Recreational Water Quality

This paper investigates the relationship between illnesses commonly associated with swimming, and faecal indicator bacteria as determined by qPCR methods, by comparing estimates of faecal indicators with incidences of illness. The study took place at three marine beaches, located in Mississippi, Rhode Island and Alabama, USA. Each of the beaches is sited within 7 miles of a treated sewage discharge outfall, and although water quality varies at each, they all comply with water quality guidelines. The beach in Mississippi was sampled in 2005 whereas the other two beaches were sampled in 2007.

Water samples were collected in waist-high and shin-high water from three transects, which included the swimming area. Environmental conditions were also recorded at the time of sampling, as well as information relating to use of the beach (numbers of people and animals etc.). The samples were tested for total Enterococcus spp. and total Bacteroidales spp. by qPCR, and samples taken in 2007 were also tested for subgroups of Bacteroides and Clostridium spp. Samples from 2007 were also tested for F+ coliphage using two methods; a culture and latex agglutination (CLAT) assay, and a culture-based EPA method.

Visitors to the beach on sampling days were asked to participate in the study. Eligible participants were asked to complete questionnaires regarding contact with water and sand, and ten to twelve days later a telephone interview was conducted to determine whether gastro-intestinal (GI), skin, respiratory, eye or ear problems had occurred following their visit. 6350 visitors participated.

The occurrence of GI illness was associated with an increase in exposure to estimates of Enterococcus spp. and Bacteroidales spp. When highest levels of Bacteroidales were present, 12% of swimmers reported illness compared to 6% of non-swimmers. Positive relationships were found between incidences of GI illness and estimates of subgroups of Bacteroides and Clostridium spp., and estimates of F+ coliphages, but these were not statistically significant.

The study is the first to find a relationship between incidences of GI illness and estimates of faecal indicator organisms, as estimated using qPCR, at marine beaches. The findings suggest that qPCR methods give a good indication of poor water quality and the effects on the health of visitors. They also support the need for more rapid testing of bathing waters.

A review of: Wade, T.J., Sams, E., Brenner, K.P., Haugland, R., Chern, E., Beach, M., Wymer, L., Rankin, C.C., Love, D., Li, Q., Noble, R. and Dufour, A.P. (2010) ‘Rapidly measured indicators of recreational water quality and swimming-associated illness at marine beaches: a prospective cohort study’, Environmental Health, 9:66.

the blue mussel as indicator of human faecal pollution

Marine benthic organisms are capable of filtering large amounts of water and hence of accumulating substances and other organisms, especially the micrometer range size, from the surrounding seawater.

In this study the blue mussel M. edulis has been used for investigations on faecal pollution.

The aim was to detect the uptake, accumulation and persistence of two human specific Enterococcus species -that are often the most abundant species in wastewater- in the blue mussels.

To this purpose, the authors investigated the presence of E. faecium and E. faecalis in a natural population of mussels collected at 4 sites potentially affected by human faecal pollution, as they were located less then 1km from discharges of local wastewater treatment plants.

To study the uptake and accumulation of bacteria mussels have been exposed to a culture of E. faecalis in a microcosm experiment.

To estimate the persistence of bacteria, contaminated mussels were tranferred to aquaria with flowing seawater without bacteria for depuration and then transplanted to coastal sites to study in situ persistence of human associated Enterococcus.

The presence of the two Enterococcus species was detected targetting two specific molecular markers:

• esp gene which encodes the enterococcal surface protein, a putative virulence factor in human associated E. faecium strains
• M66 gene which encodes a protein in E. faecalis with unknown function but it may serve a functional role during survival in human hosts

The esp and M66 markers were targetted in sampling of water and mussels at three coastal sites; interestingly, at all sites bivalves were enriched in E. coli and E. species compared to the surrounding water, and in some case esp was not detected in water samples at all.

In laboratory experiments the authors found that after 24h of exposure to contaminated water, the culturability of the E. faecalis declined exponentially, and culturable cells were not detected after 21 days of incubation. At the same time the concentration of E. faecalis in mussels was a twofold greater than in the surrounding seawater, suggesting that the uptake and bioaccumulation of bacteria was massive.

A substancial decline in the concentrations of E. faecalis in M. edulis occured after the initial 24h of depurationin aquaria.

The in situ experiment on depuration showed that the human associated E. species in mussels were detectable for at least 96h.

These results suggest that M. edulis can accumulate and retain elevated concentration of putative source specific E. species, hence these filtrating bivalve molluscs may be considered as additional targets in microbial source tracking studies.

Additionally, noncontaminated mussels may be transplanted to contaminated sites for time-integrated accumulation of markers relevant for microbial source tracking.

A review of: Roslev, P., Iversen, L., Sønderbo, H., Iversen, N. and Bastholm, S. (2009), Uptake and persistence of human associated Enterococcus in the mussel Mytilus edulis: relevance for faecal pollution source tracking. Journal of Applied Microbiology, 107: 944–953. doi: 10.1111/j.1365-2672.2009.04272.x

Apoptosis and Coral Bleaching

A review of: Tchernov D, Kvitt H, Haramaty L, Bibby TS, Gorbunov MY, Rosenfeld H, Falkowski PG (2011) Apoptosis and the selective survival of host animals following thermal bleaching in zooxanthellate corals. Proceedings of the National Academy of Sciences, 108 (24): 9905-9909.

Coral bleaching is a global phenomenon that occurs when environmental factors such as increases in temperature and/or solar irradiance causes the expulsion, digestion or loss of pigmentation of zooxanthellae which reside in the endoderm of symbiotic cnidarians. Periodic bleaching episodes can cause cnidarian mortality and have significantly reduced coral abundance in tropical reef ecosystems, however the molecular and physiological mechanisms that lead to death is not fully understood.

Apoptosis or programmed cell death has been suggested to be the pathway that leads to the breakdown of symbiosis and ultimately to the death of the host. Apoptosis is characterised by morphological changes to cells which include plasma membrane blebbing, cytoplasmic shrinkage, chromatin condensation and DNA degradation, brought about by the activity of a number of intracellular proteins in which caspases play a central role. Caspases can be up regulated by reactive oxygen species (ROS) predominantly produced from the photosynthetic activity of the symbiont and/or the mitochondrial activity of the host. Tchernov et al (2011) set out to investigate the relationship between elevated temperature and irradiance, as well as the concomitant increase in ROS production and caspase activity in a variety of symbiotic corals under controlled laboratory conditions and in situ natural conditions.

Seriatopora hystrix and Stylophora pistillata specimens were grown for several months at 26 °C and were transferred to aquaria with a temperature of 32 °C, to test for thermal stress under controlled conditions. Both corals eventually bleached but S. pistillata exhibited apoptotic morphology along with an increase in caspase activity while S. hystrix retained morphological integrity and had a decrease in caspase activity. In field experiments, Pocillopora damicornis and Montipora capitata were exposed to full natural light, low light, ambient temperature and elevated temperature conditions. An increase in caspase activity for both corals was detected at elevated temperatures in both light conditions however caspase activity was more enhanced with full sunlight exposure. Moreover, bleaching and apoptosis was curtailed upon the introduction of caspase inhibitors thus confirming the role of apoptosis in coral bleaching events.

To determine the role and source of ROS production, aposymbiotic and symbiotic M. capitata colonies were kept at 32 °C while ROS levels were measured after 11 hours. In symbiotic corals ROS production and caspase activity was higher than in aposymbiotic corals. Hydroxyl anion radicals were subsequently introduced to the symbiotic and aposymbiotic colonies kept at 26 °C under low light and 32 °C under high light, to elucidate whether ROS induces caspase activity. There was a significant increase in caspase activity observed in both colonies, although caspase activity was detected earlier in the aposymbiotic colonies while the highest caspase activity was detected in symbiotic colonies in both temperature and light regimes.

The results of the study provide insights into the physiological developments associated with bleaching and mortality processes. Different zooxanthellae clades have different lipid compositions which can impart resistance to environmental stressors that cause the uncoupling of thylakoid membranes and an associated increase in ROS production. It is hypothesised that under low light conditions, the symbiont is able to support the antioxidant defence of the host, whereas an increase in ROS production beyond a threshold can lead to the activation of apoptotic processes. Due to the different response to stress between S. pistillata and S. hystrix, thermally induced bleaching and mortality may be a host and symbiont species specific process which might explain why some corals bleach and recover, some bleach and die while some do not bleach at all. Interestingly, caspase inhibitors were able to prevent the death of the host even under high ROS production, therefore it can be speculated that therapy involving caspase inhibitors and/or anti apoptotic proteins like Bcl-2, may provide future strategies for preserving reef systems under threat from environmental perturbation.

HABs: causes and possible strategies to control them

In 2003 the USA EPA sponsored a roundtable discussion in order to develop a consensus on the relationship between HABs and eutrophication. Although the committee focused on cases study along the US coasts, the statements can be adopted also by other countries. Briefly, it has been unanimously established that :

• both cronic and episodic nutrient pollution promote the development and persistence of many HABs
• the composition -not just the quality-of nutrient pool impacts HABs
• high biomass blooms must have exogenous nutrients to be sustained
• further experimental studies are necessary to better understand the role of nutrients in HABs: the reduction in nutrient input decreases HABs, although this strategy is challenge (for many species these relationships can be complex) and extremely costly, hence developing other bloom control strategies would be better.

Many studies focusing on bloom control strategies development occurred after this meeting, and at the present day chinese institutions are strongly contributing in increasing the knowledge of this matter.

Tang and Gobler in 2011 demonstrated that the green macroalga Ulva lactuca can have a strong growth-inhibiting effects on multiple HAB species. In this experiment both fresh talli and extracts of the dried powder of macroalga were capable of lysing or strongly inhibiting the growth of 7 common HAB species through the release of heat-stable allelochemicals. Furthermore a field-based experiment proved that U. lactuca can reduce the densities of field population of A. anophagefferens.

In late 2011 a research team of Xiamen University pusblished a study showing the algicidal activity of two marine bacteria. Molecular characterization classified them to the gamma-proteobacteria subclass and the genus Vibrio and Pseudoalteromonas. These strains have been isolated and used to purify a protein (P7) that showed algicidal activity against the toxic dinoflagellate A. tamarense. P7 algicidal activity was well evident as the algal cells were lysed and cellular substances were released under visual fields of microscope.

All of these studies contribute in increasing the knowledge of toxic microalgae ecology, biology and interaction with other organisms. Two possible strategies in controlling HABs blooms were provided although more investigations are necessary in order to state their real application and possibility to be used in field: methods were very complicated and should be improved.

References:

Heisler, J., P. M. Glibert, et al. (2008). "Eutrophication and harmful algal blooms: A scientific consensus." Harmful Algae 8(1): 3-13.

Tang, Y. Z. and C. J. Gobler (2011). "The green macroalga, Ulva lactuca, inhibits the growth of seven common harmful algal bloom species via allelopathy." Harmful Algae 10(5): 480-488.

Wang, B., X. Yang, et al. (2012). "A marine bacterium producing protein with algicidal activity against Alexandrium tamarense." Harmful Algae 13(0): 83-88.

Stimulating ideas

Smith V, Brown J,
Hauton C (2003). Immunostimulation in crustaceans: does it really
protect against infection? Fish & Shellfish Immunology. 15, 71–90

This paper gives an extensive insight into the subject of
using immunostimulants to control disease. It focuses on the crustacean immune
system, how it works and how it is affected by immunostimulants to protect
against infection. After 15 years of research and compounds available
commercially as immunostimulants the authors felt that the time was right
for an appraisal of the current state-of-the-art of these substances for
crustaceans to assess how well they live up to their claimed benefits.

There is no evidence that crustaceans, or any other
protostome invertebrate, have clonally derived subsets of cells that permit
specific, adaptive and ‘memory-based’ immunity. Therefore the primary
administration of a non-virulent or attenuated antigen to provoke a protective
pool of recognition and effector cells or molecules in the event of secondary
exposure (vaccine) cannot work in these animals. Therefore immunostimulants are
an interesting way forward.

Immunostimulants receiving most attention comprise of
five main types: live bacteria; killed bacteria (bacterins or bacterial
antigen); glucans; peptidoglycans; and lipopolysaccharides (LPS). Usually these
are administered by immersion, as a dietary supplement or by injection and this
may be an important factor in determining the success of the treatment. Many of
the compounds have direct effects on various aspects of the crustacean immune
system. Glucans, LPS, bacteria and other non-self agents are known to invoke
various in vivo responses, such as change in haemocyte counts and induction of
encapsulation reactions. They also are known to induce prophenoloxidase
activation and melanisation reactions while in vitro glucans have been shown to
initiate cell degranulation and to enhance Phagocytosis. Research has further
demonstrated that phenoloxidase activation by glucans or other non-self
molecules generates a range of immunoactive agents and activities, including
peroxinectin and reactive oxygen species. However administration of immune
stimulants might, at best, be ineffective because of the natural regulatory
molecules within the immune system, or, at worse, be detrimental as has been
reported.

A large number of articles have been published which
report the beneficial effects of immunostimulatory substances. However the
authors of this paper have rigorously inspected these and found that, in a
number of instances, the presented data do not support the conclusions reached.
In several cases poor experimental design and the absence of any statistical
analysis limits the validity of some conclusions. Also, acknowledged by many
authors, is the poor reproducibility of the response for individuals of the
same species within the same treatment group. The inconsistent nature of any
response exhibited by individuals of a single species will severely impede the
efficacy of any marketed stimulant as well as the definition of appropriate
dosing regimes and any ‘booster’ treatments. The problems with the literature
pointed out by the authors did not seem to me that problematic, just that the
authors of the studies had exaggerated results; as in the case of most
scientific literature these days in order to get published.

In conclusion there is a reasonable degree of doubt about the
efficiency of stimulants due to the lack of unequivocal evidence and convincing
reasons why the use of such compounds over prolonged periods might be
detrimental to the host. There is an urgent need for proper large-scale
evaluation of the efficacy of putative immunostimulants through standardized
trials where analyses should include assessment of their ability to up-regulate
the expression of genes encoding defense proteins, as such studies are
necessary to elucidate the mode of action of any promising compounds and
thereby help to refine their use. Findings must be related to functional
end-point assays. Multi-gene analyses, for example, by micro-array technology
are desirable. The optimal dose, route and treatment time for each stimulant at
all appropriate life stages of the target species still need to be found. These
analyses will reveal if treatment with immunostimulants is appropriate for crustaceans,
and for how long protection might reasonably be expected to last.

Detection of the Hepatitis A virus in marine coastal waters

Very often waterborne illnesses are due to viruses of human faecal origin. One such virus, and the topic of this paper, is the Hepatitis A virus (HAV), which is commonly present in raw sewage and the cause of acute hepatitis worldwide. Its primary transmission is through the faecal-oral route where outbreaks occur from contaminated food or water. Studies have shown that HAV is capable of surviving outside of a host and can stay infectious for at least nine months in an aquatic environment – so it is important to understand as much as possible about it.

Despite frequent incidence worldwide, little literature is available on HAV levels in marine waters because detection of HAV is very challenging and time-consuming, due to the poor growth of wild-types in vitro. The aims of the authors were to i) develop a sensitive and rapid HAV detection method to monitor possible sources of contamination during sporadic HAV infection outbreaks and ii) identify the major HAV genotype present in Hong Kong coastal waters.

Seawater samples were collected from seven sewage-polluted sites in Hong Kong. The authors developed a sensitive and quantitative TaqMan-based PCR method (which targeted the 5’-noncoding region of HAV), this quantified the HAV particles present in the samples. Of the seven sites, HAV was detected in four; the levels detected ranged from 416 – 1028 HAV particles/L. Total Escherichia coli counts for each sample were also enumerated on TBX medium. The E. coli densities observed showed no correlation of any significance between HAV and E. coli counts. For example, at the Kwun Tong Pier, there was a high count of 118,800 E. coli/100mL, but HAV particles were barely detectable.

Use of phylogenetic analysis of 10 PCR clones from each of the HAV-positive sites indicated that the human HAV genotype IB is the most widely distributed type in Hong Kong waters.

In conclusion the stand alone use of total E. coli counts as a faecal indicator needs to be addressed. However, HAV quantification together with the use of total E. coli counts may serve as a tool to assess water quality.

This paper is short and to the point, the only problem I had with reading it was the lack of sections (introduction, methods, discussion etc.). I felt I was getting bogged down in a lot of the small details, which I would have preferred to read at the end. The conclusions from this paper are the complete opposite to another paper (Gersberg et al., 2006), which suggests E. coli can be used to indicate the presence of HAV; I feel more research needs to be done to understand why such different results were obtained. But on the whole this is a paper that will lead to further research on the matter of faecal contamination indicators.


A review of:

Yang, N., Ling Ho Chu, D., Man Lai Wong, M., Qi, H., Shui Sun Wu, R. and Yuen Chong Kong, R. (2011) Major human Hepatitis A virus genotype in Hong Kong marine waters and detection by real-time PCR. Marine Pollution Bulletin 62 2654-2658.

Probiotics: challenges and outlook

A Review of: 'Probiotics in Aquaculture: Challenges and Outlook'. 2008. By Wang, Li and Lin. Aquaculture 281, pp. 1-4

Many modern aquaculture operations require hydrobionts to be cultured at high densities, resulting in elevated stress levels, disease and deterioration of environmental conditions, leading to economic losses. In recent decades, disease prevention and control have led to substantial increase in the use of chemical additives and Vetinary medicines, however their wide spread use is now recognized as causing environmental problems and so the need for alternative techniques is increasing. The contribution of probiotics is considerable and widely accepted, having been introduced to molluscan, shrimp and fish farming as feed additives.

Intensified research efforts in recent years confirm the importance of microbials in the digestive tract. Compared to water, the digestive tract of hydrobionts is an ecosystem far richer in nutrients and therefore more favorable for growth of the majority of bacteria. Gastrointestinal bacteria take part in the decomposition of nutrients, provide the macroorganisms with physiologically active materials, such as enzymes, amino acids, and vitamins, and play an  important role in the resistance to infectious diseases, producing antibacterial materials pathogenic to 'harmful' bacteria. Antiviral effects have also been reported although the exact mechanism by which these bacteria do this is not known.

To be effective and confer health benefits, probiotic cultures must be able to retain their probiotic properties after processing, and with sufficient numbers surviving during shelf life/ storage for efficacy. The retention of high viability during preparation and storage presents particular challenges and can be regarded as a ‘major bottleneck’ in commercial probiotic production. Stability is also critical to guarantee the efficacy and can be influenced by various factors, including species, strain biotype, water activity, temperature, pH, osmotic pressure, mechanical friction and oxygen. Consequently, special attention and techniques are needed during probiotic production.

Safety considerations have been neglected for a long time but are now taken into account for the development and marketing of probiotics: new strains are carefully assessed and evaluated for both safety and efficacy prior to incorporation into products and not assumed to share the historical safety of tested or traditional strains. The safety profile of a potential probiotic strain is of critical importance in the selection process and its testing should include the determination of strain resistance to a wide variety of common classes of antibiotics such as quinolones and macrolides, and subsequent confirmation of non-transmission of drug resistance genes of virulence plasmids. Modern molecular techniques should be applied to ensure that the species of probiotics used in aquaculture are correctly identified, for quality assurance as well as safety.

I thought this was a great little paper. The authors clearly support the use and need for probiotics in aquaculture especially instead of antibiotics. However they are clearly concerned about a few main points and discuss them clearly, such as their safety, thorough testing before application and the growing need to further identify the mechanisms by which they work and further understanding of the composition and interactions that occur between the microbial populations in the GI tract.

[Could be quite a useful paper for an exam question asking for a critical evaluation of probiotics??]

more ideas to control disease in aquaculture including quorum sensing

Defoirdt, T., Sorgeloos,
P., Bossier, P. (2011) Alternatives to antibiotics for the control of
bacterial disease in aquaculture. Current Opinion in Microbiology, 14:251–258

As we know,
aquaculture is the fastest-growing food-producing sector worldwide and disease
outbreaks are considered to be a significant constraint to the development of
the sector. This paper reviews some ideas for disease control that are needed
because the use of antibiotics has been constrained due to concerns of resistance
and contamination. The main problem is resistance of useful antibiotics passing
to human pathogens.

An alternative strategy is the use of bacteriophages and
there have been many blogs on this subject. The advantages of using
bacteriophage therapy include their specificity (although this paper mentions
that some phage are effective to different family microbes) and therefore the
reduced risk of compromising the natural microbiota of the aquaculture
ecosystem. A further concern, that will be important to test,
is whether they carry any virulence genes and whether they would be safe to
use.

Another potential strategy is to target specific genes of
pathogenic bacteria by inhibiting their replication. This paper has one example
indicating the idea has not been fully researched yet but has potential.
Bacteria of Vibrionaceae all contain RctB as an initiator of
replication of their smaller second chromosome, which lacks any similarity to
characterised initiators. In a recent study, Yamaichi and colleagues identified
a compound, vibrepin, that inhibited RctB and had potent cidal activity against
V. cholerae and inhibited growth of all Vibrio species tested, including the
aquaculture pathogens Vibrio parahaemolyticus and Vibrio vulnificus.

Inhibiting growth of pathogens rather than killing them
may reduce the risk of resistance to a treatment. Short-chain fatty
acids are currently used in commercial (terrestrial) animal diets to control
pathogens such as Salmonella. They decrease the
intestinal pH, resulting in a more beneficial microbial community in the
digestive tract such as lactic acid bacteria. Recent research showed that these
compounds also inhibit the growth of pathogenic Vibrios. A major disadvantage
of using short-chain fatty acids in aquaculture feeds is that they would leach
into the culture water and consequently, high doses would be needed to retain
sufficient activity. To combat this, this paper hypothesizes that the
poly-b-hydroxybutyrate (PHB) polymer from Brachymonas bacteria can be added to
the rearing water. This is then degraded into b-hydroxybutyrate in the gut of
the aquacultured organisms and the release of this short-chain fatty acid
protects the host from pathogenic bacteria.

Another disease
control method is to reduce the virulence of pathogens by inhibiting virulence
gene expression. Quorum sensing is involved in the activation of virulence of
pathogens. Detection of signal molecules can be disrupted and stopped using
compounds found in nature from algae such as
D. pulchra, corals and sponges. Bacteria, such as Bacillus species, can degrade signal molecules in
order to control the levels. Results obtained so far indicate that organisms
with quorum sensing-inhibitory activity can be isolated from the aquatic environment
and that they show promise as novel biocontrol agents. In addition to
quorum sensing, other regulatory mechanisms control the expression of virulence
genes as well. So far, and not yet tested, is a molecule, called virstatin, that has been reported to
inhibit the transcriptional regulator ToxT, which controls the expression of
virulence genes of the ToxR regulon in Vibrios.

Overall this paper concludes that a number of different strategies must
be developed and used in rotation to prevent resistance development. The paper
is rather brief but puts forward further research directions to aid disease
control and probiotic choice.

Diarrhoea and the Sea

Diarrheal diseases derived from enteric bacteria are one the most prominent health risks in the developing world. Enteric bacteria such as enterotoxigenic Escherichia coli (ETEC) and Salmonella enterica serovar Typhimurium (S. enterica) are known as two of the leading causes of diarrheal disease. They are noted to be of particular importance in cases involving children. An increase in pathogen abundance in coastal zones has been described in recent history. This is suggested to be attributed to anthropogenic activities such as sewage discharge and agricultural runoff.

Diarrheal pathogens such as ETEC and S. enterica are able to colonise numerous diverse environments, resulting in the widespread contamination of drinking water and food. Bivalves, such as mussels, clams and oysters are popular food stuffs and are frequently associated with the transmission of pathogens from the sea to humans. This occurs by the accumulation of pathogens by a bivalve, which is then transmitted to a human through consumption (oral-faecal route). In an attempt to break this transmission pathway, efforts have been made to determine how a population of enteric bacteria is sustained within an environment. By evaluating the behaviour and adaptation of enteric bacteria, as well as the complex factors which act upon them, a comprehensive understanding of the biocomplexity of a pathogen can be achieved.

Limited literature is available describing the incidence and diligence of ETEC in marine habitats. In order to determine its possible prevalence and influence in the marine environment, this study compared the growth response to various factors of ETEC to the better studied fecal contaminant S. enterica and the marine bacteria Vibrio parahaemolyticus. An evaluation of the uptake and behaviour of the bacteria in blue mussels is provided, along with an analysis of the effect of temperature, nutrient supply, bivalve immune response and UV illumination on the viability of the bacteria.

Most parameters were assessed by culturing each species on agar plates and incubating in seawater for 8 weeks. However, to investigate the influence of bivalve immune response, an in vitro experiment was devised where the bactericidal capacity of haemocytes was measured. Haemocytes were harvested from the haemolymph of four different bivalves.

Noted results that should be highlighted from this paper include; V. parahaemolyticus was negatively affected by low temperature, whilst ETEC was positively affected. The growth of ETEC was observed in high-nutrient conditions, whilst no affect was noted in any other bacteria. In samples where blue mussels were nourished with each species of bacteria, lower ingestion rates were observed for V. parahaemolyticus when compared to other enterobacteria. Additionally in these conditions, V. parahaemolyticus is advocated to reduce the filtration activity of blue mussels.

Investigation of the influence of bivalve immune response suggested that the bactericidal capacity of bivalve hemocytes differed both on a species level as well as between each species. Results indicated that V. parahaemolyticus showed increased resistance to haemocytes from different bivalves, whilst ETEC and S. enterica were efficiently terminated by the haemocytes. It is advocated that mussels are less proficient at eliminating V. parahaemolyticus, as the haemocytes of three different species of bivalve were unable to eliminate the strain of V. parahaemolyticus used in this analysis. However, this pathway is advocated as a successful method of eradicating ETEC and S. enterica.

This paper emphasizes that the role of ETEC in the transmission of enteric diseases from an oceanic source is currently underestimated. Subsequently, it is suggested that ETEC, V. parahaemolyticus and S. enterica should be considered and measured as analogous sources of infection, when assessing the role of the marine environment as a source in the spread of enteric diseases. Nevertheless, the transmission of these pathogens to human populations is dependent upon a great number of factors, to which a lot of details are unknown. In order to successfully manage waterborne infections, a profound understanding of the relative sources and pathways of transmission are required. Further studies should concentrate on the persistence of ETEC in the marine environment.

Lactic acid bacteria and their inhibitory activity against L. gervieae

A review of, Perez-Sanchez, T., Balcazar, J.L., Garcia, Y., Halaihel, N., Vendrell, D., De Blas, I., Merrifield, D.L. & Ruiz-zarzuela, I., (2011), Identification and characterisation of lactic acid bacteria isolated from rainbow trout, Oncorhynchus mykiss (Walbaum), with inhibitory activity against Lactococcus garvieae, Journal of Fish Diseases, Vol: 34(7): 499-507

Lactococcus garvieae, a Gram-postiive pathogen is known to cause haemorrhagic septicaemia and meningoencephalitis in various fish species, as well as in mammals and has recently been isolated in humans, determining its potential as a zoonotic agent. Outbreaks of L, garvieae are common in aquaculture, where they are normally treated with antibiotics. However, this is often ineffective and has also led to antibiotic resistance.

Probiotics are now used to prevent pathogenic microorganisms such as L. garvieae causing disease in fish. Lactic acid bacteria (LAB), such as Lactococcus spp., Pediococcus spp. or Lactobacillus spp. are especially used as biological control agents in aquaculture.

This study aimed to identify and characterize the properties of the endogenous microbiota of rainbow trout, Oncorhynchus mykiss, against L. garvieae by studying their antibacterial activity, pH and bile tolerances, adherence characteristics and antibiotic resistance, in order to determine their potential as probiotics.

60 rainbow trout were collected from two fish farms. Sample mucus was obtained from the gills and intestines. Serial dilutions were then placed on tryptic soy agar, where 335 bacterial strains were isolated. They were screened for antagonistic activity against L.gervieae using agar spot assay. Then the strains were grouped using PCR and identified using 16s rRNA gene sequence analysis.

Eight of 198 isolated strains from the intestinal mucosa were antagonistic against L.gervieae. These strains were identified as Lactobacillus plantarum, Lactococcus lactis and Leuconostoc mesenteroides.

L. mesenteroides showed greater growth rate and doubling time compared to the other bacteria. They all showed a good hydrophobicity percentage, which acted as an indicator of adherence ability, with the highest in Lb. plantarum (31.59%) and significantly lower (P<0.05) in L. M=mesenteroides (11.42%) and L. lactis (24.99%). All strains showed relatively high resistance to bile, (resistant to 1.0% bile) with no significant difference in sensitivity between the bacteria. Similarly, they showed resistance to low pH, up to pH 2. L. lactis showed the highest levels at pH 1 whereas L. mesenteroides showed highest levels at pH 7.4.

The bacteria were also resistant to a majority of antibiotics tested, with little variation in susceptibility between strains. This is advantageous for establishing the beneficial microbiata in the intestines for a prolonged time, however it must be ensured that this resistance is not later passed on through plasmids.The antimicrobial substances exhibited by the three strains were also found to be sensitive to proteinase K. Moreover, after incubation of L.garvieae on the plates, measurable clearing zones were detected around the wells filled with supernatants from the strains, showing their antagonistic efficacy and that proteinaceous extracellular inhibitory compounds were at least partly responsible for the antagonism.

The clear antagonistic effects of these indigenous trout microbiota on the pathogen as shown by the results support previous studies which explain that establishing normal or protective microbiota in particular fish will constitute a key component of defensive barrier function against pathogens. This is also the first report to demonstrate clear and consistent antagonism of indigenous rainbow trout microbiota against L.garvieae, mediated in part at least, by extracellular antimicrobial peptides. Considering the good pH and bile tolerance, positive indications towards epithelial adherence and the ability to suppress pathogen growth under in vivo conditions, L.lactis, Lb, plantarum and L mesenteroides should be further studied in vivo for their potential use as probiotics for the control of L.garvieae, as it may provide an alternative to the current use of antimicrobial compounds.

Antarctic Sewage

This paper investigates waste water disposal in Antarctica and shows that more than half (52%) of the 71 research stations located in the largest pristine wilderness of the earth, still lack any form of waste and water treatment system (i.e. the process of removing physical, chemical and biological contaminants). A questionnaire, asking about the handling of waste and grey water, the type of sewage treatment and installation and operational costs, was sent out by the authors, to all 28 nations currently active in Antarctica. The results showed that 37% of the permanent stations and 69% of the summer stations completely lack any form of treatment facility and most of the stations that have a treatment system, installed it only during the past decade. The most common type is biological treatment (used by 20% of the stations), 10% of stations use macerations (which only removes solids such as waste, fats, oils and grease), another 10% use secondary biological treatment (i.e. the removal of dissolved organic matter using waterborne microorganisms) and only two stations (5%) use septic tanks. The study also shows that the treatment methods used at many stations are not efficient enough to reduce the likelihood of spreading introduced human-derived microbial pathogens to the environment. In fact, to handle the problem of microbial agents, only four stations use chemico-physical tertiary treatment systems, such as microfiltration, UV sterilization, chlorination, electric fields or ozone disinfection. Moreover, to be effective in Antarctica, depuration practices need to be adapted to local conditions, because some technologies, developed and commonly used in temperate areas, do not function in the harsh Antarctic environment. For this reason, stations with existing treatment systems often report operational problems and malfunctions, especially during periods of a high influx of people, when the plants are unable to cope with the high sewage input level (~100 litres per person per day). However, the summer climatic conditions in Antarctica, are no more extreme than conditions in Scandinavia, Greenland, Alaska and Canada, which have large populations and efficient waste water treatment.

In recent years there has been a continued growth in the number and size of stations and semi-permanent field camps in Antarctica. In the winter season approximately 1000 scientists and technicians carry out research and monitoring activities, while during the austral summer, which is also the most biologically sensitive period for terrestrial biota, the scientific community is estimated to comprise around 4000 people, to which must be added approximately 37500 tourists which visit Antarctica every summer. Most Antarctic stations are situated along the coastline, where accessibility is better and both research and tourism can noticeably increase the risk of introducing new humans infectious agents to the Antarctic flora and fauna. When waste and grey water containing microorganisms are released in the environment, microbes can remain viable in the low-temperature Antarctic conditions for prolonged periods and may have the potential to infect, cause disease, or become part of the gut flora of local bird, mammal, fish and marine invertebrates. Many studies have already detected well-known pathogens that may have been introduced by humans, such as Escherichia coli, Salmonella enteritidis, Salmonella typhimurium, Campylobacter jejuni and Pasteurella multocida, in both seal and bird populations. In addition to microorganisms, grey water, which comes from laundry, showers, sinks and washing dishes, may also contain fat, oil, detergents and other residues from personal hygiene products with surfactants that may be environmentally harmful. The organic enrichment by the sewage water from American McMurdo Station has already had a significant negative impact on the benthic community structure and high densities of coliform bacteria have been found along the shoreline outside the base.

Although the Protocol on environmental protection to the Antarctic Treaty states that the protection of the Antarctic environment is in the interest of mankind as a whole and it obliges all countries to apply and develop responsible waste management principles, much can still be done by the 28 nations operating the 82 research stations in Antarctica. The costs for installation and operation of waste water treatment systems are reasonable if compared with the overall cost of Antarctic research. The technology already exists for effective sewage treatment in the challenging polar conditions and thus, their use, coupled with a more efficient environmental monitoring, is necessary at all permanent and summer Antarctic research stations, in order to minimize the human impact on the most pristine continent on Earth and to protect it from human-derived infectious agents.

Reference:

Gröndahl, F., Sidenmark, J., Thomsen, A. (2008). Survey of waste water disposal practices at Antarctic research stations. Polar Research 28: 298-306.

All At Sea : The Comparison of Antibiotic Resistance and Human Intervention

This paper investigates the antibiotic resistance of pollution indicators within three coastal areas in India. This is a major concern as antibiotic resistance is expected in any environment were antibiotics are implemented. The improper and unnecessary use of antibiotics by humans could provide the selection of resistant pathogenic bacterial strains not indigenous to the local environment. However, due to domestic and commercial sewage, this change in the marine environment poses not only a threat to the marine community but also creates a hazard to public health.

Samples of water and sediment were taken from three different sites: The coastal areas of Chennai port and Kasimeda fishing harbour and the Coovum estuary. Four indicator genera were isolated - Escherichia coli, Vibrio spp., Salmonella spp. and Enterococcus spp. from both the water and the sediment samples taken from each site between march and may 2010.

These sample were then tested for antimicrobial sensitivity and challenged with ten different antibiotics. The antimicrobial resistance patterns were not due to specific bacteria species or taxonomic groups but among isolates among different samples and sample sites.

In the sea water samples, Salmonella and E. coli stains were predominantly resistant to vancomycin followed by penicillin, whereas Vibrio and Salmonella isolates were resistant to all of the antibiotics challenged. Kasimeda Harbour samples had the highest percentage resistance to antibiotics challenged.

In sediment samples higher resistant frequencies were observed in the Coovum estuary. Vibrio spp. and Salmonella spp. were found to be resistant to all of the antibiotics challenged whereas E. coli and Enterococcus indicators were resistant to all of the antibiotics tested.

The antibiotic resistance index (ARI) for all indicator organisms present in the sea water samples was highest in the Kasimeda harbour whereas the ARI was highest in the Coovum estuary among indicator organisms found in sediment samples.
The authors thought this was likely due to the Coovum estuary and the Kasimeda harbour having slums nearby meaning the greater population will be causing a greater volume of sewage to be present in the samples.

The authors found Vibrio spp. to be the most resistant to antibiotics and faecal indicators resistant to antibiotics were greater in the sediment samples due to the higher nutrient concentrations. This paper flags up the issue that the improper use of antibiotics may be more widespread and on a much larger scale than we currently believe.

A Review of: Vignesh, S., et al.; 2012; Antibiotic resistant pathogens versus human impacts: A study from three eco-regions of the Chennai coast, southern India; Marine Pollution Bulletin; doi:10.1016/j.marpolbul.2012.01.015.

Phytoplankton enhances inhibition of pathogen

Phytoplankton cultures are commonly used in aquaculture, where they are known as ‘green water’. The cultures are usually grown from natural seawater, and are added to larvae tanks to provide nutrition, improve water quality, and make the water less clear thereby reducing larval light stress. Most importantly they alter the bacterial composition of the water, leading to dominance by alphaproteobacteria and the Cytophaga-Flavobacterium cluster. A number of Roseobacter clade members (alphaproteobacteria), have been found to suppress growth of the fish pathogen Vibrio anguillarum. This paper aims to explore this inhibitory activity in the presence of the phytoplankton Nannochloropsis oculata, so that the benefits to aquaculture can be better understood.

Roseobacter clade strains were isolated from a culture of N. oculata, and identified using FISH and PCR. Six Roseobacter strains (Sulfitobacter sp. (two strains), Thalassobius sp., Stappia sp., Rhodobacter sp. and Antarctobacter sp.) were inoculated with V. anguillarum strain psh-9019 into three different types of media. After incubation at 20°C for 3 days, colonies were counted. Cultures of each strain on each media served as controls.

On the first medium, an organic nutrient-rich medium for heterotrophic marine bacteria (VNSS), the control V. anguillarum culture grew to 1.6x109 CFU/ml, and similar results were obtained for V. anguillarum cultured together with the Roseobacter strains. However, on the second medium, a phytoplankton culture medium (ESM), the viability of V. anguillarum decreased approximately ten-fold when cocultured with the Roseobacter strains.

The final medium was NCF, which contained substances excreted by the phytoplankton N. oculata grown in ESM. Control V. anguillarum CFUs were similar to the results obtained on ESM medium, but when cocultured with Roseobacter strains on NCF medium, V. anguillarum was completely eradicated after 5 days. These results strongly suggest that Roseobacter clade members can exhibit a much greater antibacterial effect upon V. anguillarum when cocultured with a phytoplankton species such as N. oculata.
The authors suggest that N. oculata may excrete substances such as dimethylsulfoniopropionate (DMSP) and amino acids which more optimally support growth of the Roseobacter isolates. They go on to say that under oligotrophic conditions mimicked by the ESM and NCF media, the Roseobacter strains are more efficient at obtaining small amounts of nutrients than V. anguillarum. The authors also believe that the Roseobacter strains directly inhibit the growth of the pathogen, with N. oculata releasing compounds which act as signaling molecules for communicating with Roseobacter isolates, affecting the inhibitory activity.

This paper has very extensive methods and results sections, which makes it quite complex at times. However it is one of the first to show that these particular Roseobacter strains isolated from phytoplankton cultures can affect growth of the fish pathogen V. anguillarum, and the investigation into the effects of coculture with N. oculata adds another dimension to disease control in aquaculture.

A review of: Sharifah, E.N. and Eguchi, M. (2011) ‘The phytoplankton Nannochloropsis oculata enhances the ability of Roseobacter clade bacteria to inhibit the growth of fish pathogen Vibrio anguillarum’, PLoS ONE 6(10): e26756. doi:10.1371/journal.pone.0026756.

The use of Probiotics in Shrimp Aquaculture

Over the years aquaculture has grown into a successful industry accounting for almost 50% of fish sales worldwide in 2006. This increasing demand has lead to mass husbandry resulting in the presence of pathogenic disease and therefore the overuse of antibiotics and chemotherapeutics. This overuse of antibiotics has caused the rapid evolution of resistance mechanisms and selected for resistance genes, hence leading to the presence of resistant and more virulent microorganisms. Asian aquaculture has been particularly hard hit by disease, for example shrimp industries in Bangladesh which provide the second largest exportable commodity for the country, have had devastating incidences of disease, which has been no doubt facilitated by the cramped and overcrowded rearing ponds and tanks used.

Over the years several methods of disease control have been looked into; however the addition of large amounts of antibiotics and chemotherapeutics remains the current method of choice for disease control. Recently the emergence of non-pathogenic bacteria as bio-control agents has received a lot of attention, as these probiotics help to strengthen already existing microbial communities in the environment and gut of the organisms in question. Several studies have backed up the use of probiotics claiming that even pathogens which have become unaffected by antibiotics could be controlled by the use of probionts (E.g. V.harveyi).

The aims of this study were to isolate pathogenic micro-organisms and probiotic bacteria from shrimp culture ponds in a region of Bangladesh. The authors tested these pathogens and probionts to see if the use of probiotics would have a beneficial and sustainable effect in aquaculture. The results of their study indicated that a good number of probionts challenged pathogens successfully, giving evidence of beneficial properties, although it was indicated that the effectiveness of the probionts is dependent on which pathogens were present. Further studies were also carried out to ensure the safety of the probionts in mammalian systems (an important food safety issue) with results showing that 19/37 of the probionts would be safe for mammals and therefore potentially humans if ingested.

The authors state that further research is required to develop suitable probiotics along with full scale trials and appropriate monitoring tools; however the future of this developing science seems promising. Overall I think that this paper had a good purpose, although I found throughout that it was very badly written. The English used was often incorrect and the description of the methods and results were more complicated than they needed to be. The paper was quite short and I found the structure of it a bit all over the place, however I think that the overall point of the study was a good one with encouraging results that can be expanded on in the future.

Review of: Rahman, S. Khan, S. N. Naser, M. N and Karim, M. (2009) Application of probiotic bacteria: A novel approach towards ensuring food safety in shrimp aquaculture. Journal of Bangladesh Academy of Science. 33(1) 139-144.

Microbial Fuel Cells; An Alternative Energy Source for In-Water Equipment

Staying on the same theme as my previous post of alternative sources of energy, this post looks at microbial fuel cells (MFC) and the production of electricity. As the name suggests, MFCs rely on microbes to generate a movement of electrons (electricity) without the need of an initial electrical input. Organic compounds are catabolised by the microbes in an anaerobic chamber but instead of using an atom or compound as a terminal electron acceptor, the microbes transfer electrons to a conductive electrode which runs through a circuit, generating electricity.

A very current area of research for the applied use of MFCs is in marine sediments. For example, the US Navy is interested in setting up marine sediment MFCs to maintain the batteries of in-water sensors and navigation equipment positioned in oceans around the world. The main advantage of this is a decreased need for human maintenance (Arias-Thode et al 2011). How electricity can be harvested in this way involves two graphite plates. One is buried in the marine sediment (anode) and has to be completely anaerobic. Connected to the anode via an electrical circuit is the second plate which is held in place in the aerobic water column (cathode). Microbes use the anode as a terminal electron acceptor and at the same time releases protons (H+) into the sediment. The electrons are attracted through the circuit to the cathode by the presence of the highly oxidising molecular oxygen in the water column and combines with free protons to yield H2O. This movement of electrons can be used to trickle charge a battery.

Despite the clever way in which marine sediment MFCs work little is known on the microbes that grow on the anode plates. Holmes et al (2004) conducted the first detailed study looking into these communities. A number of MFCs were placed in different marine sediments around the USA. Sediment samples were collected adjacent to the anodes 3 months after initial placement. A control was also set up which consisted of just the anode plate with no cathode attached, which meant no electrical circuit. PCR amplification of 16rRNA was used for comparison to GenBank for species identification, it was also used to get an indication of cell numbers. Similar samples were taken from the cathode.

Results showed the anodes were dominated by bacteria of the Geobacteraceae family, within the δ-Proteobacteria class. When comparing to the control anode, Geobacteraceae were 100 times more abundant on the electrically active anodes. Geobacteraceae are normally iron reducers but in this case appear to be using the graphite anode as an electron acceptor instead. A possible explanation for this is that unlike other electron acceptors like oxygen or nitrate, iron is insoluble in water and a large insoluble graphite plate seems to be acting as a good alternative. The fact that cell numbers were far higher on the electrically active anodes indicates that this must attract and promote growth more than their normal iron reduction method. The cathode also showed unique bacterial colonisation. Nitrogen oxidising γ-Proteobacteria covered the surface of the cathode. Although not an uncommon group of bacteria they were significantly different from microbes normally found in the water column and nearby biofilms. Once again showing that active MFCs positively influence the bacterial populations.

Although this is not the most current paper on this topic, I reviewed it because it appeared to be the first study trying to specifically identify the microbes involved in marine sediment MFC electricity production. I had heard of the hydrogen fuel cell before which is a very similar concept but I had not heard of MFC before.

A Review of:
Holmes DE, Bond DR, O’Neil RA, Reimers CE, Tender LR and Lovley DR (2004) Microbial Communities Associated with Electrodes Harvesting Electricity from a Variety of Aquatic Sediments. Microbial Ecology. 48: 178-190.

Additional Reference:
Arias-Thode YM, Richter K, Wotawa-Bergen A, Chadwick DB, Kan J and Nealson K (2011) Development of Microbial Fuel Cell Prototypes for Examination of the Temporal and Spatial Response of Anodic Bacterial Communities in Marine Sediments.10.1109/Oceans-Spain.2011.6003535.

Antibiotic determinants in Aeromonas sp.

A review of the paper: Ndi, O.L.,Barton, M.D., (2011), Incidence of class 1 integron and other antibiotic resistance determinants in Aeromonas spp . from rainbow trout farms in Australia, Journal of Fish Diseases, 34, 589-599

Aeromonas sp. are a gram negative group of bacteria that are pathogenic across a large range of hosts including many in fish disease and humans. The treatment and prevention of many diseases have been implemented extensively using antimicrobial agents, which has resulted in an increase in resistant strains. It is thought that this resistance may come from integrons, which are unusual DNA elements. The objective of this research was to identify some resistant determinants associated with Aeromonas sp. including those linked with integrons.

48 Aeromonas sp. were isolated from healthy fish and 42 from sediments from 9 freshwater trout farms. These strains were identified using a commercial MICROBACT 24E ID kits and also using a PCR for 16s ribosomal RNA gene. The detection of integrons and tetracycline resistance genes was also carried out PCR and other genes were detected using a range of primers. These genes were then sequenced.

The results from the identification showed discrepancies between the different methods however overall 5 species were identified with both methods showing A. hydrophila as the most common. 31% of the strains were positive for class 1 integrons. Other resistant genes were also found that code for specific enzymes involved in resistance. The final conclusion of this paper was that there were antibiotic-resistant species of Aeromonas sp. in Australian fish farms. This would have implications for human health when fish are consumed and if farm run-off gets into the environment.

Do Atlantic Herring emit sulfur-rich flatulences?

Microbial dimethyl sulfide (DMS) production is very important in the global sulfur cycle, being the major route of sulfur transfer from oceans to atmosphere and then back to the land.

Microbes convert dimethylsulfoniopropionate (DMSP), mainly produced by marine phytoplankton, to DMS by at least three different enzymes that cleave DMSP. One of these, DddL, is a DMSP lyase, whose products are acrylate and DMS, and has thus far only been found in marine α-proteobacteria. Another enzyme, DddP, occurs in α-proteobacteria and in some fungi, but its mechanism of action is unknown. 

The third system involves an acyl CoA transferases encoded by dddD, a gene that occurs in marine γ-proteobacteria and in other bacterial taxa who acquired it by horizontal gene transfer. 

DMSP-catabolizing bacteria have been already isolated from several DMSP-rich environments, including corals, algal blooms, copepods and salt marshes. But this was the first time that such bacteria have been isolated from the gut microflora of a fish, the Atlantic Herring (Clupea harengus) in this case, which feeds on copepods, which in turn consume DMSP-containing phytoplankton.

In this study, after a series of enrichments in DMSP-containing liquid medium, the gut samples were cultured by the authors, on minimal agar plates with DMSP as the sole carbon source. Two different colony types were identified and assayed for DMS production by gas chromatography. The 16S rRNA genes were cloned and then sequenced and the closest (99% identical) relatives were found to be Pseudomonas guineae LMG 24017 and Psychrobacter arenosus R7T. These newly isolated strains were termed J465 and J466, respectively and both produced large amounts of DMS, whose levels were significantly enhanced when cells were pre-grown on 5-mM DMSP. The two strains were also examined for growth on acrylate and 3-hydroxypropionate (one of the earliest catabolites formed after the action of DddD on DMSP), as some other DMSP-catabolizing species can use these compounds. Pseudomonas J465 did not grow on either compound, but Psychrobacter J466 grew on 3-hydroxy-propionate as the sole carbon source. Moreover, acrylate and 3-hydroxy-propionate were found to induce DMS production in Psychrobacter J466, but not in Pseudomonas J465.

To identify the relevant genes in these new isolates, genomic libraries of the two strains were constructed by the authors, mobilizing a wide host-range cosmid into Escherichia coli and screening it for any gene that conferred DMS production to the host. One such cosmid was identified from each library and their inserts were sequenced. Both contained homologues of dddD, which is responsible for the initial step in DMSP catabolism. In addition, several neighboring genes resembled those that occur near dddD in Marinomonas MWYL1, such as dddT (involved in DMSP transport), dddR (DMSP regulation) or dddB and dddC, which are involved in the later stages of DMSP catabolism and which respectively encode polypeptides related to alcohol dehydrogenases and aldehyde dehydrogenases.

In another study on the Atlantic Menhaden (a close relative of the Herring), approximately 65% of the DMSP was shown to be incorporated into the animals’ flesh or excreted unmodified, with very little DMS being actually released when fish were fed DMSP-containing algae. The fate of the remaining 35% of DMSP is unknown, but it can be demethylated, through the action of dmdA (a widespread gene in marine bacteria) in a process that liberates no DMS. Or otherwise the DMS eventually formed by Dddþ bacteria could be catabolized by other DMS-using bacteria.

Dddþ microbes may occur in other marine carnivores such as seabirds, cetaceans, penguins and seals which often use DMS as a chemoattractant to find their prey. So it might even be in the fish’s interest not to emit it, especially since herrings may communicate at night by forming bubbles from their posteriors. 

Further work is thus needed to determine the full range of bacterial types, and the relative contributions of different pathways involved in DMSP catabolism in marine predators and it would be interesting to establish the contributions of such animal's microflora to the global turnover of DMSP.

Reference:

Curson, A.R.J., Sullivan, M.J., Todd, J.D. & Johnston, A.W.B. (2010). Identification of genes for dimethyl sulfide production in bacteria in the gut of Atlantic herring (Clupea harengus). ISME J 4:144–146.