (CODH) to do so. Cupriavidus sp. CV2T and Pb. terrae COX each oxidised CO specifically at stationary phase, but the conditions for induction of CODH expression were distinct. Cupriavidus sp. CV2T expressed CODH only in the presence of CO, while Pb. terrae COX expressed CODH regardless of the presence of CO. Based on metabolic and phylogenetic analyses, Cupriavidus sp. CV2T is recommended as a novel species within the genus Cupriavidus. Therefore, we propose the name Cupriavidus ulmosensis sp. nov. for the type strain CV2T (= NCIMB 15506T, = CECT 30956T). This study provides valuable insights into the physiology and metabolism of carboxydovores, which colonise volcanic ecosystems during succession. ImportanceVolcanic ecosystems harbour many bacteria that contribute to the environmentally important process of carbon monoxide (CO) oxidation. We demonstrate a modified method for isolating bacteria, which consume CO at very low concentrations as a supplementary energy source (carboxydovory), leading to the isolation of two novel strains (Cupriavidus sp. CV2T and Paraburkholderia terrae COX) from volcanic strata that formed in 1917 and 2015, respectively. The conditions under which CO consumption occurs were investigated; each strain consumed CO during stationary phase, but Pb. terrae COX consumed CO regardless of the prior growth conditions while Cupriavidus sp. CV2 was more controlled. Cupriavidus sp. CV2 is a type strain of a new species, Cupriavidus ulmosensis str. CV2, which demonstrates relatively high tolerance for CO. These strains provide the basis for further study of the physiology, metabolism, and genetics of CO oxidation by carboxydovores, and will help us to understand how bacteria colonise harsh volcanic ecosystems.

Symbiotic interactions drive species evolution, with nutritional symbioses playing vital roles across ecosystems. Chemosynthetic symbioses are globally distributed and ecologically significant, yet the lack of model systems has hindered research progress. The giant ciliate Zoothamnium niveum and its sulfur-oxidizing symbionts represent the only known chemosynthetic symbiosis with a short life span that has been transiently cultivated in the laboratory. While it is experimentally tractable and presents a promising model system, it currently lacks an open-source, simple, and standardized cultivation setup. Following the FABricated Ecosystems (EcoFABs) model, we leveraged 3D printing and polydimethylsiloxane (PDMS) casting to develop simple flow-through cultivation chambers that can be produced and adopted by any laboratory. The streamlined manufacturing process reduces production time by 86% and cuts cost by tenfold compared to the previous system. Benchmarking using previously established optimal growth conditions, the new open-source cultivation system proves stable, efficient, more autonomous, and promotes a more prolific growth of the symbiosis. For the first time, starting from single cells, we successfully cultivated the symbiosis in flow-through chambers for 20 days, spanning multiple generations of colonies that remained symbiotic. They were transferred from chamber to chamber enabling long-term cultivation and eliminating the need for continuous field sampling. The chambers, optimized for live imaging, allowed detailed observation of the synchronized growth between the host and symbiont. Highlighting the benefit of this new system, we here describe a new step in the first hours of development where the host pauses growth, expels a coat, before resuming growth, hinting at a putative symbiont selection mechanism early in the colony life cycle. With this simple, open-source, cultivation setup, Z. niveum holds promises for comparative studies, standardization of research and wide adoption by the symbiosis research community.

Phytophagous insects engage in symbiotic relationships with bacteria that contribute to digestion, nutrient supplementation, and development of the host. The analysis of shield bug microbiomes has been mainly focused on the gut intestinal tract predominantly colonized by Pantoea symbionts, and other microbial community members in the gut or other organs have hardly been investigated. In this study, we reveal that the Southern green shield bug Nezara viridula harbours a Sodalis symbiont in several organs, with a notable prevalence in salivary glands, and anterior regions of the midgut. Removing external egg microbiota via sterilization profoundly impacted insect viability but did not disrupt the vertical transmission of Sodalis and Pantoea symbionts. Based on the dominance of Sodalis in testes, we deduce that N. viridula males could be involved in symbiont vertical transmission. Genomic analyses comparing Sodalis species revealed that Sodalis sp. Nvir shares characteristics with both free- living and obligate insect-associated Sodalis spp. Sodalis sp. Nvir also displays genome instability typical of endosymbiont lineages, which suggests ongoing speciation to an obligate endosymbiont. Together, our study reveals that shield bugs harbour unrecognized symbionts that might be paternally transmitted.

Filamentous sulfide-oxidizing Beggiatoa spp. are widespread in marine coastal environments and can achieve significant biomass because of their substantial size. Their ability to store phosphates in the polymerized form of polyphosphates makes them potentially key players in altering the phosphorus (P) cycle at the sediment-water interface. This study examined phosphate uptake and polyphosphate formation in a P starved culture of the Beggiatoa sp. 35Flor strain. Remarkably, even after severe P starvation over five generations, the survival of the cultures was 46%, demonstrating considerable plasticity to different levels of phosphate availability. Under these P-depleted conditions, 23% of filaments still contained polyphosphates, underscoring its critical role in their metabolism. Upon reintroduction of phosphate to starved cultures, an extremely rapid phosphate uptake was observed within the first 10 minutes, with rates reaching up to 298 mmol P g-1 protein d-1, which is significantly higher than values previously described in the literature for similar-sized organisms. The high phosphate uptake capacity of Beggiatoa spp., estimated at 0.6 - 6 mmol m-2 d-1 for typical densities of filaments in coastal sediments, suggests that these bacteria may play an important role in buffering the phosphate flux in these environments. Thereby, they reduce primary production and subsequent oxygen consumption by other organisms, creating a negative feedback loop that helps maintain ecosystem stability. ImportanceSulfide-oxidizing bacteria of the genus Beggiatoa occur ubiquitously in marine coastal sediments and have a large potential to influence phosphate fluxes at the sediment-water interface owing to their ability to accumulate polyphosphate and their large size. However, the extent to which these bacteria can contribute to phosphorus (P) sequestration or release remains poorly assessed. The importance of this study lies in demonstrating the unusual flexibility in adaptation of the Beggiatoa sp. 35Flor strain to varying P availability, including extreme P starvation, and its capacity to rapidly uptake and store available phosphate in the form of polyphosphate. When considered at a global scale, these physiological traits could lead to P retention in shallow coastal waters, which, in turn, profoundly impacts ecological stability and ecosystem functioning.

Metamonads are a large and exclusively anaerobic clade of protists. Additionally, metamonads are one of the three clades with a proposed excavate ancestral cell morphology, characterised by a conspicuous ventral groove often accompanied by a posterior flagellum with a vane. Here, we characterise four isolates of an anaerobic bacterivorous flagellate from hypersaline and alkaline soda lake environments, which represents a novel clade. Small subunit ribosomal RNA (SSU rRNA) gene phylogenies support recent phylogenomic analyses in placing this clade as the sister group to Barthelona spp., a lineage that is itself sister to or deeply branching within Fornicata (Metamonada). The cells have a distinctive morphology comprised of a hunchbacked cell body with a narrow twisting ventral groove ending in a large opening to a conspicuous cytopharynx curving up the dorsal side of the cell. The right margin of the groove is defined by a thin lip that twists slightly to the left towards the posterior. The posterior of the cell ends in a spike up to half a cell body long. The posterior flagellum bears a wide ventral-facing vane. One isolate forms cysts with a complex wall and a single plug. The narrow ventral groove and elongate cytopharynx are shared with barthelonids. We describe one isolate as Skoliomonas litria, gen. et sp. nov. Further investigation of mitochondrial-related organelles (MRO) in Skoliomonas spp. and detailed ultrastructural studies would be important to understanding the evolution of adaptation to anaerobic conditions in Metamonads--especially fornicates--as well as the evolution of the excavate groove.

Arbuscular mycorrhizal fungi (AMF; Glomeromycotina) are symbionts of most plant species that are known to possess unique intracytoplasmic endosymbiotic bacteria with an enigmatic role. Candidatus Moeniiplasma glomeromycotorum (CaMg) was shown to be widespread along the AMF phylogeny and present in most AMF species and isolates of those species. The model AMF species, Rhizophagus irregularis, that can be cultivated in vitro and for which a lot of genomic information now exists, would be the ideal model to study the true nature of the CaMg-AMF symbiosis. However, R. irregularis was never found to host endobacteria. Here we show by DNA sequencing that R. irregularis can, indeed, host CaMg (Ri-CaMg). However, this appears rare as only one R. irregularis isolate out of 58 hosted CaMg. In that isolate, the endosymbiotic bacterial population was genetically homogenous. By sequencing the complete genome of the bacteria, we found that its genome is among the smallest of all known CaMg and Mycoplasma-like genomes, with a highly reduced gene repertoire, suggesting a strong adaptation to the intracellular life. We discuss our findings in the light of previous literature on CaMg and on the same AMF isolates and suggest that these endosymbionts are more likely parasites than non-obligatory mutualists.

Endomyxans are a poorly sampled and incompletely resolved aggregate of Rhizarian lineages that fall outside Filosa and Retaria. Among them, "Novel Clade 12" (NC12; Bass et al. 2009) is an environmental clade comprised primarily of sequences derived from anoxic sediments, hitherto lacking a morphologically-characterised representative. We have cultivated a marine anaerobic eukaryotroph, SSF, that we identify as the first representative of NC12. SSF is a teardrop-shaped cell with two unequal flagella emerging a third of the way down the cell behind a distinctive row of refractile globules. The posterior end of the cell is filled with food vacuoles. There is a surface thickening discernible in light microscopy. We also describe another distinct anaerobe eukaryotrophic lineage, also cultivated from marine sediment: PG. It consists of large pyriform cells with a substantial trailing "tail" and two unequal flagella, the posterior exceptionally long. In small subunit ribosomal RNA gene phylogenies, it falls outside the characterised clades and forms a distinct novel rhizarian lineage in its own right. Together, SSF and PG represent two additional independent adaptations to anoxic conditions within Rhizaria.

Protistan predators are key regulators of microbial food webs, yet most are considered to occupy relatively narrow trophic niches. Here, we demonstrate that Mayorella spp. (Amoebozoa), isolated from marine and freshwater environments, exhibits exceptional trophic breadth spanning multiple trophic levels. Live-cell imaging revealed predation on bacteria, algae, dinoflagellates, diatoms, flagellates, ciliates, and multicellular prey including rotifers. Large or filamentous prey were engulfed whole or mechanically fragmented during ingestion. Notably, Mayorella consumed both trophozoites and cysts of free-living amoebae (Naegleria and Acanthamoeba), with clear digestion of cyst contents. Dense cultures showed aggregation around large prey and facultative cannibalism. Ingestion of microplastic-like particles occurred without evidence of digestion. Predator cell size and population density increased markedly when feeding on protist or mixed prey relative to bacterial diets, indicating pronounced trophic plasticity. These findings establish Mayorella as a broad-spectrum, cross-trophic predator with the capacity to exert top-down effects across microbial food webs and suggest a previously underappreciated role in the suppression of pathogenic free-living amoebae.

In the context of the soil food web, the transfer of plant-fixed energy and carbon to higher trophic levels has traditionally been attributed to two main energy channels: the fungal energy channel and the bacterial energy channel. Historically, protists were overlooked in the fungal energy channel, which was believed to be controlled by fungivorous microarthropods and nematodes. In this study, we investigated fungivorous protists in the rhizosphere of Arabidopsis thaliana. Our findings revealed a notable abundance and diversity of protists that have developed specialized strategies to overcome the protective cell wall of fungi. Among the identified species were two Vampyrellida (Rhizaria) species, namely Theratromyxa weberi and Platyreta germanica, as well as one Arcellinida (Amoebozoa) species, called Cryptodifflugia oviformis. While T. weberi typically consumed entire fungal cells, the other two species perforated fungal cell walls and extracted the cellular contents. We elucidate the feeding strategies and dietary ranges of the amoebae, highlighting the non-uniform nature of fungivory in protists, as different taxa have evolved distinct approaches to access fungi as a food source. Moreover, we provide publicly available cultures of these protists to facilitate further experimental investigations within the research community.

Ammonia oxidizing archaea (AOA) are widespread and highly abundant in nature. Despite their typical aerobic metabolism, they can be abundant in ecosystems where oxygen is scarce. Recent observations revealed that the AOA isolate Nitrosopumilus maritimus produces oxygen and dinitrogen at nanomolar concentrations, upon oxygen depletion through nitric oxide (NO) dismutation. Here, we explore NO dismutation capability in other ammonia oxidizers with different phylogenetic affinities and from different environmental settings. The organisms explored include three marine AOA, one soil AOA and two soil ammonia-oxidizing bacteria (AOB). Upon oxygen depletion all isolates accumulated oxygen. In incubations with 15N tracers with ongoing oxygen accumulation, the AOA strains Nitrosopumilus adriaticus and Nitrosopumilus viennensis produced 46N2O from nitrite. Transient 46N2O accumulation followed by 30N2 production was detected in the AOA strains Nitrosopumilus piranensis and Nitrosopumilus sp. CCS1, supporting the earlier observation that NO-dismutation is a common metabolism in AOA, albeit with physiological variations between different strains. An important physiological variable is the capability to reduce N2O to N2. The finding of oxygen production in several AOA, as well as AOB, indicates that this process is widely distributed among the tree of life and adds an explanation for their abundance in oxygen-depleted environments.

Labyrinthulomycetes are a class of fungus-like heterotrophic protists from the Stramenopiles lineage, recognized for their ecological role as decomposers and contributors to nutrient cycling. They colonize various substrates, from seaweed to terrestrial environments, utilizing ectoplasmic networks for nutrient absorption. This study characterized a novel Labyrinthula strain associated with the marine diatom Biddulphia. Phylogenetic analysis of the full-length 18S rRNA gene positioned this strain as a new species, Labyrinthula merlionensis sp. nov. Scanning electron and light microscopy observations revealed bi-flagellated zoospores and spindle-shaped vegetative cells with ectoplasmic networks. Time-series observations of the interactions between L. merlionensis and Biddulphia were categorised into different phases: establishment, infection, and aggregation. Scanning electron and confocal microscopy observations during the infection phase established the use of ectoplasmic nets to target the marginal ridge regions between diatoms, and the detection of labyrinthulid cells within diatom frustules. These findings enhance the understanding of the diversity, morphology, and ecological roles of Labyrinthulomycetes, particularly their intra- and extra-cellular interactions with diatom hosts.

Marine non-cyanobacterial diazotrophs (NCDs) are recognized as globally distributed, however, few representatives have been isolated in pure cultures. As a result, understanding the physiology, growth rate, substrate preference and dinitrogen (N2) fixation capabilities proves difficult. Thalassolituus haligoni. sp. nov., BB40 was isolated from a fjord-like inlet within Kjipuktuk (Halifax), Nova Scotia. The fully sequenced genome displayed all necessary genes required for N2 fixation, and various carbon uptake pathways. The gram-negative flagellated rod shape bacterium displayed significantly higher growth rates in medium amended with nitrate (NO3-) or ammonia (NH3), compared to dissolved N2, as the sole nitrogen source. Biological N2 fixation rates were detectable across all conditions, measuring a range from 9.34 x 10-6 to 1.4 x 10-1 fmol N cell-1 day-1. Growth of the isolate was successful between 4 {degrees}C up to 35 {degrees}C, with a Topt of 20 {degrees}C for N2, and between 27 - 30 {degrees}C for fixed nitrogen (NO3- and NH3). The closest relatives to T. haligoni, were found to be the uncultured Arc-gamma-03 (99% average nucleotide identity (ANI)) and Oceanobacter antarcticus (81% ANI). T. haligoni also displays versatile capabilities for growth on various carbon, and nitrogen sources, and antibiotics. Collectively this study provides an in-depth physiological assessment of an Oceanospirillales diazotrophic species which we presently have limited knowledge of.

Life in Movile Cave (Romania) relies entirely on carbon fixation by bacteria. The microbial community in the surface water of Movile Caves hypoxic air bells is dominated by large spherical-ovoid bacteria we identified as Thiovulum sp. (Campylobacterota). These form a separate phylogenetic cluster within the Thiovulaceae, consisting mostly of freshwater cave bacteria. We compared the closed genome of this Thiovulum to that of the marine strain Thiovulum ES, and to a genome we assembled from public data from the sulfidic Frasassi caves. The Movile and Frasassi Thiovulum were very similar, differing greatly from the marine strain. Based on their genomes, cave Thiovulum can switch between aerobic and anaerobic sulfide oxidation using O2 and NO3- as electron acceptors, respectively. NO3-, is likely reduced to NH3 via dissimilatory nitrate reduction to ammonia using periplasmic nitrate reductase (Nap) and hydroxylamine oxidoreductase. Thus, Thiovulum, is likely important to both S and N cycles in sulfidic subterranean aquatic ecosystems. Additionally, we suggest that the short peritrichous flagella-like structures typical of Thiovulum are type IV pili, for which genes were found in all Thiovulum genomes. These pili may play a role in veil formation, connecting adjacent cells and the exceptionally fast swimming of these bacteria.

In a field experiment we investigated the influence of the environmental filters soil type and plant species identity on rhizosphere community assembly of Cercozoa, a dominant group of (mostly bacterivorous) soil protists. The experiment was set up with two plant species, lettuce and potato, grown in an experimental plot system with three contrasting soils. Plant species (14%) and rhizosphere origin (vs. bulk soil) with 13%, together explained four times more variation in cercozoan beta diversity than the three soil types (7% explained variation in beta diversity). Our results clearly confirm the existence of plant species-specific protist communities. Network analyses of bacteria-Cercozoa rhizosphere communities identified scale-free small world topologies, indicating mechanisms of self-organization. While the assembly of rhizosphere bacterial communities is bottom-up controlled through the resource supply from root (secondary) metabolites, our results support the hypothesis that the net effect may depend on the strength of top-down control by protist grazers. Since grazing of protists has a strong impact on the composition and functioning of bacteria communities, protists expand the repertoire of plant genes by functional traits, and should be considered as protist microbiomes in analogy to bacterial microbiomes. HighlightMicrobiomes of rhizosphere protists are plant species-specific and tightly co-evolving with their bacterial prey, thereby extending and modifying the functional repertoire of the bacterial-plant symbiosis.

Bacteria in aquatic environments are a principal food source for predatory protists. Whereas interactions between bacteria and protists are recognized to play important roles in determining the pathogenesis and epidemiology of several human pathogens, few studies have systematically characterized the interactions between specific aquatic bacteria and protists beyond the prey-predator relation. We therefore surveyed the outcome of individual co-cultures between 18 different genome-sequenced marine bacteria with known virulence gene repertoires and three model protist species widely used for assessing bacteria-protist interactions. Strikingly, ten, five, and three bacterial isolates were capable of lysing the protists Acanthamoeba polyphaga, Tetrahymena pyriformis and Euglena gracilis, respectively. A majority of the bacteria were able to grow and/or maintain viable populations in the presence of viable protists. Some bacteria survived longer in the presence of viable protists but not heat-killed protists, and were observed in protist vacuoles. In this respect, thus, marine bacteria are similar to several protist-dependent human pathogens, including Legionella. Analyses of growth patterns in low-nutrient media showed that co-cultivation with A polyphaga allowed one bacterial strain to overcome nutritional stress and obtain active growth. Five isolates depended on viable amoebae to grow, notwithstanding nutrient media status. The remarkable capability of surviving encounters with, and even actively killing, bacterivorous protists, indicates that diverse (and possibly novel) bacterial defense strategies and virulence mechanisms to access nutrients are widespread among marine bacteria. The diversity of interactions uncovered here has important implications for understanding ecological and evolutionary consequences of population dynamics in bacteria and protists. IMPORTANCEThe microbiome constitutes the base of food webs in marine waters. Its composition partly reflects biotic interactions, where bacteria primarily are considered as prey of predatory protists. However, studies that focus on one or a few species have shown that some bacteria have abilities to escape grazing and may even be capable of lysing their protist predators. In this study, we substantially extend these findings by systematically investigating interactions among multiple taxa of both bacteria and protists. Our results show that marine bacteria display a wider and more complex range of interactions with their predators than generally recognized - from growth dependency to protist lysis. Given that such interactions play key roles in the pathogenesis and epidemiology of several human pathogens, our findings imply that bacterial virulence traits can contribute to defining the structure and ecology of the marine microbiome.

Microalgal fitness in nature is shaped by interactions within a diverse microbial community, yet most experimental studies have examined algal-bacterial interactions in pairwise systems. It is well established that bacteria can exhibit growth promoting or inhibiting effects on co-existing algae. Comparatively little information is available about how additional partners can alter the outcome of diatom-bacteria interactions. In the present study, we screened the pairwise interaction of the marine diatom Skeletonema marinoi with ten different bacteria. This screening identified Marinobacter adhaerens as a growth promoting and Vibrio cyclitrophicus HSW24 as growth inhibiting partner. Growth inhibition of V. cyclitrophicus was associated with cell lysis, chain fragmentation and altered pigmentation whereas M. adhaerens supported increased chlorophyll a fluorescence, uniform pigmentation, intact chains and healthy cell morphology. In a tripartite community containing both bacteria and the alga, M. adhaerens protected S. marinoi from the inhibitory effect of V. cyclitrophicus in a density dependent manner. Comparative metabolomics revealed distinct metabolic profiles between the pairwise and tripartite interactions. This allowed to identify metabolites that were up-regulated in the tripartite community and therefore candidates for the observed protection. Among these, kynurenic acid and N-acetyltyramine were identified in bioassays as protective molecules, thus clearly highlighting the importance of secondary metabolites in this interaction. The present findings demonstrate that a third bacterial partner can alter the outcome of an antagonistic algal-bacterial interaction by means of chemical mediators. This work has implications for our understanding of microbial community functioning that cannot only be derived from the investigation of pairwise interactions.

Rock-inhabiting fungi thrive in subaerial oligotrophic environments such as desert rocks, solar panels and marble monuments where organic carbon and nitrogen are scarce. We tested whether the rock-inhabiting fungus Knufia petricola showed a preference regarding nitrogen ([Formula] or [Formula]) and carbon (glucose or sucrose) sources and whether it was sensitive towards carbon and nitrogen limitation. As this fungus produces the carbon-rich, nitrogen-free 1,8-dihydroxynaphthalene (DHN) melanin, we tested whether a melanin-deficient mutant would be less sensitive to carbon limitation. The carbon and nitrogen concentrations were the primary predictors of growth, with a broad optimum partially explained by an optimal fungal C:N ratio. Limiting carbon or nitrogen supply decreased biomass formation, CO2 production and biofilm thickness but promoted substratum penetration through filamentous growth. The nitrogen content of the biomass was flexible within limits, increasing upon increasing nitrogen supply or decreasing carbon supply. The carbon use efficiency was fairly constant, whereas melanization correlated with a higher nitrogen content of the biomass despite melanin being nitrogen-free. In conclusion, in vitro, K. petricola switches to explorative growth under nutrient limitations, like fast-growing fungi, revealing universal fungal resource-acquisition patterns. Graphical abstract text and imageCarbon and nitrogen availability affect biofilm growth and morphology of the extremotolerant fungus Knufia petricola Abolfazl Dehkohneh, Julia Schumacher, Bastiaan J. R. Cockx, Karin Keil, Tessa Camenzind, Jan-Ulrich Kreft, Anna A. Gorbushina, Ruben Gerrits Growth of the rock-inhabiting fungus Knufia petricola was studied by varying carbon and nitrogen sources and concentrations. Overall, growth was best predicted by the carbon and nitrogen concentrations. Carbon and nitrogen limitation promoted substratum penetration through filamentous growth. O_FIG O_LINKSMALLFIG WIDTH=158 HEIGHT=200 SRC="FIGDIR/small/712823v1_ufig1.gif" ALT="Figure 1"> View larger version (44K): org.highwire.dtl.DTLVardef@6d98bdorg.highwire.dtl.DTLVardef@146aac5org.highwire.dtl.DTLVardef@757fa8org.highwire.dtl.DTLVardef@ff709_HPS_FORMAT_FIGEXP M_FIG C_FIG