A new vector of a changing marine environment: the seagrass holobiont

As marine environments are changing under the pressure of ocean acidification (OA), it is important for better understanding of this process to study the effect of decreased pH on crucial ecosystem components like seagrasses. Seagrass meadows act as nurseries for marine fauna, as sediment stabilizers and as primary producers for example. The plants colonize large areas of our tropical marine reefs and its leaves are covered with a biofilm community of epiphytes that further extend the ecological role of the plants. As seagrasses are photosynthetic oganisms, it was generally thought that an increasing CO₂ availability would positively affect the plants. However, we know that for corals – in addition to a weakened skeleton - OA causes a change in the coral mucus, which in turn results in coral diseases for example. It is thus interesting to look at the holobiont of seagrasses to understand the OA impact. Especially since data on this topic is scarce and the literature available describes research that did not address the total epiphyte community accurately. Hassenrück and collegues (2015) investigated how the epiphytic biofilm of Enhalus aroides was affected by increased CO₂ pressure, using molecular tools to avoid the limitations of earlier research.

The effect of OA was simulated with a comparison between seagrass biofilm communities at a natural CO₂ vent in Papua New Guinea (pH 7.8) and a control site (pH 8.3). At both sites, the taxonomic composition of the epiphytes at both sites was determined by 16S and 18S rRNA gene sequencing, leaf age was taken into account and data on carbon and nitrogen content of the leaves was acquired. The latter is additional information needed to distinguish life stage development from the effect of OA on the epiphytes. As E. acroides produces new leaves on a monthly basis and the seagrass shoots used represented the first four life ages, which comes down to a plant settlement period of 4-5 month that is covered in this study. However, growth rates might have increased due to low pH conditions, in which case the shoots represented older life stages as well.

Both carbon and nitrogen content decreased with leaf age, from 33% to 26% dry weight and 2% to 1% respectively. Epiphyte cover increased however, albeit with values three times lower at the vent site than at the control site. It is suggested that this difference is caused by the supression of pH-sensitive organisms at the vent site. Although the epiphyte cover increased less at the vent site, the biofilm communities of E. acroides showed the same richness at both sites, which could imply an increase in scarce epiphytes at the vent site and thus less increase in epiphyte abundance. The results however, show no significant difference in the amount of rare bacterial species per site. The authors don’t say anything about this with regards to eukaryote composition at both sites. Furthermore, three other patterns were found in the epiphyte composition, which differed a lot between the two sites. Only around 30% of the epiphytic OTUs were shared between any two of the samples. This variation was dominantly explained by sampling site, but also by leaf age, albeit four times less. Besides, the composition shifted succesionally with leaf age, which could be explained by the changing organic matter transfer from the leaves to the biofilm. Lastly, also a trend in pairwise similarity was shown between bacteria and eukaryotes. The authors suggested this as a sign of shaping and impacting responses of both communities on one and another, since the correlation found was stronger than could possibly be explained by solely abiotic factors.

The last part of the research done focuses on the taxonomic details of the community structure of the epiphytic biofilms. Most findings are supported by earlier studies and the extensive comparison with other literature characterizes this part of the research paper. This enhances the strength of the article in particular, since the topic of OA impact on seagrass epiphytes is so understudied and thus needs more context. The article covered several research questions and therefore provides the field of environmental microbiology with a lot of new insights. I was especially suprised by the way the seagrass holobiont might be a source of information to indicate the occurance of coral diseases due to OA, but there are many more clues for further research that can be derived from this study.

Article reviewed:
Hassenrück, C., Hofmann, L. C., Bischof, K., & Ramette, A. (2015). Seagrass biofilm communities at a naturally CO2‐rich vent. Environmental microbiology reports, 7(3), 516-525.

Selective enrichment of saccharides and ions in sea spray aerosols

Sea spray aerosols (SSAs) are formed by bubble bursting at the ocean surface. These aerosols can reduce solar radiation and act as cloud condensation and ice nuclei. Rising bubbles scavenge surface-active organic matter from the surface micro layer (SML), thus leading to a different composition of SSAs compared to the SML or subsurface water. As microbes and viruses are the main regulators of organic carbon in the marine environment, it is thought that their activity might also change the composition of SSAs. In their paper, Jayarathne et al. (2016) investigated the enrichment of budding SSAs with organic carbon (OC), especially saccharides, and inorganic ions. Moreover, the paper also looked at the selective transfer from water to air and the influence of the microbial loop. 

The study was conducted with coastal ocean water in a wave flume at the University of California, two phytoplankton blooms occurred during the experiment. SSAs were generated by wave-breaking and samples were collected from the covered wave flume with quartz fibre filters. Furthermore, SSAs were divided into smaller and larger SSAs. Enrichment Factors were used to evaluate enrichment relative to Na⁺. Thermal optical analysers, ion-exchange-chromatography and high-performance anion-exchange-chromatography were used to detect OC, ions and saccharides respectively. 

The values and temporal variations of biological activity and dissolved organic carbon (DOC) were similar to data obtained in field studies. Both smaller and larger SSA were mostly composed of sodium (18 and 22%) and chloride (33 and 41%). While all particles showed the same components, larger SSA showed more inorganic ions and smaller SSAs were more enriched with OC (8 compared to 1.2%). 

SSAs were significantly enriched with OC, indicating a selective enrichment process. Overall, the enrichment factors did not change with biological activity. However, the authors suggest that differences in the influence of biological activity between different sized SSAs were undetectable in this experiment.

Saccharides formed 11 and 27% of OC on smaller and larger SSAs respectively, with glucose being the most abundant saccharide throughout the experiment. Enrichment factors for SSAs were higher for smaller aerosols but both factors were still significantly higher than for the SML. Smaller SSAs mainly contained saccharides used for energy storage and associated with DOM. However, larger SSAs also harboured structural saccharides which are associated with POM. This difference may arise during the formation of the SSAs. While smaller SSAs are formed by film drops, larger SSAs are formed by jet drops from older, less-enriched parts of the SML. Moreover, the abundance of saccharides also changed rapidly during the experiment, presumably due to changes in biological activity. 

SSAs were also significantly enriched with cations compared to the SML. Divalent cations such as, Ca²⁺and Mg²⁺ were more abundant than single charged cations. The observed selective enrichment of cations corresponded to the binding affinities of the cations to fatty acids and anionic surfactants. Thus, cations may be selectively transported to the SSAs when organic molecules are present. Furthermore, while the mechanisms of enrichment are not known, charged organic compounds may also transport anions to SSAs. Nitrate and sulfate were enriched in SSAs and also showed temporal variation, presumably due to changing biological activity. In contrast, chloride was not enriched on SSAs.

In conclusion, the study provides evidence that enrichment in SSAs is a dynamic process and is affected by biological activity. Moreover, the authors suggest that the established method of estimating salt concentrations using Na⁺ concentrations is biased because of the selective enrichment of divalent cations. The paper doesn’t name any potential ecological impacts, but in my opinion selective enrichment of SSAs could at the very least provide airborne microbes with important nutrients. 

Reference:

Jayarathne, T., Sultana, C. M., Lee, C., Malfatti, F., Cox, J. L., Pendergraft, M. A., ... & Bertram, T. H. (2016). Enrichment of Saccharides and Divalent Cations in Sea Spray Aerosol During Two Phytoplankton Blooms. Environmental Science & Technology. http://pubs.acs.org/doi/abs/10.1021/acs.est.6b02988

The Microbiome of Corals

Corals are reef building animals that are inhabited by bacteria, fungi and archeae. The microbes can live within the coral in their tissue or the skeleton or on their surfaces within mucus. This association allows that corals colonized different habitats. Corals are exposed to changes e.g. in temperature and have to deal with diseases like the white band or the white plague disease. They are caused by microbes but it is hard to identify the causal agents. This study by Chu & Vollmer (2016) aims to indentify bacterial species that are associated with healthy corals and how specific these relationships are. They studied the changes in coral microbiomes over time and space by analysing 16S rRNA from coral-associated bacterial communities. Chu & Vollmer used 100 coral colonies near Bocas del Toro, Panama. The samples were taken from Acropora cervicornis, A. palmata, Diploira labyrinthiformis, D. Strigosa, Porites astreoides and P. furcata in December, April and October 2012-2013.

The results show that the bacterial community composition within the corals depends mostly on coral host species while site and time are less important. Furthermore, the phylogeny of the corals is reflected by the similarity of the composition of their associated bacteria.

Chu & Vollmer found that Acropora spp. incorporate high abundances of Campylobacteraceae which are associated with the white band and white plague disease in Monastrea spp. That leads to the conclusion that healthy corals of one species can host potentially pathogenic microbes.

The core microbiomes seem to be shared between the species. Campylobacteraceae and Endozoicimonaceae were found as the core microbiomes in Acropora. Porites were dominated by Endozoicimonaceae and Gammaproteobacteria while Dilpoira showed high abundances of Alteromonadales. The dominant phylum in the OTU’s are Proteobacteria with high abundances of the order Flavobacteriales, Alteromonadales, Oceanospirillaler and Burkholderiales. Thus, the core microbiome ‘includes shared and host-specific bacterial phylotypes’ (Chu & Vollmer, 2016).

All in all this study shows the complexity of corals and their symbionts and provides information about potential coral pathogens that are incorporated in healthy species which do not seem to be effected by the pathogens. Furthermore the colonizing bacteria seem to have different niches and nutrient demands and are therefore found in different coral species and are host specific.

In my opinion this study is a base for further research on identifying the potential pathogens for a better understanding of coral diseases and their mechanisms. The methodology part is missing and at some points it is not clear what the authors aim to state. Additionally, it might be easier to split up the results and discussion part and not to put all together like Chu & Vollmer did. It would be interesting to see how the bacteria found in the corals effect their hosts and to know more about their ecological function. Last but not least I would like to see how global warming effects the pathogens in the healthy coral hosts and if they can become pathogenic for their hosts, too.

Reviewed paper:

Chu, N. D., & Vollmer, S. V. (2016). Caribbean corals house shared and host-specific microbial symbionts over time and space. Environmental microbiology reports.

Merry Microlayer

The sea surface microlayer (SML) forms an important boundary between the sub surface layer (SSL) and the atmosphere, and may have an important function in the cycling of different nutrients. Atmospheric deposition is one of the main external sources of nutrients, and the SML is the first point of contact for these aerosols. The south east Mediterranean Sea (SEMS) is a low nutrient area dominated by small sized oligotrophs. Previous studies have shown the importance of atmospheric deposition in supplying the SEMS with nutrients, but none have looked at whether this input of nutrients affects the SML and SSL differently. Astrahan et al have attempted to remedy this using a mesocosm set up to look at biomass and activity of various groups in the neuston.

Water samples of the SML and SSL were collected, as were aerosols from a dust storm. Samples were incubated in outdoor pools either with aerosol addition or without. Samples from each replicate were taken at various intervals over the next 44 hours, and primary production and bacterial production were measured. Before aerosol addition, heterotrophic bacterial abundance was similar in both the SML and SSL, but bacterial production was roughly 50% higher in the SML. Following the addition of aerosol to the samples, heterotrophic bacterial abundance increased 2-fold in both the SML and SSL water, but the increase in bacterial production was much higher in SML than the SSL. The authors suggest that the added nutrients may have been used to fulfil metabolic needs rather than for growth and cell division, which explains the larger increase in production rather than abundance.

A mesocosm setup can only go so far in showing us how different processes occur in the ocean, especially when only considering a very small 44 hour window post aerosol deposition. The authors did not look at the diversity of bacterial groups, but I think this would have been really interesting to see if the bacterial community changed after aerosol addition and how quickly this change occurred. I also think it would have been better to increase the length of the study, as the first 44 hours shows us the initial changes, but gives us no indication of how long this enrichment lasts or when the community abundance and production returns to normal.

Astrahan P., Herut B., Paytan A., Rahav E.(2016) The Impact of Dry Atmospheric Deposition on the Sea-Surface Microlayer in the SE Mediterranean Sea: An Experimental Approach. Frontiers in Marine Science. 3:222

Jingle bell…shaped ciliates

Peritrichs are a subclass of ciliates that are usually bell or disc shaped and have a prominent paroral membrane starting in the oral cavity and circling around the anterior of the cell in a counter clockwise direction. Pseudovorticella is a genus within this subclass that was established to differentiate the organisms within this genus from Vorticella, who have differently patterned silverline system and have different oral infraciliature. This genus is rarely investigated for a few reasons, firstly Pseudovorticella has relatively few species compared to Vorticella and receives little attention despite its wide distribution. Secondly the two genus are very similar in vevo and the known species of Pseudovorticella are insufficiently described. Thirdly the two genus are similar in living morphology, but the taxonomic ranking of many Vorticella can be doubted due to the lack of silver staining data on them.

Sun et al. (2013) sampled coastal areas of the Yellow Sea in Shandong, China, by placing an artificial substrate and leaving it for 10 days to allow for colonisation. Once collected the ciliates were isolated and identified. Five species from the genus Pseudovorticella were isolated, three of which were already known species (Pseudovorticella plicata, P. banatica and P. anomala), and the two other species were not thought to have been seen before, named Pseudovorticella dingi and P. wangi. They then go on to describe the two newly discovered species in great detail and giving comparisons of other similar species. With the three already known species thy simply add more detail to the already known information.

This paper is a simple identification paper, they do not talk about or hazard a guess as to any possible niches the new or old species may fulfil. This paper simply contributes to the known biodiversity of the ciliates and adds more detail to help the taxonomic ranking of the current known species.

Merry Christmas

Referenced paper:

Sun P., Ma H., Shin M.K. and Al-Rasheid K.A., (2013). Morphology of two new marine peritrich ciliates from Yellow Sea, Pseudovorticella dingi nov. spec. and P. wangi nov. spec., with supplementary descriptions of P. plicata, P. banatica and P. anomala (Ciliophora, Peritrichia). European journal of protistology. 49(3). pp.467-476.

Better Out than In – or why superoxide in coral may be misunderstood

Reactive oxygen species (ROS) such as superoxide are created through the reduction of molecular oxygen to water and play a critical role in coral bleaching. The internal accumulation of Superoxide have previously been linked to physiological stress. The build-up of these toxic chemicals within the coral are thought to damage the photosynthetic capability of Symbiodinium cells and impair mitochondrial electron transport in the coral host. This eventually leads to the expulsion of the algae companions, resulting in the loss of coral colour, energy production and organic carbon.

Previously, internal superoxide has been considered a detrimental molecule, however newly discovered pathways and sources of superoxide have revealed that a diverse group of heterotrophic bacteria enzymatically produce extracellular superoxide at the corals surface. Unlike the traditional detrimental effects of having intercellular superoxide, external production may aid the corals resilience to disease and increase thermotolerance.

Despite the bad reputation of superoxide, very little is known about the origins, distributions and ecological underpinning of superoxide production in natural coral communities. Previous studies utilized indirect evidence of oxidative stress based on observations, gene expression and proteomics resulting in often inaccurate and biased results. However Diaz et al. utilized recent developments in non-invasive chemiluminescent techniques allowing the team to capture the first in-situ measurements of external superoxide production by several species of thermally stressed and bleaching corals.

Observations during a bleaching event saw corals that were susceptible to bleaching lacked the production of external superoxide, while resistant corals had high concentrations of external superoxide. Additional observations were also carried out on corals that weren’t undergoing physiological stress, these corals still carried out the production of superoxide. These findings were then reinforced by a parallel study which examined corals in a lab environment. The combination of these results has resulted in scientists realizing that superoxide may be vitally misunderstood and instead of a being a detrimental molecule, it may be an essential molecule for the well-being of coral. Future lab experiments are essential for investigating the relationship between superoxide and physiological stressors without the natural variability of the coral reef. Focus should also be directed to the handful of other stressors corals experience such as pathogens.

The measurement of superoxide isn’t without its hardships, due to the volatile 30 second lifespan of superoxide, even these successful measurements shown will be seeded with unavoidable bias. However the research team have carving their way through previous bias methodologies and unreliable data using novel techniques to reveal important discoveries that go against the grain of studies on the area. This initial discovery may lead to future research that is essential to achieve a better understanding of the physiology of corals, leading us towards future developments in coral management and help develop bleaching mitigation strategies.

Diaz, J., Hansel, C., Apprill, A., Brighi, C., Zhang, T., Weber, L., McNally, S. and Xun, L. (2016). Species-specific control of external superoxide levels by the coral holobiont during a natural bleaching event. Nature Communications, 7, p.13801.

Breaking the Water Mould: An oomycete pathogen of an Atlantic seagrass

Oomycetes have well-described ecological roles as the saprophytes and pathogens of terrestrial plants. For a long time, the oomycetes were recognised as a fungal taxon but subsequent genetic analyses have demonstrated them to be heterotrophic stramenopiles – the same group as diatoms and kelp. However, much like their fungal look-a-likes, marine oomycetes are very poorly understood relative to their terrestrial cousins. Despite the devastating effects oomycetes can have on terrestrial plants, virtually no work has been carried out to understand their role in the pathogenesis of marine angiosperms. Here, Govers and colleagues (2016) present the first report of a widespread oomycete infection in the seagrass Zostera marina.

Seeds of this species were collected from countries on both Atlantic coasts and visually inspected for oomycete sporangia. Infected seeds were then cultured on an oomycete-selective agar known as ‘ParpH’ and colonies were sequenced for ITS rRNA gene signatures. Strikingly >99% of all the collected seeds were infected by the oomycetes Halophythophthora sp. Zostera and Phytophthora gemini. To understand this phenomenon more comprehensively, Z. marina seeds were subjected to a germination experiment under laboratory conditions in the presence (or absence) of oomycete pathogens. Perhaps most surprisingly, following 110 days of incubation in sediment, two thirds of the infected seeds were no longer infected; they had shrugged off the oomycete pathogen. Those that remained infected, however, exhibited a sixfold lower germination rate than the non-infected control seeds, indicating a deleterious effect of oomycetes on seagrass reproduction.

The authors also introduced physiochemical variation into their experimental design. This was achieved by growing infected seeds in differential sediment treatments (mud or sand) and at low or high temperatures (5 vs 12^oC). These conditions were selected to be ecologically realistic of cool or warm winters in the Dutch Wadden Sea. The authors found that infection rates were higher in sandy sediment and at cooler temperatures. The authors postulate that both organic-rich, muddy sediments and warmer temperatures favour bacterial sulphide production and anoxia (corroborated by physiochemical profiling of sediments from field study sites), which could hinder oomycete growth and reproduction.

These findings centre around a robust hybrid methodology, combining in situ surveys with controlled laboratory experimentation. The molecular work, however leaves much to be desired as the samples sequenced were culture-dependant, having been isolated from colonies grown on agar. As with a multitude of other marine microbial taxa, the oomycetes may suffer from poor culturability and these samples may not be representative of the potentially pathogenic oomycete community. I would have been keen to see metabarcoding conducted on environmental samples from the sample field sites, to gain a holistic oomycete community profile. It is too important to note that Koch’s postulates have not yet been fulfilled, as with many cases of marine disease, and therefore further work is needed.

The identification of marine oomycete pathogens, particularly in a species of conservation interest, is an eye-opening discovery. Previously, the study of seagrass disease was focussed on the Labyrinthulomycetes (also stramenopiles), which cause a devastating wasting disease. This study provides evidence that other taxa may be key players, and a proper understanding of their biology could reap serious conservation benefits. The physiochemical results already suggest that sediment and climate optimisation could be important in mitigating disease in seagrass conservation efforts.

Reviewed Paper: Govers, L. L., Man, W. A., Meffert, J. P., Bouma, T. J., van Rijswick, P. C., Heusinkveld, J. H., ... & van der Heide, T. (2016, August). Marine Phytophthora species can hamper conservation and restoration of vegetated coastal ecosystems. In Proc. R. Soc. B (Vol. 283, No. 1837, p. 20160812). The Royal Society. http://rspb.royalsocietypublishing.org/content/283/1837/20160812

CCA 0, CFD 1

Corals are important bio-engineers, depositing calcium carbonates and creating vast expenses of reefs which are visible from space. Many of these reefs are colonised by crustose coralline algae (CCA), which can cover up to 50% of living reefs, and consequently act as important sites for coral larval settlement. Although CCA are an important part of reef ecology, very little is known about them or their susceptibility to disease. Disease could potentially cause drastic reductions in CCA populations, with known on effects which promote community shifts to fleshy macro-algal dominance (DDAMnation – Rohwer, 2010). The authors of this paper aim to look at the effects of ocean warming and OA on CCA, and the interaction with coralline fungal disease (CFD), a previously described CCA disease from the Pacific. A combination of field observations and experimental manipulations were used.

Histopathological changes were examined using fragments of CCA which displayed signs of CFD, with Grocott’s methanamine silver being used to confirm the presence of fungal hyphae. To culture the fungus associated with CFD, cellulose agar was used with media containing antibiotics to stop bacterial growth, and a control without antibiotics to preclude any negative effects on fungal growth. Cultures were incubated at 25, 27 or 30 °C, aerated, and left for 21 days. Frozen, ground CFD lesions were thawed, and individual fungal filaments were isolated. DNA was extracted from filaments, and fungal specific 18s rRNA PCR amplification was used from mixed-environmental samples. Phylogenetic analysis was assessed using the National Centre for Biotechnology Information (NCBI), which was used to align the 653bp 18S rRNA gene sequences. A phylogenetic tree was constructed and evolutionary distances were computed.

The quantification of CFD occurrences was done using photo-quadrats, from 12 permanent sites between July- August and October-November, 2008. 40 of the 59 transects surveyed became permanent as part of the Palmyra disease monitoring programme and were surveyed again in 2009 for CFD occurrences and CCA cover. CFA was also measured during El Nino, with 13 CFD cases being photographed weekly, with CFD vital rates (lesion surface area and linear progression rate) were calculated using Image J software.

Temperature and acidification experiments were conducted using C0₂ bubbling and heating, to examine independent and interactive effects of OA and waring on CFD disease dynamics as well as CA growth (net calcification).  OA conditions were mimicked by bubbling pre-mixed air enriched with C0₂ (reflecting atmospheric conditions projected for 2100). Four independent water baths (two ambient and two warmed) held experimental aquaria that were randomly assigned to elevated or present-day C0₂ conditions, in order to create every combination of warming and OA treatment. Aquaria were covered to prevent evaporation and rainwater from affecting salinity, and placed under a shade cloth to mimic the natural light found at 10m on the forereef (where the CCA is found). Controls were also conducted for algal metabolism. Net calcification rates of CCA were quantified to the nearest mg, and CFD disease rates were calculated using ImageJ (e.g lesion progression)

The paper demonstrated that ocean warming and acidification can have complex interactive effects on marine disease dynamics. Habitat difference seemed to have an important role, with some reefs (Palmyra’s forereef) having a CFD occurrence independent of CCA abundance; suggesting that host abundance alone does not explain observed spatial variation in CFD. The authors mention that the results for the experiments may differ if they were in-field (because CFD variation could be a result of the abundance of preferential host species of CCA) but note that CCA taxonomy is difficult and requires microscopic examination making it impossible in the field. The overall results from the experiments indicate that less acidic but warmer conditions characterise the most favourable conditions for CFD occurrence. Regardless of the mechanisms behind fine-scale CFD variations, it is clear that the disease is more abundant and virulent under elevated temperatures. This suggests that the prevalence and virulence of CFD may increase in line with future climate projections. Unsurprisingly, the experiment also showed that elevated pCO₂ contributes to increasing acidity, which leads to the degradation and structural loss of CCA, suggesting net dissolution. This is likely to increase the vulnerability of CCA, and the virulent effect of CFA is only going to exacerbate this.  However, the authors did discover a slowed growth of lesions in more acidic water, but this minor positive is  swamped by the overriding negative effects that increased acidity has on the whole CCA organism.

All in all, this paper is a fascinating look at the effects of OA and increased temperature on the relationship between parasitic CFD and CCA. Unfortunately, it seems that the parasitic fungi is better suited to the future projected oceanic conditions, and therefore is likely to thrive, resulting In potential major losses in CCA ecosystems, and damages to the coral reefs they inhabit.

Reviewed: Williams GJ et al. 2014 Ocean warming and acidification have complex interactive effects on the dynamics of a marine fungal disease. Proc. R. Soc. B 281: 20133069.

http://dx.doi.org/10.1098/rspb.2013.3069

How low can your poo go?

Microplastics in the marine environment is a globally anthropogenic issue and can threaten ecosystems in a variety of ways. It is well understood that these plastics are consumed by marine life and can build up through the food chain when higher trophic organisms consume prey who have already ingested microplastics. This can have detrimental effects on the health of organisms, such as reduced feeding habits, less energy and increased immune response. Microplastics can also facilitate the transportation of organic pollutants and toxic materials such as polycyclic aromatic hydrocarbons and hydrocarbon petroleum residues into deeper waters.

Zooplankton, such as copepods are ecologically significant with respect to their abundance throughout the world’s oceans and the biological processes they perform. They can also provide important information on ocean pollutants and acidification in a wide spectrum. Copepods faecal pellets are important for transportation of particulate organic matter (POM), carbon and nutrients from the surface to the benthos. Copepods are marine nutrient recyclers and play an important role in consuming, digesting, repackaging and excreting POM. However, when copepods ingest microplastics as prey, this paper asks the important ecological question, what are the impacts microplastics have on copepod faecal pellets and the environment around them?

Previous literature into microplastics in zooplankton has been limited, with papers mainly focusing on the interactions in surface waters. However this paper explores for the first time, by demonstrating sinking rates, that there can be an impact in deeper waters.

This study used marine copepods Calanus  helgolandicus and Centropages typicus, allowing them to consume either natural prey (consisting of phyto-flagellates, diatoms and coccolithophores); cultured prey (the unicellular haptophyte Isochrysis galbana) and microplastics. The representative microplastics used in this paper are 20.6 μm polystyrene and 7.3 μm fluorescent polystyrene beads which is one of the most commonly manufactured polymers in the world. The concentrations used (1000 microplastics mL-1) are higher than in previous literature, however these concentrations were decided based on concentrations consistent with areas of high contamination in open water.

Both species readily ingested the microplastics, which were encapsulated in the gut and all pellets, including those with microplastics sank. The study showed that incorporation of microplastics into the pellets did not impact the size or volume of the pellets; however, plastic treatments showed to have a much lower density and significantly slower sinking rates (38.3 ± 2.6 m day−1) allowing pellets to become more susceptible to being eaten or fragmented. The paper also showed that pellets containing microplastics can be transferred to larger organisms, C. typicus pellets were shown to be transferred to the larger copepod C. helgolandicus via coprophagy, emphasising the issues previously discussed.

Zooplankton pellets are an important source of food for a wide variety of marine organisms globally, this highlights that the problem is not confined to a few species but can have serious wider impacts on communities. I would be interested to understand more about the different types of plastics impacting on faceal pellets, as this study used polystyrene which is used for most packaging eg plastic cutlery, yoghurt pots etc. However, polyethylene (plastic bags, single-use plastic bottles etc) is manufactured in a variety of denisities so different plastics may have differing effects.

Paper reviewed:  

Cole, M., Lindeque, P. K., Fileman, E., Clark, J., Lewis, C., Halsband, C., & Galloway, T. S. (2016). Microplastics alter the properties and sinking rates of zooplankton faecal pellets. Environmental science & technology, 50(6), 3239-3246. http://pubs.acs.org/doi/abs/10.1021/acs.est.5b05905

Environment vs geographic location in influencing the biogeography of fungi on a global scale

Marine fungi are both a diverse and important part of ocean ecosystems, however compared to many other microbes, fungi are unstudied, due to this understudy much less is known about fungi biogeographic patterns and the main factors which influence them both geographical distance and habitat type have been shown to be important in influencing distribution, a recent paper by Tisthammer et al (2016), was the first to look at these factors on a global scale to assess their relative importance in determining community composition the paper also looked at if there was a different in fungi assemblages between sediment and water column samples.

Methods

·         The authors used data from the International Census of Marine Microbes Dataset, including information about environmental parameters (e.g. temperature, salinity, and concentrations of phosphate, nitrate, dissolved oxygen and silicate) for the sample sites. For some sites there was incomplete data for environmental conditions, this was overcome by using information from the world ocean atlas to estimate these environmental parameters. To ensure accurate analysis, when reliable data was not available for sample these samples were excluded.

·         Samples were separated into pelagic or benthic in order to compare the community composition between the habitats.

·         The International Census of Marine Microbes Dataset used primers to amplify the regions of the ribosomal small subunit V9 which the authors used in the study.

·         Operational taxonomic units were determined at 97% similarity.

·         In total 42 pelagic and 14 benthic samples which contained 10,793 fungal sequences were used.

Findings 

The authors found a clear difference in the marine fungal community composition between the pelagic and benthic samples. They found that very few OTU were shared between these two realms of the 739 OUT present in the dataset only 114 were found to occur in both the water column and sediment site.

As hypothesised by the authors the environmental variables between sites were a significant predictor in determining the marine fungal community structure and played a much stronger role then geographic location (73% compared to 18%). Furthermore by looking at the effect of individual environmental variables it was found that depth was the best explanatory variable, explaining 24.3% of the total explainable variance dissolved oxygen was ranked second (23.5%) and nitrate third (22.9%).

The study used one of the most extensive datasets to date to examine the global distribution of marine fungal communities, although previous studies showed that both geographic distance and habitat are important predictors of fungal community composition this study was the first to address which of the two contributed more to community differences, previous studies also had limited geographic scales (<1000 km). However even studies that did examine larger geographic ranges tended to focus on specific habitats such as mangroves, or hydrothermal vents in contrast with the present paper which investigated a range of habitats to allow differences between environmental parameters to be more widely accessed. There were still limits to the study for example the microbial techniques used were insufficient for capturing a number of rare OUT, it is noteworthy to mention that even with this limit the conclusions and community similarity and divergence are unlikely to be significantly impacted, however I think that future studies could focus on addressing this bias in sampling by using techniques that are able to detect these rare OUT using primers with a higher specificity for fungal taxa to provide a more complete picture. The authors used secondary data and therefore had no control of where the data was collected which lead to sampling locations being unevenly distributed making it harder to draw conclusions. The authors also run into trouble with the sequence region they used due to the conserved nature of the SUU v9 region there were limits to the resolution to lower taxonomic ranks. The study also only included free living fungi I think it would be interesting if fungi associated with macroscopic host were considered in future studies as this would allow us to see the implications to the ecological dynamic this association facilitates. Overall even with the limitations of the study this study has helped in our understanding of fungal distribution and the factors that influence it on a global scale and contributed a new understanding of factors affecting fungal distribution.

Tisthammer, K., Cobian, G. and Amend, A. (2016). Global biogeography of marine fungi is shaped by the environment. Fungal Ecology, 19, pp.39-46.

A sponge bath the fun-gi deserves

Sponges are a filter feeding, stationary organism that can be found in a wide range of marine habitats. They can harbour a wide range of microbes which may account for up to 40% of the sponge tissue. One of these microbes is filamentous fungi which play a role in carbon and nutrient recycling in the environment.

Passarini et al. (2013) aimed to taxonomically rank filamentous fungi found on the sponge Dragmacidon reticulatum to assess the fungal diversity. To achieve this they collected samples from the along the coast of Brazil, from which samples of internal tissue were removed and grown on a medium to isolate the fungi present. The genetic material was then extracted and amplified, from then Matrix-assisted laser desorption/ionisation mass spectrometry was used to spectrally identify and sequence the genetic material. In total they isolated 98 different filamentous fungi from two samples of D. reticulatum, noting 64 distinct fingerprints amongst 24 genera.

I think this paper is useful in identifying what organisms can live inside sponges, and also shows how using a polyphasic approach is useful when it comes to identifying species, but I think the paper has a few draw backs. Firstly is the poor grammar, there are multiple sentences that don’t make sense simply because they haven’t proof read the paper correctly. Secondly is the use of the term ‘prokaryotic’, it was 1977 when the three domain system was first proposed and I think that almost 40 years later scientists that are getting papers published really should be using the current terminology. In the introduction that marine derived fungi and bacteria may have significant biotechnological uses, but that is almost the last they say if it, the word biotechnology is not mentioned again and they rarely allude to it in discussion. They also mention nothing about why these fungi are living inside the sponge and do not hint towards any relationship there may be, whether it symbiotic or parasitic.

Referenced paper:

Passarini M.R., Santos C., Lima N., Berlinck R.G. and Sette L.D., (2013). Filamentous fungi from the Atlantic marine sponge Dragmacidon reticulatum. Archives of microbiology. 195(2). pp.99-111.

Antifungal Coral Fungi

Corals and their holobionts are important organisms and are vulnerable to disease. They lack a cell based immune responses, thus it is possible that microbial associations provide some defence. Previous research has showed that marine-derived fungi associated with invertebrates do produce metabolites that have antibiotic properties. Therefore, it is possible fungi associated with corals produce secondary metabolites that act in a similar way, including acting as a fungicide. If this was found to be true, there is huge potential for industrial applications. It is hard to tell from Putri et al (2015) paper if previous research has been done into coral associated fungi and their antimicrobial properties. Although it seems there is little evidence of coral symbiont fungi producing antifungals against fungi which are infectious to humans. Trying to answer this question was Putri and his team who used the soft coral, Sinularia sp.

Samples of Sinularia sp. were collected from the North Java Sea, homogenised and spread on MEA medium. Colonies of fungi, identified by morphological features, were purified. After incubation of these colonies, Candida albicans or Aspergillus flavus were added to the plates and left for further incubation. It is unclear from the methods, whether the infectious fungi were separate or together. To see if antifungal activity had occurred, the formation of inhibition zones was monitored. Extracts from fungi colonies were sequenced and the constituents identified by phytochemical tests.

Only one fungal associate, of 15, was found to inhibit the growth of both infectious fungi. The symbiotic fungi had 99% similarity to Aspergillus unguis. Phytochemical tests revealed the presence of phenols which can act as fungicide, flavonoids which can disrupt membrane function and biosynthetic processes, as well as terpenoids which have antimicrobial properties. All suggesting that there could be potential applications of coral associated fungi in pharmaceuticals, but a lot more research is necessary.

This paper was hard to read and the introduction gave little background knowledge, most of it is given in the discussion. The justification was based largely on pharmaceuticals and little else. I feel like they are jumping the gun in looking into human pathogenic fungi; that they are trying to be the first in a much underexplored area. I feel as though there should be more marine research to act as a basis for further research on human fungal diseases. This paper is littered with mistakes, including incorrect spellings. Despite all these issues with writing, it doesn’t seem to be bad science and does look into an area that basically nothing is known about.

Putri, D. A., Radjasa, O. K., & Pringgenies, D. (2015). Effectiveness of marine fungal symbiont isolated from soft coral Sinularia sp. from Panjang Island as antifungal. Procedia Environmental Sciences, 23, 351-357.

Haemocyanin – the underdog of the squid-vibrio symbiosis.

Euprymna scolopes are known for their light organ symbiosis with the luminous bacterium Vibrio fischeri. The bobtail squid harvest planktonic symbiont cells from seawater each generation and juvenile squid’s light organ exposes two pairs of prominent ciliated epithelial appendages. These appendages secrete mucus, which creates currents that facilitates recruitment of symbionts.  Gram-negative bacteria all adhere to the cilia however after 3 hours V. fischeri are the only bacteria able to persist. V fischeri then travel down ducts and reach internal crypts where they multiply, which causes the light organ to undergo irreversible morphogenesis. The mechanisms behind the specificity of V. fischeri symbionts has been speculated and a study by Kremer et al. 2014, suggests the oxygen binding mechanism, haemocyanin may be involved in the establishment and persistence of the V. fischeri symbiosis.

Adult Hawaiian bobtail squid (E. scolopes) were collected from Oahu, Hawaii and maintained and bred in a recirculating seawater system. Juveniles were incubated overnight with either V. fischeri wild type, a light production defective mutant or with the absence of V. fischeri. Full length cDNA sequences of haemocyanin isoforms were obtained by rapid-amplification of cDNA ends (RACE) and aligned with sequences from a variety of mollusc species for phylogenetic reconstruction. Haemocyanin transcripts from both isoforms were then quantified by qRT-PCR. Affinity-purified polyclonal antibody of haemocyanin was produced and immunocytochemistry experiments were carried out to visualise and localise haemocyanin transcript and protein in the squid tissues. The functional properties, including oxygen affinity of the hemocyanins were measured with a diffusion chamber. Oxygen affinity was determined from whole-blood samples, collected either at night or in the day.  A pH sensitive fluorescent probe (SNARF) was used to identify the flow between haemolymph and the light organ crypts. Associated pH was measured at both of these sites using a pH- sensitive probe. Phenol oxidase and antimicrobial activities of haemocyanin were also determined.

Results showed Euprymna scolopes synthesises two distinct isoforms of haemocyanin. Localisation and expression of haemocyanin transcripts showed that both isoforms were expressed in similar levels in all tissues. The main site of synthesis of both haemocyanin isoforms was in the gills. Mantle and gut within the juveniles, eyes and central core of the light organ of adults, were also secondary sites of synthesis. Fine-scale distribution of haemocyanin protein also showed high levels in the gills, pore and epithelium regions of the light organ and within the mucus adjacent to these regions. Haemolymph passes from the vascular system into the crypt spaces, which has a lower pH than the haemolymph. This suggests high affinity haemocyanin takes up oxygen in the gills and circulates until it reaches the crypt space where the pH is significantly lower due to bacterial metabolism. V. fischeri luciferase has a higher affinity for the oxygen, which causes the oxygen to dissociate from the haemocyanin.  Therefore bacteria metabolism over a diel cycle in relation to their behaviour influences the delivery of oxygen by altering the pH of the crypt spaces and haemocyanin is an important component of this cycle. V. fischeri showed resistance to the phenol oxidase antimicrobial activity of the haemocyanin, whereas growth was inhibited in other marine gram negative and positive bacteria. This therefore infers the haemocyanin within the mucus may play a role in the specificity of symbiont selection.

I found this paper to be interesting as the specificity of V. fischeri in this symbiosis was briefly mentioned in a previous piece of literature I read, which speculated that the specificity was due to the high oxidative stress the bacteria have to endure within the crypt space. As this paper suggests the specificity is in fact determined before the symbiont enters the host, it provided an alternative and conflicting theory, which was interesting to study. Overall this study highlights the emerging importance of haemocyanin in the role of specificity in selecting V. fischeri for the symbiosis as well as their importance in the control and adjustment of the physiology involved in the maintenance of the squid-vibrio symbiosis. I believe this paper and its findings provide a good basis for future research regarding the antimicrobial properties of haemocyanin.

Kremer, N., Schwartzman, J., Augustin, R., Zhou, L., Ruby, E., Hourdez, S. and McFall-Ngai, M. (2014). The dual nature of haemocyanin in the establishment and persistence of the squid-vibrio symbiosis. Proceedings of the Royal Society B: Biological Sciences, 281(1785), pp.20140504-20140504.