Do Viruses play a vital role in the food chain of marine ecosystems?

In marine ecosystems viruses are known to cause phytoplankton cells to lyse which releases the cell contents into the water as dissolved organic matter (DOM), this in turn provides nutrients for heterotrophic bacteria which may then be consumed by dinoflagellates, linking in to the traditional ocean food chain. Some nutrients from lysed cells may also become mineralised, therefore providing nutrients to phytoplankton of the same species that are resistant to the virus and other species of phytoplankton. This study looks at marine microorganism population dynamics in more detail and puts the role of viruses in nutrient cycling into practice. They performed two experiments within their study. In the first one they studied two phytoplankton species, Phaeocystis pouchetii and Rhodomonas salina, and the effects that a virus specific to P. pouchetii (PpV) had on both populations. In the second the populations of bacteria and heterotrophic nanoflagellates (HNFs) were also recorded, along with the nutrient levels.

In the first experiment it was found that the populations of both species of algae didn’t start growing until 3 or 4 days into the experiment, at which point the populations started increasing exponentially until it levelled off due to nutrient availability or viral infection. The results showed that the population of P. pouchetii was negatively impacted even when low concentrations of the virus were present, with the population decreasing to zero after just 13 days. The population of R. salina on the other hand was effected much less drastically by the virus and rather than killing off the species completely it levelled off the population more quickly, having a lower population at the end of the experiment than had the viruses not been there. On top of this the levels of R. salina were in fact higher than when the algae had to compete with P. pouchetii.

The results of experiment two showed that the bacterial population increased considerably after lysis of the P. pouchetii took place. This increase was higher when P. pouchetii was the only algal species (culture 2) than when R. salina was present too (culture 3). After peaking the bacterial populations fell and then stabilised with the bacteria in culture 2 stabilising at a higher population than the bacteria in culture 3. The HNF population initially rose more quickly in culture 3 than in culture 2 and as the HNF numbers rose the bacterial levels fell as there were larger numbers of HNFs to consume the bacteria. The populations of HNFs in culture 3 fell and stabilised at a lower population than that in culture 2 but the reason for this was that there was initially a higher concentration of dinoflagellates in the R. salina than the P. pouchetii. The measurements of nutrient levels showed why the increase in the bacteria population took place as levels of nutrients generally rose due to being released during lysis of the P. pouchetii. As there was a higher concentration of P. pouchetii to lyse in culture 2 the nutrient levels rose higher than in culture 3, explaining the differences in bacteria numbers between the 2 cultures.

This experiment has shown that viruses are able to release nutrients into the water, by lysing algal cells, to allow bacteria to grow more quickly as well as allowing other species of resistant algae to grow without competition, showing that the presence of viruses can allow two species that occupy a similar niche to coexist in some cases.

Reference: Haaber, J and Middelboe, M (2009), Viral lysis of Phaeocystis pouchetii:Implications for algal population dynamicsand heterotrophic C, N and P cycling. International Society for Microbial Ecology, Vol 3, Issue 4, pg 430-441

Ocean acidification: the end for corals?

Coral reefs are massive structures, covering around 620,000km², and have given rise to incredibly diverse community assemblages. Included in this diversity are the photosynthetic symbionts associated mutualistically with the corals themselves and the other microorganisms which inhabit the coral tissues and structures. The interactions between these organisms are important for coral health and despite many reports focussing negatively on the relationship between microorganisms and corals; it is likely that most of the interactions are in fact positive and very influential as the microorganisms may provide nutrients and antimicrobial agents for protection. This has lead to the development of the coral probiotic hypothesis by Reshef et al. (2006) which suggests that by changing the coral-resident community, the coral holobiont can adapt more quickly and effectively to the changes in environmental conditions that we impose on them (increased temperatures, CO² concentration etc.).

This study focuses on the importance of reduced pH through ocean acidification and the effects it has on the scleractinian coral Acropora eurystoma and its microbial community. This was achieved by taking coral samples from the Red Sea which were split into two groups and subjected to two different pH’s: 8.2 (ambient seawater) and 7.3 (acidified). This was done for 10 weeks, after which the samples were fractionated and the bacteria isolated from each fractionation: skeleton, tissue and mucus. The DNA were extracted and PCR amplification of the 16S rRNA was carried out, the products of which were put through DGGE analysis in order to construct clone libraries to enable identification of the OTU’s present in each coral fractionation for each pH treatment. Finally, antibacterial screening was carried out on five common indicator bacterial strains to determine whether the coral bacteria had any beneficial antibacterial properties.

The DGGE analysis showed that in all fractionations, community composition was affected by pH. This was supported by the clone libraries which found a total of 103 OTU’s for the reduced pH of 7.3, compared to 74 for the ambient pH of 8.2. This increase was seen across several bacterial groups including the two dominant ones: Gammaproteobacteria and Cyanobacteria. Differences were also found in the antibacterial screening tests as the corals maintained at pH 7.3 showed a marked increase in bacteria producing antibacterial activity than those maintained at 8.2. The most noteworthy of these were the Vibrionaceae and Rhodobacteraceae which comprised 50 and 29% of the activity, respectively.

The increased diversity of bacteria in the reduced pH treatment could initially suggest that ocean acidification may not be as seriously damaging to some corals as has been previously thought. In fact, where the coral probiotic hypothesis is concerned, a greater diversity would seemingly allow corals more flexibility to adapt to changing environmental factors, as there would be more options of potentially beneficial bacteria to inhabit their tissues. However, as the authors note that the reduced pH used was not representative of the predicted levels for the next 100 years, it may be that such diversity would not be available in reality and so corals could still have trouble adapting to changing conditions and still be caused damage. Furthermore, it appears that several of the OTU’s found at the lower pH were genetically similar to strains that have been associated with coral disease previously; therefore this greater diversity may well be harmful to corals. Although, it must be noted that the abundance of some harmful bacteria such as Desulfobacter, a sulphate-reducing group partially responsible for black band disease, diminished in the reduced pH. This shows the need for more detailed and realistic research as in order for it to be useful, we must be able to apply it properly to real world scenarios by using real world parameter levels.

While several groups of the OTU’s found in the pH 7.3 fractionations are usually considered to be harmful to corals, none of the coral specimens in either treatment showed any sign of bleaching or disease. However, other authors have reported that just a few slight changes to the coral bacterial community can dramatically change the health of the coral and so here, in the reduced pH treatment, the shift to a more opportunistic set of OTU’s known to be associated with stressed or diseased corals seems to be an important biomarker for the risk that reduced pH may impose on corals and perhaps, over a longer time period signs of disease may have begun to show.

Consequently, this study appears to be of great importance as it highlights the need to monitor coral microbial communities more closely and to understand the relationship within the holobiont. With more detailed research of the area we may be able to protect corals and the mass of life they support better in the future, even though we cannot turn back time and undo the damage we have already inflicted upon them.

A review of: Dalit Meron, Elinor Atias, Lilach Iasur Kruh, Hila Elifantz, Dror Minz, Maoz Fine and Ehud Banin (2010) The impact of reduced pH on the microbial community of the coral Acropora eurystoma. The ISME Journal; 5, 51-60.

Rethinking the N cycle

New processes and players in the nitrogen cycle: the microbial ecology of anaerobic and archaeal ammonia oxidation

This paper is relevant to our next lectures and is hyper-linked to on the lecture slides. The paper has a lot of details but here I have taken the main points.

Nitrogen is fundamental to the structures and biochemical processes that define life. Our understanding of how this element is cycled on Earth has changed drastically in just the last few years. First by the discovery of anaerobic ammonium oxidation in natural systems, and more recently by the discovery of aerobic ammonia oxidation within the domain Archaea.

In the conventional view of nitrification (NH3-NO2_-NO3_), the metabolic labour is divided between two distinct groups of organisms, ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria (NOB). Denitrification (NO3_-NO2_-NO-N2O-N2) is primarily heterotrophic, facultative, occurs under low oxygen conditions and is widespread among over 50 different genera. In this mini review, the authors focus on recent developments related to the microbial ecology of anaerobic and archaeal ammonia oxidation.

Heterotrophic denitrification was considered the only sink for fixed nitrogen under anoxic conditions in natural systems. But then anaerobic ammonium oxidation (anammox) to N2 gas was obtained from anoxic (denitrifying) bioreactors of wastewater treatment plants by ‘anammox’ bacteria that have unique features. These features include; the use of hydrazine (N2H4, i.e., rocket fuel) as a free catabolic intermediate, the biosynthesis of ladderane lipids and the presence of an anammoxosome (intracytoplasmic compartment).

Anammox has been found to be ubiquitous and will likely to be found in virtually any N-containing ecosystem with a pronounced suboxic zone or chemocline. The process has been found to be going on in a number of environments using a combination of 15N-based tracer studies, analysis of ladderane lipid biomarkers, fluorescent in situ hybridization and phylogenetic and quantitative PCR analysis of 16S rRNA sequences. More sophisticated tracer methods and/or gene marker studies are needed to assess the importance of this process in the environment and research is underway. However studies so far indicate that anammox is probably responsible for 30–50% of all marine N loss.

The review also summarizes studies that have found ‘new players in the N cycle’. They are among the mesophilic Crenarchaeota which are mesophilic archaea that are now recognized to be an ubiquitous component of marine plankton. The genes and associated enzymes of ammonia oxidation are being found in many archaea clades. The review concludes that ammonia oxidizing archaea are much more abundant than ammonia oxidizing bacteria.

The review has a whole section discussing these new paradigms. It says that the processes can only be characterized by employing multiple, complimentary approaches, to address a growing number of questions. One such question is; if N loss in oxygen minimum zones is principally driven by the autotrophic process of anammox rather than heterotrophic denitrification, what happens to organic carbon, if it is not remineralized via denitrification? There are many other questions posed that ask us to rethink the N cycle.

It is clear that the microbial ecology of anammox bacteria and AOA will be an area of active research for years to come, and will be essential to our understanding of the global N and carbon cycles going forward.

Do viruses protect phytoplankton from UV stress?

As recent evidence suggests, ultraviolet radiation (and most significantly ultraviolet radiation B or UVB) is on the increase in north and south temperate latitudes. The detrimental effects of this have been emphasised in many previous studies regarding mutagenesis, primary production rates, loss of pigmentation and many others. All of this is in spite of defensive strategies employed by many systems (avoidance, repair etc.).

Marine viruses are the most abundant biological particles in the sea, and they are known for playing a significant role in many essential marine cycles. Ultraviolet radiation (UVR) is considered responsible for the loss of infectivity and destruction in bacteriophages and cyanophages, though some viruses have been shown to possess a repair gene for such eventualities. The effects of UVR on the interactions between viruses and phytoplankton are currently not investigated due to the lack of available cultured samples.

In this investigation, 5 important marine phytoplankton-virus systems were cultured onto F2 medium and kept in water at a constant temperature (10-15^o C depending on their place of origin). These were regularly mixed and exposed to 14 hours of light to 10 hours of dark, with UVR lamps (UVA and UVB) on for 4 hours of each light period. These conditions were to mimic normal surface water conditions, with normal recorded levels of UV radiation.

3 experiments were conducted. The first contained only phytoplankton and was exposed to photosynthetic active radiation (PAR) (normal light) only, PAR+UVA and PAR+UVA+UVB. The second consisted of the same tests, but with the addition of viruses to fresh cultures of the same strain. These phytoplankton were previously infected by the virus, allowed to recover to ensure immunity against it and then co-cultured with the same virus. The third involved isolating the viruses from the second experiment and adding them to more fresh culture to test their ability to re-infect phytoplankton after exposure to UVR.

The results showed that there was no difference at all in the cultures when exposed to PAR or UVA. However, there was varied change in response to the addition of UVB – in experiment 1 some species showed no change while others showed intermediate responses and some even died. In experiment 2 some of the cultures seemed to be less sensitive to UVB than they were in experiment 1. In experiment 3, viruses exposed to UVB tended to take longer to infect the phytoplankton cells, suggesting that infectivity was decreased.

It seems that in this experiment, the viruses indirectly protected the phytoplankton from UVB damage. It is possible that during infection, the viruses actually transferred protective genes to their host. Though this theory is yet to be proven, it could demonstrate an ecological advantage for previously infected phytoplankton, as they are not only resistant to the virus, but also seemingly less sensitive to UVB. There are however further implications for these results – during periods of high levels of UVB, infectivity of viruses could decrease which could lead to long lasting algal blooms, having a knock-on effect on the wider marine ecosystem.

Reference: Jacquet, S. and Bratbak, G. (2003) Effects of ultraviolet radiation on marine virus-phytoplankton interactions, FEMS Microbiology Ecology, 44, 279-289

The bacteria formerly known as . . .

Halophilic Bacteriovorax (Bx) were formerly known as marine Bdellovibrio. They are obligate Gram-negative predatory bacteria which can be found in various environments. Alike Bdellovibrio, Bx have a very unique biphasic life cycle which is reminiscent of viruses. They swim at very high speeds (160 um s-1) and can detect prey using a chemosensory system which is not very well understood. They then attach to their prey and enter the periplasmic space. Once inside they can lay dormant however usually they consume the cytoplasm of the prey, elongate and then differentiate into several motile cells which lyse the prey cell and start a new cycle. Its therefore quite possible they have an important role in contributing to bacterial mortality and nutrient cycling.

They differ from Bdellovibrio and freshwater Bx species mainly due to their salt tolerance, %G+C ratio, fatty acid profiles, antigenic variations and habitat. Phylogenetic trees have shown that Bx have two major branches of which one contains two freshwater species and the other two marine species. However previous studies of Bdellovibrio and Bx have more or less completely ignored the marine strains. The aim of this study is to gain some perspective of the diversity and geographical distribution of halophilic Bx by analysing the ssu-rRNA sequences from isolates collected from a very wide range of geographical and environmental sites.

The method I'm about to give is an extremely shortened, general version of what they actually did in the study. Samples were collected from marine, salt lakes and estuarine environments from literally all over the world (27 sites). They were mainly pelagic water samples however some from biofilms from oyster shells and a few from gills of aquarium fish. There don't seem to be any sediment samples taken which I thought would probably lower the diversity a bit but I'm sure they had their reasons. The water samples were filtered to leave the small Bx which were then enriched in a prey-seawater broth. The prey organism was Vibrio parahaemolyticus which had been shown through previous studies to be the best prey. They were then plated out using a double agar overlay method and the plaques derived where confirmed to contain Bx using fluorescent microscopy. Several rounds of sub culturing were performed to produce pure cultures. Culture suspensions were then filtered to get rid of any prey cells and centrifudged leaving Bx cells in the pellet. The supernatant was removed and a DNeasy Tissue Kit was used to extract DNA from the pellet. The ssu-rRNA gene sequences were then amplified using PCR with universal rRNA gene primers. They then used various programs to produce a phylogenetic tree.

One hundred and eleven novel marine strains of Bacteriovoracaceae were isolated from the marine environments. A phylogenetic tree based on the ssu-rRNA gene sequence data and using Geobacter as an outgroup revealed two main branches. One branch included one hundred and ten of the isolates and other branch included the other isolate, several freshwater isolates and two soil isolates. Analysis showed eleven distinct clusters (isolates with >96.5% similarity) however only eight contained two or more isolates. Almost half of the isolates fell into two clusters (3 and 9) which were also the most widespread geographically. Although most clusters were found in many different locations some were specific to certain areas. Cluster 5 was only found in estuarine waters and cluster 12 was only found in warmer tropical waters. Isolates from many different clusters could also be found in the same site.

The paper very convincingly suggests that Bx are incredibly more diverse and geographically dispersed in marine environments that previously thought. The experiment itself is going to be a huge underestimation of the true diversity of these bacteria. For instance Bx have been shown to be quite particular about choosing their prey and therefore it is quite possible that many are prey-specific to bacteria which cannot be cultured or are unknown. It is also suggested that this prey-specificity may be what drives their distribution. The divergent boundaries for clusters (>3.5%) were intentionally quite high so its possible that there could have been many new species isolated. I quite liked the paper as it looked at the phylogenetic tree of these marine bacteria which is incredibly important for understanding them and future studies. There doesn't seem to be many papers at all like this one as most studies seem to neglect the marine strains. I strongly agree that these bacteria could have a much more important role in nature than first thought. They are also being extensively studied for obvious reasons in the medical field as a possible solution to antibiotic-resistant Gram-negative pathogens.

A review of Pineiro, S. A. et al.(2007) Global survey of diversity among environmental saltwater Bacteriovoracaceae, Environmental Microbiology, 9(10), 2441-2450

Salt: A microbial preference?

A review of Hollister, B. E. et al (2010) Shifts in microbial community structure along an ecological gradient of hypersaline soils and sediments, The ISME Journal. 4, 829-838

Across the globe, microbes can be found adapting and living in numerous extreme environments. One example is hypersaline environments, such as salt lakes, hypersaline springs and solar salterns, which sustain diverse microbial communities. Lozupone and Knight 2007, established that levels of salinity are a major factor in determining the composition of microbial communities. Moreover, recent global surveys have confirmed that sediment environments sustain more phylogenetically diverse bacterial communities than any other environment. Despite this, the majority of research published on microbial diversity in hypersaline ecosystems has been focused on aquatic environments only. This has allowed for a better understanding of the biology of extreme environments while also discovering novel organisms as well as enzymes with potential for various biotechnological applications, but has also pointed out the necessity for similar research to be done on sediment and soil communities.

The research was focused at La Sal del Ray, a naturally occurring salt lake in southern Texas, whose diversity and microbial communities have not been characterised. 8 samples were taken along a transect placed across the shoreline and lake sediment, The samples were chemically and physically analysed, recording properties such as pH, water content and salinity, and along with quantitative PCR, 16s rRNA tag-pyrosequencing and multivariate statistics, they were used to identify the microbial communities present and to establish the relationships they share with their environment.

The tests indicated that bacteria accounted for 97% of all the rRNA copies detected. Despite the limited abundance of Archaea, distribution patterns were found, as they were absent in the terrestrial sites and slowly increased in population (up to 8%) along the transect to the aquatic sites. Limited correlation between their abundance and sodium content was also found, suggesting a strong correlation between Archaeal abundance and water content. Significance tests confirmed correlation between community diversity and the sample water, pH and total inorganic carbon content and the Shannon diversity index value supported this showing diversity and community size decreasing as conditions became more water-logged and salt-rich. To establish the diversity and richness, 16s rRNA cloning and sequencing was conducted and found 16596 unique OTUs overall. Around 24 bacterial and 2 Archaeal phyla were detected, with Proteobacteria and Bacteroidetes being the most frequently encountered.

The authors concluded that the distribution of OTUs in the lake were either site-specific, occurring in only one of the sites, or cluster-specific, the clusters being terrestrial, intermediate or aquatic. They were also able to show which communities preferred higher or lower levels of salinity. For example, Acidobacteria and Rhodobacteraceae were detected in all the sites, however their abundance varied as Acidobacteria abundance declined as the conditions became more salt-rich and water-logged and instead, Rhodobacteraceae abundance increased. Similarly, many other site-specific or cluster-specific organisms were found, increasing our understanding of the salt lake microbial distribution and specifically what preferences they each have to salinity.

However, by assessing their results overall, it was concluded that although salinity has a major function in determining community composition at global scales, in regard to this study area and in salt-rich systems, they have little influence. Instead, the composition along the transect is based on microbial requirements for oxygen, carbon substrates that they metabolise and the range of pH they can tolerate. This is supported by the high correlation between microbial communities and water-logging, as although oxygen levels were not measured directly, water content is a good indicator of oxygen and thus supports the theory that microbes ultimately distribute to areas based on the oxygen availability.

The study has produced significant and detailed results. The researchers expanded their study by introducing variables such as different conditions for example, pH, organic carbon and calcium content and how microbial distribution correlates with them. This makes the study more representative by assessing many aspects of the ecosystem instead of just focusing on one, also making data more reliable and less bias. They have also taken into consideration possible errors when using PCR and pyrosequencing and have tried to reduce these as much as possible by using a combination of techniques which reduces individual errors and bias of each method and gives us a more detailed picture of the communities. The study also compared their results to previous studies focusing on hypersaline environments and found that the relative abundance of many of the dominant species they detected were very similar to the results of the other studies, further supporting the accuracy of the data and expanding the field of knowledge regarding global hypersaline microbial communities.

The authors established that around half of the estimated diversity of the communities were detected through sequencing and that some bacterial species found could not be identified. It would be interesting to find out a more accurate figure of the different species present by further PCR and sequencing, and through larger gene libraries, identify the unknown bacteria.

Other references:

Lozupone C.A, Knight R, (2007). Global patterns in bacterial diversity. Proc Natl Acad Sci USA 104: 11436-11440

Another threat to coral reefs, are human pathogen’s causing coral diseases?

Acropora palmata was once one of the most abundant corals found in the Caribbean; however in 2006 it was listed under the United State endangered species act. The cause of the decline in coral numbers is mostly due to a disease that is unique to this coral species, acroporid serratiosis (APS), or more commonly known was white pox. The bacterium which causes this disease is Serratia marcescens, it is also known to cause respiratory and urinary tract infections, meningitis and pneumonia in humans. Following a major outbreak of the disease in corals in 2003 it was found that the strain PDR60 of the bacterium was found in untreated human waste water and diseased A.palmata, this suggests that human sewage could be a cause of the increase in the prevalence of the coral disease.

Because of the endangered status of A.palmata only 8 treatments and 3 controls could be performed, healthy corals not infected with APS were collected and inoculated S.marcescens of different strain and origin, strains were determined using a three step method, culturing on MCSA then on DNase with Toluidine Blue agar and then PCR.

1. Strain PDR60 from APS affects A.palmata

2. Strain PDR60 from APS affects A.palmata

3. Strain PDR60 from APS affects A.palmata

4. Strain PDR60 from an apparently healthy non host coral Siderastrea siderea

5. Strain PDR60 from the snail Coralliophila abbreviata

6. Strain PDR60 from wastewater

7. Strain PDL100 ( confirmed APS pathogen) APS affected A.palmata

8. Strain ww131 from wastewater

Controls include : E.Coli, sterile CaCO₃ sediment (vehicle control) and seawater.

After inoculation the number of days taken until tissue loss occurred was measured, once tissue loss occurred the surface mucopolysaccharide layer (SML) was collected using a needle, the SML contained necrotic snail tissue. The SML was centrifuged and then spread onto MCSA agar, positive bacterial colonies appear red/pink which is characteristic of a S.marcescens culture. The positive bacterial cultures were plated onto DTC plates, DTC presumably stands for the type of agar however this is not explained. If the DTC plates were positive for S.marcescens, this appears as a red halo on the DTC plate they were then plated onto non selective TS agar to form pure cultures.

Isolates from APS affected coral, which after growth on MSCA and DTC plates were presumed to be S.marcescens, underwent a Seratia-specific PCR, band pattern were used to assess the relatedness between strands. This allowed the confirmation that the bacteria collected from the APS legions was in fact the same strain that the coral fragments had been inoculated with. This method of proving disease is known as Koch’s postulates, Koch’s postulates has identified strain PDR60 in host and non-host corals and also snails, the aim of the paper was to assess whether the strains from the non-host coral S.sidera and the snail C.abbreviata along with wastewater strains could cause disease in a healthy A.palmata.

Strain PDR60 from wastewater and diseased A.palmata caused disease within four and five days respectively, this shows that wastewater is a source of APS, furthermore the human strain was shown to be a coral pathogen through Koch’s postulates.

The inoculates were considered virulent if tissue loss occurred in the corals before day 15, as this was when the control vehicle (sediment) isolate caused tissue loss. No S.marcescens was detected in the water or in the coral fragments before exposure. The strains collected from C.abbreviata and S.sidera caused disease after the 15 days period observed in the control (CaCO₃ sediment).Therefore it is possible another stressor may have caused the tissue loss, or however the strains isolated from non-host coral and snails could in fact be virulent but didn’t show signs because the A.palmata fragments may have been resistant to APS. This hypothesis is supported by the fact that the isolate 3 which was extracted from APS affected coral did not cause disease in the corals which were exposed to it. Strain PDL100 of the bacterium S.marcescens had limited pathogenicity and the pathogenicity has been shown to of decreased in recent years. Along with V.shiloi , a bacteria which causes coral bleaching, and A.corallicida, which causes white plaque. The author states that this may be because of development of host resistance to the bacterial strains. Examples of further studies would need to assess the susceptibility of host corals and also the genetic variation between S.marcescens strains and isolates.

The importance of understanding that bacterial strains from human wastewater can cause disease in coral is high; the majority of the sewage from around the Florida Keys and the Caribbean is not treated and is disposed of within a limestone substrate that allows leaking of sewage into coastal waters. Coral reefs are already threated in a variety of different ways, this study outlines the need for waste water disposal reforms as advanced wastewater treatments can reduce S.marcescens to undetectable levels. The transfer of a human disease to another animal is known as reverse zoonosis, however in this case the pathogen has passed from a vertebrate to an invertebrate and from terrestrial environment to a marine one. Further research into the tolerance of the human strain to salinity may be important as if this strain is able to tolerate high salinity levels it may become prevalent on reefs causing reoccurring infections.