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My Research

Opening up the black box of marine
phototroph-heterotroph interactions


open the black box


Numerous different interactions between marine organisms have been described - for example cnidarians and foraminifera with endosymbiotic unicellular algae (Lee, 2006; Rowan, 1998), sponges with bacteria (Althoff et al., 1998), bryozoans with proteobacteria (Lim & Haygood, 2004), cleaner shrimps operating cleaning stations (Bunkley-Williams & Williams, 1998), anemones and anemonefish (Fautin, 1991). Many of these relationships are mutualistic and involve, if any, only one microbe.

Marine microorganisms are responsible for about 50% of global carbon fixation (Field, 1998), play a crucial role in nutrient cycling (Arrigo, 2005) and are abundant in surface ocean waters with about 106 microbial cells per ml (Cole, 1982; Giovannoni & Stingl, 2005). However, relatively little is known about the relationships between these microbes.

The interactions between marine microorganisms that have been studied most extensively are between microalgae and bacteria (Amin et al., 2012; Cole, 1982). Bacteria provide nutrient regeneration and vitamins to algae, facilitate iron uptake, as well as modify the environmental conditions, such as oxygen concentration, in close proximity to algal cells (Amin et al., 2009; Cole, 1982). This may enhance the growth of both organisms, as microalgae can provide a source of carbon for heterotrophs. In the case of vitamin B12 supply, these interactions may be different even between closely related species (Croft et al., 2005), as symbiosis arises through loss of the metE gene (encoding for vitamin B12-independent methionine synthase) in response to an available environmental pool of vitamins (Helliwell et al., 2011). Although it is usually considered to be a mutualistic relationship in laboratory cultures, there are arguments that it is not a direct symbiosis in the natural environment, but indirect scavenging for available molecules present in seawater (Droop, 2007).

The Black Queen Hypothesis

This loss of genes otherwise essential for survival is well-described by The Black Queen Hypothesis, which states that gene loss may provide an advantage to the organism, as long as the function is leaky in other organisms providing an available public pool of the product (Morris et al., 2012). Unlike Hamilton’s rule, this is not based on altruistic action, but just a simple, selectively favoured, reduction in living costs. Examples supporting the hypothesis include: hydrogen peroxide removal (Morris et al., 2011), iron and reduced sulphur acquisition (D'Onofrio et al., 2010; Tripp et al., 2008) and algae-bacteria biofilm formation (Lubarsky et al., 2010; Roeselers et al., 2007). Helper strains are always present in the community, although in significantly smaller numbers than the organisms requiring help and thus may be considered to be keystone species (Morris et al., 2012). If removed, the organism with reduced genome may not be able to grow due to toxicity or lack of nutrients. However, apart from explaining the hypothesis and examples supporting it, no data was presented to explain how these relationships form and evolve, how specific they are and whether this is the reason why axenic cultures are difficult to grow.

Prochlorococcus - heterotrophic bacteria interactions

Focusing on phototroph-heterotroph bacterial interactions, the discovery of the role of helpers in removing hydrogen peroxide was made as part of a research project aimed at improving axenic cultivation of Prochlorococcus (Morris et al., 2008). Prochlorococcus are highly abundant, globally distributed phototrophic bacteria (Partensky et al., 1999), that make up 51% of biomass and are responsible for about 45% of carbon dioxide fixation in the northeast Atlantic (Jardillier et al., 2010). As it is difficult to grow Prochlorococcus in axenic culture, especially on semi-solid media, a method of growing the phototrophic bacteria together with heterotrophic bacteria and subsequently removing these “helper” bacteria to obtain a pure culture was proposed (Morris et al., 2008). The authors proposed that these “helper” bacteria removed oxidative stress, though they did not investigate the precise mechanism nor whether it happens in the natural environment. Subsequently, these authors confirmed the relationship in the open ocean – if helper species are absent, the hydrogen peroxide concentration increases to levels lethal for Prochlorococcus causing cell envelope damage and loss of photosynthetic activity) (Morris et al., 2011). As the authors suggest, the helper strain may benefit from the organic matter leaking from the phototroph, but more research should be done to establish whether this relationship is commensalism or mutualism.

A study on interactions between Prochlorococcus and 344 different heterotrophic bacterial isolates revealed that the response of cells to coculture is different between Prochlorococcus ecotypes adapted to low or high light conditions (Sher et al., 2011). Positive interactions, resulting in faster growth or higher final chlorophyll fluorescence, were observed only in the low-light Prochlorococcus ecotype and were more common than inhibitory interactions. They were possibly caused by small, diffusive molecules, not cell to cell contact, in contrast to interactions that caused a delay in growth, which were observed only when cell to cell contact was possible. In the case of Alcanivorax sp. HOT7G9 and Rhodobacter sp. HOT5F3 strains, the peak chlorophyll fluorescence was higher for co-cultures separated by a membrane than those grown together (Sher et al., 2011).

Synechococcus - Roseobacter interactions

Synechococcus is a phototrophic picocyanobacteria, highly abundant in almost all marine ecosystems, with a relatively small genome (Scanlan et al., 2009). In the northeast Atlantic it forms about 20% of phytoplankton biomass and contributes about 21% to primary production (Jardillier et al., 2010).

In a study using atomic force microscopy (Malfatti & Azam, 2009), 6-42% of Synechococcus cells were found to be conjoint with heterotrophic bacteria. Some of these were further connected to other Synechococcus-heterotrophic bacterial cells through fine pili or cell- surface gel matrices, forming networks of up to 20 connections. Although the percentage of cells conjoined with heterotrophs in coastal and offshore samples was the same, more networks were observed in coastal than offshore samples (55% and 4% respectively).

Roseobacter is a diverse clade, estimated to form about 20% of all bacteria in coastal waters and about 15% in mixed-layer open ocean systems (Buchan et al., 2005), which has metabolically versatile cells able to compete well with other microorganisms (Moran et al., 2007). Ruegeria pomeroyi, basonym Silicibacter pomeroyi (Yi et al., 2007), was the first major marine heterotrophic bacterial clade to have its genome sequenced (Moran et al., 2004).

Roseobacter genes suggest frequent interactions with neighbouring cells and possibly direct capture of organic matter from eukaryotes - for example vir-related genes encoding a type IV secretion system for translocating DNA or proteins to other cells and close homologs of non-ribosomal peptide synthases genes, which may encode a novel peptide responsible for signaling or host-microbe interactions (Moran et al., 2007). Many interactions between Roseobacter strains and other organisms have been described - including with red and green macroalgae, diatoms, bryozoans, dinoflagellates, cephalopods, oysters (Wagner-Dobler & Biebl, 2006). Confocal laser scanning microscopy showed Roseobacter cells living intracellularly or as epiphytes with Pfiesteria-like heterotrophic dinoflagellate associated with harmful algal blooms (Alavi et al., 2001).


To summarise, despite the major role that marine bacteria play in the global ecosystem, little is known about their interactions with other organisms. The Black Queen hypothesis can explain why these interactions form, but except for the Prochlorococcus-heterotrophic bacteria relationship based on hydrogen peroxide removal, it is unclear how they form and are maintained in the natural environment, including the molecular basis of how different members of the microbial community rely on one another. The interactions between the second most abundant phototrophic picocyanobacteria - Synechococcus - and a dominant heterotroph - Roseobacter - are possible and can be studied as a model relationship to improve our understanding of the physiology of marine microbes, interactions between them, as well as the implications these relationships may have for global biogeochemical cycles.

Project objectives

Focusing on the model Synechococcus-Roseobacter interaction, this PhD project aims to identify and characterise the molecules produced and consumed by the phototroph and heterotroph growing in co-culture and describe the pathways involved in these processes. The project is divided into three major objectives:

  1. To identify the most-frequently occurring heterotrophic bacterial partners present in non-axenic Synechococcus sp. cultures. This work will be used to assess the prevalence of Synechococcus-Roseobacter interactions and to identify which other marine bacterial genera are “selected” during Synechococcus isolation.
  2. To study the evolution of phototroph-heterotrophic bacteria co-cultures. This will focus on following clonal purification of new Synechococcus sp. isolates and the heterotrophic bacteria that co-occur with them, assessing if specific genera are selected.
  3. To characterise the specific metabolite(s) present or absent in the milieu during Synechococcus-Roseobacter co-culture compared to growth of the axenic Synechococcus sp. culture. The generality of Synechococcus-Roseobacter co-culture metabolite production will be assessed in several Synechococcus sp. strains (e.g. Synechococcus spp. WH8102, CC9311, WH7805, WH5701). In every case, metabolite production will be assessed in co-culture compared to axenic controls and, where appropriate, specific metabolites will be purified and identified and in the case of novel molecules - characterised. Purified metabolites will be assessed for their toxicity to Synechococcus, if they are consumed by Roseobacter and how and when they are made.

Synechococcus spp

Synechococcus spp. WH8102, CCMP2515 (CC9311), WH7805, WH5701


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Amin, S.A., Parker, M.S., Armbrust, E.V. (2012) Interactions between diatoms and bacteria, Microbiology and Molecular Biology Reviews, 76 (3), 667-684.

Amin, S.A., Green, D.H., Hart, M.C., Kupper, F.C., Sunda, W.G., Carrano, C.J. (2009) Photolysis of iron-siderophore chelates promotes bacterial-algal mutualism, Proceedings of the National Academy of Sciences of the United States of America, 106 (40), 17071-17076.

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D'Onofrio, A., Crawford, J.M., Stewart, E.J., Witt, K., Gavrish, E., Epstein, S., Clardy, J., Lewis, K. (2010) Siderophores from neighboring organisms promote the growth of uncultured bacteria, Chemistry & Biology, 17 (3), 254-264.

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Jardillier, L., Zubkov, M.V., Pearman, J., Scanlan, D.J. (2010) Significant CO2 fixation by small prymnesiophytes in the subtropical and tropical northeast Atlantic Ocean, The International Society for Microbial Ecology Journal, 4 (9), 1180-1192.

Lee, J.J. (2006) Algal symbiosis in larger foraminifera, Symbiosis, 42, 63-75.

Lim, G.E., Haygood, M.G. (2004) "Candidatus Endobugula glebosa", a specific bacterial symbiont of the marine bryozoan Bugula simplex, Applied and Environmental Microbiology, 70 (8), 4921-4929.

Lubarsky, H.V., Hubas, C., Chocholek, M., Larson, F., Manz, W., Paterson, D.M., Gerbersdorf, S.U. (2010) The stabilisation potential of individual and mixed assemblages of natural bacteria and microalgae, PLoS one, 5 (11), e13794.

Malfatti, F., Azam, F. (2009) Atomic force microscopy reveals microscale networks and possible symbioses among pelagic marine bacteria, Aquatic Microbial Ecology, 58, 1-14.

Moran, M.A., Belas, R., Schell, M.A., Gonzalez, J.M., Sun, F., Sun, S., Binder, B.J., Edmonds, J., Ye, W., Orcutt, B., Howard, E.C., Meile, C., Palefsky, W., Goesmann, A., Ren, Q., Paulsen, I., Ulrich, L.E., Thompson, L.S., Saunders, E., Buchan, A. (2007) Ecological genomics of marine Roseobacters, Applied and Environmental Microbiology, 73 (14), 4559-4569.

Moran, M.A., Buchan, A., Gonzalez, J.M., Heidelberg, J.F., Whitman, W.B., Kiene, R.P., Henriksen, J.R., King, G.M., Belas, R., Fuqua, C., Brinkac, L., Lewis, M., Johri, S., Weaver, B., Pai, G., Eisen, J.A., Rahe, E., Sheldon, W.M., Ye, W., Miller, T.R., Carlton, J., Rasko, D.A., Paulsen, I.T., Ren, Q., Daugherty, S.C., Deboy, R.T., Dodson, R.J., Durkin, A.S., Madupu, R., Nelson, W.C., Sullivan, S.A., Rosovitz, M.J., Haft, D.H., Selengut, J., Ward, N. (2004) Genome sequence of Silicibacter pomeroyi reveals adaptations to the marine environment, Nature, 432 (7019), 910-913.

Morris, J.J., Lenski, R.E., Zinser, E.R. (2012) The Black Queen Hypothesis: evolution of dependencies through adaptive gene loss, mBio, 3 (2), 1-7.

Morris, J.J., Kirkegaard, R., Szul, M.J., Johnson, Z.I., Zinser, E.R. (2008) Facilitation of robust growth of Prochlorococcus colonies and dilute liquid cultures by "helper" heterotrophic bacteria, Applied and Environmental Microbiology, 74 (14), 4530-4534.

Morris, J.J., Johnson, Z.I., Szul, M.J., Keller, M., Zinser, E.R. (2011) Dependence of the cyanobacterium Prochlorococcus on hydrogen peroxide scavenging microbes for growth at the ocean's surface, PLoS one, 6 (2), e16805.

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Wagner-Dobler, I., Biebl, H. (2006) Environmental biology of the marine Roseobacter lineage, Annual Review of Microbiology, 60, 255-280.

Yi, H., Lim, Y.W., Chun, J. (2007) Taxonomic evaluation of the genera Ruegeria and Silicibacter: a proposal to transfer the genus Silicibacter Petursdottir and Kristjansson 1999 to the genus Ruegeria Uchino et al. 1999, International Journal of Systematic and Evolutionary Microbiology, 57 (4), 815-819.

Main Supervisor:

Prof. David Scanlan

Marine Microbiology 
School of Life Science
University of Warwick


Dr Christophe Corre

Chemical Biology - Natural Products
Department of Chemistry
and School of Life Sciences
University of Warwick

Dr Joseph Christie-Oleza

Marine Molecular Microbiology
School of Life Sciences
University of Warwick