Saturday 31 December 2011

Northern Elephant Seals in danger from pathogenic and antimicrobial –resistant bacteria!

Stoddard, R. A., Atwill, E. R., Gulland, F. M. D., Miller, M. A., Dabritz, H. A., Paradies, D. M., Worcester, K. R., et al. (2008). Risk factors for infection with pathogenic and antimicrobial-resistant fecal bacteria in northern elephant seals in California. Public health reports Washington DC 1974, 123(3), 360-370. Association of Schools of Public Health.

Stoddard et al. took on a tricky experiment design. The aim of their study was: to identify potential risk factors in northern seals, which relate to increased odds of bacterial infection (Campybacter jejuni, Salmonella spp. Escherichia coli- antimicrobial resistant). The study took place off the cost of California (nice), 34 million inhabitants 72% of who are coastal residents. Pollution can enter the coastal waters from many different sources e.g. municipal, industrial and agricultural effluent, storm runoff, and sewage outlets. Most of these pollutants are transported into the ocean via rivers eventually.

The Centre for Disease and Control and Prevention, claim that water transported bacterial infections are diluted to half of the 3-5 billion cases of diarrhoea in the U.S. each year. CDCP reports also implicate that the bacteria under investigation in Stoddard et al. research are responsible for the majority of the infections. Direct pathogen exposure increase is not the only concern of their research, antimicrobial resistance strand influxes are also pivotal to their research.

The study took place from July 2003-2004. The study consisted of: 165 juvenile northern elephant seals from their natal beach and 196 juvenile seals that were stranded on the California coat. Stranded seals were sampled after 24-48 hrs. of being admitted to a sanctuary rescue centre. Bacteria were sampled from all study specimens and culture, antimicrobial analysis was carried out. The complexity to their design arose during analysis when calculating risk factor coefficients. They were obtained by using parameters: gender, weight, county of standing, month, human pop. Freshwater outflow and accumulative precipitation levels at regular prior intervals. The weather data that was used was not always reliable e.g. the confidence in the historical precipitation records were not always correct, due to equipment malfunction during documentation.

Their findings were:

· Odds of C jejuni and antimicrobial resistance of E.coli were higher in faeces of seals from sites of higher freshwater outflow.

· Odds of contracting salmonella spp. in faeces is 5.4 times greater of seals stranded in locations with low levels of 30 day cumulative precipitation.

· Odd of juvenile seals having antimicrobial resistant E.coli in its faeces increase substantially, in relation to corresponding increases of freshwater outflow. The same trend was found for C jejuni.

The authors concluded: juvenile northern elephant seals are contracting antimicrobial resistant faecal bacteria and pathogenic bacteria, which are most likely acquired from terrestrial river outflow. Therefore the terrestrial locations can impact the ecology of the marine environment, and the health of the animals and humans that rely on it. Demonstrating a critical need to deepening the understanding of the complexities of the ecology of the terrestrial/marine interface.

This research highlights the validity of keeping accurate weather records and records of effluence of pollution. This information is integral to producing models for understanding the feedback loops and mechanisms that are occurring. Due to this, this research is of significant importance. Studies like these may also highlight the complexities that may be occurring in other coastal regions and indicate further anthropogenic pollutant negative feedback relationships on both microbial community composition and acquired antimicrobial resistance. Further monitoring of just how prolific antimicrobial resistance is becoming in coastal waters would indicate the extent of this problem. Making this vital research.

Aspergillus flavus: Marine organisms or terrestrial migrants?

When thinking about marine organisms we very rarely think about fungi. They do occur and can have a detrimental effect on other marine organisms. Although fungi are present in marine environments a lot of the time these species have also been found in terrestrial environments. A prime example of this is Aspergillus sydowii, which has been attributed to causing aspergillosis in the sea fans Gorgonia ventalina although there is much debate on how the organism reaches the Caribbean as the fungus is also found in Africa. The most accepted theory is that spores are blown over from Africa in dust clouds. Evidence has however shown that fungi taken from terrestrial environments are unable to cause aspergillosis and that there are differences between the marine and terrestrial strains.
This study addresses this debate using another fungus which is thought to have a role in aspergillosis, Aspergillus flavus. The fungus is an opportunistic pathogen which may cause disease in humans if infected plant matter is consumed. The fungus is known to have a high tolerance of salt which would clearly make it well adapted to living in marine environments. The researchers have examined the relationship between isolates from marine environments to isolates from terrestrial environments using Amplified Fragment Length Polymorphisms (AFLP-PCR). They tested A.flavus found in many different terrestrial environments as well as that found in diseased and healthy sea fan tissue and seawater.

The results clearly showed the genetic differences between A.flavus found in all of the different environments. Despite expectations there was always some differentiation between individuals of the fungus within isolates implying that cloning of the fungus was not occurring in any of the environments. Although there were clear differences between the fungi found in different environments there does not appear to be any specific clade that is more common in sea fans (figure 2 in the article clearly illustrates this). It also shows that there are no distinct clades separating A.flavus found in diseased tissue than in healthy tissue implying that the fungus found in diseased does not have any more particular adaptations for pathogenesis. As well as this marine and terrestrial isolates do not seem to form different populations (as shown in figure 1). This suggests that there is not a particular clade of A.flavus that is better adapted to living in marine environments but that it is able to survive in a wide range of environments.

It is still not known whether Aspergillus sporulates in the water or whether the populations come solely from terrestrial environments. Due to the lack of differences between terrestrial and marine Aspergillus it is thought that the Aspergillus colonies in the sea are dispersed from the land but we know that Aspergillus spores are able to survive in deep sea conditions so it is a possibility that dispersal may occur from colonies in the sea as well, although more research is needed to confirm this.

Anabella Zuluaga-Montero, Luis Ramírez-Camejo, Jason Rauscher, Paul Bayman, Marine isolates of Aspergillus flavus: Denizens of the deep or lost at sea?, Fungal Ecology, Volume 3, Issue 4, November 2010, Pages 386-391

When chitin meets Vibrio cholerae

A review of: Pruzzo, C., Vezzulli, L. and Colwell, R. R. (2008) Global impact of Vibrio cholerae interaction with chitin. Environmental Microbiology 10(6. 1400-1410

The paper gives an in depth review of the interaction between Vibrio cholerae and chitin. This beneficial substrate-bacteria interaction is readily observed in the marine environment. This relationship is significant in microbial ecology because of the complicated and important impact it has upon the lifestyle of the bacterium. This review documents any recent discoveries as well as an evaluation of the current literature. This allows the author to draw attention to the multiple lifestyles of V. cholerae and present a global perspective on V. cholerae-chitin interactions.

V. cholerae has been a recent focus of research, with extensive investigation regarding its genetics, ecology and physiology. However, research has largely concentrated on bacterial strains that are pathogenic to humans. Chitin is one of the most abundant biopolymers to be found in nature and is suggested to be the most prevalent in the marine environment. The relationship between chitin and V. cholerae is one the most readily evaluated incidents in microbial ecology and an example of a successful bacteria-substrate connection. The interaction provides V. cholerae with various benefits which include the adaption to nutrient availability gradients in the environment, stress tolerance and protection from predators. This interaction has a significant effect upon the lifestyle of the bacterium, influencing its ecological function in nature as well as its function inside and outside a human host.

The interaction between V. cholerae and chitin can be identified at several hierarchal levels in the environment. This hierarchal scale helps to describe the influence of the interaction on a cellular, multicellular, community and ecosystem level outlining it as a biological system that spans from the cell to the global environment. The properties of the interaction characterise different hierarchical levels are all related to one another. These include physiological responses (e.g. chemotaxis, cell multiplication, induction of competence and chitin utilization), the formation of biofilms at a multicellular level and an effect upon commensal and symbiotic relationships of higher organisms in a community. These parameters combine to globally influence the cycling of biogeochemical nutrients (e.g. C and N) and the pathogenicity of humans and animals. For further detailed descriptions regarding any of the hierarchal levels mentioned here, the paper should be consulted.

The hierarchal perspective utilised in this paper helps the reader to understand the multiple lifestyles of V. cholerae. It concisely demonstrates the role of the V. cholerae-chitin connection in the environment, showing its significance on both a cellular level as well as its influence on a larger global scale. This approach advocates that the various lifestyles of V. cholerae and its pathogenicity-related properties are derived from selective environmental pressures in its primary marine habitat. It could be argued that if diligence of V. cholerae in its primary habitat is essential, then V. cholerae has been successful in acquiring the necessary adaption’s to ensure survival. This suggests that its fitness and function outside its natural environment is dependent upon its ability to acquire the crucial properties that are specific to its new habitat or environment (e.g. acquiring fundamental adaptations to survive in the human gut).

In review of the current literature, it is proposed that the processes involved with cholera in the human gut have resulted from a specific adaptation, accumulated by V. cholerae, associated in supporting osmoregulation in the host. This outlines the V.cholerae-chitin interaction, characterised at each level in the hierarchal scale, as a possible model to aid the investigation of the role of primary habitat selection in regards to the emergent pathogenicity attributes of bacteria of which the marine environment is their primary habitat. I think that this field is always going to be of interest to safe guard our global population and it may be possible to use the conclusions drawn from this paper in understanding and identifying other virulence factors in human diseases.

Microbes takes on the largest oil spill in history...

A review of: Bethanie R Edwards et al (2011). Rapid microbial respiration of oil from the Deepwater Horizon spill in offshore surface waters of the Gulf of Mexico. Environ. Res. Lett. 6 (July-September 2011) 035301.


The Deepwater Horizon disaster is known to be the largest offshore oil spill in history. It happened in April 2010 when the drilling platform suffered a methane explosion and fire, resulting in the platform sinking. Over 600 million litres of oil seeped out into the sea before the well could be finally plugged. The fate of the oil is still not fully understood and this study explores the key role that microbes play in the degradation of the surface water oil.

Microbes can aid in the degradation and remediation of oil as they respond quickly to the addition of petroleum hydrocarbons. However, in doing this they are limited by the availability of phosphate, and in the case of this study, the offshore waters near the site of the spill are oligotrophic. This lack of inorganic nutrients would lead to microbes giving a more minimal response to the oil and at a slower rate.

In order to prove that this is the case for microbes, the researchers collected surface-water samples at twelve locations around the site of the spill. They used the same sites as previous studies researching the effects of the oil spill so as to correlate results. Measurements were taken for community respiration rates, lipase enzyme activities, microbial abundances, microbial and microbial biomass. To investigate the limitations that phosphate provides measurements of phosphate concentrations and activities were also taken. Hydrocarbon degradation rates were also measured and correlated with community respiration rates in order to further ascertain the fate of the oil.

The results of the study revealed that phosphate was scarce at all twelve sites and there was also no difference between areas inside and outside of the oil slick. Alkaline phosphatases (enzymes used by the microbes in order to access DOP when phosphate is scarce) were however significantly higher within the slick, indicating enhanced phosphorus stress. This shows that microbes within the slick were under higher stress due to the higher demand for phosphorus caused by the availability of organic carbon in the oil.

Community respiration rates were found to be five times greater within the oil slick compared to outside of it, indicating enhanced oxidation of organic carbon. Microbes clearly were able to readily respond to the addition of hydrocarbons despite the phosphorus stress, which is against what was predicted.

Lipase activity was also enhanced inside the slick. It is thought that this must be due to the addition of DOSS (Dioctyl sodium sulfosuccinate) which was in the dispersants used in response to the oil spill. This chemical contains many for ester linkages than in crude oil and so is thought to have encourages the activity. It is also possible that the enhanced respiration in microbes was due to the oxidation of dispersants in addition to hydrocarbon degradation.

Bacterial and phytoplankton biomass were suggested to be relatively constant throughout the study, whether within slicks or not. Microbial growth was stunted within the slick, despite increased lipase activity, phosphatase activity and enhanced respiration. It is thought from looking at previous studies that top-down processes such as grazing are the cause of this. An increase in respiration in response to nutrient changes is consistent with past studies and concludes that the addition of nitrogen and phosphorus to oil contaminated seawater results in faster degradation of hydrocarbons.

The study very much highlights the underestimated abilities of microbes. The observations have shown just how rapid the response of microbes can be, even in the face of huge stress and limitation. It also explains the unknown interactions between the microbes and the human defence of using dispersants which helped to break down the oil to a state which the microbes could possibly more easily degrade it.

Factors to Consider in Biogeochemistry

A review of: Strom SL (2008) Microbial Ecology of Ocean Biogeochemistry: A Community Perspective. Science, 320: 1043-1045.

Marine biogeochemical processes which in turn influence atmospheric climate are regulated predominantly by marine microbes that cycle organic carbon, nitrogen, phosphorus and sulphur. Microbial ecology and its influence on resource availability and biogeochemistry is central to many ocean based studies, although some methods are deemed to provide limited perspective due to ‘top-down’ and ‘bottom-up’ approaches. A good example of a’ top-down’ approach would be the ocean iron fertilisation experiments conducted in the last 20 years.

Ecological data and genomic findings indicate a broader view is needed to understand organism interactions and the impact it has on process regulation and microbial distribution and function. Factors such as heterotrophic grazing, lytic viruses, allelopathy and symbioses have a strong influence in shaping communities and must be considered when applying modern research methods in elucidating whole-community structure and function, to gain greater insights into biogeochemical cycling as well as the derivation and roles of the significant genetic diversity contained within a given community.

Biomass accumulation in the marine environment is insubstantial when compared to terrestrial ecosystems because microbial production turns over in days to weeks from mortality processes which consequently apply selective pressure and continually reshape a population, resulting in adaptations in favour of reducing mortality. This is supported by metagenomic surveys that have identified numerous genes related to community interactions, such as genes for antibiotics and polysaccharide synthesis which may impart cell surface defences.

Strom (2008) describes various examples which underscore the boundaries in our understanding of environmental gradients, environmental tolerance, nutrient limitations, nutrient uptake and storage, competitive ability, susceptibility to mortality, etc. of organisms involved in cycling carbon, silicon, nitrogen and dimethyl sulphide (DMS), including Prochlorococcus, Synechococcus, Emiliania huxleyi, diatoms, nitrifying and denitrifying bacteria.

DMS is produced by marine microbes through cleavage of dimethyl sulfoniopropionate (DMSP) which is then released into the atmosphere and act as cloud condensation nuclei, resulting in increased cloud cover. DMS fluxes have been thoroughly investigated but difficult to predict. Community interactions play a role in DMS production, such as in E. huxleyi blooms which produce DMSP to inhibit grazing by protists. DMSP is metabolized by bacteria and the propensity for a bacterial community to produce DMS or demethylate DMSP is dependent on community composition, hence knowledge of community interactions is essential for a mechanistic and predictive understanding of broad-scale processes.

The role of the cell surface in mediating resource acquisition, defence and mortality is poorly understood in marine microbes. Host specificity of viral infections depends on cell surface recognition and accordingly impact mortality therefore further investigations may explain how organisms adapt to selective pressures. In addition, allelopathic interactions in marine microbial communities are not well defined and may also be an effective variable in shaping community structure and function.

The genomic diversity that underlies microbial activity, microbial distribution, community processes and ecosystem function amalgamated into a holistic approach is required to comprehend the complexity of microbial ecology and biogeochemistry.

New Yeti Crab discovered using a Novel form of Symbiosis

A review of: Thurber AR, Jones WJ, Schnabel K (2011) Dancing for Food in the Deep Sea: Bacterial Farming by a New Species of Yeti Crab. PLoS ONE

Symbiosis is a way of life for many creatures and a survival technique for others. We heard about the symbiosis between V. fischeri lighting up its host squid in Colins lectures and have seen many examples of corals becoming a home for a large collection of bacteria. The symbiosis here is shown between the newly described, Kiwa puravida, a methane - seep crab and the bacteria that grow on its chelipeds (claws). Although there has been many studies completed on epibiont symbiosis, exactly how the collection of bacteria is attained remains a mystery and their benefit to the host is unclear.

Firstly the bacteria found associated with K. puravida was investigated. Interestingly it seems that much of the bacteria found to be growing on this new species are similar to that found on other decapods whose habitat includes vents and seeps, only two were unique to K. puravida. This backs up hypotheses that suggest horizontal transmission events by vent species epibionts have occurred. As well as this, organisms found at vents and seeps do not often co – exist (although similar species occupy both as the same chemical reactions seem to fuel the vents) suggesting the symbiotic bacteria have free living stags which allow them to disperse among communities.

K. puravida were seen to be waving its chelipeds at methane seeps which have a high nutrient content. Carbon isotopes provide evidence that the crab uses the bacteria as its main food source. The carbon isotope composition found in K. puravida is lighter than that found in phytoplanktonic production and so the energy source for the crab does not come from filtered phytoplankton. The fatty acid composition of the crabs muscle tissue reflected that which is also found in the bacteria which cover the crab’s setae and the abundant fatty acid 16:2 was not found in the phytoplankton sample. This however does not completely reject the idea that some nutrition may be scavenged by the crabs. Shrimp found in the same habitat are known to scavenge free living microbes from surrounding rocks. Although the rocks where Kiwa was found were not tested, rocks from the same seep did not contain any 16:2 fatty acids either; this plus the lack of scavenging behaviour support the hypothesis that the main food source for K. puravida comes from the bacteria found on its setae. The morphological features and behaviour of the crab shows us how it has become perfectly adapted to its habitat. It has specific mouthparts which allow it to scrape the harvested bacteria from its setae to its mouth. It also facilitates the growth of its epibiotic symbionts by waving its chelipeds to remove boundary layers which form around active bacteria and can stunt growth by reduced productivity. The waving allows the bacteria to access oxygen in the water column and the sulphide or methane it needs from the seeps, therefore increasing the chemoautotrophic yield and in turn the food source for the crab. It is farming its epibiotic bacteria.

The role of epibionts remains unclear. There doesn’t seem to be a regular pattern of behaviour between the bacteria and their many hosts found around the vents. Rimicaris exoculata has not been seen farming its bacteria and does not have the correct features to do it either, others do not have similar fatty acid composition to the bacteria found covering it. Kiwa puravida has been shown to farm its bacteria and it consumes it with adapted mouthparts whilst ensuring its symbionts get the nutrition need to produce a large yield. It’s a new and interesting form of symbiosis. It would be interesting to see if the rocks where Kiwa were found contain any fatty acid composition similar to what is found in its muscle tissue, and if not, scavenging for food could be eliminated as a source of nutrition for this crab.

Bloomin' Great : The use of algal compounds to treat infections of commercial crops.

This paper investigates the use of compounds found in extracts of macro-algae to limit injury and disease in plant tissues, particularly in commercial crop species during different seasons. Macro-algae produces a vast variety of complex natural products that could be a promising source of novel bioactive compounds to protect plants from the stress imposed by pathogens. This paper evaluates the effectiveness of aqueous and ethanolic crude extracts from nine different seaweeds collected on the coastline of Chile in different seasons on different plant pathogens (bacterial, fungal and viral).

Samples were taken from nine Chilean marine macro-algae collected at different seasons and ground into powder after liquid nitrogen treatment. Aqueous extracts were prepared by reconstituting the powder in water and raising the temperature to 85-90'C for 1 hour. This suspension was then filtered and centrifuged at 6000rpm for 20 minutes. The supernatant was then removed and evaporated under reduced pressure using a 50'C rotary evaporator until 1/4 of the original volume. Ethanolic extracts were performed by grinding seaweed as before but suspended in 50% ethanol at room temperature without light for 24 hours. The suspensions was then filtered and then stored in bottles protected from light. A second extraction was also prepared from the remaining suspension and evaporated using a 40'c rotary evaporator. These extracts were then dried in a vacuum desiccator using silica gel and then stored at 4'C until use.

Of the nine samples taken only four of the macro algae were effective against the pathogens tested. The bacterial pathogens tested were P.syringae and E.carotovora with high extract concentrations (10,000ppm) of the brown macro-algae Lessionia trabeulata inhibiting P.syringae and E.carotovora growth by 40-60% in comparison to the control. Macrocystis integifolia partially reduced growth of P.syringae by 50% in comparison to the negative control. These results suggest that the activity of these compounds on bacterial growth is dose and season dependent.

Experiments to determine potential anti fungal activity was carried out in vitro and in vivo. The in vitro antifungal activity of all algae samples was tested using P.cinnamomi and B.cinerea. Only extracts from G.chilensis led to a reduction of growing capabilities of P.cinnamomi and were only effective in high concentrations (10000ppm).

The authors then examined whether the extracts have some properties to protect plant leaves against infection with B.cinerea. To do this tomato petioles were treated with ethanolic and aqueous extracts at different concentrations before pathogen challenge. None of the aqueous extractions caused a reduction in injury severity in leaves after pathogen infection, however ethanolic extracts from L.trabeculata reduced damage in tomato leaves caused by B.cinerea infection resulting in a reduction of both the number and the size of lesions caused.

The results from the in vivo anti viral experiment did not show any protecting effect on tobacco leaves from damage caused following tobacco mosaic virus (TMV) infection. However, extracts from the algae Durvilla antarctia did lead to a reduction of the number and size of necrotic lesions. The addition of Durvillea antarctica extracts led to a 90% reduction in injury compared to those detected in negative controls. This protective effect provided by the extracts is more effective than those gained by using commercial antiviral Ribavirin.

This study proves its importance because traditional pesticides are no longer effective due to resistance, new agricultural practices rendering old pesticide techniques obsolete and a greater human consciousness of the wellbeing of the environment. As a parting note, the authors also indicate there have been reports that macro-algae compounds have been shown to be effective against human pathogens.

Review of : Jiménez. E, Dorta. F, Medina. C, Ramírez. A, Ramírez. I, Peña-Cortés. H; 2011; Anti-Phytopathogenic Activities of Macro-Algae Extracts; Mar. Drugs 2011, 9, 739-756.

Vibrio shiloi successfully inhibits free radicals and causes coral bleaching

This paper looks into more detail at how Vibrio shiloi, the most well studied bacterium known to cause coral bleaching, is able to become a successful virulent at raised temperatures. The genes which code for some of the virulent factors are only expressed at high temperatures explaining why the coral and algae are able to co-exist with the bacteria during lower temperatures of 15-20oC. In temperatures between 26oC and 31oC the bacteria is thought to adhere to methyl β-D-galacto pyranoside receptors of the coral mucus and penetrates into the host cell, where the corals symbiotic zooxanthellae live. It then produces a toxin that transports ammonia, which the coral has produced, into the algal cells in order to inhibit photosynthesis and lyse algal cells, which causes the coral bleaching. The zooxanthellae produce a high concentration of free radicals as a by-product of photosynthesis. These are extremely toxic to bacteria and act as a defence for the coral. The V.shiloi must have some sort of mechanism for overcoming the free radicals and this paper looks at how Superoxide Dismutase (SOD), a free radical inhibitor, may play an important role in the bacterial infection of the coral.

The researchers grew V.shiloi at 23oC and at 30oC and tested for the SOD activity at both temperatures as well as the type of SOD which was produced by the bacteria. The findings showed that V.shiloi is better adapted at surviving and growing at higher temperatures (30oC rather than 23oC). They also showed that there was an increase in SOD activity in V.shiloi grown at a higher temperature. Their results clearly showed that the V.shiloi grown at 23oC was not as good at surviving in a stressful environment as V.shiloi grown at 30oC as number of viable cells was significantly fewer.

They also tested for the specific SOD that V.shiloi produces and found it to be a Manganese SOD (Mn-SOD) which converts superoxide anion radicals into hydrogen peroxide. They came to this conclusion by testing whether the SOD was inhibited by either H2O2 or KCN. If it were Fe–SOD it would have been inhibited by H2O2 and if it were CuZn–SOD KCN would have inhibited it but neither H2O2 nor KCN inhibited SOD so it must have been Mn-SOD.

In conclusion, one of the reasons that V.shiloi becomes virulent at high temperatures is that they have increased expression of genes that code for Mn-SOD, which allows it to overcome the corals natural defences and penetrate the coral by inhibiting the free radicals produced by the symbiotic zooxanthellae.

Reference: Murali, M, Raja, S, & Devaraj, S 2010, 'Neutralization of radical toxicity by temperature-dependent modulation of extracellular SOD activity in coral bleaching pathogen Vibrio shiloi and its role as a virulence factor', Archives Of Microbiology, 192, 8, pp. 619-623

Friday 30 December 2011

Glycogen: a vital aid to the persistence and transmission of Vibrio cholerae?

A review of: Lori Bourassa and Andrew Camilli (2009). Glycogen Contributes to the Environmental Persistence and Transmission of Vibrio cholerae. Mol Microbiol, 72(1): 124–138.

Cholera is an acute intestinal infection caused by toxigenic strains of the gram-negative bacterium Vibrio cholerae. Cholera is a well known infection and frequently occurs in many of the developing regions. This report explores the possibility that the ability to store carbon as glycogen can aid the transition of pathogenic Vibrio cholerae between the nutrient rich human intestinal tract and the nutrient poor aquatic environments. Much is still unknown concerning the changes that V. cholerae makes in order to successfully transition between its host and the aquatic environment. Few factors are known to facilitate bacterial fitness or the transmission to new hosts but they are beginning to come to light...

Vibrio cholerae is now known to promote the expression of genes which are needed in the aquatic environment during the late stages of infection. These genes are brought about in preparation for transition to aid success. Another survival strategy is the use of chitin once in the environment to support the bacterium’s carbon and nitrogen needs. In order to investigate the benefits of accumulating glycogen in V. Cholerae, ‘mutants’ were created which lack some genes needed for glycogen synthesis and degradation.

Glycogen has already been shown to accumulate in bacteria as a result of limited required nutrients and an excess of carbon. The exact role of glycogen storage in bacteria is not thoroughly understood, but it is thought that the stores can be used for survival during periods of carbon starvation such as when they are excreted and exist in low nutrient environments for a considerable time.

The results of the study support that Vibrio cholerae stores glycogen during human infection and that these stores can prolong survival in difficult conditions faced during the bacterium’s life cycle (such as in rice-water stool and the resulting low nutrient aquatic environments). The significant results are as follows;

- Nitrogen limitation induces glycogen accumulation in V. cholerae.
- Glycogen synthesis mutants are impaired for growth during the glycogen accumulation phase.
- Vibrio cholerae utilises glycogen stores to prolong survival in nutrient poor environments.
- Glycogen stores are protective and prolong survival in rice-water stool.
- Glycogen–rich Vibrio cholerae are more virulent in a transmission model of cholera infection.

The results to the study are all successful in showing that glycogen is an important contributing factor to the fitness and transmission of Vibrio cholerae. Survival during the stresses that the human body and the aquatic environment provide is exceptionally challenged and many pathogens have evolved mechanisms to enhance their fitness, especially during the transition stage. This study has shown that glycogen stores play an important role not only in the transition of the bacterium from host to environment, but also in the environment itself and in transition to new hosts. This study is the first to highlight this ability within Vibrio cholerae, and it would be interesting to see this research furthered. This may be in terms of how glycogen can be used to protect the bacterium against the stresses it faces, or with the use of other limiting factors which could occur within the host or the external environment.



The study itself is presented in great detail and I have tried to simplify this in terms of results as best I can in my review. However, this has meant that a lot of the details from the methodology have been lost, so if anyone is interested in that aspect, it may be best to refer to the rather long and complex section of the paper!

Sunscreens makes corals pale


More than 10,000 tons of UV filters are produced annually for the global market and consumption of cosmetic sun products associated with tourism in marine coastal areas is rapidly increasing worldwide. To evaluate the potential impact of sunscreens on hard corals and their symbiotic algae, the authors conducted in situ and laboratory experiments in several tropical regions (Atlantic, Indian, and Pacific Oceans and the Red Sea) supplementing coral branches of Acropora spp., Stylophora pistillata, and Millepora complanata with different concentrations (10, 33, 50, and 100 μL/L) of several common ultraviolet filters contained in sunscreens formula.

In all replicates and at all sampling sites, sunscreen addition, even in very low concentrations, resulted in the release of large amounts of coral mucous (composed of zooxanthellae and coral tissue) within 18–48hr and complete bleaching of the coral within 96hr. Bleaching was faster in systems subjected to higher temperature, suggesting synergistic effects with this variable. The coral response to sunscreen exposure was not dose dependent, as the same effects were observed at low and high sunscreen concentrations. Therefore, UV filters can have potentially negative impacts even at concentrations lower than those used in the study.

Sunscreens typically comprise up to 20 or more chemical compounds. To identify the organic UV filters or preservatives possibly responsible for coral bleaching, seven compounds typically present in sunscreens were selected by the authors (i.e., butylparaben, ethylhexylmethoxycinnamate, benzophenone-3, 4-methylbenzylidene camphor, octocrylene, ethylhexylsalicylate, 4-tert-butyl-4-methoxydibenzoylmethane and propylene glycol) and each single ingredient was tested on Acropora spp. The results showed that sunscreens containing parabens, cinnamates, benzophenones, and camphor derivatives noticeably contribute to hard-coral bleaching even in very low concentrations.

In addition, TEM and epifluorescence microscopy analyses revealed a loss of photosynthetic pigments and membrane integrity in the zooxanthellae released from treated corals, whereas zooxanthellae membranes from untreated corals were intact suggesting that sunscreens were somehow capable of damaging the symbiotic algae. Furthermore, after the addition of sunscreens, viral abundance in seawater surrounding coral branches increased by a factor of 15 than in controls. Because, prior to any treatment, the hard corals were washed and incubated in virus-free seawater and because TEM analysis of treated corals showed the presence of roundhycosahedral virus-like particles around and inside the zooxanthellae, the authors concluded that viruses were directly released  from the symbiotic algae and that the bleaching effect of sunscreens was due to organic ultraviolet filters, which are capable to induce the lytic viral cycle in symbiotic zooxanthellae with latent infections.

Hence sunscreens, by promoting zooxanthellae viral infection, are likely to play an important role in coral bleaching. Considering a rough estimate of 78 million of tourists per year in areas hosting reefs, two daily applications per tourist traveling on a 5-day tourist package, an average usage of 20g per application and given that at least 25% of the amount applied is washed off during swimming and bathing, the authors calculated that between 4,000 and 6,000 tons of sunscreens are released every year in tropical reef areas. Because human recreational use of tropical ecosystems is steadily increasing worldwide, the global impact of sunscreens on coral bleaching will considerably grow in the future and actions are therefore needed to stimulate the research and utilization of UV filters that do not threaten the survival of coral reefs.

Reference
Danovaro R, Bongiorni L, Corinaldesi C, Giovannelli D, Damiani E, Astolfi P, Greci L, Pusceddu A (2008). Sunscreens cause coral bleaching by promoting viral infections. Environ Health Perspect 116:441–447

The top 150 words in our blog

For a bit of fun, I made a Wordle "word cloud" showing the most common words in the posts up to today (common words excluded). You can view it here. Bacteria and coral on top ... bit of a disappointing show for viruses, I think! Should we get some T shirts printed?

http://www.wordle.net/show/wrdl/4617541/BIOL3309_bLOG_ANALYSIS


In first week of January I will send you all individual feedback and your grade for this term. Well done to everyone - you have found some very interesting papers and I have enjoyed reading your reviews and the accompanying dialogue.

Don't spoil NYE by blogging until 23.59 .... Happy New Year!

Krill that might kill?

Algal blooms or Harmful Algal Blooms (HABs) are becoming more and more prevalent over the past two decades in both frequency and size, which as we are aware can be to the detriment of the ecosystems in which they occur. They consist of phytoplanktonic organisms with the most harmful being dinoflagellates and diatoms which can be toxin producing and can effect a multitude of marine organisms. In 1991 the first major event (for Monterey Bay, California) occurred where more than 200 brown pelicans and cormorants were found dead on the beaches. The cause of all these deaths was found to be a neurotoxin called domoic acid (DA) which was transferred from the diatom Pseudo-nitzschia and there have been similar incidents involving birds and sea lions since.

In 2000 the authors of this paper set out to understand if Euphausiids (krill) were a potential vector for DA in marine food webs. Krill are important members of the zooplankton grazer community and are also the primary diet of squid, baleen whales and many seabirds, if they are potential vectors DA could also affect many other marine organisms.

The sample location was Monterey Bay, California between March and August 2000 with six collection sites within the area. A 0.7m Bongo net with a 333µm mesh was used to collect the krill and the samples were frozen immediately after collection and stored at -20°C. Scanning electron microscopes (SEM) were used to determine the gut contents of the krill and to see if Pseudo-nitzschia were present. To determine if DA was present receptor binding assays were used. The authors also took samples of the water column to see if the diatoms and or DA were present and at what concentrations using cell counts and receptor binding assays. As krill were a new matrix for measuring DA measurement the authors had to also test the efficiency of the toxin extraction process before running any field samples.

The authors found that DA in Euphausia pacifica ranged from 0.1-44µg DA equiv. g-1 tissue. The DA content of krill varied with the concentration of Pseudo-nitzschia species found in the water column. The highest concentration reaching 106 cells L-1 of Pseudo-nitzschia australis.

Given the strong correlation between DA concentration found in krill and the concentration of the diatoms in the water column, the authors have provided compelling evidence for the role of krill as a potential transfer agent of the phycotoxin DA to higher trophic levels. It is difficult to record all deaths because not all of the victims will wash ashore and the authors do point out that there is still much to learn about the transmission of DA to higher level consumers and the affects DA may have to krill behaviour, development and or mortality.

A Review of: Sibel Bargu, Christine L. Powell, Susan L. Coale, Mark Busman, Gregory J. Doucette, Mary W. Silver. (2002). Krill: a potential vector for domoic acid in marine food webs. Marine Ecology Progress Series. 237, 209-216.

Shading Reduces Progression of Coral Disease

Many studies attribute the global phenomenon of increased coral disease outbreaks to an increase in novel pathogens and their virulence in warmer waters. Alternatively, the compromised-host hypothesis (CHH) considers that coral disease outbreaks are becoming more frequent because environmental stressors are compromising the health of the coral host, leaving it more susceptible to infection from pre-existing pathogens as well as novel ones. In both explanations thermal stress is key as the CHH suggests that bleached corals (bleaching being a response to thermal stress) are far more susceptible to infection than those which still have their zooxanthellae; and pathogens generally rely on increased temperatures to increase their virulence. The paper makes this link between bleaching and infection with relation to high irradiance stress as this can upset the photosystems of the symbiotic zooxanthellae, consequently leading to bleaching. Therefore the authors hypothesise that when irradiance stress is reduced; the corals will be less prone to bleaching and so less prone to disease, thus disease progression will be slower.

The massive coral Colpophyllia natans was used to test this hypothesis and several colonies suffering from white plague were shaded. This shading reduced UV radiation by 80% and photosynthetically active radiation (PAR) by 40%, which was thought to reduce stress by allowing corals to down-regulate photoinhibition; a process which builds up damaging reactive oxygen microbe species of coral symbiont. Therefore, overall oxidative stress was reduced in the coral holobiont. I think it is important to mention here that the authors refer to the coral holobiont, not just the microbial community or the coral itself. Many of the papers I have read have focussed in on one aspect of the holobiont, which I understand in necessary for a detailed investigation, yet have failed to properly relate their findings back to the symbiosis as a whole.

Significant results were produced from this study as disease progression was markedly reduced in shaded corals, whereas it increased in unshaded corals. However, the authors note that in no case did disease progression stop all together. Even so, this is still a really important finding as it shows us that there could be a way of helping corals to recover and at the very least, reduce the effects of disease. Furthermore, the corals were only observed for 10 days after shading began so perhaps if the trials were run for longer, disease progression would have ceased and perhaps regressed.

The mechanisms behind these findings were unexplained although several ideas were suggested. The reduced radiation could have alleviated stress, in accordance with the compromised-host hypothesis, therefore reducing disease susceptibility. Following on from this, it may be that reduced irradiance reduced thermal stress also, and so the antimicrobial agents produced by the coral and its symbionts (which are usually retarded by heat) may have been able to keep being produced continue working. Finally, it may be that pathogen virulence is coupled with irradiance and therefore the lack of it reduced their activity.

This list of possible explanations demonstrates that far more work could be done to better our understanding of this topic as it could be replicated and run for longer and with other diseases. It could also ease our minds on the subject of rising sea levels damaging corals as if the water were deeper, less light would penetrate to the corals, perhaps mimicking the results found here.

A review of: E. M. Muller and R. van Woesik (2009) Shading reduces coral disease progression. Coral Reefs 28: 757–760

Thursday 29 December 2011

Kill A Whale?

This paper demonstrates the broad applicability of using pan viral microarray-based diagnostics to determine potential pathogens that were not initially considered in the deaths of marine mammals this paper demonstrates this by the discovery of West Nile virus (WNV) as the cause of death in a killer whale.

In 2007 a 14-year-old male killer whale at the Marine Park in San Antonio died without notable premonitory signs. After conventional diagnostic assays were performed the final diagnosis was septicaemia secondary to a primary viral infection that caused swelling of the brain. To determine the organism responsible, DNA micro-arrays with highly conserved sequences from > 1000 viruses were selected to screen for known and novel viruses. The results from these assays demonstrated a significant homology with viruses of the family Flaviviridae; in particular West Nile virus. Reverse transcription PCR primers targeting WNV were then used to confirm the microarray results which yielded a sequence with a 99% identity and hundred percent acid identity to the WNV strain OK 03. This diagnosis of WNV was further supported by performing an immunohistochemical staining on brain tissue which demonstrated abundant WNV antigen. These findings broaden the known host tropism of WNV to include cetaceans in addition to the previously known pinnipeds.

The authors also evaluated WNV exposure within the same cohort as well as a geographically distant cohort of whales by using serologic testing. They found the serum from the affected whale and its five cohort killer whales from the San Antonio Park which have regular contact with each other gave positive results. Five other whales were tested in Orlando which had no contact to the whales in San Antonio and were found to be negative for West Nile.

The authors of this paper suggest that health evaluations of free ranging and captive cetaceans should include WNV serology to assess exposure rates. Although this report focuses on killer whales' "loafing" behaviour (which would allow a mosquito bite) it is also seen in many coastal dolphins thus exposing them to possible WNV infection. Potential viral shedding can occur in many ways and, until we know the implications of this infection in Marine animals, WNV should be considered as a cause of death in marine mammals.

Review of : St. Leger J, Wu G, Anderson M, Dalton L, Nilson E, Wang D.; 2011; West Nile virus infection in killer whale, Texas, USA, 2007; Infect Dis; Accessed DEC/2011; http://dx.doi.org/10.3201/eid1708.101979

Dancing for Food in the Deep Sea: Symbiosis in a New Species of Yeti Crab

A review of: Thurber A.R., Jones W.J., Schnabel K. (2011) Dancing for Food in the Deep Sea: Bacterial Farming by a New Species of Yeti Crab. PLoS ONE 6(11): e26243. doi:10.1371/journal.pone.0026243

Hydrothermal vents and cold seeps are a good place to look for examples of chemoautotrophic bacterial symbiosis. As understanding of these reducing systems widen, new species are being constantly revealed. In 2005, a new family of crab were discovered at a hydrothermal vent. The individual was named Kiwa hirsuta and it had chelipeds, or claws, that were covered in dense setae with epibiotic bacteria which led to the common name, ‘Yeti crab’. In 2006, a second species of the Yeti crab, Kiwa puravida n. sp, was discovered, in a methane seep in Costa Rica, which is formally described in this paper. However, I will focus on the interesting example of symbiosis that is described in this paper.

Despite these discoveries, how epibiont bearing crustaceans harvest their symbionts has remained largely elusive. Only the crab, Shinkaia crosnieri, has been observed scraping off its epibiotic bacteria which it transfers to its mouth. Yet, even the importance of this is unknown. A key aspect of this type of mutualistic symbiosis (farming) is the direct transfer of energy from the symbionts to the host. This can be shown through biomarker analysis, and carbon isotopic and fatty acid biomarker analysis are used in this paper.

Morphological and molecular data suggest that Kiwa puravida n. sp is a new species of the Kiwaidae family with high similarity in 18s rRNA sequence (98%) to the related Kiwa hirsuta. Additionally, 16s rRNA bacterial gene sequences were collected from Kiwa puravida n. sp including ε, δ and γ-proteobacteria. γ-proteobacteria were similar to the other epibionts collected from Kiwa hirsuta, Shinkaia crosnieri and Rimicaris exoculata (shrimp), being 97% and 98% similar across the four taxa. Phylogenetic analysis of the proteobacteria found that two clades of ε-proteobacteria were unique to Kiwa puravida n. sp and that, interestingly, phylotypes of some ε-proteobacteria were more similar to phylotypes of other species than those from the same host.

Phylogenetic analysis found that there is an epibiotic fauna that specialises in these reducing systems and the presence of closely related ε and γ-proteobacteria on a variety of hosts suggests multiple horizontal transmissions by these epibionts. Additionally, as the hosts are likely to inhabit more than one site, there is the suggestion that these similarities mean free-living bacterial stages of these symbionts.

This new species, Kiwa puravida n. sp was not observed scavenging food as was suggested for its relative Kiwa hirsuta. Biomarker analyses found an abundance of 16:1 FA (monounsaturated fatty acids) alongside the presence of 16:2 FA which along with an isotopic composition that indicates chemosynthetic nutrition, suggests that Kiwa puravida n. sp ’s main food source is its epibiotic bacteria.

Kiwa puravida n. sp has both morphological and behavioural adaptations to harvest its symbionts. A set of specialised comb row setae on its 3rd maxilliped, a mouth appendage, is used by the crab to scrape bacteria off the whip-like barbed setae which adorn its chelipeds, sternum, and pereopods (legs) and transfer them to its mouth. But, a species must also facilitate the growth of its epibionts in order to farm them. Kiwa puravida n. sp does this by providing an attachment substrate in the form of these setae but it also moves its chelipeds continually which increase the epibiont productivity. How? Chemoautotrophic symbionts require access to oxygen and reduced compounds such as sulphide or methane from the seep. During carbon fixation, a boundary layer is formed resulting in the depletion of one or more of these solutes, limiting productivity. The behaviour of the crab moving its chelipeds removes these boundary layers and so restores productivity.

Wednesday 28 December 2011

Returning to life after Christmas

.... eventually emerged from Christmas torpor and impressed to see some of you have been blogging right through the festive season. (Although pleased to see there were none on Christmas Day itself, that would be rather sad). I will get round to reading these soon.

I got an alert about this intersting new paper on the Yeti Crab. Anyone want to review this one?
Thurber AR , Jones WJ , Schnabel K , 2011 Dancing for Food in the Deep Sea: Bacterial Farming by a New Species of Yeti Crab. PLoS ONE 6(11): e26243. doi:10.1371/journal.pone.0026243

Colin

Bringing to attention the importance of extracellular enzymes; and their role in the carbon pump

Cunha, A., Almeida, A., Coelho, F. J. R. C., Gomes, N. C. M., Oliveira, V., & Santos, A. L. (2010). Bacterial Extracellular Enzymatic Activity in Globally Changing Aquatic Ecosystems. Applied Microbiology, 124-135.

This paper is not strictly on marine microbes, but relates to extra cellular enzyme and their potential change in activity due to globally changing aquatic ecosystems (as the title suggests). After my last blog about the discovery of unique properties of Em2L8 it spurred my thinking in relation to bioremediation and the potential of extra cellular enzymes in this context. This intrigue led to the discovery of the above paper, so yes, I am doing a review, of a review.

Heterotrophic microorganisms are key to the microbial loop process; nutrient cycling and carbon flow through aquatic food webs. The main sources of organic matter to the microbial loop are exudates from phytoplankton, algae and bacterial cell material due to grazing. In addition to this river flow and terrestrial deposition also provide organic matter input. These factors combined are a large reservoir of energy for heterotrophic microbes.

Bacterial cell membranes are semi permeable by passive transport, and are mainly restricted to the passage of simple chemical compounds (low molecular weight). The architectural properties of gram negative and gram positive cell wall constructs are different, but are also similar in regards to their membrane selectivity of specific chemical compounds. Having stated this it is known that gram positive cell wall is not as discriminating as gram negative. Particular organic matter and dissolved organic matter consist mainly of high molecular weight compounds. To allow transport across outer membranes of cells large molecular substrate complexes must be hydrolysed outside of the cell. This step is mediated by extracellular enzymes allowing heterotrophic bacteria to gain nutrition from the diverse range of organic substrates that would otherwise be in an impermeable form. Taking this into consideration any disruption to the functioning of extracellular enzymes would potentially detrimentally impact the rate of metabolism of bacteria, consequentially limiting nutrient and remineralisation cycling. If this occurred to the carbon cycle, it could increase the Co2 in the atmosphere and potentially accelerate global climate change.

The extracellular enzymes production and activity is specific to the hydrolytic action e.g. oxygenases and peroxidases. Some extracellular enzymes are not truly separate from the host and are bound to the exterior wall or contained in the peri-plasmic space. True extracellular enzymes catalyse reactions in a detached form from their producers.

Cunha et al. highlight that the identities of particular members of mixed assemblages of microbes are capable of producing mixtures of extracellular enzymes and their structure and action is largely unknown. This is due to the fact most molecular analysis focuses on rRNA sequences, that provide little information about the degradable capabilities of uncultured microorganisms. This indicates to me that further methods need to be developed for investigating the degradable faculties of microbes or methods of analysing bacteria and substrate complexes such as marine snow. It is known that marine snow aggregates are colonized by heterotrophic microbes which express high levels of hydrolytic processes. As it is thought that they are epicentres of carbon remineralisation. Ocean acidification may have an effect on bacterial extracellular activity and negatively affect primary producers, which would affect the heterotrophs and consequentially inhibit the vertical transport of particular organic carbon to deeper ocean layers (biological carbon pump). Due to the severe implications of such an event, it seems obvious it is pivotal to further investigate the complexities of biogeochemical cycling and the consequences climate change may have on extra-cellular enzyme activity. Keeping people up to date on the developments in understanding of biogeochemical cycling responses to global climate change, in theory would lead to more effective environmental management; therefore I would advocate further research in this filed.

Disease in Marine Systems

Parasites and pathogens use many marine organisms as hosts; the resulting mortalities can lead to a range of changes, not only in the host population but in the habitat, resultantly altering community structure. Human impact escalates the number of stressors affecting marine ecosystems, making it increasingly important to understand these diseases and the timing of their outbreaks. Anthropogenic changes, such as overfishing and the introduction of terrestrial diseases into the marine environment alter community structures. Furthermore global issues, such as climate change and pollution are thought to increase disease prevalence, in both terrestrial and marine ecosystems. Previous reviews have suggested that over the past three decades, the increase in disease outbreaks has been coupled with climate change; however, lack of data on marine communities is preventing direct testing of this hypothesis.

The authors of this paper developed a proxy method to test a prediction of the increasing disease hypothesis: that reports of disease in scientific literature have increased since 1970. All online literature from ISI web of science, and quantified reports of disease in natural populations of marine organisms were searched from 1970-2001. The investigation looked at nine different taxonomic groups: turtles, corals, mammals, urchins, molluscs, sea grasses, decapods, sharks/rays and fishes, and detected important trends of disease. This method was the first, unbiased, quantitative use of normalised trends in literature, to investigate an ecological hypothesis.

The method showed an increase in total disease reports for all groups, however when the data was normalised an increase was only shown in turtles, corals, mammals, urchins and molluscs. No significant trend was shown in sea grasses, decapods, sharks/rays and counter to the hypothesis a decrease in disease was shown in fish. The results were tested for author bias, and no significant changes occurred in the data reviewed. The proxy method works on the assumption that: actual change in disease overtime will be accompanied by a corresponding change in publishing frequency by scientists. The method has been shown to work before; however it is limited by the inability to distinguish between an event that did not occur and an event which was not reported.

Overall two implications can be taken from the data collected. Firstly, the reported increases of disease were not due to increased study by marine biologists; and secondly, factors such as global change can have complex effects on disease. Temperature change has been linked to an increase in turtle and mollusc disease; along with coral bleaching, while bioaccumulation of toxins in mammals, due to pollution, has been shown to increase susceptibility to disease. Finally the decline in fish diseases has been found to correspond with the reduction of the fish population, due to overfishing. This is thought to have reduced the abundance of disease transmission and lead to the documented and observed decline in fish parasites. Finer scale investigations into disease and its impacts on each taxonomic group are required to fully understand the impacts and causes of marine diseases.

Review of: Ward, J.R. Lafferty, K.D. The Elusive baseline of marine disease: Are disease in ocean ecosystems increasing? PLOS Biology, 2 (4) pp. 542-547.

Not Just Zooxanthellae...

Scleractinian (or hard corals) are an integral part of the worlds’ coral reefs, and are thought to be formed as a result of the interactions between these corals and their symbiotic zooxanthellae and other microorganisms which are numerous but their function is yet to be confirmed. It has been suggested that microorganisms residing on the surface of the coral may provide protection against pathogens through inter-specific competition or by the secretion of antibiotic substances. They may also be providing the coral with nutrients that are not being supplied by the symbiotic zooxanthellae, such as nitrogen and phosphorous.

A mucous-rich micro-layer extends a few millimetres above the surface of Scleractinian corals where nutrient transfer occurs. This layer is also thought to provide a growth medium for these microorganisms, and white patches have been recorded covering the surface of some of the larger species. These patches are caused by unicellular heterotrophs belonging to the stramenopile group.

Favia Corals from the gulf of Eilat and the red sea were sampled in 2005-2006 from water 1-6 metres deep. The mircobiota from these samples were cultured, cloned and their DNA amplified using PCR. The DNA was then sequenced to determine the species of microbes present in the white patches.

DNA profiling revealed that 90% of the microbes present in these patches were stramenopiles of the Thraustochytriidae family, while several other families shared the remaining 10%. From the 90% majority, 50% were unable to be identified to genus level, while the other 50% were found to be one of 3; Aplanochytrium (23%), Thraustochytrium (17%) and Labyrinthuloides (10%).

In contrast to previous studies that found negative coral-protist associations, the results of this study suggests that these white patches do not cause any visible damage to the coral hosts. The microorganism family Thraustochytriidae are known to degrade a wide variety of organic substrates. It is possible that this process aids symbiosis with the corals by providing them with smaller, more easily absorbed organic compounds.

The authors of this paper consider the relationship between the Favia coral and the microorganisms that cause the white patches to be mutualistic, as the microbes are thought to live off the coral mucus, and in turn provide the coral with pre-processed small organic compounds. The relationship is however poorly understood at present, and is in need of considerable further study.

A review of N. Siboni, D. Rasoulouniriana, E. Ben-Dov, E. Kramarsky-Winter, A. Sivan, Y. Loya, O. Hoegh-Guldberg, A. Kushmaro (2010) Stramenopile Microorganisms Associated with the Massive Coral Favia sp., The journal or Eukaryotic Microbiology, 57 236-244

Monday 26 December 2011

A Lake Under the Sea: Anoxic Animals and Anoxic Protists

A couple of posts by Jelena and myself have looked at the deep hypersaline anoxic basins (DHABs) of the Mediterranean and the prokaryotic and eukaryotic life found within them. This paper, although not purely microbiology, does make some interesting comparisons between the first obligate anaerobic animals and single cellular anaerobic protists. These animals were not just the first anoxic animals to be discovered in the DHABs, but also in the world in general.

In a number of expeditions to a DHAB in the Mediterranean Sea called L’Atalante, Danovaro et al (2010) discovered and examined three new species of animal that belonged to the phylum Loricfera. These three metazoans were found living in the sediments of the brine lake and were less than a millimetre in size. Two other phyla were found but staining methods suggested that they had been dead for some time, possible victims of the DHAB’s unforgiving environment. This study concluded that this was the first example of purely oxygen independent animals to have been discovered. Other metazoans have been found living in anoxic environments before but only at certain stages of their lifecycle, these three species however appear to live their entire lives in 0% oxygen.

Mentel and Martin (2010) examined the results of the previous study in more detail and made many cellular comparisons with unicellular protists. Transmission electron microscopy (TEM) revealed a complete lack of traditional eukaryotic mitochondria, instead there were organelles which resembled hydrogenosomes. Instead of using oxygen as a terminal electron acceptor, hydrogenosomes pass electrons from pyruvate to 2H+ to produce H2. These organelles have never been seen in animals before but are found in members of the unrelated unicellular phyla Amoebozoa and Metamonada. Surrounding the hydrogenosomes were what appeared to be rod shaped cells. The authors suggested that they might be methanogen achaea in a endosymbiotic relationship with the cells as this is common with protists that contain hydrogenosomes. Sadly, most of the research has so far been carried out with TEM and so the details about the hydrogenosomes and the rod shaped cells remain a mystery.

Mental and Martin (2010) discuss the fact that recent evidence shows that deep sea environments were anoxic until about 580 million years ago, about the same time that large animals start to appear in the fossil record. They passionately suggest that anaerobic animals might have evolved from anaerobic protests similar to the ones seen today but when oxygen levels increased it gave way to more productive aerobic animals that rapidly increased in size during the Cambrian explosion. They concluded from this idea that the animals discovered in L’Atalante have evolved from anaerobic unicellular eukaryotes millions of years ago and are not descended from aerobic animals that adapted to the DHAB after its formation.

I think it will be fascinating to see what research into the metabolism and genomes of these organisms will uncover. I imagine such studies will indeed take place as it is a very significant discovery but I cannot find any articles on it just yet.

A Review of:
Mentel, M and Martin, W (2010) Anaerobic animals from an ancient, anoxic ecological niche. BMC Biology. 8, 32.

Additional Reference:
Danovaro, R. Dell’Anno, A. Pusceddu, A. Gambi, C. Heiner, I and Kristensen, R (2010) The first metazoan living in permanently anoxic conditions. BMC Biology. 8, 30.

Using the ‘Snot sucker’ method to discover bacterial assemblages within compartments of the coral holobiont

It is accepted that corals have a diverse, and host-species specific microbiota; however it is poorly documented as to how they are organised within the coral holobiont. A number of studies have been carried out, all using a variety of techniques such as crushed coral, scraping of surface, airbrushing, swabs and milked mucus to name a few, which, although they have given some fascinating results are not highly regarded by the authors of this article as they believe that these methods are easily contaminated. For this reason, when trying to understand the differences, and organisation of assemblages within the coral holobiont, they used a new method for removing layers of bacterial habitats within the coral naming the apparatus used the ‘snot sucker’.

Corals were collected, mounted on a screw cap system and returned to the reef until collection. Sample collection took place at a reef flat by Heron Island (23o27’S, 151o55’E).Samples of the surrounding water column and sediment were also collected as a potential supply of coral associated bacteria.

The snot sucker is a ‘50ml falcon tube, with two 3-way valves grafted onto it, one at the bottom and one at the top. A 60ml syringe, with tubing, was attached to the bottom stopper valve allowing filtered water to be flushed over the coral and loosely attached surface mucus layer (SML) collected by the top valve.’ The snot sucker was used both in situ and in the lab, and previously used methods, mentioned earlier (airbrushed-tissue layer, collection of tissue slurry etc.) were tested by the authors too.

DNA fragments were visualised using DGGE with Sybr ® Gold and a UV transilluminator. Results were statistically analysed using a one-way permutation analysis of variance (PERMANOVA), analysis of contribution to similarities (SIMPER), and results are laid over a 2-D plot using a non-metric multidimensional scaling (MDS) and based on a similarity profile (SIMPROF). They looked for differences in both, bacterial assemblages within compartments of the coral holobiont and also the techniques used. From the Shannon-Weiner diversity index, the diversity varied significantly between different techniques; the snot sucker being the greatest in diversity and swabs and milked mucus being the lowest. The worst technique employed appeared to be the swabs, with average similarity between that and other sample types seldom exceeding 25%; they came to the conclusion that it may include contamination from other sources as there was a 35% similarity to sediment for example.

The study demonstrated that bacterial communities do differ between compartments of the holobiont; with significant differences being shown between all the coral compartments and the surrounding environment samples; supporting the idea that coral harbour and maintain a distinct microbial flora. Within their discussion the authors explain where possible sources of bacteria, which may colonise the SML arrive from e.g./ faecal matter, water column etc. and how the fact that the SML is significantly different in bacterial diversity compared to the surrounding environment showed that bacterial communities will occur even on what may be considered as the harshest environment offered by the coral, as it is constantly affected by hydrodynamic processes and the chemical make-up differs from coral to coral.

A review of:

Sweet, M. J., Croquer, A. And Bythell, J. C. (2010). Bacterial assemblages differ between compartments within the coral holobiont. Coral Reefs. Vol. 30. (1) pp. 39-52.

Found at: http://www.springerlink.com/content/e26284260u337u4h/export-citation/

Accessed on: 22/12/2011